Sweet Potato Processing Technology [1 ed.] 978-0-12-812871-8, 0128128712

Table of contents :
Content: 1. Sweet Potato Starch and its Series Products 2. Sweet Potato Proteins 3. Sweet Potato Dietary Fiber 4. Sweet Potato Pectin 5. Sweet Potato Granules 6. Sweet Potato Anthocyanins 7. Chlorogenic Acids From Sweet Potato

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Sweet Potato Processing Technology

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Sweet Potato Processing Technology

Taihua Mu, Hongnan Sun, Miao Zhang and Cheng Wang Key Laboratory of Agro-Products Processing, Ministry of Agriculture Institute of Food Science and Technology, Chinese Academy of Agricultural Sciences, Beijing, P.R. China

Academic Press is an imprint of Elsevier 125 London Wall, London EC2Y 5AS, United Kingdom 525 B Street, Suite 1800, San Diego, CA 92101-4495, United States 50 Hampshire Street, 5th Floor, Cambridge, MA 02139, United States The Boulevard, Langford Lane, Kidlington, Oxford OX5 1GB, United Kingdom Copyright © 2017 China Science Publishing & Media Ltd. Published by Elsevier Inc. All rights reserved. No part of this publication may be reproduced or transmitted in any form or by any means, electronic or mechanical, including photocopying, recording, or any information storage and retrieval system, without permission in writing from the publisher. Details on how to seek permission, further information about the Publisher’s permissions policies and our arrangements with organizations such as the Copyright Clearance Center and the Copyright Licensing Agency, can be found at our website: www.elsevier.com/permissions. This book and the individual contributions contained in it are protected under copyright by the Publisher (other than as may be noted herein). Notices Knowledge and best practice in this field are constantly changing. As new research and experience broaden our understanding, changes in research methods, professional practices, or medical treatment may become necessary. Practitioners and researchers must always rely on their own experience and knowledge in evaluating and using any information, methods, compounds, or experiments described herein. In using such information or methods they should be mindful of their own safety and the safety of others, including parties for whom they have a professional responsibility. To the fullest extent of the law, neither the Publisher nor the authors, contributors, or editors, assume any liability for any injury and/or damage to persons or property as a matter of products liability, negligence or otherwise, or from any use or operation of any methods, products, instructions, or ideas contained in the material herein. British Library Cataloguing-in-Publication Data A catalogue record for this book is available from the British Library Library of Congress Cataloging-in-Publication Data A catalog record for this book is available from the Library of Congress ISBN: 978-0-12-812871-8 For Information on all Academic Press publications visit our website at https://www.elsevier.com/books-and-journals

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CONTENTS

Introduction....................................................................................... xiii Chapter 1 Sweet Potato Starch and its Series Products ........................1 Section 1: Overview of Sweet Potato Starch and its Series Products .............................................................................3 1.1 Sweet Potato and its Starch ...............................................3 1.2 The Structure and Morphology of Sweet Potato Starch ....4 1.3 Chemical Composition of Sweet Potato Starch .................7 1.4 Characteristics of Sweet Potato Starch ..............................9 1.5 Sweet Potato Starch Noodles and Vermicelli...................14 1.6 Sweet Potato Resistant Starch .........................................15 Section 2: Production Technology of Sweet Potato Starch and its Series Products .....................................................16 2.1 Production Technology of Sweet Potato Starch...............16 2.2 The Production Process of Sweet Potato Starch Noodles and Vermicelli....................................................19 2.3 The Sweet Potato Resistant Starch Production Process ...21 Section 3: Physicochemical Properties of Sweet Potato Starch and its Products ...............................................................24 3.1 The Structure and Physicochemical Properties of SLPS and CFS.............................................................24 3.2 Comparison of the Quality of SLPS and CFS .................36 Section 4: Applications of Sweet Potato Starch................................41 4.1 Application of Sweet Potato Starch in Food ...................42 4.2 Applications of Sweet Potato Resistant Starch ................42 References...........................................................................................43 Further Reading .................................................................................48 Chapter 2 Section 1: 1.1 1.2

Sweet Potato Proteins ........................................................49 Overview of Sweet Potato Proteins ..................................52 The Sources and Structures of Sweet Potato Proteins......52 Research Status on the Technologies Used to Produce Sweet Potato Proteins ......................................................55 1.3 The Biological Activity of Sweet Potato Protein..............55 1.4 Physicochemical Properties of Sweet Potato Protein........56

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Section 2: Production Technologies of Sweet Potato Protein ...........57 2.1 Effects of Solvent on the Extraction of Sweet Potato Protein..................................................................58 2.2 Salting-Out Method .........................................................60 2.3 Isoelectric Precipitation Method ......................................62 2.4 Foam Separation Method................................................64 2.5 Ultrafiltration Method .....................................................70 2.6 Thermal Denaturation Method........................................75 2.7 Purification Method of Sweet Potato Protein ..................78 Section 3: Biological Activity of Sweet Potato Protein.....................80 3.1 Antioxidant Activity ........................................................80 3.2 Trypsin Inhibitory Activity of Sweet Potato Protein........83 3.3 Anticancer Activity of Sweet Potato Protein ...................86 3.4 Obesity Prevention and Weight Loss ...............................88 Section 4: Functional Properties of Sweet Potato Protein ................92 4.1 Solubility of Sweet Potato Protein ...................................92 4.2 Emulsifying Properties of Sweet Potato Protein...............95 4.3 Gelling Properties of Sweet Potato Protein .................... 105 4.4 Structural Properties of Sweet Potato Protein................ 107 4.5 The Foaming Properties and Foam Stability of Sweet Potato Protein ................................................. 110 4.6 Water-Holding and Oil-Holding Capacities of Sweet Potato Protein ................................................. 110 Section 5: Applications of Sweet Potato Protein ............................ 111 5.1 Edible Protein Powder ................................................... 112 5.2 Emulsifier....................................................................... 113 5.3 Humectant ..................................................................... 114 5.4 Raw Material of Active Peptides ................................... 114 5.5 Biological Medicine ....................................................... 115 References......................................................................................... 115 Further Reading ............................................................................... 118 Chapter 3 Section 1: 1.1 1.2 1.3 1.4 1.5

Sweet Potato Dietary Fiber.............................................. 121 Introduction of Dietary Fiber ........................................ 124 The Definition of Dietary Fiber..................................... 124 The Composition of Dietary Fiber ................................ 124 The Classification of Dietary Fiber................................ 125 Dietary Fiber Extraction Methods................................. 125 The Physicochemical and Functional Properties of Dietary Fiber ............................................................. 127 1.6 The Mechanism of Dietary Fiber in Preventing Obesity ........................................................................... 128

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Section 2: Sweet Potato Dietary Fiber Extraction Technology .................................................................... 131 2.1 Sieve Method ................................................................. 132 2.2 Sieve Combined Enzymatic Hydrolysis Method ............ 134 2.3 Biological Method ......................................................... 135 2.4 Chemical Separation Method ........................................ 139 Section 3: The Physiological Properties of Sweet Potato Dietary Fiber ................................................................. 140 3.1 Effect of Sweet Potato Dietary Fiber on Preventing Obesity ........................................................................... 140 3.2 Effects of Sweet Potato Dietary Fiber on Treating Obesity ........................................................................... 151 3.3 Other Functional Properties of Sweet Potato Dietary Fiber .............................................................................. 160 Section 4: The Physicochemical Properties of Sweet Potato Dietary Fiber ................................................................. 161 4.1 Effects of Different Factors on the Water-Holding Capacity of Sweet Potato Dietary Fiber ........................ 161 4.2 Effects of Different Factors on the Water-Swelling Capacity of Sweet Potato Dietary Fiber ........................ 162 4.3 Effects of Temperature on the Oil-Holding Capacity of Sweet Potato Dietary Fiber ........................ 164 4.4 Effects of Different Factors on the Viscosity of Sweet Potato Dietary Fiber............................................ 166 4.5 Water-Holding Capacity of Dietary Fibers From Different Sweet Potato Varieties .................................... 167 4.6 Water-Swelling Capacity of Dietary Fiber From Different Varieties of Sweet Potato................................ 168 4.7 Oil-Holding Capacities of Dietary Fibers From Different Varieties of Sweet Potato................................ 169 Section 5: The Applications of Sweet Potato Dietary Fiber ........... 170 5.1 The Applications of Sweet Potato Dietary Fiber in Bread ......................................................................... 170 5.2 The Applications of Sweet Potato Dietary Fiber in Beverages ....................................................................... 174 5.3 The Applications of Sweet Potato Dietary Fiber in Meat Products ........................................................... 174 5.4 The Applications of Sweet Potato Dietary Fiber in Staple Foods .............................................................. 174

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5.5 The Applications of Sweet Potato Dietary Fiber in Condiments ...................................................... 175 5.6 The Applications of Sweet Potato Dietary Fiber in Health Foods ............................................................. 175 References......................................................................................... 175 Further Reading ............................................................................... 179 Chapter 4 Section 1: 1.1 1.2 1.3 1.4 1.5 Section 2: 2.1 2.2 2.3 2.4 Section 3: 3.1 3.2 3.3 3.4 3.5 3.6 Section 4: 4.1 4.2 4.3 Section 5: 5.1 5.2

Sweet Potato Pectin ......................................................... 183 An Overview of Pectin ................................................... 186 The Distribution of Pectin ............................................. 186 The Structure of Pectin .................................................. 188 The Research Methods Determining Pectin Structure ... 189 Extraction of Pectin ....................................................... 191 Functional Properties of Pectin...................................... 193 Production Technology of Sweet Potato Pectin ............. 201 Extraction Process of Pectin From Sweet Potato........... 201 The Determination of the Galacturonic Acid Content in the Pectin Solution ....................................... 202 Factors Affecting the Yield and Galacturonic Acid Content of Sweet Potato Pectin............................. 203 Optimization of Pectin Extraction From Sweet Potato Pulp .................................................................... 206 Biological Activities of Sweet Potato Pectin .................. 208 The Preparation of pH-Modified Pectin ........................ 208 The Preparation of Thermal-Modified Pectin ................ 209 The Cell Cultures ........................................................... 209 Effects of Sweet Potato Pectin on Cancer Cell Survival Rates................................................................ 209 Effects of Sweet Potato Pectin on Cancer Cell Proliferation ................................................................... 210 Effects of Sweet Potato Pectin on Cancer Cell Metastasis ...................................................................... 214 Physicochemical Characteristics of Sweet Potato Pectin.................................................................. 227 A Viscosity Analysis of Sweet Potato Pectin.................. 227 The Gelation Properties of Sweet Potato Pectin ............ 231 The Emulsifying Properties of Sweet Potato Pectin.................................................................. 240 Applications of Sweet Potato Pectin .............................. 255 Candied Fruit ................................................................ 255 Bread ............................................................................. 255

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5.3 Frozen Food .................................................................. 256 5.4 Yogurt Products............................................................. 256 5.5 Beverages ....................................................................... 256 5.6 Others ............................................................................ 257 References......................................................................................... 257 Further Reading ............................................................................... 260 Chapter 5 Sweet Potato Granules..................................................... 263 Section 1: Background for Developing Sweet Potato Granules ........................................................................ 264 Section 2: Technologies and Key Points in Manufacturing Sweet Potato Granules................................................... 265 2.1 Manufacturing Technologies.......................................... 265 2.2 Key Points ..................................................................... 266 Section 3: Applications of Sweet Potato Granules ......................... 270 3.1 Applications of Sweet Potato Granules in Bread ........... 270 3.2 Applications of Sweet Potato Granules in Biscuits ........ 272 3.3 Applications of Sweet Potato Granules in Noodles ....... 275 3.4 Applications of Sweet Potato Granules in Thick Slurries ........................................................................... 276 References......................................................................................... 277 Chapter 6 Section 1: 1.1 1.2 Section 2: 2.1 2.2 Section 3: 3.1

Sweet Potato Anthocyanins.............................................. 279 Review of Sweet Potato Anthocyanin............................ 282 Introduction to Anthocyanins........................................ 282 Status of Sweet Potato Anthocyanins ............................ 285 The Preparation of Sweet Potato Anthocyanins ............ 297 The Extraction of Sweet Potato Anthocyanins .............. 297 Purification of Sweet Potato Anthocyanins ................... 309 The Stabilities of Anthocyanins From Sweet Potato ..... 312 The Effects of Temperature on the Thermal Stabilities of Anthocyanins ............................................ 312 3.2 The Effects of High Pressure on the Thermal Stability of Anthocyanins............................................... 313 3.3 The Effects of Different pH Levels on the Thermal Stability of Anthocyanins............................................... 314 3.4 Effects of Different Solvent Treatments on the Thermal Degradation of Sweet Potato Anthocyanins.... 316

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Section 4: Biological Activity of Sweet Potato Anthocyanin.......... 324 4.1 The Effects of Sweet Potato Anthocyanin on Acute Alcoholic Liver Damage and Dealcoholic Effects ......... 324 4.2 The Effects of Sweet Potato Anthocyanin on Subacute Alcoholic Liver Damage................................. 334 Section 5: Applications of Sweet Potato Anthocyanins .................. 343 5.1 Pharmaceutical Industry ................................................ 343 5.2 Food Industry ................................................................ 343 5.3 Cosmetics Industry......................................................... 346 References......................................................................................... 347 Further Reading ............................................................................... 353 Chapter 7 Section 1: 1.1 1.2 1.3 1.4 Section 2: 2.1 2.2 2.3 2.4

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Section 3: 3.1 3.2 3.3

Chlorogenic Acids From Sweet Potato ............................. 357 Overview of Chlorogenic Acids From Sweet Potato...... 359 Composition and Structure of Chlorogenic Acids.......... 360 Biological Activities of Chlorogenic Acids..................... 360 Extraction, Separation, and Purification Methods of Chlorogenic Acids ..................................................... 365 Qualitative and Quantitative Analyses of Chlorogenic Acids.......................................................... 368 Technology to Prepare Chlorogenic Acids From Sweet Potatoes ..................................................... 370 Pretreatment of Sweet Potato Leaves............................. 371 Total Polyphenol Contents ............................................ 371 Extraction of Chlorogenic Acids From Sweet Potato Leaves ................................................................ 371 Purification of Chlorogenic Acids From Sweet Potato Leaves Using AB-8 Macroreticular Adsorbent Resin ............................................................ 371 Qualitative and Quantitative Analyses of Chlorogenic Acids From Sweet Potato Leaves by HPLC ............................................................................ 378 Biological Activities of Chlorogenic Acids From Sweet Potatoes ............................................................... 381 In vitro Antioxidant Activity of Chlorogenic Acids From Sweet Potatoes ..................................................... 382 Antimicrobial Activities of Chlorogenic Acids From Sweet Potatoes ..................................................... 388 Aldose Reductase Inhibitory Activities of Chlorogenic Acids From Sweet Potatoes ....................... 389

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3.4 Anticancer Activities of Chlorogenic Acids From Sweet Potatoes ..................................................... 390 3.5 Other Biological Activities of Chlorogenic Acids From Sweet Potatoes ..................................................... 391 Section 4: The Stability of Chlorogenic Acids From Sweet Potatoes ......................................................................... 391 4.1 Effect of pH on Chlorogenic Acids From Sweet Potato Leaves ................................................................ 392 4.2 Effects of Heat Treatments on Chlorogenic Acids From Sweet Potato Leaves ............................................ 393 4.3 Effects of Light on Chlorogenic Acids From Sweet Potato Leaves ...................................................... 396 Section 5: The Applications of Chlorogenic Acids From Sweet Potatoes ............................................................... 398 5.1 Food Industry ................................................................ 398 5.2 Medicine and Health-Protection Industry...................... 398 5.3 Daily Chemical Industry................................................ 399 References......................................................................................... 400 Further Reading ............................................................................... 403 Appendix 1: Production Line of Sweet Potato Protein....................... 405 Appendix 2: Production Line of Sweet Potato Dietary Fiber ............. 407 Appendix 3: Production Line of Sweet Potato Pectin ........................ 409 Index................................................................................................. 411

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INTRODUCTION

Sweet potato, a spiral flowering annual or perennial herb classified into the Ipomoea category of the Convolvulaceae section, commonly known as Hongshu, Baishu, Digua, Fanshu, Hongyu, and Hongshao, was introduced from Latin America into China in the Wanli Period of the Ming Dynasty, and has a cultivation history in China of over 400 years. The cultivation of sweet potato is characterized by low-input, high-output, and drought and nutrient-stress resistant. Sweet potato is a major food crop only behind wheat, rice, and corn in China. It is mainly used for making starch and related products, such as starch noodles. However, in these production processes, a large number of by-products are produced, such as sweet potato juice, residues, peel, and cirrus. These by-products contain protein, dietary fiber, pectin, anthocyanin, chlorogenic acid, and many other functional components, which play important roles in regulating the functions of human body. It is important to support research and development of the nutritional and functional components extracted from sweet potato and its byproducts, as well as their application. This will be of great significance in promoting a healthy industrial structure for the sweet potato processing industry and developing human dietary habits. In recent years, our research team undertook several research projects such as The earmarked fund for China Agriculture Research System, The national science-technology support plan projects: study on Chinese sweet potato varieties for special-use and its suitability on processing, research and development of deep processing technology of sweet potato protein, Pilot technology research and demonstration of protein extraction from the waste water in sweet potato processing, Key technology research and industrialization demonstration of deep processing of sweet potato, and The introduction and utilization of high value utilization technology of agricultural processing by-product. Over 10 years of intensive study were performed in the sweet potato processing field, and our research team has solved numerous technical difficulties, accomplished a number of scientific achievements, and educated lots of highly qualified talents. The book Sweet Potato Processing

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Introduction

Technology was compiled based on our research achievements. This book includes seven chapters, contents summarized as follows. Chapter 1, Sweet Potato Starch and Its Series Products, introduces the structure, morphology, chemical components, physicochemical characteristics, and series products of sweet potato starch. Chapter 2, Sweet Potato Proteins, focuses on sweet potato protein, extraction technologies, and introduces its biological activity, functional properties, and applications in food, health food, and other fields. Chapter 3, Sweet Potato Dietary Fiber, focuses on the dietary fiber and its production technology, as well as its biological activity, physicochemical properties, and applications in food, health food, and other fields. Chapter 4, Sweet Potato Pectin, introduces sweet potato pectin and its production technology, as well as its biological activity, physicochemical properties, and applications in food, and other fields. Chapter 5, Sweet Potato Granules, introduces sweet potato particle powder and its background, processing technology, and applications in the food field. Chapter 6, Sweet Potato Anthocyanins, focuses on sweet potato anthocyanin and introduces its research, extraction, and purification technologies, stability, biological activity, and applications in food, medical, cosmetics, and other fields. Chapter 7, Chlorogenic Acids From Sweet Potato, introduces the research, extraction and purification technologies, biological activity, stability, and application prospects of sweet potato’s chlorogenic acid. This book was written based on the research of our team. The purpose of this book is to provide a reference and guidance for the deep processing and comprehensive utilization of sweet potato, and to provide technical support for scientific innovations in the domestic sweet potato industry. Chapter 1, Sweet Potato Starch and Its Series Products, was edited by Miao Zhang and Fuming Deng. Chapter 2, Sweet Potato Proteins, was edited by Jingwang Chen, Penggao Li, Zhongkai Zhao, and Zhidong Xiong. Chapter 3, Sweet Potato Dietary Fiber, was edited by Xiaomei Wang and Mengmei Ma. Chapter 4, Sweet Potato Pectin, was edited by Xin Mei, Xiaoyan Peng, Jinshu Yang, Zheng Zhao, and Yanyan Zhang. Chapter 5, Sweet Potato Granules, was edited by Cheng Wang and Weizhong He. Chapter 6, Sweet Potato Anthocyanins, was edited by Xingli Liu and Taihua Mu. Chapter 7,

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Chlorogenic Acids From Sweet Potato, was edited by Hongnan Sun and Lisha Xi. We sincerely acknowledge the work presented in the related books and papers referenced here, which was performed by many local and foreign experts and scholars. In view of the limitations of the author’s level and the rapid development of science and technology in sweet potato processing and comprehensive utilization, the contents of the book may inevitably contain some outdated information or omissions. Comments and suggestions are most welcomed.

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CHAPTER

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Sweet Potato Starch and its Series Products

SECTION 1: OVERVIEW OF SWEET POTATO STARCH AND ITS SERIES PRODUCTS 1.1 Sweet Potato and its Starch 1.2 The Structure and Morphology of Sweet Potato Starch 1.2.1 Amylose and Amylopectin 1.2.2 X-Ray Diffraction Type 1.2.3 Morphology and Size of Sweet Potato Starch 1.3 Chemical Composition of Sweet Potato Starch 1.3.1 Amylose Content 1.3.2 Lipid Content 1.3.3 Phosphorus Content 1.3.4 Moisture Content 1.4 Characteristics of Sweet Potato Starch 1.4.1 Gelatinization Temperature 1.4.2 Gelatinization Enthalpy 1.4.3 Swelling Power 1.4.4 Solubility 1.4.5 Retrogradation Rate 1.4.6 Viscosity 1.5 Sweet Potato Starch Noodles and Vermicelli 1.5.1 Brief Introduction to Starch Noodles and Vermicelli 1.5.2 Research on Starch Noodles and Vermicelli 1.6 Sweet Potato Resistant Starch

Sweet Potato Processing Technology. DOI: http://dx.doi.org/10.1016/B978-0-12-812871-8.00001-5 Copyright © 2017 China Science Publishing & Media Ltd. Published by Elsevier Inc. All rights reserved.

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Sweet Potato Processing Technology

SECTION 2: PRODUCTION TECHNOLOGY OF SWEET POTATO STARCH AND ITS SERIES PRODUCTS 2.1 Production Technology of Sweet Potato Starch 2.2 The Production Process of Sweet Potato Starch Noodles and Vermicelli 2.2 1 Brief Introduction of the Production Process of Sweet Potato Starch Noodles and Vermicelli 2.2.2 The Sweet Potato Starch Noodle and Vermicelli Production Process without Alum 2.3 The Sweet Potato Resistant Starch Production Process 2.3.1 Sweet Potato Esterified Starch 2.3.2 Sweet Potato Acetylated Starch 2.3.3 Sweet Potato Microporous Starch 2.3.4 Sweet Potato Starch-based Super Absorbent Resin SECTION 3: PHYSICOCHEMICAL PROPERTIES OF SWEET POTATO STARCH AND ITS PRODUCTS 3.1 The Structure and Physicochemical Properties of SLPS and CFS 3.1.1 Proximate Composition 3.1.2 Structural Analysis 3.1.3 Physicochemical Characteristics Analysis 3.2 Comparison of the Quality of SLPS and CFS 3.2.1 Color of the Sweet Potato Starches and Noodles 3.2.2 Retrogradation of Starch and Noodles 3.2.3 Cooking Quality 3.2.4 Textural Properties 3.2.5 Microstructure SECTION 4: APPLICATIONS OF SWEET POTATO STARCH 4.1 Application of Sweet Potato Starch in Food 4.2 Applications of Sweet Potato Resistant Starch 4.2.1 Applications of Sweet Potato Esterified Starch 4.2.2 Application of Sweet Potato Acetylated Starch 4.2.3 Application of Sweet Potato Starch-Based Superabsorbent Resin References Further Reading

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Abstract This chapter introduces the overview, production technology, physicochemical properties, and applications of sweet potato starch and its series products. It starts by presenting the structure and morphology, chemical composition, and characteristics of sweet potato starch, as well as the research on sweet potato starch noodles and vermicelli and resistant starch. It then explains production technology of sweet potato starch, starch noodles and vermicelli, and resistant starch and followed a briefly introduction on physicochemical properties of sweet potato starch, starch noodles, and vermicelli. By the end of the chapter, the applications of sweet potato starch and resistant starch are explained, suggesting the deep-processing products of sweet potato starch can play important roles in wide areas and industries.

SECTION 1: OVERVIEW OF SWEET POTATO STARCH AND ITS SERIES PRODUCTS 1.1 SWEET POTATO AND ITS STARCH Sweet potato (Ipomoea batatas Lam) is an annual herb of the family Convolvulaceae. According to the FAO statistics, as of 2010, there were more than 100 countries planting sweet potato worldwide. The production in Asia is the greatest, accounting for 91.4%; Africa is second, accounting for 5.1%, followed by Latin America, and then Europe. In Asia, the countries with large growing areas are China, Japan, South Korea, Vietnam, and Indonesia (Zhu et al., 2011). Since the 16th century (Ming Dynasty), sweet potato has become an important crop in China, with a large planting area that runs from the southern part of Hainan Province to the northern part of Heilongjiang Province (Zhu et al., 2011). Presently, China’s annual sweet potato production is 70.96 million tonnes, accounting for about 68% of the world’s sweet potato production and genetic resources (FAOSTAT, 2016). Starch is the main component of sweet potato, accounting for about 50% to 80% of its dry weight (Aina et al., 2009; Zhu et al., 2011). Therefore, sweet potato is an ideal starch resource and energy crop. Sweet potato starch plays an important role in the food, chemical, and pharmaceutical industries. Raw materials can produce natural starch and modified starch by primary processing. After the deep processing of natural starch and modified starch, varied starch products

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can be produced, such as glucose, maltogenic amylase, sugar substitutes, citric acid, sorbitol, vitamin C, and so on (Cai et al., 2008). In the food industry, sweet potato starch cannot only be used as a processing material, in addition to being used in the manufacturing of vermicelli, jelly, and other consumable products, but also as a food additive, which can be used in food as a thickener, stabilizer, or tissue reinforcing agent to improve foods to retain water, control water flow, and maintain food storage quality (Cai et al., 2008; Aina et al., 2009). Dutch Rabobank published an international food and agricultural research report on starch production in the world in 2003 (Chang, 1993), which pointed out that starch had become an important way for grain consumption. Among the numerous processed agricultural products, starch has the widest range of applications. Starch and its products are not only used for soups, meats, flavorings, breads, and drinks, but they can also be used for the production of textiles, paper, fuel, adhesives, plastics, and paints. The structure, composition, and characteristics of starch are important indices that determine its application (Chen et al., 2003).

1.2 THE STRUCTURE AND MORPHOLOGY OF SWEET POTATO STARCH 1.2.1 Amylose and Amylopectin Starch is a kind of high molecular polysaccharide that is composed of a single type of sugar unit. The basic structure of starch is D-glucose, and the starch molecules form covalent polymers through the linkage of the glycosides after the D-glucose excises the water molecules. The structure of sweet potato starch is mainly composed of two kinds of polymers, amylose, and amylopectin, which are found in starch from other plant species. 1.2.1.1 The Structure of Amylose The structure of amylose is usually expressed by the average degree of polymerization (DP), and the number-average DP (DPn) and the weight-average DP (DPw) are the most common representational methods used for DP. In addition, the range of DP is called the apparent polymerization degree distribution. The DPn of sweet potato amylose is between 4400 and 3025, DPw is 5400, and the distribution range of DP is between 19,100 and 840 (Gao, 2001). The DPw/DPn ratio of sweet potato starch is 1.3, which indicates that the molecular

Sweet Potato Starch and its Series Products

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weight distribution of sweet potato starch is narrow, the homogeneous degree of DP distribution is high, and most sweet potato starch has a molecular weight less than the DPn (Gao, 2001). There are many reports on the molecular weight of sweet potato amylose. Namutebi (2003) found that the average molecular weight of sweet potato starch amylose is from 367,000 to 521,000, which was much higher than that of 83,000 to 141,000 reported by Zhang and Oates (1999). These differences may be due to the different varieties of sweet potato and also may be caused by different determination methods. 1.2.1.2 The Structure of Amylopectin The structural model of amylopectin indicates that starch molecules have complex branches. To facilitate the structural analysis, the structures of the starch molecules were divided into three types, A, B, and C, and associated characteristics were determined, including the A chain’s reducing end is connected through α-1,6 linkages with the B or C chain; the B chain is connected with one or a plurality of A chains, its reducing end is connected through α-1,6 linkages with the C chain; the C chain is the main chain containing reducing ends. Amylopectin molecules have one C chain; therefore, one end of the C chain is the reducing end, and the other end is the nonreducing end. The chain length of amylopectin is generally reported as the average chain length. Chain length refers to the number of glucose residues in each of the nonreducing ends. Fig. 1.1 shows the structure of sweet potato

Figure 1.1 Structural schematic diagram of sweet potato amylopectin (Aina et al., 2009).

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amylopectin. The sweet potato amylopectin is mainly composed of three kinds of branched chains, A, B, and C, which is similar to the structure in other starches. The C chain, as the main chain, is located at the reducing end of the amylopectin molecule, the reducing end group is connected through 21,6 linkage, and the B chain is connected with one or a plurality of A chains. Sweet potato amylopectin is generally dominated by its B chain (.11 glucose residues per long chain), and the A chain (#11 glucose residues per short chain) is shorter than the B chain. The average chain length of sweet potato amylopectin was between 6 and 45 glucose residues/chain, where the proportion of average chain length from 6 to 10, from 11 to 15, from 16 to 20, from 21 to 30, and from 30 to 45 glucose residues/chain out of the total average chain length distribution of amylopectin were 12.4%
15.2%, 33.2%
33.8%, 22.9%
24.6%, 21.1%
23.0%, and 6.3%
7.1%, respectively (Namutebi et al., 2003).

1.2.2 X-Ray Diffraction Type Starch has a stable crystal region, which is considered to be an ordered arrangement of amylopectin molecules in the starch granule. Different starches can be divided into A, B, or C, with C being a hybrid between A and B types. Sweet potato starch is mainly A type (Cai, 2008) but also has some C type (Zhu et al., 2011) or CA type, which is between the A and C types (Ahmed et al., 2010; Noda et al., 1996, 2001). Fig. 1.2 shows the X-ray diffraction patterns of three kinds of sweet potato starch (Namutebi et al., 2003). These three kinds of sweet potato starch showed diffraction peaks at 15.0, 17.0, 17.8, and 23 (2), which are the typical of A-type starch (Noda et al., 1996). The crystal

Figure 1.2 X-ray diffraction patterns of three types of sweet potato starches.

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type of sweet potato starch might also change under certain conditions. Thus, the A type of sweet potato starch could be transformed into the B-type after being treated by high hydrostatic pressure technology (Ong et al., 1994; Gao et al., 2001). The C-type of sweet potato starch can also be gradually transformed into the B type by decreasing the cultivation temperature to an ambient temperature for sweet potato (Noda et al., 2001). The crystallinity of sweet potato starch was usually 38%, which was higher than that of cassava (37%) or potato (28%) (Gao et al., 2001).

1.2.3 Morphology and Size of Sweet Potato Starch The morphologies and sizes of natural starch granules are mainly determined by the starches’ origin, and the differences in these properties can be determined by optical or electron microscopy. The granule shape of sweet potato starch is polygonal or circular, partly oval, and bell shaped. The particle size distribution ranges from 3.4 to 27.5 μm, and the average particle size is between 8.4 and 15.6 μm. Different sweet potato starches have different particle sizes, and the particle sizes for the same kinds of sweet potato starches can also increase gradually with the growth and maturation of the sweet potato (Kitahara et al., 2002). However, the soils’ fertility, and the sweet potato planting and harvesting dates, have little effect on the particle size (Noda et al., 1996, 1997). In addition, the swelling power, solubility, and digestibility characteristics were affected by the granule size of sweet potato starch. These starch granules are larger; therefore, the swelling power and solubility are greater, but the digestibility decreases significantly (Zhang et al., 2001).

1.3 CHEMICAL COMPOSITION OF SWEET POTATO STARCH 1.3.1 Amylose Content Starches from most crops contain B25% amylose (Chen et al., 2003). The amylose content of sweet potato starch is between 15.3% and 28.8% (Table 1.1). The amylose content of sweet potato is affected by the variety and the processing method. Heat treatments slightly increase the sweet potato amylose content (Lilia et al., 1999), while α-amylase treatments significantly reduce the amylose content, suggesting that amylose is susceptible to enzymatic attack and then degradation (Rocha et al., 2010). However, the amylose contents in the same sweet potato varieties grown in different areas did not change

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Sweet Potato Processing Technology

significantly (Garcia et al., 1998), and sweet potato planting and harvesting times, as well as the fertilization status, also have no significant effects on the sweet potato amylose content (Noda et al., 1997). The amylose level also directly affects the retrogradation rate, gelatinization temperature, and swelling property of the sweet potato starch. Generally, the higher the amylose content, the faster the retrogradation rate, the higher the gelatinization temperature, and the lower the swelling power (Noda et al., 1996).

1.3.2 Lipid Content Lipids influence the properties of sweet potato starch. The formation of a starch
lipid complex can improve the textural properties of various foods (Chen et al., 2003). Compared with the lipid content of cereal starch, the lipid content in sweet potato starch is low, so the effect of the starch
lipid interaction on the starch properties is not significant (Gao et al., 2001). The lipid content in sweet potato starch ranges from 0.14% to 0.21% (Table 1.1), but its content is also different in the starch of different sweet potato varieties (Collado et al., 1999). The presence of a high lipid content inhibits the swelling and dissolution of sweet potato starch granules, increases the opacity of the starch paste and membrane, influences the thickening and adhesive abilities, and leads to the oxidative rancidity of starch (Chen et al., 2003). However, the appropriate addition of lipids can also reduce the viscosity of sweet potato starch and improve the stability of the starch paste (Gao et al., 2008).

Table 1.1 Chemical Composition of Sweet Potato Starch Component

Content (%)

References

Amylose

15.3
28.8

Zhu et al. (2011); Aina et al. (2009); Noranizan et al. (2010); Chen et al. (2003); Kitahara et al. (2005); Ahmed et al. (2010); Namutebi et al. (2003); Zaidul et al. (2007)

Moisture

8.0
11.8

Zhu et al. (2011); Aina et al. (2009); Chen et al. (2003); Jangchud et al. (2003)

Protein (db)

0.1
0.23

Aina et al. (2009); Rocha et al. (2010); Chen et al. (2003); Jangchud et al. (2003); Kitahara et al. (2005)

Ash (db)

0.1
0.5

Aina et al. (2009); Rocha et al. (2010); Jangchud et al. (2003)

Fat (db)

0.14
0.21

Rocha et al. (2010); Chen et al. (2003); Jangchud et al. (2003); Kitahara et al. (2005)

Phosphorus (db)

0.014
0.022

Chen et al. (2003)

Sweet Potato Starch and its Series Products

9

1.3.3 Phosphorus Content There was a large difference in the phosphorus content of different sweet potato varieties, and the phosphorus content in sweet potato starch ranged from 0.014% to 0.022% (Table 1.1). Most of the phosphorus in the sweet potato starch bound with starch molecules through covalent bonds. The phosphorus content ranged from 290 to 320 g/g, and its content in amylose (3
6 μg/g) is lower than that in amylopectin (117
144 μg/g; Chen et al., 2003). There are no significant effects of different fertilizer levels on the phosphorus content of sweet potato starch (Noda et al., 1996). Usually, phosphorus increases the viscosity and improves the gel strength of sweet potato starch. At the same time, phosphorus reduces the gelatinization temperature of sweet potato starch, accelerates the hydration and swelling, and increases the transparency (Chen et al., 2003). Thus, sweet potato starch with high a phosphorus content is suitable for the production of starch noodles/ vermicelli.

1.3.4 Moisture Content The moisture content of sweet potato starch ranges from 8% to 11.8% (Table 1.1). The moisture level of sweet potato starch mainly depends on the degree of starch dryness and also depends on the starch and water-binding capacity (Chen et al., 2003). The higher the moisture content of starch, the easier microbes grow, resulting in a lower quality. For most of the starch production areas, the safe storage moisture content is below 13% (Gao, 2001). China’s national standards for the moisture content of starch indicates that the desired moisture contents of potato starch and cassava starch were less than 14% and 18%, respectively, and the maximum moisture contents should not exceed 15% and 20%, respectively. However, there are no national standards for the moisture content of sweet potato starch.

1.4 CHARACTERISTICS OF SWEET POTATO STARCH 1.4.1 Gelatinization Temperature Differential scanning calorimetry (DSC) determined the pasting parameters, Tonset (To), Tpeak (Tp), and Tend, of sweet potato starch as being 66.2
71.3 C, 69.5
79.78 C, and 80.4
88.5 C, respectively, with Tend rarely decreasing to 75.29 C (Table 1.2), whereas the gelatinization temperature range of sweet potato starch, as determined by rapid

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Sweet Potato Processing Technology

Table 1.2 DSC Properties, Swelling Power and Solubility of Sweet Potato Starch Related Parameters

Data

References

To ( C)

66.2
71.3

Zhu et al. (2011); Ahmed et al. (2010); Noda et al. (1996, 2002); Kitahara et al. (2005); Alummoottil et al. (2005); Garcia et al. (1998); Collado et al. (1999)

Tp ( C)

69.5
79.78

Zhu et al. (2011); Rocha et al. (2010); Ahmed et al. (2010); Noranizan et al. (2010); Noda et al. (1996); Chen et al. (2003); Moorthya (2010); Alummoottil et al. (2005); Garcia et al. (1998); Collado et al. (1999)

Tend ( C)

75.29
88.5

Zhu et al. (2011); Rocha et al. (2010); Ahmed et al. (2010); Garcia et al. (1998); Collado et al. (1999)

WH (J/g)

7.8
15.5

Zhu et al. (2011); Rocha et al. (2010); Ahmed et al. (2010); Moorthya (2010); Kitahara et al. (2005)

Swelling power (85 C, mL/g)

32.3
50

Ahmed et al. (2010); Moorthya (2010)

Solubility (85 C, %)

1.5
13.65

Zhu et al. (2011); Aina et al. (2009); Moorthya (2010)



Note: To—Tonset, initial pasting temperature; Tp—Tpeak, peak pasting temperature; Tend—pasting termination temperature; WH—gelatinization enthalpy.

Table 1.3 Paste Parameters of Sweet Potato Starches Viscosity and Relative Parameters

Data

References

Peak viscosity

143
469

Zhu et al. (2011); Aina et al. (2009); Noda et al. (1996)

Minimum viscosity

91
214

Zhu et al. (2011); Aina et al. (2009)

Breakdown viscosity

29.4
255

Zhu et al. (2011); Aina et al. (2009); Noda et al. (1996)

Final viscosity

82.9
284

Zhu et al. (2011); Aina et al. (2009)

Retrogradation viscosity

15
78

Zhu et al. (2011); Aina et al. (2009); Noda et al. (1996)

Gelatinization temperature

65.9
87.7

Aina et al. (2009); Noda et al. (1996); Moorthya (2010)

Peak time

3.4
7.44

Zhu et al. (2011); Aina et al. (2009)

viscosity analyzer, mostly ranged from 65.9 to 79.9 C, with a small peak at 87.7 C (Table 1.3). The differences in the pasting temperature values were obtained by DSC and the rapid viscosity. In addition, the gelatinization temperatures of sweet potato starch from different varieties and regions are also different. During sweet potato cultivation, earlier planting times can significantly improve the To, while later harvest times can reduce the To (Noda et al., 1997). However, the fertilizer concentration has little effect on the gelatinization temperature of sweet potato starch (Noda et al., 1996).

Sweet Potato Starch and its Series Products

11

1.4.2 Gelatinization Enthalpy The gelatinization enthalpy of sweet potato starch is related to its amylopectin, intramolecular bonds, genes, and environmental factors (Gao, 2001). The enthalpy value of sweet potato starch is between 7.8 and 15.5 J/g (Table 1.2). Because sweet potato amylopectin is the main component of its crystalline region, the higher the amylopectin content, the greater the enthalpy (Zhang and Oates, 1999). The gelatinization enthalpy is also significantly affected by the different varieties and the different planting conditions (Noda et al., 1996; Garcia et al., 1998). In addition, the gelatinization enthalpy of starch from sweet potato during the early growth period is low, between 11.8 and 13.4 J/g (Kitahara et al., 2002), whereas postponing the harvest date increases the gelatinization enthalpy of sweet potato starch (Noda et al., 1997).

1.4.3 Swelling Power

The swelling power of starch was determined at 85 C. The swelling power of sweet potato starch was between 32.5 and 50 mL/g (as shown in Table 1.2), showing strong intramolecular forces. Sweet potato starch has two section expansions, suggesting that there are two kinds of binding forces (Chen et al., 2003). The swelling power is different for sweet potato starch from different varieties and is also significantly different for starch from the same sweet potato varieties under different temperature conditions (Aina et al., 2009; Chen et al., 2003). Recent research showed that the swelling power of sweet potato starch is greatly influenced by amylose, which plays the role of diluent in the starch expansion process, especially when amylose and lipid compounds form complexes. Then, the swelling of starch granules can be significantly inhibited (Gao, 2001). However, some studies showed that there is no significant correlation between the amylose content and swelling power (Collado et al., 1999). In addition, the swelling power of sweet potato starch is also affected by the molecular weight and molecular shape of amylopectin. The more amylopectin with 6
9 glucose residues/chain, the greater the swelling power of starch, whereas the more amylopectin with 12
22 glucose residues/chain, the smaller the swelling power (Gao, 2001).

1.4.4 Solubility The solubility of sweet potato starch is between 1.5% and 13.65% (as shown in Table 1.2). The lower solubility of sweet potato starch might

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Sweet Potato Processing Technology

be because of the smaller size of the starch granules, stronger internal binding capabilities, and less glucose containing phosphate groups. Starches from different sweet potato cultivars showed different solubility levels. The solubility of starch increased with the temperature increase, and the highest solubility reached 13.65% at temperatures above 85 C (Moorthya, 2010). The solubility of some of the commercial sweet potato starch produced in Peru reached 28% (Garcia et al., 1998). In addition, when the temperature was below 60 C, the solubility of starch from sweet potatoes of different regions showed no significant differences. However, when the temperature was higher than 60 C, the difference was significant (Nuwamanya et al., 2010). The greater the swelling power, the greater the solubility of the starch, but at the same temperature treatment, the dissolution rates of sweet potato starch from different varieties were not exactly the same (Gao, 2001).

1.4.5 Retrogradation Rate During the short-term retrogradation after starch gelatinization, the amylose molecules can be rearranged into parallel straight lines, and further gelatinization requires temperatures of up to 130 C (Tan et al., 2008). In addition, during the long-term retrogradation after starch gelatinization, the external branches of amylopectin also slowly recrystallize, producing recrystallized amylopectin, which can also regelatinize to the gel state at temperatures below 55 C. Under the same conditions, compared with other starches, the retrogradation rate of corn starch is the fastest, which is mainly due to the higher content of amylose (28%) and lipids (0.8%) (Gao, 2001). The amylose and lipid contents of sweet potato starch are relatively low. By contrast, sweet potato starch shows a low-to-moderate retrogradation rate (Collado et al., 1999). The retrogradation rate and degree of retrogradation of sweet potato starch increase with the increase in the amylose content. An increase of short-branched chains (CL 12
14 glucose residues/ chain) in amylopectin causes the retrogradation rate to accelerate, whereas an increase of short-branched chain (CL 9
11 glucose residues/chain) in amylopectin causes the retrogradation rate to decelerate (Miyazaki, 2000; Zhang and Oates, 1999). In addition, the retrogradation of sweet potato starch is also dependent on the concentration, storage temperature, pH value, and chemical compositions of starch. At high starch concentrations, low storage temperatures, and a suitable range of pH conditions, the retrogradation rate accelerates,

Sweet Potato Starch and its Series Products

13

whereas the retrogradation rate decreases significantly at higher ionic concentrations (Gao, 2001; Ishiguroa et al., 2003).

1.4.6 Viscosity The peak, lowest, breakdown, final, and setback viscosity ranges of sweet potato starch (concentration of 10%, w/w) were from 143 to 469 Rapid Visco Unit (RVU), from 91 to 214 RVU, from 29.4 to 255 RVU, from 82.9 to 284 RVU, and from 15 to 78 RVU, respectively (Table 1.3). The viscosity properties of sweet potato starch are greatly influenced by the variety of sweet potato starch, the starch content, and the interactions between different components (Gao, 2001; Nuwamanya et al., 2010). There are significant differences between the peak viscosities and the peak times in different cultivars of sweet potato starch, and the peak viscosity and the amylose content have a significant negative correlation (Collado et al., 1999). However, other studies reported that the peak viscosity and the amylose content are irrelevant (Chen et al., 2003). In addition, the higher the starch concentration, the greater the viscosity (Yuan et al., 2008). The presence of lipid compounds in starch reduces the peak viscosity of the starch paste and improves its stability (Gao et al., 2008). The breakdown viscosity of starch is an important parameter when measuring the stability of starch paste, which reflects the ability to resist mechanical shearing during heating. The lower the breakdown viscosity, the higher the shearing resistance. Similar to other starches, the breakdown value of sweet potato starch is not only affected by the various starch components but is also affected by the fine structure of sweet potato amylopectin. It is also affected by the fine structure of sweet potato starch. The higher the proportion of long chains in the amylopectin, the higher the stability. This was mainly because the amylopectin long chains can intertwine with other amylopectin molecules, reducing its decentralized trend, increasing the radius of gyration, and maintaining the starch viscosity at the same time (Zaidul et al., 2007; Yuan, 2008). The retrogradation viscosity is different between the breakdown of viscosity and final viscosity, which is an index that measures the starch retrogradation rate. For different varieties of sweet potato starch, the retrogradation rate is different. This is mainly dependent on the amylose content and amylopectin properties of

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Sweet Potato Processing Technology

sweet potato starch. An ideal starch widely used in food should have characteristics that maintain its stability and form a smooth structure at low concentrations, keeping it soft and flowing at low temperatures and resistant to high shearing at high temperatures (Garcia et al., 1998).

1.5 SWEET POTATO STARCH NOODLES AND VERMICELLI 1.5.1 Brief Introduction to Starch Noodles and Vermicelli Starch noodles and vermicelli, with thousands of years of history, are traditional foods generally preferred in China and the majority of Asian countries and regions (Zhao et al., 2009). The noodles are made by sweet potato starches isolated during sour liquid processing (SLPS), the traditional technology in China. The starch and other impurities are separated by sour liquid produced by natural fermentation, but the method is complicated, highly empirical, and the extraction time relied on nature, including the season, climate (air humidity, temperature, etc.), geography, and bacteria. It was not suitable for mechanization or automation for continuous production. Therefore, the noodles and vermicelli have been made by sweet potato starches by centrifugation (CFS) in recent years.

1.5.2 Research on Starch Noodles and Vermicelli Starch noodle and vermicelli have been produced for a long time, but relevant research reports on their production factors are limited and were mainly from China, Japan, and Southeast Asian countries. Wei (1987) studied mung bean dipping, the isolation and identification of Lactococcus lactis in the sour liquid, and the relationship between starch extraction and metal ions during Longkou vermicelli production. They then promoted a new method to improve the recovery rate of starch. Qian (1990) reported systematic research on the properties of raw starch and the physicochemical properties of vermicelli. Yuan (1991) studied the physicochemical properties, including X-ray diffraction, amylose and amylopectin content, relative molecular weight, starch swelling power, and the solubility and its related characteristics, of vermicelli made by the starches from mung bean, broad bean, potato, and sweet potato. Zhang (2001) introduced this Chinese specialty food in the German authoritative academic journal Starch, causing widespread concern. Chen et al. (2003) studied the physicochemical properties of starches from three sweet potato varieties found in China and the

Sweet Potato Starch and its Series Products

15

quality of the noodles they produced and found that sweet potato starch is not only suitable for producing starch noodles, but also that the sensory qualities of the starch noodles made by “SuShu8” starch are better than those produced by mung bean starch. In addition, the smaller the sweet potato starch particles’ size, the better the starch noodles’ texture. Huang et al. (2008) found that the sweet potato amylose content and gel strength are significantly positively correlated with shear stress and deformation, and tensile strength and deformation and were significantly negatively correlated with the breaking rate of starch noodles. The lower the starch gelatinization temperature, the more obvious the long-term retrogradation and the better the noodles quality. Similar results were also found in the study of Zhao et al. (2009).

1.6 SWEET POTATO RESISTANT STARCH In 1992, Englyst et al. (1992) classified sweet potato starches. Rapidly digestible starch is the starch that is digested within 20 min by α-amylase and fungal glucoamylase; slowly digestible starch is the starch that is digested between 20 and 120 min; and resistant starch is the starch that has not been digested after 120 min. In 1996, resistant starch was further defined and standardized by the European Resistant Starch Association. It is the general name of starch and its hydrolysate, which are not absorbed by the small intestines of a healthy human body. Eerlingen et al. (1995) divided starch into four categories, the physical embedding starch RS1, resistance starch granules RS2, retrograded starch RS3, and chemical modified starch RS4, according to the resistant starch morphology and physicochemical properties. RS1 is formed by the starch buried in the starch food matrix. These starch granules could not be fully expanded and dispersed in the aqueous solution due to cell wall or protein separation, which made the amylase and starch granules unable to contact them, producing the phenomenon of antienzymatic hydrolysis. Under the influence of mastication or food processing, the RS1 content in food was very easily changed, being mainly found in fully- or partially ground grains, beans, and wheat. RS2 contains resistant starch granules and nongelatinized starch granules. Usually, when starch granules are not gelatinized, they have a high degree of resistance to α-amylase digestion. Gallant et al. (1992) believed that the high density and partial crystallinity of starch could reduce its sensitivity to the enzyme. This kind of starch mainly exists in the raw starch of potatoes and peas, green banana, and high

16

Sweet Potato Processing Technology

amylose corn starch. Due to its structural integrity, high density and the natural crystal structure of high amylose corn starch, the resistance to digestion was formed in RS2 (Ranhotra et al., 1996; Guraya et al., 2001). RS3 refers to partially recrystallized gelatinized starch that forms more stable hydrogen bonds during cooling and storing processes, and thus has digestion-resistant characteristics, which often occur in the cooled rice, bread, potato, and canned pea products after cooking. RS4 is starch with molecular structural changes caused by chemical crosslinking methods and some introduced chemical functional groups. Thus, digestion-resistant characteristics (Woo et al., 2002), such as acetyl starch, hydroxypropyl starch, heat denatured starch, and phosphorylated starch, are produced, and they can be used as food-processing ingredients.

SECTION 2: PRODUCTION TECHNOLOGY OF SWEET POTATO STARCH AND ITS SERIES PRODUCTS In Section 1, a preliminary introduction to sweet potato starch, starch noodles, vermicelli, and sweet potato resistant starch was provided. The following contains a brief introduction to the production technology of sweet potato starch, starch noodles, vermicelli, and sweet potato resistant starch.

2.1 PRODUCTION TECHNOLOGY OF SWEET POTATO STARCH In our country, there are currently two main sweet potato starch production methods: Traditional sour liquid and centrifugation (Song, 2003; Wang, 2008). The production technology of sweet potato starch using the sour liquid method is as follows: First, sweet potatoes are ground, and the starch slurry is collected after filtering to remove residues. After a natural fermentation period, the slurry slowly becomes more acidic, reaching pH B4, changing the slurry to a sour liquid that can precipitate starch. The sour liquid is the “waste” during the production of sweet potato starch by the sour liquid method and is an indispensable “additive” for the sour liquid method. Thus, it is formed during production and is then returned to the production. The chemical components of sour liquid mainly included starch, water, protein, cellulose, lactic

Sweet Potato Starch and its Series Products

17

acid, peptides, amino acids, soluble oligosaccharides, monosaccharides, and ash. For different sour liquids, the chemical components are also different. The role of sour liquid is mainly to obtain pure starch using L. lactis, lactic acid, and other metabolites in the sour liquid to separate the other impurities and starch. The quality and quantity of the sour liquid is directly related to the extraction rate of the starch and the quality of the related products. After sweet potato has been ground, the starch slurry is obtained, which contains starch, protein, fiber, and other components. To more efficiently separate starch, fiber, and protein, the sour liquid is added using the traditional technology. It includes L. lactis, which has the ability to agglutinate starch granules. Thus, the starch particle settling speed is greatly accelerated, and the adsorption effects of protein and fiber are determined. Thus, the starch is separated from protein and fiber. In a word, the production technology of sweet potato starch by the sour liquid method is a traditional starch production method using the addition of naturally fermented sour liquid (relying on natural microorganisms, mainly the acidic slurry fermented by Lactobacillus) to make the starch rapidly settled. This method requires less investment is economical and has other advantages. It is widely used in our country, and its processing flow is shown in Fig. 1.3. The main shortcomings of this method include the requirement for experienced workers to produce starch, which is constrained by many factors, such as the bacterial ratio and type in the air, the pH value of the sour liquid, the temperature of the workshop, human error during worker operation, and the hardness and metal species in the water. These factors can easily lead to fluctuations in starch production using the sour liquid method at different times and within different batches.

Figure 1.3 Flow chart of the factory-based production of sweet potato starch using sour liquid.

18

Sweet Potato Processing Technology

The centrifugation technology, which was rapidly developed in recent years, is a method that relies on high-speed centrifugation to rapidly separate starch. The grinding and filtration in centrifugation technology were similar to those used in the sour liquid method, but the centrifugation technology separated the starch, protein and fiber in the slurry using a dish-type centrifuge or differential grade cyclone washing technology, or a combination of these. The combined processing technology isolated the crude starch slurry using a dish-type centrifuge to separate the protein, fiber, and other components and then obtained the purified starch using a differential grade cyclone scrubber. The new method was suitable for modern, continuous, large-scale production. Its disadvantage was that the investment in production equipment was large, and the color and appearance of the starch was not ideal. Therefore, further research was required to improve and perfect the production equipment and technology. The production processes and operation flows of the sour liquid and centrifugation methods showed significant differences, which may affect or change the structures and physicochemical properties of sweet potato starch and directly led to quality differences in the starch derivatives produced. Liu and Shen, (2007a) found that mung bean starch produced using the sour liquid method had higher protein, fat and amylose contents, higher gel transmittance and brightness, and lower swelling powers and solubility levels than those produced by the centrifugation method. Gao et al. (1998) believed that the vermicelli produced from starch by the sour liquid method showed more desirable levels of transparency, flexibility, and cooking resistance than those of the starch produced by the centrifugation method. However, Liu and Shen, (2007b) found that there were no significant differences in the polarized property, microstructure, and X-ray diffraction of mung bean starch by the sour liquid and centrifugation methods. In addition, relevant reports also pointed out that the starches extracted by lactic acid fermentation (sour liquid method) and the centrifugation method showed significant differences in their physicochemical properties, such as rice starch (Lu et al., 2003), tapioca starch (Numfor et al., 1995), corn starch (Yuan et al., 2008), and so on. It may be that the Lactobacillus, enzyme, and acid changed the composition and structure of starch during lactic acid fermentation processing and then affected the physicochemical properties of the starch (Chang et al., 2006; Numfor et al., 1995).

Sweet Potato Starch and its Series Products

19

2.2 THE PRODUCTION PROCESS OF SWEET POTATO STARCH NOODLES AND VERMICELLI 2.2.1 Brief Introduction of the Production Process of Sweet Potato Starch Noodles and Vermicelli Starch noodles are a kind of traditional food having pliable, refreshing, and lubrication characteristics, which are very popular among Chinese consumers. The processing of sweet potato starch materials into noodles and vermicelli was divided into two types, one used fresh sweet potato as a raw material, first processed into starch and then processed into starch noodles and vermicelli, and the other one directly used starch as the raw material for starch noodles and vermicelli processing. The traditional processing techniques of sweet potato starch noodles/ vermicelli generally included: Starch processing, gelatinization and dough making, extruding, hot water gelatinization, drying, and packaging. To ensure the quality of sweet potato starch noodles/vermicelli, a certain amount of alum was usually added during gelatinization. The modernization of the processing techniques of sweet potato starch noodles/vermicelli generally used the starch as a raw material, and the production processing steps were slurry mixing, gelatinization, dough making, extrusion and shaping up, heating and gelatinization, cooling, freezing and thawing, drying (air drying and oven drying), and packaging.

2.2.2 The Sweet Potato Starch Noodle and Vermicelli Production Process without Alum During the production of sweet potato starch noodles and vermicelli, alum was used as a traditional additive, even though the excessive intake of aluminum could cause potential harm to human health. The Sanitary Standards of Using Food Additives (GB 2760-2007) in China authorized that the aluminum content in products should not exceed 100 mg/kg of dry sample, which means that the alum dosage should be less than 1m. However, for a long time, to obtain the ideal sweet potato starch noodles and vermicelli, a higher alum concentration was added to the sweet potato starch noodles and vermicelli during processing, and the excessive aluminum situation was very serious. In recent years, looking for alum substitutes and developing sweet potato starch noodles and vermicelli without alum has become a hot research topic.

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Sweet Potato Processing Technology

2.2.2.1 The Production Process of Pure Sweet Potato Starch Noodles and Vermicelli Ma (2012) invented a pure sweet potato starch noodle processing method using the following specific steps: Slurry mixing, stirring, the first gelatinization (the sweet potato starch slurry was added to the starch noodle processing machine by screw extrusion, with an adjusted motor speed of 600
900 rpm and gelatinization temperature 70
90 C, which made jelly sheets reach a medium-well state), the secondary gelatinization (the medium-well jelly sheets were placed in 90
100 C water to complete gelatinization), retrogradation, cutting, and air drying or oven drying to obtain pure sweet potato starch noodles. 2.2.2.2 The Sweet Potato Starch Noodle and Vermicelli Production Process Zhang et al. (2011) studied the following production process of pumpkin-sweet potato starch noodles: raw material (sweet potato starch, pumpkin juice), ground (mass fraction of pumpkin juice was 35%), boiling and gelatinization (96
98 C, 58
60 s), extruding and shaping (diameter 1 mm, length 150 cm), natural cooling, freezer storage (26 to 28 C, 6
10 h), water thawing (20
30 C), loosening, natural drying (final water content 10%
13%), packaging, and storage. Zhao (2009) researched and developed the following formula for the nutritional sweet potato starch noodles without alum: 40-g cooked sweet potato mud with white flesh, 35-g pea starch, 25-g sweet potato starch, 4-g modified potato starch, 1-g salt, 0.5-g composite thickening agent, and 0.4-g compound phosphate. 2.2.2.3 Effects of Different Additives on the Quality of Sweet Potato Starch Noodles and the Vermicelli Chen (2006) found that Artemisia sphaerocephala Krasch. gum was favorable for starch retrogradation, improving the edible quality of sweet potato vermicelli and could be used as a processing aid instead of alum with an additive amount of 0.2%
0.3%. However, the addition of soybean protein decreased the retrogradation of sweet potato starch and had a negative impact on the texture of sweet potato starch-based vermicelli. Tan et al. (2008) studied the effects of different additives on the quality of sweet potato vermicelli and found that when the proportion of konjac gum and Artemisia sphaerocephala Krasch. gum was

Sweet Potato Starch and its Series Products

21

0.95:0.05, and the total addition amount was 1%, the quality of sweet potato starch vermicelli was close to that of both mung bean vermicelli and sweet potato vermicelli with alum. Wang et al. (2008) added the synthetic sweet potato starch phosphate ester, konjac powder, and compound phosphate to sweet potato starch, producing sweet potato starch vermicelli without alum. When the amount of sweet potato starch phosphate ester added was 6%, the Pro proportion and added amounts of konjac powder having compound phosphate was 2/3% and 1%, respectively. The breaking rate and cooking loss were better than those of sweet potato starch vermicelli with alum.

2.3 THE SWEET POTATO RESISTANT STARCH PRODUCTION PROCESS 2.3.1 Sweet Potato Esterified Starch Esterified starch is a type of modified starch, which is used as a raw material and is crosslinked using sodium trimetaphosphate, hexametaphosphate, and other phosphates, as crosslinking agents, and sodium hydroxide as the catalyst being widely used in food, medicine, paper making, and textile industries. He et al. (2004) treated sweet potato starch with an orthophosphate in an aqueous slurry to produce sweet potato starch phosphate monoesters. Compared with sweet potato starch, the paste transparency of sweet potato starch phosphate monoesters was significantly improved. The retrogradation tendency of the paste was weakened, and the freeze
thaw stability was improved. The technology of sweet potato starch phosphate monoesters in an aqueous slurry was as follows: starch, phosphate solution, pH value (using HCl or NaOH) adjusting, stirring, filtering, predrying, sieving, esterification reaction, washing (water or alcohol), and production. Ma et al. (2012) used sodium trimetaphosphate to react with sweet potato starch to prepare sweet potato starch phosphate ester and indicated that the gelatinization temperature of sweet potato starch phosphate ester increased. The hot and cold viscosity stabilities, freeze
thawing stability, and salt tolerance capability were higher than those of raw starch, but the transparency and retrogradation showed no obvious advantages. Acetic acid ester starch is a common chemically modified starch. It has the characteristics of weak coagulation, high transparency, low pasting temperature, and stable storage, and thus, could be used as an

22

Sweet Potato Processing Technology

ideal food thickener. Yuan (2006) prepared the acetic anhydride esterification starch and determined its properties, using sweet potato starch as the raw material. The transparency, solubility, swelling power, and freeze
thaw stability of esterified starch were determined and obviously improved compared with those of raw sweet potato starch. The preparation of the sweet potato acetic anhydride esterification starch was as follows: Starch, addition of water, stirring, adjusting of pH value (3% NaOH), adding acetic anhydride, reacting, adjusting pH value to 6.5, filtering, washing by distilled water, vacuum drying, crushing, and sieving (200 mesh).

2.3.2 Sweet Potato Acetylated Starch Acetylated starch has the characteristics of easy gelatinization, good freeze
thaw stability, and high paste transparency, but it lacks heat, acid, and shearing resistance. Gao (2010) studied the properties of crosslinked starch, acetylated starch and crosslinked acetylated starch paste, along with sweet potato starch, as raw materials, and found that sweet potato starch, after crosslinking modifications, enhanced the hot paste stability but decreased the transparency. In addition, freeze
thaw stability showed no obvious improvement, and modifying acetylation reduced the pasting temperature, and improved the transparency and freeze
thaw stability of starch, but the hot paste stability was poor. The crosslinked acetylated composite modifications reduced the pasting temperature of starch but improved the hot paste stability, transparency, and freeze
thaw stability of starch paste. The preparation process of sweet potato acetylated starch was as follows: Sweet potato starch slurry (40%), pH value adjusted to 8.0 (3% NaOH), acetic anhydride (6% based dry starch quality) added, 45 C water bath incubation for 1 h, and pH value adjusted to neutral. This was followed by washing, drying, crushing, sieving through 100 mesh screen, and obtaining sweet potato acetylated starch.

2.3.3 Sweet Potato Microporous Starch Microporous starch is a type of modified starch. The starch is treated by acid or enzymatic hydrolysis, which causes erosion and micropore formations on the surface of starch granules and then changes the adsorption properties. Xie et al. (2008) prepared crosslinked microporous sweet potato starch using phosphorus oxychloride as a crosslinking agent, high-temperature resistant α-amylase as a crosslinking enzyme, and the hydrolysis method of crosslinking. Compared with

Sweet Potato Starch and its Series Products

23

those of the sweet potato starch, the water absorption rate, oil absorption rate, retrogradation, shear resistance, acid resistance, and freeze
thaw stability were improved greatly. Huang et al. (2011) studied the production process of sweet potato microporous starch by fermentation, and the specific process flow was as follows:

Activation on inclined plane - seed culture - enzyme production by fermentation - partial hydrolysis - microporous starch - continued fermentation by adding supplementary materials - filtration washing - drying - crushing - sweet potato microporous starch.

2.3.4 Sweet Potato Starch-based Super Absorbent Resin Super absorbent resin is a new kind of functional polymer material that contains a large number of hydrophilic residues. Due to its unique spatial network structure, swelling (they can absorb several hundred times their own weight or even tens of thousands times of water) could occur in a short period of time after contact with water. Then, the gel was formed, having a strong water retention ability that would not drip even under pressure. The performance of superabsorbent resin was far superior to that of traditional absorbent materials (such as cotton, paper, etc.) and could be used as a water holding agent, dehydrating agent, soil improvement agent, etc. Thus, it was widely used in food, hygiene, agriculture, petroleum, chemical, materials, and other industries (Xu, 2004). At present, starch-based super-absorbent resins use corn and cassava starch as their main raw materials, and the preparation of superabsorbent resin using sweet potato starch has also attracted interest. Jiang et al. (2004) studied the influence of initiator concentrations, catalyst concentrations, and other factors, on the water absorption of superabsorbent resin using potassium permanganate as an initiator with sweet potato starch as the raw material, and the optimum conditions of the reaction were determined. Yuan et al. (2006) prepared superabsorbent resin through grafted copolymerization between sweet potato starch and acrylic acid, initiated by ammonium persulphate under the protection of nitrogen (N) using the N,N0 methylene double acrylic amide (monomer) as a crosslinking agent.

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Sweet Potato Processing Technology

The effects of the amount of the crosslinking agent, concentration of sweet potato starch slurry, starch variety, the amount of monomer and initiator, and reaction temperature on the water absorption properties of the resin were investigated. The absorbent resin obtained had a strong absorption capability, with a water absorbing quality that could be as high as 500 times its own weight. Wu et al. (2005) prepared superabsorbent resin through the copolymerization of sweet potato starch and acrylic acid, with sweet potato starch as the main raw material, N,N0 -methylene bisacrylamide as the crosslinking agent, potassium persulfate as the initiator, and glycerol as the crosslinking agent. The specific process was as follows:

SECTION 3: PHYSICOCHEMICAL PROPERTIES OF SWEET POTATO STARCH AND ITS PRODUCTS In Section 2, we discussed the main production methods of sweet potato starch, traditional sour liquid, and centrifugation. Sweet potato starch is mainly used for the production of sweet potato starch noodles and vermicelli. The following is a briefly introduction to the physicochemical properties of sweet potato starch, starch noodles, and vermicelli.

3.1 THE STRUCTURE AND PHYSICOCHEMICAL PROPERTIES OF SLPS AND CFS Here, we systematically compare the structural differences in SLPS and CFS. The differences in their physicochemical characteristics are explained from the point of view of structure, thus providing fundamental data that will be useful in adapting sweet potato starch to food products.

3.1.1 Proximate Composition The proximate composition of SLPS and CFS are shown in Table 1.4. The purity levels of the starches produced by the two methods were relatively high, and there was no significant difference between them.

Sweet Potato Starch and its Series Products

25

Table 1.4 Chemical Composition of Sweet Potato Starch (%, w/w, db) Samples

Starch Purity

LCFS

Amylose

Protein

26.01 6 0.21

a

0.08 6 0.00a

24.27 6 0.21

b

0.11 6 0.00a

96.09 6 1.25a

24.27 6 0.19a

0.16 6 0.00a

CSLPS

95.23 6 0.62

22.21 6 0.19

0.17 6 0.00a

Samples

Lipid

LCFS

96.22 6 0.62

a

LSLPS

97.27 6 1.84

a

CCFS

a

Ash

b

Phosphorous

0.13 6 0.01

b

0.010 6 0.00b

0.26 6 0.03

a

0.023 6 0.00a

0.07 6 0.02a

0.15 6 0.01b

0.011 6 0.00b

0.11 6 0.11

0.18 6 0.02

0.029 6 0.00a

0.10 6 0.07

a

LSLPS

0.16 6 0.04

a

CCFS CSLPS

a

a

Different letters in the same column indicate significant differences (P , 0.05). LCFS, laboratory sweet potato starch made by centrifugation; LSLPS, laboratory sweet potato starch made by sour liquid processing; CCFS, commercial sweet potato starch made by centrifugation; CSLPS, commercial sweet potato starch made by sour liquid processing.

The amylose contents of laboratory sweet potato starch made by SLPS (LSLPS), laboratory sweet potato starch made by centrifugation (LCFS), commercial sweet potato starch made by SLPS (CSLPS), and commercial sweet potato starch made by centrifugation (CCFS) were 24.27%, 26.01%, 22.21%, and 24.27%, respectively. The amylose content of SLPS was lower than that of CFS, which might be due to the partially degradation of the amylose of SLPS. Liu et al. (2006) indicated that extracellular enzymes produced by Lactobacillus in the sour liquid method could decompose partial starch during starch precipitation, which reduced the starch yield. Therefore, the degradation effects of the enzymes in the sour liquid method on starch might be attributed to the decrease in the amylose content. In addition, the protein, fat, ash, and phosphorus contents in SLPS were significantly higher than those in the starch produced by centrifugation. Some Lactobacillus may be precipitated with the starch during fermentation, resulting in the increases in the protein, ash, and phosphorus contents (Liu and Shen, 2007a). Compared with SLPS, the contents of Al, Mg, Fe, and Zn of CFS were significantly higher, whereas the K and Ca contents were significantly lower. Other heavy metal elements, such as Cu, Mn, Mo, Pb, As, Cd, and Cr, were not detected in either starch (Table 1.5).

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Table 1.5 Mineral Compositions of Sweet Potato Starches (db, w/w, mg/kg) Samples

Al

K

LCFS

30.24 6 0.34

LSLPS

2.42 6 0.21

CCFS

19.82 6 0.38a

CSLPS

2.84 6 0.23

Samples

Fe

5.98 6 0.12

a

Ca 173.10 6 0.87

31.59 6 0.44a

238.91 6 0.55

a

28.59 6 0.67b

15.50 6 0.58b

304.93 6 2.10b

55.19 6 0.87a

35.62 6 0.34

827.18 6 0.98

7.28 6 1.12b

10.70 6 0.98

b

b

Mg b

b a

b

Zn

LCFS

5.01 6 0.37

a

16.15 6 0.99

LSLPS

3.89 6 0.78b

7.40 6 0.31b

CCFS

6.20 6 0.44a

6.35 6 0.45a

CSLPS

4.72 6 0.56b

2.87 6 0.32b

a

Cu Mn Mo Pb As Cd Cr a

Not detected

Different letters in the same column indicate significant differences (P , 0.05). LCFS, laboratory sweet potato starch made by centrifugation; LSLPS, laboratory sweet potato starch made by sour liquid processing; CCFS, commercial sweet potato starch made by centrifugation; CSLPS, commercial sweet potato starch made by sour liquid processing.

Table 1.6 Size Distributions of Sweet Potato Starch Granules Samples

Size Range (μm)

Mean Size (d43, μm)

LCFS

2.60
30.66

13.15a

LSLPS

2.32
26.16

11.44b

CCFS

2.62
30.51

13.13a

CSLPS

2.49
25.98

11.57b

Different letters in the same column indicate significant differences (P , 0.05). LCFS, laboratory sweet potato starch made by centrifugation; LSLPS, laboratory sweet potato starch made by sour liquid processing; CCFS, commercial sweet potato starch made by centrifugation; CSLPS, commercial sweet potato starch made by sour liquid processing.

3.1.2 Structural Analysis 3.1.2.1 Particle Size Distribution The particle size distributions and mean granule sizes of SLPS and CFS are shown in Table 1.6. LSLPS, LCFS, CSLPS, and CCFS exhibited granule size ranges of 2.32
26.16, 2.60
30.66, 2.49
25.98, and 2.62
30.51 μm, respectively. The granule size ranges of SLPS were much narrower than those of CFS. In comparison with centrifugation, SLPS also produced a smaller average granule size (P , 0.05); LSLPS, LCFS, CSLPS, and CCFS had average granule sizes of 11.44, 13.15, 11.57, and 13.13 μm, respectively. 3.1.2.2 Scanning Electron Microscopy Both the CFS and SLPS granules were present in different shapes, including round polygonal, polygonal, spherical, oval, and round.

Sweet Potato Starch and its Series Products

27

Figure 1.4 SEM of sweet potato starches produced by sour liquid and centrifugation methods (LCFS, laboratory starch made from centrifugation; LSLPS, laboratory starch made from sour liquid processing; CCFS, commercial starch made from centrifugation; CSLPS, commercial starch made from sour liquid processing; 2000 3).

There was no significant difference in the overall morphologies of the starch granules obtained from the two methods. Smooth granule surfaces were observed in LCFS and CCFS, whereas LSLPS and CSLPS granules exhibited obvious fissures or holes on their surfaces (Fig. 1.4). Chinsamran et al. (2005) observed similar phenomena in cassava, sweet potato, and rice starches prepared by natural fermentation, and the pores were probably caused by the hydrolysis of -amylase in fermentation broth. In addition, Rocha et al. (2010) also found similar pore structures on the surface of sweet potato, cassava, and potato starch after -amylase hydrolysis. In conclusion, compared with CFS, the starch granules of SLPS were partially degraded during SLPS. 3.1.2.3 Polarizing Microscopy Under a polarizing microscope, the starch granules show black polarization crosses that divide the granules into four white areas. The polarization crosses are generated from spherulitic structures with double refraction, and the light passing through the crystal produces polarized light. Starch granules are observed and identified by polarizing microscopy. When the starch grains are fully expanded, crushed, or heat dried, the crystal structures disappear. Fig. 1.5 is the polarized

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Sweet Potato Processing Technology

Figure 1.5 Polarization crosses of sweet potato starches produced by sour liquid and centrifugation methods (LCFS, laboratory starch made from centrifugation; LSLPS, laboratory starch made from sour liquid processing, CCFS, commercial starch made from centrifugation; CSLPS, commercial starch made from sour liquid processing; 500 3).

microscopy of two kinds of sweet potato starches. Under the polarizing microscope, we observed that the LCFS, LSLPS, CCFS, and CSLPS had obvious polarization crosses, indicating that the spherulitic structure of sweet potato starch granules was not changed significantly during extraction by these two methods. In addition, some small particles failed to display the distinct polarization phenomenon, which might be due to the focal lengths of the small particles, whereas the large particles were inconsistent or overlapped. The cross-polarization properties of mung bean starches extracted by sour liquid and centrifugation methods showed no significant differences (Liu and Shen, 2007a). 3.1.2.4 Fourier Transform Infrared Spectrometry The differences in the Fourier transform infrared spectrometry fingerprints of SLPS and CFS are mainly distributed in 3400
3600 cm21. The relatively broad peak (3420 cm21) in 3400
3600 cm21 is caused by the O
H stretching vibration, which is related to the connections of hydrogen bonds between glucoside chains or intramolecular chains of starch molecules, including free hydroxyl groups and hydroxyl

Sweet Potato Starch and its Series Products

29

groups from the formed hydrogen bond. A wide peak reflects the complex stretching vibration between the hydroxyl groups of the starch molecules, whereas a sharp peak reflects the concertina movement of intramolecular hydroxyl and the free hydroxyl groups (Jiang et al., 2010). The peak of SLPS at 3420 cm21 peak is sharp, whereas that of the CFS is broader, suggesting that more intramolecular hydroxyl and free hydroxyl groups formed in SLPS, which produces weaker hydrogen-bonding interactions among the starch molecules and lower internal stabilities. The peaks at 1158 cm21 and 1081 cm21 reflect the C
O stretching vibrations of C
O
H in glucopyranoside rings, which decrease in the SLPS, indicating breaks in the ordered structure (Fang et al., 2002). In addition, the peak of LCFS is at 990 cm21, whereas that of LSLPS shifted to 992 cm21, which might be due to higher levels of intermolecular and intramolecular •OH that form in the starch produced by SLPS (Wang et al., 2010). 3.1.2.5 X-Ray Diffraction The X-ray diffraction (XRD) patterns of SLPS and CFS are presented in Fig. 1.6. They both have single broad peaks at 15 and 23 (2) and a dual peak at 17
18 (2), which indicate the typical A-type starches (Jiang et al., 2010), suggesting that the starch crystals remained unchanged during the extraction by SLPS. Liu and Shen (2007b) found that the crystals of mung bean starch also remain unchanged during extraction processes in the sour liquid and centrifugation methods. However, the XRD peak strength of the SLPS was weaker than that of the CFS, which suggests that the amylopectin structure is broken, decreasing the crystallization peak strength of the starch. The decrease in the crystalline peak of the SLPS might be attributed to the

Figure 1.6 The XRD spectra of sweet potato starches produced by the sour liquid and centrifugation methods (LCFS, laboratory starch made from centrifugation; LSLPS, laboratory starch made from sour liquid processing; CCFS, commercial starch made from centrifugation; CSLPS, commercial starch made from sour liquid processing).

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Sweet Potato Processing Technology

preferential hydrolysis of the crystalline structure during sour liquid processing. Chang et al. (2006) found that the crystallinity of mung bean starch extracted from lactic acid fermentation broth is lower than that of water and other extracts (distilled water, NaOH solution, and Na2SO3 solution). Rocha et al. (2010) reported that A-type starches are more susceptible to enzymatic attacks than B-type starches because the A and B1 short chains of the A-type are less stable in the crystalline structure and more susceptible to rearrangements. 3.1.2.6 Thermal Properties The thermal properties of SLPS and CFS are presented in Table 1.7. The Tp values of LSLPS and CSLPS are not significantly different from those of LCFS and CCFS, whereas the R and WH values of LSLPS and CSLPS are significantly lower than those of LCFS and CCFS (P , 0.05), respectively. The increase in the peak gelatinization temperature indicates that the amorphous regions of the starch granules are degraded during the SLPS (Sandhu et al., 2007). R is strongly influenced by the degree of amylopectin branching (the lower the degree of branching, the narrower the melting temperature range), which suggests that the degrees of LSLPS and CSLPS amylopectin branching are lower than those of the LCFS and CCFS amylopectins, respectively (Li et al., 2001). The decrease in the WH value indicates a disruption in the crystalline region and in the helical structures of the SLPS granules (Sandhu et al., 2007). These results are in accordance with the results of the XRD analysis. Similar results have been reported for cassava starch prepared by lactic acid fermentation (Numfor et al., 1995). When the starch was degraded by an enzyme, Table 1.7 DSC Analysis of Sweet Potato Starches Produced by Sour Liquid and Centrifugation Methods Samples

To ( C)

Tp ( C)

Tc ( C)

LCFS

72.02 6 0.00a

77.69 6 0.25b

83.23 6 0.02a

11.21 6 0.02a

7.46 6 0.07a

LSLPS

71.41 6 1.08

a

78.66 6 1.17

a

81.73 6 0.49

b

10.33 6 1.58

b

4.49 6 0.11b

CCFS

71.33 6 0.35

a

77.78 6 0.19

b

83.01 6 0.33

a

11.45 6 0.68

a

4.48 6 0.06a

CSLPS

72.83 6 0.54a

10.19 6 0.22b

2.67 6 0.08b

78.74 6 0.17a

R ( C)

82.78 6 0.33b

WH (J/g)

Different letters in the same column indicate significant differences (P , 0.05). LCFS, laboratory sweet potato starch made by centrifugation; LSLPS, laboratory sweet potato starch made by sour liquid processing; CCFS, commercial sweet potato starch made by centrifugation; CSLPS, commercial sweet potato starch made by sour liquid processing. To, Tp, Tc, WH, and R represent onset temperature, peak temperature, conclusion temperature, enthalpy, and gelatinization range (Tc
To), respectively.

Sweet Potato Starch and its Series Products

31

the enzyme could attack the amorphous regions mainly composed of amylose and crystalline regions (mainly composed of amylopectin), resulting in the degradation of amylose and amylopectin. This affected the DSC properties of the starches (Rocha et al., 2010; Jane et al., 1999). Zhou et al. (2004) also found a slight increase in the gelatinization temperature and a decrease in the enthalpy of the gelatinization of legume starches after an -amylase treatment.

3.1.3 Physicochemical Characteristics Analysis 3.1.3.1 Color Any adverse starch color affects the color of starch noodles and vermicelli, which reduces the quality of the products and thus affects the product’s consumer appeal (Galvez and Resurreccion, 1992). A high lightness value and a low chroma value are desired for starches. As shown in Table 1.8, the lightness of SLPS was noticeably higher than that of CFS. A similar result was found for mung bean starches prepared by SLPS and centrifugation (Liu and Shen, 2007a). Lactic acid or some other soluble and thermo-sensitive substances present in sour liquid may act as inhibitors of polyphenoloxidase activity to improve the starch color (Qin et al., 1997). In addition, lactic acid in sour liquid also can improve the color of starch (Rani et al., 1998). 3.1.3.2 Paste Transparency The transparency of starch paste is an important factor affecting food processing, which is highly correlated with retrogradation. If the retrogradation of starch paste was poor, the transparency was low. Table 1.8 Color of Sweet Potato Starches Produced by the Sour Liquid and Centrifugation Methods Samples

Color La

aa

ba

LCFS

92.43 6 0.18c

2 0.61 6 0.00c

3.83 6 0.057b

LSLPS

95.01 6 0.33

b

2 0.75 6 0.014

2.97 6 0.057c

CCFS

90.50 6 0.59

2 0.77 6 0.00

c

2.61 6 0.014c

CSLPS

92.93 6 0.03b

b c

2 0.91 6 0.007b

2.67 6 0.021b

Different letters in the same column indicate significant differences (P , 0.05). L , lightness, 100 for white and 0 for black; aa, red-green, positive values indicate red and negative values indicate green; bb, blue-yellow, positive values indicate yellow and negative values indicate blue; LCFS, laboratory sweet potato starch made by centrifugation; LSLPS, laboratory sweet potato starch made by sour liquid processing; CCFS, commercial sweet potato starch made by centrifugation; CSLPS, commercial sweet potato starch made by sour liquid processing. a

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Table 1.9 Light Transmittance of Sweet Potato Starches Produced by the Sour Liquid Processing and Centrifugation Methods (%) Samples/Concentration

0.1

0.5

1.0

29.3 6 0.02

2.0

Ba

10.3 6 0.78

Bb

8.3 6 0.34

LCFS

76.3 6 0.56

Aa

LSLPS

78.2 6 0.27

Aa

27.5 6 0.11

CCFS

79.3 6 0.78

Aa

27.8 6 0.34

CSLPS

79.4 6 0.68Aa

Ba

24.8 6 1.24Bb

Ca

Cb

10.3 6 0.21

Ca

9.4 6 0.72Ca

6.4 6 0.23Da 5.8 6 0.65Da 7.1 6 0.44Da 6.7 6 0.33Da

Note: Different capital letters, A
D, indicate significant differences in the same row (P , 0.05); different lowercase letters, a
d, indicate significant differences in the same column (P , 0.05). LCFS, laboratory sweet potato starch made by centrifugation; LSLPS, laboratory sweet potato starch made by sour liquid processing; CCFS, commercial sweet potato starch made by centrifugation; CSLPS, commercial sweet potato starch made by sour liquid processing.

Figure 1.7 The swelling power of sweet potato starch produced by the sour liquid processing and centrifugation methods (LCFS, laboratory starch made from centrifugation; LSLPS, laboratory starch made from sour liquid processing; CCFS, commercial starch made from centrifugation; CSLPS, commercial starch made from sour liquid processing).

The light transmittance is commonly used to indicate the level of starch paste transparency, indicating its ability to combine with water (Gao, 2008). As shown in Table 1.9, the transparency of sweet potato starch paste decreases with an increase in concentration. At a concentration of 0.1%, the transparency of all of the starches is close to 80%, with no significant differences. When the starch concentration rises to 0.5%
1.0%, the transparency of LSLPS is significantly lower than that of LCFS. The transparency of the CSLPS is also lower than that of CCFS, but the difference is not significant. At a concentration of 2%, none of the transparency levels of any of the starches is significantly different. 3.1.3.3 Swelling Power and Solubility The swelling powers and solubility patterns of SLPS and CFS are shown in Figs. 1.7 and 1.8, separately. The swelling power and

Sweet Potato Starch and its Series Products

33

Figure 1.8 Solubility of sweet potato starch produced by the sour liquid processing and centrifugation methods (LCFS, laboratory starch made from centrifugation; LSLPS, laboratory starch made from sour liquid processing; CCFS, commercial starch made from centrifugation; CSLPS, commercial starch made from sour liquid processing).

solubility increase with increasing temperature (55
95 C), showing two-stage swelling characteristics. Although the swelling powers of LSLPS and CSLPS are greater than those of LCFS and CCFS in the 55
95 C range, there are no significant differences at temperatures below 80 C (P . 0.05). LSLPS and CSLPS have significantly higher solubilities than LCFS and CCFS, respectively, in the temperature range of 65
95 C (P , 0.05). Tester and Morrison (1990) reported that the swelling power of starch primarily depends on the swelling properties of amylopectin, for which amylose acts as both a diluent and an inhibitor. Therefore, compared with CFS, the lower amylose content in starches produced by the sour liquid is likely to be the main reason for its greater swelling power. The increase in the swelling power of the SLPS makes the amylose more soluble. Thus, its solubility was also higher than that of CFS. 3.1.3.4 Pasting Properties The effects of SLPS and centrifugation on the pasting properties of the sweet potato starches are shown in Table 1.10. The pasting temperatures (Ptemp) of LSLPS and CSLPS are higher than those of LCFS and CCFS (P , 0.05), respectively. LSLPS and CSLPS exhibited noticeably lower peak viscosity (PV) levels compared to those of LCFS and CCFS (P , 0.05), respectively. The lower PV of SLPS may be due to the low pH induced by lactic acid or enzymatic action during the fermentation process. However, Numfor and Schwartz (1995) found that acidification of the native starch with citric acid has no effects on its PV between pH 6.9 and 4.5, suggesting that the differences between

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Sweet Potato Processing Technology

Table 1.10 Paste Viscosity Profiles (Using 6% Starch, Dry Basis) of Sweet Potato Starches Sample

LCFS

Ptemp ( C)

Tp (min)

HPV

CPV

BDV

Stability Ratio

Setback Ratio

78.2a

8.60a

178a

85b

138b

93a

0.48b

1.62a

a

a

b

a

a

b

73

0.57

a

1.52b

LSLPS

80.6

CCFS

81.0b

8.83a

a

a

CSLPS

Pasting Properties (RVU) PV

82.3

8.90

8.83

168

95

148

181a

89a

137a

92a

0.49b

1.54a

b

b

b

b

a

1.48b

159

86

127

73

0.54

Different letters in the same column indicate significant differences (P , 0.05). LCFS, laboratory sweet potato starch made by centrifugation; LSLPS, laboratory sweet potato starch made by sour liquid processing; CCFS, commercial sweet potato starch made by centrifugation; CSLPS, commercial sweet potato starch made by sour liquid processing. Ptemp, Tp, PV, HPV, CPV, stability ratio, and setback ratio represent peak temperature, peak time, peak viscosity, hot paste viscosity, cold paste viscosity, HPV/PV, and CPV/HPV, respectively.

SLPS and CFS may be mainly attributed to enzymatic action. Moorthya (2010) found that a slight decrease in PV, as well as a lower paste viscosity, for fermented cassava starch may be due to its greater solubility and the presence of a fairly high percentage of small granule sizes. Thus, compared to CFS, the higher solubility and smaller granule sizes of SLPS, caused by the enzymatic degradation of the starch granules, were likely to be the main reason for the lower PV. In addition, a decrease in PV was also observed in the starches from cassava, sweet potato, peruvian carrot, and potato hydrolyzed by bacterial α-amylase (Chinsamran et al., 2005). Rocha et al. (2010) also found the same phenomenon in sweet potato, cassava, and potato starch after -amylase hydrolysis. The stability ratio of the starches is defined as the ratio of hot PV (HPV) to PV (HPV/PV). The stability ratios of LSLPS and CSLPS are higher than those of LCFS and CCFS, respectively (Table 1.10), indicating a more stable paste viscosity during heating and shearing. The setback ratio, defined as the ratio of cold paste viscosity (CPV) to HPV (CPV/HPV), reflects the extent of starch retrogradation. The higher the setback value, the greater the tendency to starch retrogradation. LSLPS and CSLPS have lower setback ratios compared with LCFS and CCFS (P , 0.05, Table 1.10), indicating that SLPS exhibits a lower retrogradation extent than CFS. The lower setback ratio of SLPS might be related to its lower amylose content and the lower degree of amylose fraction polymerization in the starch (Putri et al., 2011; Zhang et al., 1999).

Sweet Potato Starch and its Series Products

35

3.1.3.5 Freeze
Thaw Stability (Retrogradation) Starch retrogradation is related to the syneresis of starch gel; therefore, syneresis can be used to measure the degree of starch retrogradation (Hoover et al., 1997). The retrogradation degree of sweet potato starch is determined by the amount of water after centrifuging the starch gel after seven freeze
thaw cycles. After freezing at 218 C, the starch gel becomes spongy, resulting in the rapid resorption of water after centrifugation, making the measurement more difficult. The syneresis of SLPS and CFS after a freeze
thaw treatment at 218 C is shown in Fig. 1.9. The syneresis of SLPS is greater than that of CFS. In general, the greater the syneresis, the greater the retrogradation of starch. The syneresis capabilities of SLPS at 4 C without a freeze
thaw treatment are shown in Fig. 1.10. The syneresis capability of starch produced by the sour liquid method is significantly lower than that of CFS. After comparing the results with and without a freeze
thaw treatment, it was found that they were not consistent. Hoover et al. (1997) observed that the large amount of water that precipitates during a freeze
thaw treatment is due to the intramolecular and intermolecular hydrogen bonding interactions (amylose
amylose, amylose
amylopection, and amylopection
amylopection). Yuan and Thompson (1998) found that the degree of syneresis increases as the freeze
thaw cycle number increases, because the retrogradation degree of starch amylose increased. In fact, none the starch in the frozen state underwent retrogradation; however, the amylopectin underwent

Figure 1.9 Syneresis of sweet potato starch produced by the sour liquid and centrifugation methods, followed by a 218 C freeze
thaw cycle (LCFS, laboratory starch made from centrifugation; LSLPS, laboratory starch made from sour liquid processing; CCFS, commercial starch made from centrifugation; LSLPS, commercial starch made from sour liquid processing).

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Sweet Potato Processing Technology

Figure 1.10 Syneresis of sweet potato starch produced by the sour liquid and centrifugation methods, and then stored at 4 C (LCFS, laboratory starch made from centrifugation; LSLPS, laboratory starch made from sour liquid processing; CCFS, commercial starch made from centrifugation; CSLPS, commercial starch made from sour liquid processing).

retrogradation only during the freeze
thaw treatment. The long chain ratio of amylopectin in starch produced by the sour liquid method is greater than that in starch produced by the centrifugation method, whereas the retrogradation rate of the long chain ratio is greater than that of the short chain ratio (Jane et al., 1999). The syneresis of the starch gel without a freeze
thaw treatment is mainly caused by amylose because the amylose is easily recrystallized at 4 C. These results are consistent with the retrogradation results obtained using a Brabender viscometer. In addition, syneresis stimulated by a freeze
thaw cycle may also reflect the interactions between amylose and amylopectin.

3.2 COMPARISON OF THE QUALITY OF SLPS AND CFS Starch noodles are a widely popular traditional food in China and the majority of Asian countries and regions, with a history of thousands of years (Zhao et al., 2009). Starch is the main constituent of starch noodles, and the properties of starch play important roles in the processing and quality of starch noodles. Owing to differences in the processing of the two kinds of sweet potato starches, differences in the physicochemical properties of starch may occur, affecting the quality of starch noodles. Here, the SLPS and CFS were selected to make starch noodles. The physicochemical characteristics of the starches and the quality of the

37

Sweet Potato Starch and its Series Products

starch noodles were compared and analyzed to provide a theoretical basis for the production of sweet potato starches and starch noodles.

3.2.1 Color of the Sweet Potato Starches and Noodles The colors of SLPS and CFS are shown in Table 1.11 and Fig. 1.11. The brightness scores of LCFS, CCFS, NLCFS, and NCCFS were significantly lower than those of LSLPS, CSLPS, NLSLPS, and CSLPS. The research suggests that any adverse starch color affects the color of starch noodles and vermicelli and thus reduces the quality levels of starch noodles and vermicelli products (Galvez and Resurreccion, 1992). The brightness of CFS is significantly lower than those of SLPS; therefore, the color of the noodles made from CFS is not as good.

3.2.2 Retrogradation of Starch and Noodles The syneresis of a starch gel is related to, and can be used to measure, the retrogradation of the starch and its noodles. The greater the amount of water after syneresis, the greater the rate and degree of retrogradation (Hoover et al., 1997). The retrogradation of starch and its noodles from SLPS and centrifugation is shown in Fig. 1.12. The syneresis of starch noodles from SLPS and CFS present linear upward trends for 24 h, increasing as the treatment time increases. After 4 h, Table 1.11 Colors of the Sweet Potato Starches and Noodles (Uncooked) Samples

Color L

a

LCFS

92.43 6 0.18

LSLPS

95.01 6 0.33

CCFS

b

b

2 0.61 6 0.00

b

2 0.75 6 0.01

3.83 6 0.06a

a

2.97 6 0.06b

90.50 6 0.59b

2 0.77 6 0.00b

2.61 6 0.01b

CSLPS

92.93 6 0.03

a

2 0.91 6 0.00

2.67 6 0.02a

NLCFS

66.12 6 0.03

b

2.79 6 0.03

NLSLPS

a

a

a

6.42 6 0.02a

68.72 6 0.13a

2.91 6 0.06a

6.51 6 0.05a

NCCFS

63.61 6 0.11

2.25 6 0.15

8.54 6 0.07a

NCSLPS

65.13 6 0.04

b a

b

2.42 6 0.03

a

8.13 6 0.07a

The different letters in the same column indicate significant differences (P , 0.05). LCFS, laboratory starch made from centrifugation; LSLPS, laboratory starch made from sour liquid processing; CCFS, commercial starch made from centrifugation; CSLPS, commercial starch made from sour liquid processing; NLCFS, starch noodles made with laboratory starch produced by centrifugation; NLSLPS, starch noodles made with laboratory starch produced by sour liquid processing; NCCFS, starch noodles made with commercial starch produced by centrifugation, NCSLPS, starch noodles made with commercial starch produced by sour liquid processing.

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Sweet Potato Processing Technology

Figure 1.11 The appearances of the uncooked and cooked noodles: (A) Starch noodles made with starch produced by centrifugation, uncooked; (B) starch noodles made with starch produced by centrifugation, cooked; (C) starch noodles made with starch produced by sour liquid processing, uncooked; and (D) starch noodles made with starch produced by sour liquid processing, cooked.

Figure 1.12 Syneresis of sweet potato starches noodles made using a centrifugal cyclone separator and sour liquid processing (0
24 h) (NLCFS, starch noodles made with laboratory starch produced by centrifugation; NLSLPS, starch noodles made with laboratory starch produced by sour liquid processing; NCCFS, starch noodles made with commercial starch produced by centrifugation; NCSLPS, starch noodles made with commercial starch produced by sour liquid processing).

Sweet Potato Starch and its Series Products

39

the syneresis of noodles made from SLPS and CFS were significantly higher than those of noodles made from starch produced by SLPS. At 24 h, the syneresis of the noodles is as follows: LCFS (67.43%) . LSLPS (63.65%) . CCFS (60.17%) . CSLPS (54.01%). The research shows that the retrogradation of starch is related to the quality of starch noodles. The faster the starch undergoes retrogradation, the more easily it forms a dense structure capable of forming starch noodles (Kasemsuwan et al., 1998; Tan, 2008). In addition, the sweet potato amylose easily undergoes retrogradation at 4 C, which leads to a high degree of recrystallization, and results in an increase in syneresis capability after starch gel formation (Tan et al., 2008). Therefore, the syneresis of noodles made from both processes might also change because of different amylose contents.

3.2.3 Cooking Quality The parameters reflecting the cooking quality of starch noodles are swelling index, cooking loss rate, and breakage rate. When boiling, the starch noodles absorb water, which the internal starch regelatinize. After heating, the starch noodles begin swelling, slowly becoming soft, elastic, smooth, and transparent. During this process, small debris and soluble components of starch noodles continuously enter the water due to swelling, constituting the “cooking loss” (Lee et al., 2006; Yang et al., 2009). The swelling index, cooking loss, and breakage rate of sweet potato starch noodles are shown in Table 1.12. There are no significant differences in the contents of dry matter, cooking loss rate, or breakage rate among the noodles made from SLPS and CFS. However, the swelling indices of NLCFS and NCCFS are much lower Table 1.12 Swelling Indices, Cooking Loss Rates, and Breakage Rates of Sweet Potato Starch Noodles Samples

Dry Matter Content (%)

Swelling Index (%)

Cooking Loss Rate (%)

Broken Rate (%)

NLCFS

87.44 6 0.89

a

616.74 6 17.83

0.24 6 0.09

a

10.00 6 0.00a

NLSLPS

87.41 6 0.34

a

687.37 6 12.75

0.33 6 0.08

a

7.50 6 3.53a

NCCFS

87.23 6 0.56a

666.75 6 24.59b

0.16 6 0.04a

7.50 6 3.53a

NCSLPS

87.78 6 0.21

714.36 6 16.99

0.18 6 0.06

10.00 6 0.00a

a

b a

a

a

The different letters in the same column indicate significant differences (P , 0.05). NLCFS, starch noodles made with laboratory starch produced by centrifugation; NLSLPS, starch noodles made with laboratory starch produced by sour liquid processing; NCCFS, starch noodles made with commercial starch produced by centrifugation; NCSLPS, starch noodles made with commercial starch produced by sour liquid processing.

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Sweet Potato Processing Technology

than those of NLSLPS and NCSLPS, which might be due to the lower swelling power of CFS.

3.2.4 Textural Properties The textural properties of starch noodles are the key indicators for measuring their quality. The greater the tensile strength and tensile deformation of starch noodles, the better the elasticity (Wang et al., 2010; Smewing, 1997), whereas the higher the shear stress, the better the chewiness (Tan et al., 2006). As shown in Table 1.13, the maximum breaking stresses of NLCFS and NCCFS are slightly less than those of NLSLPS and NCSLPS, whereas the tensile strength and stretch forming are much greater than those of NLSLPS and NCSLPS. The high shear stress may be related to the lower swelling index (Table 1.12). The lower the swelling index of starch noodles, the lower the water contents during boiling, and the greater the hardness.

3.2.5 Microstructure The SEM micrographs of noodles made using starch produced by both processes are shown in Fig. 1.13. When placed in the low magnification (50 or 70 3), the surfaces of the two kinds of noodles are smooth, whereas there were small holes in the cross sections. There were no obvious differences. In summary, the swelling power and solubility of CFS are significantly lower than those of the SLPS, whereas the retrogradation is Table 1.13 Textural Properties of Sweet Potato Starch Noodles Samples

Maximum Breaking Stress (g)

Tensile Strength (g/mm2)

Stretch Forming (%)

NLCFS

39.96 6 4.82

18.64 6 2.02

104.52 6 5.38a

NLSLPS

43.47 6 8.72a

17.11 6 0.44a

98.44 6 4.72b

NCCFS

30.22 6 6.49

a

15.73 6 3.07

a

97.05 6 2.83a

NCSLPS

32.14 6 5.14

a

14.23 6 5.54

a

94.13 6 4.55b

Samples

Maximum Shear Stress (g)

Shear Stress (g/mm2)

Shear Deformation (%)

NLCFS

58.94 6 3.69

a

28.60 6 3.47

a

20.38 6 2.50a

NLSLPS

55.45 6 5.68

a

25.38 6 1.59

b

19.47 6 2.06a

NCCFS

40.79 6 3.37a

17.14 6 0.64a

46.24 6 5.13a

NCSLPS

36.55 6 1.42

16.32 6 0.69

24.90 6 3.62b

a

a

a

b

The different letters in the same column indicate significant differences (P , 0.05). NLCFS, starch noodles made with laboratory starch produced by centrifugation; NLSLPS, starch noodles made with laboratory starch produced by sour liquid processing; NCCFS, starch noodles made with commercial starch produced by centrifugation; NCSLPS, starch noodles made with commercial starch produced by sour liquid processing.

Sweet Potato Starch and its Series Products

41

Figure 1.13 SEM of surfaces of starch noodles prepared using centrifugation-processed (NCFS) and sour liquidprocessed starch (NSLPS).

greater. Because the brightness score for CFS is lower than that of the starch produced by SLPS, the color and transparency of the noodles made with CFS are significantly lower than those of noodles made with sour liquid-processed starch after boiling. In addition, the swelling index of the former was significantly less than that of the latter, whereas the retrogradation, stretch forming, and shear stress were significantly greater. The differences among the properties of the two starches and the qualities of their noodles were related to their different amylose contents.

SECTION 4: APPLICATIONS OF SWEET POTATO STARCH In the 1950s and 1960s, sweet potato played an important role by resisting natural disasters and solving the food crisis. During this period, the planting sweet potato area reached its peak in China. With economic, scientific and technological advancements, people’s

42

Sweet Potato Processing Technology

standards of living have gradually improved, the food structure has changed greatly, and the popularity of sweet potato has gradually declined, accompanied by a decrease in the planting area. Since the beginning of the 21st century, with an adjustment in the industrial structure and the development of the processing industry in China, sweet potato is again becoming popular because of its special economic value. At present, the sweet potato in China is an important raw material for food- and feed-related industries, the chemical industry, and other light industries, which are gradually developing a comprehensive utilization and deep processing of the sweet potato. In addition, sweet potato starch is being utilized more and more widely. This section describes the applications of sweet potato starch.

4.1 APPLICATION OF SWEET POTATO STARCH IN FOOD At present, the application of sweet potato starch in food is mainly in the production of sweet potato starch noodles and vermicelli. We have introduced the detailed production processes of sweet potato starch noodles and vermicelli (Sections 1.5 and 2.2). We had learned that sweet potato starch noodle production mainly uses sweet potato starch as a raw material, but this cannot meet the demands of human nutrition and health. The special purple sweet potato is rich in anthocyanins, which have many physiological functions, such as anticarcinogen, antihypertension, and free radical scavenging. Thus, an array of purple sweet potato products has presently appeared. Many enterprises use purple sweet potato powder as an additive to make purple sweet potato starch noodles, which improves their nutritional value, but also increases the breakage rate, and other problems. These have attracted the attention of experts and related research is being carried out. We believe that these problems will be solved in the near future.

4.2 APPLICATIONS OF SWEET POTATO RESISTANT STARCH 4.2.1 Applications of Sweet Potato Esterified Starch Crosslinked esterified resistant starch can be used as an excellent thickening agent and stabilizer in food, replacing expensive food additives. Wu et al. (2009) used sodium trimetaphosphate as a crosslinking agent and acetic anhydride as an esterifying agent to modify the starch, forming crosslinked esterified resistant starch, and studied its applications in tomato sauce. The tomato sauce containing crosslinked

Sweet Potato Starch and its Series Products

43

esterified resistant starch was bright red in color, had good brightness, a delicate lubricated texture, a sweet and sour taste, and maintained the flavor, as well as good storage stability.

4.2.2 Application of Sweet Potato Acetylated Starch Zou et al. (1996) studied the rheological characteristics of sweet potato acetylated starch and its applications in food and found that sweet potato acetylated starch can be used as a thickening agent in juice, fruit sauces, and cold foods, and also as a sugar substitute foaming agent.

4.2.3 Application of Sweet Potato Starch-Based Superabsorbent Resin Wu et al. (2009) studied a cotton seed coating agent using a sweet potato starch-based superabsorbent resin, and found that the sweet potato starch superabsorbent resin significantly increases the cotton seed germination rate, seedling stem length, stem weight, leaf weight and chlorophyll content, and decreases the content of the membrane lipid oxidation product malondialdehyde. It also improves the activity of polyphenol oxidase, so as to effectively alleviate the plant drought stress because it is an excellent drought-resistant coating agent. Jiang et al. (2004) used sweet potato starch-based superabsorbent resin as a corn seed coating reagent, and the results showed that it could effectively improve the corn seed germination rate. Thus, sweet potato starch with its unique characteristics is widely used in industries related to food, medicine, chemicals, energy, and textiles, and the deep-processing products of sweet potato starch can play important roles in wider areas and industries.

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Zhou, Y., Hoover, R., Liu, Q., 2004. Relationship between α-amylase degradation and the structure and physicochemical properties of legume starches. Carbohydr. Polym. 57, 299
317. Zhu, F., Yang, X.S., Cai, Y.Z., et al., 2011. Physicochemical properties of sweet potato starch. Starch/Stärke 63, 249
259. Zhao, J.Y., Jia, D.Y., Yao, K., et al., 2009. The development of the sweet potato vermicelli without alum. Grain Oil Deep Proc. Food 5, 12
14 (in Chinese). Zou, X.X., Chen, L.Q., 1996. Study on the rheological properties and application of acetylation sweet potato starch. Nat. Sci. J. Xiangtan Univ. 18 (2), 58
61 (in Chinese).

FURTHER READING EURESTA, 1992. European Flair Concerted Action on Resistant Starch. Physiological implications of the consumption of resistant starch in man. Eur. J. Clin. Nutr. 46 (Suppl 2), 1
3. Zhu, Y.B., Shi, F.Y., Wu, Y.P., et al., 2012. Study on production of fuel ethanol by simultaneous fermentation of sweet potato starch. Jiangsu Agric. Sci. 40 (5), 246
249 (in Chinese).

CHAPTER

2

Sweet Potato Proteins

SECTION 1: OVERVIEW OF SWEET POTATO PROTEINS 1.1 The Sources and Structures of Sweet Potato Proteins 1.2 Research Status on the Technologies Used to Produce Sweet Potato Proteins 1.3 The Biological Activity of Sweet Potato Protein 1.4 Physicochemical Properties of Sweet Potato Protein SECTION 2: PRODUCTION TECHNOLOGIES OF SWEET POTATO PROTEIN 2.1 Effects of Solvent on the Extraction of Sweet Potato Protein 2.1.1 Types of Protein in Sweet Potato 2.1.2 Effects of Different Solvents on the Extraction Yield and Purity of Sweet Potato Protein 2.2 Salting-Out Method 2.2.1 Definition 2.2.2 Principle of the Salting-Out Method 2.2.3 Sweet Potato Protein Extraction Technology of the Salting out Method 2.2.4 Factors Influencing Protein Salting out 2.3 Isoelectric Precipitation Method 2.3.1 Definition 2.3.2 Principle of the Isoelectric Point Precipitation 2.3.3 Extraction of Sweet Potato Protein by the Isoelectric Point Method 2.3.4 Factors Influencing Isoelectric Point Precipitation 2.4 Foam Separation Method 2.4.1 Definition 2.4.2 Principle of foam Separation Sweet Potato Processing Technology. DOI: http://dx.doi.org/10.1016/B978-0-12-812871-8.00002-7 Copyright © 2017 China Science Publishing & Media Ltd. Published by Elsevier Inc. All rights reserved.

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2.4.3 Foam Separation Process of Sweet Potato Protein 2.4.4 Factors Influencing Foam Separation 2.5 Ultrafiltration Method 2.5.1 Definition 2.5.2 Principle of Ultrafiltration 2.5.3 Ultrafiltration of Sweet Potato Protein 2.5.4 Factors Influencing Ultrafiltration 2.6 Thermal Denaturation Method 2.6.1 Definition 2.6.2 Mechanism of Thermal Denaturation 2.6.3 Thermal Denaturation Production Process of Sweet Potato Protein 2.6.4 Factors Influencing Thermal Denaturation 2.7 Purification Method of Sweet Potato Protein 2.7.1 Ion-exchange Chromatography Method 2.7.2 Sephadex G-75 Gel Chromatography Method SECTION 3 BIOLOGICAL ACTIVITY OF SWEET POTATO PROTEIN 3.1 Antioxidant Activity 3.1.1 Superoxide Anion Radical Scavenging Activity of Sweet Potato Protein 3.1.2 Hydroxyl Radical Scavenging Activity of Sweet Potato Protein 3.1.3 DPPH Radical Scavenging Activity of Sweet Potato Protein 3.1.4 Linoleic Acid Peroxidation-Inhibiting Effect of Sweet Potato Protein 3.2 Trypsin Inhibitory Activity of Sweet Potato Protein 3.2.1 Determination of the Trypsin Inhibitor Activity of Sweet Potato 3.2.2 Factors Influencing Trypsin Inhibitory Activity of Sweet Potato 3.3 Anticancer Activity of Sweet Potato Protein 3.3.1 The Inhibitory Effects of Sweet Potato Protein on Cancer Cell Proliferation 3.3.2 Inhibitory Effects of Sweet Potato Protein on the In Vitro Migration and Invasion of Cancer Cells 3.3.3 Inhibitory Effects of Sweet Potato Protein on Tumor Metastasis

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3.4 Obesity Prevention and Weight Loss 3.4.1 Effects of Sweet Potato Protein on the Proliferation and Differentiation of 3T3-L1 Preadipocytes 3.4.2 Experimental Animal Study of Obesity Prevention 3.4.3 Animal Experimental Study on Losing Weight and Reducing Lipid Levels SECTION 4 FUNCTIONAL PROPERTIES OF SWEET POTATO PROTEIN 4.1 Solubility of Sweet Potato Protein 4.1.1 Effect of pH Value on the Solubility of Sweet Potato Protein 4.1.2 Effects of pH and NaCl on the Solubility of Sweet Potato Protein 4.1.3 Effects of Salt Species on the Solubility of Sweet Potato Protein 4.2 Emulsifying Properties of Sweet Potato Protein 4.2.1 Effects of the Protein Concentration and Oil Volume Fraction on the Emulsifying Properties of Sweet Potato Protein 4.2.2 Effects of pH on the Emulsifying Properties of Sweet Potato Protein 4.2.3 Effects of NaCl on the Emulsifying Properties of Sweet Potato Protein 4.2.4 Effects of High Hydrostatic Pressure Treatment on the Emulsifying Properties of Sweet Potato Protein 4.3 Gelling Properties of Sweet Potato Protein 4.4 Structural Properties of Sweet Potato Protein 4.4.1 Effects of Heat Treatment and pH on the Structure of Soluble Sweet Potato Protein 4.4.2 Effects of High Hydrostatic Pressure Treatments on Structure of Sweet Potato Protein 4.5 The Foaming Properties and Foam Stability of Sweet Potato Protein 4.6 Water-Holding and Oil-Holding Capacities of Sweet Potato Protein SECTION 5 APPLICATIONS OF SWEET POTATO PROTEIN 5.1 Edible Protein Powder 5.2 Emulsifier

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5.3 Humectant 5.4 Raw Material of Active Peptides 5.5 Biological Medicine References Further Reading

Abstract This chapter starts by introducing the production technologies of sweet potato protein, including the methods of salting out, isoelectric precipitation, foam separation, ultrafiltration, thermal denaturation, and purification. Then the biological activities of sweet potato protein and the effects of different treating time and concentration of protein are discussed, such as antioxidant activity, trypsin inhibitory activity, obesity prevention, and weight loss. Next, it is presented that sweet potato protein has good solubility, emulsifying, gelling, and foaming properties, which are affected significantly by pH and salt, and heat and high hydrostatic pressure treatments can influence the structural properties of sweet potato. Furthermore, the functional properties of sweet potato protein are introduced. By the end of the chapter, the applications of sweet potato protein are explained, suggesting that sweet potato protein can be used as edible protein, emulsifier, humectant, raw material of active peptides, and biological medicine.

SECTION 1: OVERVIEW OF SWEET POTATO PROTEINS In general, sweet potato roots contain 1.73%
9.14% (dry basis) proteins, in addition to rich starches and soluble sugars (Purcell et al., 1972), which can easily be digested and used by the human body. The vast majority of proteins in sweet potato are water-soluble, even though it has always been treated as waste water along with the processes of producing sweet potato starch, which corrupt and pollute rivers and lakes.

1.1 THE SOURCES AND STRUCTURES OF SWEET POTATO PROTEINS Jones and Gersdoff (1931) first separated the salt-soluble proteins from sweet potato roots and named “Ipomoein” in 1931. In 1985, Maeshima

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53

found a protein that mainly existed in sweet potato roots, accounting for 60%
80% of soluble protein, and named “Sporamin.” Then, Varon et al. (1989) confirmed that “Ipomoein” and “Sporamin” are proteins with molecular weights of 25 kDa. And the main protein in sweet potato roots is Sporamin. As a kind of storage protein, Sporamin is used for supplying nitrogen during the root germination stage as a nutrient source. Generally speaking, Sporamin is mainly distributed in tuberous roots, with the root vacuoles being the most abundant, accounting for B70% of the total protein (Hattori et al., 1988). The stem only contains small amounts of Sporamin, whereas the leaf and petiole basically have no Sporamin (Hattori et al., 1985; Chen et al., 1997). As a globular protein, the primary structure of Sporamin consists of B229 amino acid residues, where two groups of S
S bonds exist between the 45th and 94th cysteines (Cys45
Cys94), and the 153th
160th cysteines (Cys153
Cys160), respectively. In nonreducing conditions, there are three molecular isomers with different molecular weights linked by S
S bonds (Mu et al., 2005). Yao et al. (2001) simulated its structural model according to the trypsin inhibitory activity of Sporamin, as shown in Fig. 2.1. Sporamin is a unique multigene-encoding protein that can be divided into two subgroups, Sporamins A and B (Hattori et al., 1989). Based on the homologies of gene sequences, Sporamins A and B are determined to encode two kinds of sweet potato storage proteins, which can be completely separated by polyacrylamide gel electrophoresis (SDS
PAGE) under nonreducing conditions. The appropriate molecular weight of Sporamin A is 31 kDa, whereas that of Sporamin B is about 22 kDa. Moreover, the amino acid sequences, polypeptide patterns, structures, and immunological characteristics are similar between the two proteins (Maeshima et al., 1985), but still different. Sporamin A contains 219 amino acid residues, whereas Sporamin B is composed of 216 amino acid residues (Table 2.1), and there is twice as much Sporamin A in sweet potatoes as Sporamin B. The structures of sweet potato proteins in different pH ranges (pH 3 to 9) are relatively stable. After heating at 100 C for 10 min, sweet potato protein molecules formed a combination polymer having S
S bonds (Xue, 2006). The β-sheet and β-turn structures are rich in sweet potato proteins, and the secondary structure’s composition changes

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Figure 2.1 Three-dimensional structure model of Sporamin.

Table 2.1 Amino Acid Sequences of Sporamin Sporamin

Sequence

A

1 mkaftlalfl alslyllpnp ahsrfnpirl ptthepasse tpvldingde vraggnyymv 61 saiwgagggg lrlahldtms kcasdvivsp ndlddgdpit itpaaadpes tvvmaltyqt 121 frfniatnkl cvnnvnwgiq hdsasgqyfl kagefvsdns nqfkievvda nlnfykltyc 181 rfgsdkcynv grfhdpmlrt trlalsnspf vfvikptdv

B

1 mkalalffll slyllpnpah skfnpirlrp ahetassetp vldingdevr agenyyivsa 61 iwgagggglr lvrldsssne casdvivsrs dfdngdpiti tpadpestvv mpstfqtfrf 121 niatnklcvn nvnwgikhds esgqyfvkag efvsdnsnqf kievvndnln aykisycqfg 181 tekcfnvgry ydpltratrl alsntpfvfv ikptdm

Note: Data from the Biotechnology Information National Center of the USA.

with the pH values. The high hydrostatic pressure processing can also change the secondary structures, promote the formation of hydrogen bonds in proteins, which then affect the functional properties of sweet potato proteins (Mehmood, 2013).

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1.2 RESEARCH STATUS ON THE TECHNOLOGIES USED TO PRODUCE SWEET POTATO PROTEINS Although there is limited research on the nutrition and health characteristics of sweet potato proteins, studies on the extraction technologies used for sweet potato proteins are still less, especially those involved in the industrialized production of sweet potato proteins. In recent years, scholars had some success in improving the glycoprotein extraction technologies employed with sweet potato, such as using inorganic ceramic membranes to separate the sweet potato glycoprotein from starch production waste water, and the extract’s biological activity and structure were also investigated (Cheng et al., 2004, 2005). Furthermore, different reagent methods are used to extract sweet potato glycoprotein, such as chloroform, ethanol, and acetone (Li et al., 2001; Zhao et al., 2005; Meng et al., 2009; Liang et al., 2009). Sweet potato protein is a type of storage protein and has essential differences compared with glycoprotein. The combination of ultrafiltration and acid precipitation methods was adopted to prepare sweet potato protein, and a preliminary study determined the physical characteristics (Li et al., 2007; Mu et al., 2005). Since then, there have also been reports on the preparation of sweet potato protein using the combination of ultrafiltration and acid precipitation methods (Hou et al., 2010). However, there is limited research on sweet potato protein industrial production technology and related equipment, especially nutritional and functional data regarding sweet potato protein in the food industry. Our group has thoroughly investigated sweet potato starch processing enterprises in China on a different scale and systematically studied the preparation technologies of sweet potato protein, such as the ammonium sulfate precipitation, isoelectric point precipitation, ultrafiltration, combined acid precipitation and ultrafiltration, and thermal denaturation methods. The functional and health-related properties of sweet potato protein were also investigated in previous works (Mu et al., 2009a,b; Guo et al., 2011; Sun et al., 2012; Mehmood et al., 2013), which laid the foundation for industrialization, development, and application of sweet potato protein.

1.3 THE BIOLOGICAL ACTIVITY OF SWEET POTATO PROTEIN A comparison of the essential amino acid composition of sweet potato protein with those of other plant proteins (Table 2.2) indicates that sweet potato protein contains 18 different amino acids, 8 of which are

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Table 2.2 Essential Amino Acid Composition of Sweet Potato Protein and Other Plant Protein Amino Acid

Sporamin A

Sporamin B

Soybean

Peanut

Sesame

Thr

7.0

6.8

4.0

2.9

4.0

Val

8.5

9.1

5.0

5.2

4.7

Met

2.2

1.2

1.5

1.1

3.7

Ile

4.8

5.9

4.7

4.3

4.1

Leu

7.0

5.6

7.1

7.9

7.1

Phe

5.3

5.7

4.6

6.2

6.0

Lys

4.2

4.1

6.4

3.6

3.8

Trp

0.9

0.7

1.2

1.2

3.3

EAA (%)

39.9

39.1

34.6

32.4

36.7

human essential amino acids that account for 39% of the total amino acid content, which is significantly higher than in soybean, peanut, and sesame proteins. The biological value of sweet potato protein is 72, higher than that of potato (67), soybean (64), and peanut (59) proteins. Therefore, sweet potato protein is a unique high-quality plant protein with a high nutritional value. The amino acid composition of sweet potato was different among the varieties (Xue, 2006; Mu et al., 2009a). According to the analysis of the amino acid composition of the protein from “Xu 55-2,” lysine was the primary limiting amino acid. In addition to good nutritional properties, sweet potato protein also has beneficial biological activities, such as the ability to scavenge hydroxyl, DPPH, and superoxide anion free radicals (Xue, 2006). At the same time, as a natural trypsin inhibitor, sweet potato protein has good trypsin inhibitory activity and can effectively inhibit the proliferation, invasion, and metastasis of cancer cells (Deng, 2009). In addition, sweet potato protein can inhibit the proliferation of preadipocytes, prevent them from dividing into mature adipocytes, and reduce the number of fat cells. In obese mice, the protein can lead to weight loss, decrease the fat coefficient, and can significantly reduce the levels of total cholesterol and triglyceride. Thus, sweet potato protein has very good lipid-lowering effects and other health-related functions (Xiong, 2009).

1.4 PHYSICOCHEMICAL PROPERTIES OF SWEET POTATO PROTEIN Sweet potato protein has good physicochemical properties. Sweet potato protein has a high solubility under both acidic and alkaline

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conditions. It also has a good emulsifying activity, especially in an alkaline environment. The high concentration of protein can stabilize the emulsion, and the pH effects the emulsification of sweet potato protein. The emulsifying ability is lowest at the isoelectric point. Furthermore, the addition of salt can reduce the pH effect on the emulsifying activity and weaken the stability of the emulsion, whereas calcium salt can significantly improve the emulsifying activity and emulsion stability of sweet potato protein (Xue, 2006; Guo, 2010). As a new food-processing technology, high hydrostatic pressure can effectively improve the structural and emulsifying properties of sweet potato protein. In addition, sweet potato protein has good gelation properties, but these are significantly influenced by the extraction and preparation methods. The hardness, elasticity, and resilience of protein gels formed using the isoelectric point precipitation method are more effective than those prepared by ultrafiltration but have less dense protein gel net structures (Arogundade et al., 2012). In addition, sweet potato protein has a good foaming property, and the oil-holding capacity is superior to other plant protein resources but is different among the varieties.

SECTION 2: PRODUCTION TECHNOLOGIES OF SWEET POTATO PROTEIN Sweet potato protein is mainly a storage protein with 60%
80% being soluble protein. During sweet potato starch processing, sweet potato protein mainly exists in the cell fluid and is often discharged as waste water. Sweet potato protein has good nutritional, functional, and health care-related characteristics and is an important component of sweet potato. Therefore, it is very important to study the extraction and processing technologies of sweet potato protein. Based on the molecular weight, solubility, charge, and adsorption properties of protein molecules, the methods used to extract sweet potato protein are mainly the salting out, isoelectric point precipitation, foam separation, ultrafiltration, and thermal denaturation methods. There are also some combination methods, such as ultrafiltration and isoelectric point, or ultrafiltration combined with isoelectric point precipitation. Based on the purity requirement for research and other applications, sweet potato protein should be purified to a high purity using DEAE cellulose anion-exchange chromatography or Sephadex filtration.

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2.1 EFFECTS OF SOLVENT ON THE EXTRACTION OF SWEET POTATO PROTEIN 2.1.1 Types of Protein in Sweet Potato Based on the distribution coefficients of a protein in different solutions, such as water, salt solutions, and alcohol, proteins are often divided into water-soluble, salt-soluble, and alcohol-soluble proteins. Using successive and continuous water, NaCl solution (0.2 M) and ethanol solution (70%) extractions, water-soluble, and salt-soluble proteins were found to be the main components of sweet potato protein, accounting for 68.17% and 12.96% of the total content, whereas the alcohol-soluble part was only B2.20 %. Moreover, other proteins were not extracted by the solutions. Thus, B80% of sweet potato protein is soluble, and about B20% of sweet potato protein is insoluble in water and salt solutions (Li, 2007). The electrophoresis results, without the addition of β-mercaptoethanol, showed that water-soluble and salt-soluble proteins had consistent molecular weights at 22 and 31 kDa, respectively. However, the alcohol-soluble protein had an extra stained band (14 kDa). With the addition of β-mercaptoethanol, all three types of proteins showed the same single-stained band at 25 kDa, but the alcohol-soluble proteins extra 14 kDa band can still be detected. This indicates that disulfide bonds exist between the 22 and 31 kDa protein molecules, whereas there is no disulfide binding between the low molecular weight molecule and these two proteins (Fig. 2.2). Glycoprotein staining was performed after the electrophoresis test, and the proteins extracted by these three solvents were not glycoprotein (Li, 2007).

2.1.2 Effects of Different Solvents on the Extraction Yield and Purity of Sweet Potato Protein Comparing the effect of different solvents, such as water, alkaline solution (0.1-M NaOH), and solutions with different concentrations of NaCl have effects on the extraction yield and purity of sweet potato protein. The research data showed that an alkaline environment can increase the extraction yield of sweet potato protein, whereas the existence of salt ions was not conducive to protein dissolution. Furthermore, the most pure protein was extracted using water as the solvent, followed by extraction using an NaCl solution. Sweet potato

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Figure 2.2 SDS
PAGE of sweet potato protein. Note: Lane 1, standard marker; Lane 2, water-soluble protein; Lane 3, salt-soluble protein; Lane 4, alcohol-soluble protein; Lane 5, alkali-soluble protein. (A) without β-mercaptoethanol; (B) with β-mercaptoethanol.

Table 2.3 Effects of Different Solvents on the Extraction Yield and Purity of Sweet Potato Protein Solvent

Water

NaCl

NaOH

0.1 M

0.2 M

0.5 M

1.0 M

Purity (%)

63.40 6 0.58

54.57 6 0.54

55.21 6 0.30

56.15 6 0.48

53.60 6 0.95

33.22 6 0.65

Extraction yield (%)

23.72 6 0.62

19.74 6 0.39

19.73 6 0.21

17.55 6 0.93

21.27 6 0.61

40.53 6 0.31

Note: All of the data are means 6 standard deviations (n 5 3).

protein was mainly dissolved in water and salt solutions. The purity and yield of sweet potato protein extracted from solutions of different NaCl concentrations were similar (Table 2.3). The extracted protein yield was the highest (40.53%) when NaOH solutions were used as solvents, but the protein powder purity was the lowest (33.22%) (Table 2.3). Electrophoresis without the addition of β-mercaptoethanol indicated that the 22-kDa band was observed, as were some extra polymers (MW . 66 kDa). In addition, the 31 kDa band had completely disappeared. With the addition of β-mercaptoethanol, only a 25-kDa band was stained after an alkali treatment, which indicated that the produced polymers were formed by disulfide bonds (Fig. 2.2), and that the 31-kDa fragment was involved in the formation of polymers (Li, 2007).

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2.2 SALTING-OUT METHOD 2.2.1 Definition Salting out is defined as the process of adding inorganic salts to solutions to precipitate dissolved substances. For example, ammonium sulfate is commonly used to aggregate and extract crude protein.

2.2.2 Principle of the Salting-Out Method The solubility of a protein in an aqueous solution is determined by the degree of the hydration between water molecules and hydrophilic group, and the electric charges of the protein molecules. The affinity of the neutral salts to water molecules is stronger than that of the proteins, which causes the weakening or disappearance of the hydrating membrane. With the addition of neutral salts, the surface charge of the protein is neutralized because of the increase in the ionic strength, which easily reduces the solubility of the protein and promotes the aggregation and precipitation of the protein molecules.

2.2.3 Sweet Potato Protein Extraction Technology of the Salting out Method The salting-out method mainly uses ammonium sulfate to extract sweet potato protein. The main operation steps begun with freshly peeled sweet potatoes being ground in a 1% citric acid solution. The slurry was then sieved and centrifuged to remove starch and other insoluble substances. Then, the lean supernatant was cooled to 4 C, and ammonium sulfate was added to a saturating concentration. After that, the slurry was kept statically to allow sweet potato protein to precipitate. Then, the protein fraction was obtained from sweet potato protein by centrifugation. After an ammonium sulfate treatment, protein precipitation, containing large amounts of ammonium sulfate, occurred. Thus, it was necessary to dissolve the precipitate in distilled water and remove the salt by dialysis or ultrafiltration. This ultimately resulted in producing sweet potato protein powder by freeze drying. The advantages of this method are that limited equipment is required, and there is a strong ability of controlling and producing protein with high yields. However, this method consumes a large amount of salt, such as ammonium sulfate. The specific process and parameters are as follows: Sweet potato is cut into small pieces (2 cm3) - color protection (1:2 ratio of sweet potato solid to 1% citric acid solution) - slurry - sieve with cheese cloth (0.15 mm pore size) - centrifugation (10,000 3 g,

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30 min) - supernatant - addition of saturated ammonium sulfate centrifugation (5000 3 g, 30 min) - dissolve precipitate in distilled water - dialysis - freeze dry - sweet potato protein powder. The method of determining the ammonium sulfate concentration required for precipitating the protein was as follows: Small amount of sweet potato protein liquid was cooled to 0
5 C, and then solid ammonium sulfate powder was mixed in until there was no precipitate in the supernatant. To produce a precipitation curve, we determined the protein concentration when the precipitate was first generated. If a satisfactory yield has been achieved, then the saturation range can be narrowed to increase the purity.

2.2.4 Factors Influencing Protein Salting out 2.2.4.1 Protein Concentration The protein concentration can limit protein precipitation but can promote aggregation. The higher the protein concentration, the lower the salt saturation limit needed. Although a higher concentration has positive effects on precipitation, it also causes the coprecipitation of other components, which affects the purity. Thus, it is necessary to select the appropriate concentration to avoid contaminating coprecipitates. 2.2.4.2 Salt Saturation During the process of salting out, different proteins have different salt saturation or ionic strength requirements. When separating mixed proteins, the salt saturation level is usually increased gradually from low to high. After centrifugation or filtration, a high salt concentration is added to the protein again until other proteins have been completely precipitated. The composition of sweet potato protein is relatively simple, so 60% saturation with ammonium sulfate is enough for sweet potato protein precipitation. 2.2.4.3 Type of Salt The most effective salt is the anionic with multiple charges. The ions commonly used for protein salting are ammonium sulfate, magnesium sulfate, sodium sulfate, sodium chloride, and sodium phosphate. Ammonium sulfate is the most widely used salt because of its low temperature coefficient and high solubility (saturated solubilities are 754 and 706 g/L at 20 and 0 C, respectively). Within the solubility range, most of the proteins and enzymes can be salted out. In addition, the

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piecewise salting-out effect of ammonium sulfate is better than that of other salts, which have difficulty causing protein denaturation. 2.2.4.4 pH Value In general, the greater the net charge of the protein, the higher the solubility. Changing pH value can change the charge nature of the protein and thus change the solubility of the protein. The solubility of the protein is lowest at the isoelectric point, and the protein is easily precipitated. Thus, except in some special cases, the pH value is often selected in the vicinity of the protein’s isoelectric point. Ammonium sulfate is a weak acid, and the pH value of its saturated solution is less than 7. To improve the effects of salting out, hydrochloride is widely used to adjust the pH value of the protein solution to 4.0. 2.2.4.5 Temperature Because of the protective effect of salt concentration, the temperature requirement is not strict, and the salting out operation can be performed at room temperature. However, temperature obviously affects salt crystallization under neutral conditions. For example, serum protein is more readily precipitated at 25 C than at 0 C, whereas sweet potato protein is sensitive to temperature and is salted out at a low temperature. To obtain and separate more proteins easily, protein solutions should be stored for more than 3 h, or overnight, at 4 C after precipitation.

2.3 ISOELECTRIC PRECIPITATION METHOD 2.3.1 Definition The isoelectric point precipitation method commonly depends on the protein having its lowest solubility level at the isoelectric point and various proteins having different isoelectric points.

2.3.2 Principle of the Isoelectric Point Precipitation Protein molecules show zwitterionic characteristics at the isoelectric point, at which the net molecular charge is zero (positive and negative charges are equal). Because of the lack of repulsive activity, the molecular force between proteins is weakened, and they are easily gathered for precipitation. Thus, the protein has its minimum solubility at its isoelectric point and easily forms precipitates. Many physical properties, such as viscosity, swelling, and osmotic pressure, are also low at the isoelectric point, which is beneficial to the precipitation of suspensions.

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2.3.3 Extraction of Sweet Potato Protein by the Isoelectric Point Method The isoelectric point method mainly uses hydrochloric acid (HCl) or citric acid to adjust the pH to near the isoelectric point to precipitate and then separate sweet potato protein. The main steps are as follows: Freshly peeled sweet potatoes are ground after color protection processing. Then, the homogenate is filtered through four layers of cheese cloth and centrifuged (10,000 3 g, 20 min) to remove starch and other insoluble substances. The isoelectric protein isolate is obtained by adjusting the pH of the clear supernatant obtained at pH 4. This suspension is allowed to stand at room temperature for 1 h, and then, the supernatant is carefully decanted and the remaining slurry centrifuged (5000 3 g, 15 min). The resulting precipitate is reconstituted in distilled deionized water twice at a 1:3 protein-to-solvent ratio and centrifuged after readjusting to pH 4 with an HCl solution. The precipitated protein is reconstituted in distilled deionized water at a 1:3 solid-to-solvent ratio, and the pH is adjusted to pH 7 with an NaOH solution. The protein is lyophilized and stored as sweet potato protein powder (Arogundade et al., 2012). This method is simple, requires less equipment, and produces a high protein yield, but it consumes large amounts of acid and alkali. The specific process and parameters are as follows: Sweet potato is cut into small pieces (2 cm3) - color protection (1% sodium bisulfite solution at 1:2 solid-to-solvent ratio) - grind sieve with cheese cloth - centrifugation (10,000 3 g, 30 min) - supernatant - adjust pH to 4.0 with 2-M HCl and stand 1 h - centrifugation and sedimentation (5000 3 g, 15 min) - dissolve precipitate (1:3 solid-to-solvent ratio) - centrifugation and sedimentation (5000 3 g, 15 min) - dissolve precipitate (1:2 solid-to-solvent ratio; adjust pH to 7.0 with 2-M NaOH) - freeze dry - sweet potato protein powder.

2.3.4 Factors Influencing Isoelectric Point Precipitation There are many factors that affect the precipitation of proteins. 2.3.4.1 Type of Acidity Regulator For the isoelectric point precipitation method, acidity regulators, including HCl, sulfuric acid, acetic acid, phosphoric acid, citric acid, and lactic acid, are commonly used to maintain a pH level. In addition, fermentation can produce some acidic components to reduce the pH value of the solution and adjust it to near the protein isoelectric point. However, this method sometimes leads to partial proteolysis or

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degeneration. The effects of different acidity regulators on protein precipitation are different. HCl is commonly used to regulate the pH value during the extraction of sweet potato protein. 2.3.4.2 Types of Proteins Different protein types have different isoelectric points. If the isoelectric points of the different proteins were similar, then the proteins would be precipitated at the same time, which can seriously influence the sedimentation effects of the proteins during precipitation. 2.3.4.3 Metal Ions The isoelectric point of a protein will shift if the protein combined with metal ions. For example, the isoelectric point of insulin is 5.3, but it increases to 6.2 when combined with Zn21. Thus, the pH value may require adjustment when the isoelectric point is chosen after the addition of metal ions to the solution.

2.4 FOAM SEPARATION METHOD 2.4.1 Definition The foam separation method is based on the principle of surface adsorption, using foam bubbles as raw carriers to concentrate and separate the surface active materials and particles from the liquid phase. The method is also known as adsorptive bubble separation.

2.4.2 Principle of foam Separation Foam separation involves inflating bubbles in the solution, with the surfactant gathering at the gas
liquid interface (the surface of the bubble). A foam layer forms at the top of the solution. Separating the foam layer from the bulk liquid phase concentrates the surfactant (in the foam layer) and purifies the bulk liquid phase at the same time. The characterization parameters of foam separation are the enrichment ratio and the recovery rate. The formulae are as follows: Enrichment ratio 5 Recovery rate 5

Cfoamate Cinitial

Cfoamate 3 Vfoamate 3 100 % Cinitial 3 Vinitial

(2.1) (2.2)

where Cinitial is the protein concentration of the sample solution (mg/ mL or g/L), Cfoamate is the protein concentration of the foam liquid (mg/

Sweet Potato Proteins

65

Figure 2.3 Schematic drawing of the foaming process.

mL or g/L), Vfoamate is the initial volume of the sample solution (mL or L), and Vfoamate is the volume of the foam (mL or L) (Liu, 2013). 1. 2. 3. 4. 5. 6. 7. 8.

Air compressor Gas rotor flow meter Three-way valve Gas distributor Sample outlet Sample inlet Collecting pipe Foam collector

At present, various proteins and enzymes can be isolated by foam fractionation, including sweet potato protein, rhamnolipid, lysozyme, amylase, soybean protein, serum protein, and pepsin (Tan et al., 2005; Fu et al., 2004) successfully isolated soluble sugars and proteins from sweet potato roots using the centrifugation method, and the contents were 3.8% and 34.8%, respectively. However, using the foam fractionation method, 98.8% sugar and 74.1% protein were obtained.

2.4.3 Foam Separation Process of Sweet Potato Protein Foam separation of proteins is mainly based on the certain surface activity of protein, and the protein can be adsorbed at the gas
liquid interface. Sweet potato protein has a good surface activity, which is suitable for protein extraction using the foam separation method. The main process steps are as follows: Sweet potato is ground after color protection treatment, the homogenate is filtrated using cheese cloth to remove the fiber composition and large particles and then centrifuged at 10,000 3 g for 30 min. The supernatant is treated with a foam separation device (Fig. 2.3), and the density of the foam liquid increases as

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the concentration of the separation solution increases. Foam liquid is eventually freeze dried. The specific process and parameters are as follows: Sweet potatoes cut into small pieces (2 cm3) - color protection (1% sodium bisulfite solution at 1:2 solid-to-solvent ratio) - grind sieve with cheese cloth - centrifugation (10,000 3 g, 30 min) - supernatant - foam separation device - collect foam - freeze dry sweet potato protein powder. Foam separation has many advantages, including the simple structure of the device, low-energy consumption, small cost, and convenient operation and maintenance. In addition, foam separation towers can be transformed into temperature controlling devices, which can be operated at normal or low temperatures. At present, the method is mainly used for the separation of cells, proteins, enzymes, and saponins, which can obtain a high enrichment ratio. Liu (2013) optimized the separation conditions of sweet potato protein from the starch waste water. They include foam-to-liquid ratio 1:4, pH 4, air-flow 0.1 L/min, bubbling time 100 min, fluid volume 500 mL, and an inclined angle of 30 , to have a recovery rate of 84.1%. Subsequently, the foam protein was concentrated by ultrafiltration, then separated and purified using an anion exchange column DEAE-52 and Sephadex G-75 gel chromatography column, respectively.

2.4.4 Factors Influencing Foam Separation 2.4.4.1 Concentration The influent protein concentration is an important factor that affects the separation efficiency of protein solution foam. As Fig. 2.4 shows, with an increase in the influent protein concentration, the recovery rate first increased and then decreased, whereas the enrichment ratio decreased gradually. The recovery rate was 54% when the influent protein concentration was 6.20 mg/mL. However, when the influent protein concentration was below 4.51 mg/mL, the recovery percentage was 67%, and it decreased with the decreasing influent protein concentration. If a higher enrichment ratio is desired, then the protein concentration of the solution can be reduced. If a high recovery percentage was needed, then the protein concentration of the solution can be increased.

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Recovery percentage

4.5 4 3.5 3 2.5 2 1.5 1 0.5 0

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Influent protein concentration (mg/mL)

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1

2

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Figure 2.4 Effects of the influent protein concentration on the recovery rate and enrichment ratio.

0

pH Figure 2.5 Effects of the pH on the recovery percentage and enrichment ratio.

2.4.4.2 pH Value The pH value of the solution is also an important factor that affects the separation efficiency of the protein-containing foam. With an increase in the pH, the recovery percentage initially increased and then decreased. The highest recovery rate was achieved at pH 4.0, whereas the enrichment ratio was relatively flat (Fig. 2.5). 2.4.4.3 Influent Volume With an increase in the influent volume, the recovery percentage showed a trend of slowly increasing and then decreasing. When the influent volume was less than 500 mL, the recovery percentage slowly

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2

100 90

1.8

80 Recovery rate (%)

60

1.4

50 1.2

40 30 20

Recovery percentage

1

Enrichment ratio

0.8

10 0

Enrichment ratio

1.6

70

0.6 200

350

500

650

800

Influent volume (%) Figure 2.6 Effect of influent volume on the recovery percentage and enrichment ratio.

increased as the influent volume increased (Fig. 2.6). When the influent volume reached 500 mL, the recovery percentage was at its maximum, and it sharply declined as the influent volume increased. This may be because the enrichment ratio was decreasing as the influent volume was increasing. In addition, the slope was smaller when the influent volume was less than 500 mL, but the largest slope rate was obtained in the 500
600 mL range. At greater than 600 mL, the slope tended to be stable. 2.4.4.4 Foaming Time With an increase in the foaming time, the enrichment ratio initially increased and then decreased. The recovery percentage showed the same trend. Because the foaming time is short, sweet potato protein gathered incompletely and a high protein concentration remained at the bottom of the tower. However, if the foaming occurred for a longer time, the protein concentration could not form the foam tower, which decreased the recovery percentage and the enrichment ratio. As shown in Fig. 2.7, the recovery percentage and enrichment ratio reached their maximum values at 100 min. 2.4.4.5 Air-Flow Rate With an increase in the air-flow rate, the recovery percentage changed slowly, and the enrichment ratio initially increased and then decreased. At lower flow rates, the air cannot effectively move the protein from

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1.6

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1.4

Recovery rate (%)

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50 40

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30

Enrichment ratio

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Recovery percentage

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0 40

60

80

100

120

Foaming time (%) Figure 2.7 Effect of foaming time on recovery percentage and enrichment ratio.

1.2

90

1

70 0.8

60 50

0.6

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Recovery percentage

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Enrichment ratio

Enrichment ratio

Recovery rate (%)

80

0.4 0.2

20 0.1

0.15

0.2

0.25

0.3

Air flow rate (L/mL) Figure 2.8 Effects of air-flow time on recovery percentage and enrichment ratio.

the bottom solution of the tower to the foam layer. With a high flow rate, the recovery percentage of sweet potato protein initially decreased slightly then tended to be stable, which may be because the protein in solution was recovered to the foam liquid. The reflux vortex brings the foam surface back to the solution, resulting in a reduction in the volume of the foam liquid, and the enrichment ratio was initially increased and then decreased (Fig. 2.8). 2.4.4.6 Inclined Angle of the Foam Separation Tower With an increase in the inclined angle, both the recovery percentage and enrichment ratio showed decreasing trends. As shown in Fig. 2.9, at an inclined angle of 30 , the recovery percentage and enrichment ratio reached their maximums of 84.07% and 1.29, respectively. There

90

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1.3

80

1.2

75

1.1

70

Recovery percentage

1

65

Enrichment ratio

0.9

60

30

40

50 60 Inclined angle (°)

70

Enrichment ratio

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Recovery rate (%)

70

0.8

Figure 2.9 Effects of inclined angle on recovery percentage and enrichment ratio.

was a large amount of liquid in the gaps of the foam layers in the vertical foam separation tower, making it difficult for the liquid to flow back into the solution. In contrast, for the inclined foam separation tower, the liquid in the foam layer gaps easily flowed back to the bottom of the tower. For the recovery percentage, the increase in the inclined angle promoted the hydraulic pressure of the foam layer, reducing the volume of the foam liquid. Therefore, the recovery percentage of the protein gradually decreased as the inclined angle increased. When the inclination angle was 70 , despite the increase in gaps between the foam, the liquid easily flowed back to the solution, leading to a reduction in the foam liquid’s volume. Thus, 30 was selected as the optimum inclined column angle.

2.5 ULTRAFILTRATION METHOD 2.5.1 Definition Ultrafiltration is a membrane separation technology. Under a certain pressure, the solute containing smaller molecules and the solvent can pass through a membrane having certain size apertures, whereas the larger molecules cannot. These are left on the side of the film, resulting in the partial purification of the larger molecular material.

2.5.2 Principle of Ultrafiltration Ultrafiltration is a membrane separation process using a pressureactivated membrane to retain the colloids, particles, and components with high molecular weight under an outer pressure, whereas the small

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Figure 2.10 Schematic drawing of ultrafiltration. Note: 1—feed trough; 2—pump; 3—filter room; 4—liquid outlet; 5—constant pressure valve; 6—concentrate outlet; 7—concentrate return pipe.

solute particles and water pass through (Fig. 2.10). The microporous membrane filter’s surface can retain a sample with a molecular weight of more than 10 kDa, and the pore diameter of ultrafiltration membrane was 0.001
0.02 μm.

2.5.3 Ultrafiltration of Sweet Potato Protein Sweet potato protein is composed of Sporamins A and B, which have molecular weights of 22 and 31 kDa. Therefore, because of the difference between sweet potato protein molecular weight and those of other components, low molecular weight components can be removed by ultrafiltration. The main process steps are as follows: Sweet potatoes are ground after the color protection process, crude fiber, and large particles are first separated out, and then high-speed centrifugation is used to remove the starch and other insoluble components and some of the large molecules. The supernatant is passed through the ultrafiltration column to extract and concentrate sweet potato protein. During the ultrafiltration process, the effective pores are easily blocked by sweet potato protein. Therefore, it is necessary to constantly add distilled water to discharge the low molecular weight components. Specific steps are as follows: Sweet potatoes cut into small pieces (2 cm3) - color protection (1% sodium bisulfite solution at 1:2 solid-to-solvent ratio) - grind sieve with cheese cloth - centrifugation (10,000 3 g, 30 min) -

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supernatant - ultrafiltration (intercepted molecular weight is 10 kDa) - freeze dry - sweet potato protein powder. The concentration ratios and ultrafiltration times directly affect the purity of sweet potato protein powder. Li et al. (2007) found that the concentration ratio has a positive linear correlation with the purity of the protein powder produced by ultrafiltration (correlation coefficient is 0.95, P , 0.01). When the concentration ratio increases, the purity of the protein powder also gradually increases (Li et al., 2007), but the recovery rate decreases after ultrafiltration treatment. The effects of different ultrafiltration times on the extracted sweet potato protein’s yield and purity were studied by Arogundade and Mu (2012). When ultrafiltration was performed four, six, and seven times, the corresponding extraction yields were 41.3%, 41.3%, and 51.3%, and the purity levels of the protein were 76.0%, 82.0%, and 82.1%, respectively. Extracting proteins by ultrafiltration has the advantages of simple operation, no chemical reagents, and ultrafiltration technology with mild experimental conditions compared with evaporation and freeze drying. There is no change in the phase, temperature or pH, and this prevents the degradation of biomacromolecules, loss of activity, and autolysis. Ultrafiltration is mainly used for desalination, dehydration, and the concentrating of biological macromolecules, but it requires a good deal of equipment and is time-consuming when used in protein production.

2.5.4 Factors Influencing Ultrafiltration Many factors influence the ultrafiltration of the permeation flux and reduce its efficiency. The main factors are as follows. 2.5.4.1 Liquid Flow Rate Increasing the flow rate of the feed liquid is beneficial to reduce the polarization and increase the flux, but it requires an increase in the pressure of the feed liquid and in energy consumption. In general, the flow rate is controlled at 1
3 m/s. 2.5.4.2 Operating Pressure The relationship between ultrafiltration flux and operation pressure depends on the properties of the membrane and gel layer, and the operating pressure of ultrafiltration should be near the limit flux. In

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the ultrafiltration process, the small molecules in the liquid can pass through the membrane only under a certain pressure. If the working pressure is too low to suit the normal operation, then the production of the filtrate would be reduced. On the contrary, a high working pressure increases the polarization layer and offsets a turbocharged growth effect. Thus, the precipitate will be firmly attached to the membrane and difficult to remove. In addition, the membrane pores will be quickly blocked, then lessened the efficiency of ultrafiltration. Each kind of ultrafiltration membrane has a pressure range in which the operation should be performed. 2.5.4.3 Temperature The operating temperature is mainly determined by the physical and chemical properties of the material. A high temperature can reduce the viscosity of the material’s liquid phase, increase the mass-transfer efficiency, and improve the flux. Thus, ultrafiltration should be performed at the maximum allowable temperature. The intermolecular forces can be offset, and the viscosity will be reduced by increasing the temperature. At the same time, it also affects the working performance of the membrane and increases the permeability. However, a high temperature shortens the life of the ultrafiltration membrane. 2.5.4.4 Operational Cycle Time During ultrafiltration, a gel layer is gradually formed on the surface of the membrane, which could lead to a decrease of the permeate flux. When the flux is at its minimum value, it requires washing. The period of time to reach this point is called the operational cycle time. Changes in the operational cycle time are closely related to the cleaning conditions. 2.5.4.5 Influent Protein Concentration During ultrafiltration, both the concentration and viscosity of the liquid gradually increase. In addition, the thickness of the gel layer also increases; therefore, the maximum allowable concentration of the liquid should be determined. The feed liquid concentration directly affects the filtration rate, and the flux of ultrafiltration has a linear relationship with the logarithm of the concentration. Generally speaking, the viscosity of the liquid will increase as the concentration increases, and the time required to form the polarization layer will be shortened, reducing the rate and efficiency of ultrafiltration. Therefore, it is necessary to control the concentration of the feed liquid during ultrafiltration.

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2.5.4.6 Pretreatment of Feed Liquid The pretreatment of the material liquid is required to improve the permeate flux of the membrane and ensure the stable operation of the ultrafiltration membrane. The pretreatment directly affects the degree of the membrane pollution, the production capacity of the system, and the service life of ultrafiltration membrane. Pretreatments are commonly performed using high-speed centrifugation, filtration, pH adjustments, heat treatment, cold storage, or a combination of methods. As a newly developed method, the flocculent method can effectively remove some organic molecules, such as tannin, pigment, pectin, and other unstable components, from solutions. 2.5.4.7 Membrane Cleaning Membranes should be washed regularly to maintain a certain level of transmission. This can also prolong the service life of the membrane. Generally, under the allowable pressure and pH values, the temperature is lower than 60 C, and the ultrafiltration membrane can be used for 12
18 months. Poor cleaning shortens the service life of the membrane. 2.5.4.8 Pore size of Ultrafiltration Membranes The membrane’s pore size should be consistent with the size of the target component in the sample solution. Large pore sizes result in poor separation and affect the filtration rate and stability. However, if the pore size is too small, the effective component’s permeability rate is low, resulting in a great loss. Ultrafiltration membranes are porous and have an asymmetric structure compared with microfiltration membranes. These asymmetric membranes include a very thin cortex (generally less than l μm) and a multihole sublayer. Thus, the ultrafiltration membrane is mainly characterized by the thickness of the cortex, pore size distribution, and surface porosity. The typical pore sizes of ultrafiltration membranes are in the 2
100-nm range. Generally, using a single method, such as salting out, isoelectric point precipitation, foam, or ultrafiltration, the purity of the extracted protein is not high. There are macromolecular components, such as pectin, soluble starch, and some sugars, which cannot be removed by a single treatment. This affects the purity of the proteins. Therefore, ultrafiltration is generally combined with another method, such as salting out or isoelectric point precipitation, to further remove impurities and obtain the protein of a higher purity level.

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2.6 THERMAL DENATURATION METHOD 2.6.1 Definition Denaturation is a process that can cause changes in the natural conformation of a protein but does not involve any peptide bond cleavage. The protein denaturation caused by heating is called thermal denaturation. The soluble proteins of the natural state are precipitated by heating to achieve separation and extraction.

2.6.2 Mechanism of Thermal Denaturation The stabilized conformation of a protein is maintained by hydrogen bonds and van der Waals forces and is affected by many factors, such as environmental temperature, salt ion concentrations, and pH values. If the ambient temperature rises to a limit point, then the hydrogen bonds and van der Waals forces can be disrupted, resulting in the protein molecule being in a denatured state. This process is the thermal denaturation. Natural sweet potato protein has a high solubility level, but heat treatments can lead to precipitation by disrupting the hydrogen bonds and van der Waals forces.

2.6.3 Thermal Denaturation Production Process of Sweet Potato Protein For thermal denaturation, sweet potato protein in slurry is initially denaturated by heating and cooled for later flocculation and sedimentation. After centrifugation to remove the water, sweet potato protein powder is obtained by a final drying treatment. The specific processing steps and parameters are as follows: Sweet potatoes cut into small pieces (2 cm3) - color protection (0.1% sodium bisulfite solution at 1:2 solid-to-solvent ratio) - grind - sieve with cheese cloth - centrifugation (10,000 3 g, 30 min) supernatant - thermal denaturation - flocculation and sedimentation - centrifugation (5000 3 g, 15 min) - precipitation - drying - sweet potato protein powder. The suitable technological parameters for extracting sweet potato protein were pH of 4.5, heating temperature of 95 C, and heating for 3 min, which resulted in a sweet potato protein purity of 59.27%, and an extraction yield that could reach 82.54%. In the actual production process, the thermal denaturation step is usually replaced by a high-temperature steam-injection process. After concentration, the

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Figure 2.11 Thermal denaturated sweet potato protein powder (left) and freeze-dried protein powder (right).

denatured sweet potato protein powder could be obtained using spray drying. As shown in Fig. 2.11, although the freeze-drying method results in a relatively high solubility level for native sweet potato protein, it is difficult to apply it to large-scale industrialization because it does not provide an economic benefit. The thermal denaturation method is only for extracting denaturated sweet potato protein, and the purity of the protein is relatively low. However, it also has some advantages, such as simple operation, lower equipment investment, and no chemical substances. Moreover, denaturated sweet potato protein is easier to digest and is suitable for mass production. At present, this technology has been industrialized and is being used by a sweet potato starch company in Shanxi Province to produce sweet potato protein products.

2.6.4 Factors Influencing Thermal Denaturation 2.6.4.1 Heating Temperature Temperature is the main factor promoting the heat-based denaturation of protein. The protein will not be denatured and precipitated until the heating temperature breaks protein hydrogen bonds and disrupts van der Waals forces. Sweet potato protein begins to undergo flocculation and precipitation when the temperature is higher than 85 C, and the precipitate content increased with the increasing temperature, as did the recovery rate and purity (Fig. 2.12). 2.6.4.2 Denaturation Time With extension of time, the recovery rate of sweet potato protein initially increased and then decreased. It reached a maximum value of

Sweet Potato Proteins

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72 70

44

68 42 66

Purity (%)

Recovery rate (%)

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40 64 38

62

Recovery rate

Purity

60

36

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95

Temperature (°C) Figure 2.12 Effects of temperature on the recovery rate and purity of heat-denatured protein.

77

Recovery rate (%)

76 75 74 73 72 71 70 0.5

1

2

3

5

7

Heating time (min) Figure 2.13 Effects of heating time on the recovery rate of sweet potato protein.

75.64% at a 5-min heating time. The recovery rates at 3 and 5 min had high values of 75.32% and 75.64%, respectively. Thus, the denaturation time is appropriately maintained at 3
5 min (Fig. 2.13). 2.6.4.3 Cooling Temperature A cooling treatment can promote the flocculation of heat-denatured sweet potato protein. In addition, the cooling temperature has a certain impact on the protein recovery rate. The recovery rate was slightly reduced as the cooling temperature increased. Moreover, the recovery rates were not significantly different when the cooling temperatures were in the 20
50 C range (Fig. 2.14).

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77 Recovery rate (%)

76 75 74 73 72 71 70

20

30 40 50 Cooling temperature (°C)

60

Figure 2.14 Effects of cooling temperature on the recovery rate of sweet potato protein.

100 80

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Purity 4.5

5.0

20

Recovery rate 5.5

6.0

Purity (%)

Recovery rate (%)

80

6.5

7.5

0

pH

Figure 2.15 Effects of the pH values on the recovery rate and purity of sweet potato protein.

2.6.4.4 pH Value The pH value has a significant influence on sweet potato protein recovery rate and purity level. Sweet potato protein has a high recovery rate and purity level near the isoelectric point, and the recovery rate and purity level gradually decrease as the pH increases (Fig. 2.15).

2.7 PURIFICATION METHOD OF SWEET POTATO PROTEIN Generally, sweet potato protein purity is less than 70% when using a single extraction method, such as salting out, isoelectric point, ultrafiltration, or foam separation. Therefore, it is necessary to combine several methods. Using the combined salting out and ultrafiltration methods, or combined isoelectric precipitation and ultrafiltration methods, the purity of

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sweet potato protein can reach more than 85%. It usually needs to be further purified by DEAE-52 ion-exchange column chromatography or Sephadex G-75 gel chromatography to produce the highest purity level of sweet potato protein, which can be higher than 95% (Xiong, 2009).

2.7.1 Ion-exchange Chromatography Method Ion-exchange chromatography mainly uses the natural reversible exchange property that occurs between ions in the mobile phase and an ion exchange agent (with the ion-exchange properties of the material) to separate the ionic compounds, which is the exchange reaction process between ions in solution and ion-exchange agent functional groups. Ionic compounds having a small charge and weak affinity are eluted first, whereas ionic compounds having a large charge and strong affinity strong are eluted later. The crude sweet potato protein powders were commonly purified using DEAE-52 ion-exchange chromatography with 1-mmol/L EDTA and 0.2-mol/L NaCl in a 50-mmol/L Tris
HCl buffer solution (pH 7.5) as the eluent. After a preliminary purification, SDS
PAGE results showed that sweet potato protein fractions mainly have three stained bands at 22, 31, and 66 kDa.

2.7.2 Sephadex G-75 Gel Chromatography Method Small molecular weight salts, sugars and other impurities in crude protein can be removed by DEAE-52 ion-exchange chromatography. However, to produce a higher purity sweet potato protein, further purification using a Sephadex column is necessary. Based on the molecular weights of the main components of sweet potato protein, Sephadex G-75 glucan was chosen for column packing. Preliminary purified and concentrated sweet potato proteins were further purified by Sephadex G-75 chromatography. Then, the main peak was collected using a 50-mM Tris
HCl buffer solution as the eluent (containing 0.1-M NaCl and 1-mM EDTA, pH 7.5). The white powder produced after freeze-drying had protein content greater than 99%. There are two stained bands at 31 and 22 kDa when β-mercaptoethanol was not added, whereas a single stained band of 25 kDa can be detected after adding β-mercaptoethanol, which indicated that high-purity sweet potato protein could be obtained using the Sephadex G-75 gel chromatography method.

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SECTION 3: BIOLOGICAL ACTIVITY OF SWEET POTATO PROTEIN The ability of active groups to combine with specific receptors and produce corresponding biological effects is termed biological activity. It can also be understood as the ability to induce changes in the normal mechanisms of cells. Sweet potato protein can effectively remove the oxidative reaction induced by free radicals and inhibit the effects of fat cells and cancer cell proliferation. Moreover, sweet potato protein also has strong trypsin inhibitory and anticancer activities, as well as positive effects on reducing blood lipid levels, enhancing immunity, and curing diabetes. Thus, sweet potato protein has a strong biological activity and can be a new health food resource.

3.1 ANTIOXIDANT ACTIVITY The metabolic processes of an organism are often accompanied by the production of a small amount of free radicals and reactive oxygen species. Under normal circumstances, the organism has a complete defense system composed of antioxidases and antioxidants, which can eliminate free radicals, prevent lipid peroxidation, and maintain the organism’s metabolic balance of free radicals. When the organism becomes old or has suffered the influence of environmental toxins, disorder occurs in the free radical metabolism, and the reactive oxygen species and free radicals have significant toxic effects on the body. Exogenous antioxidants help maintain the redox balance of the organism. The abilities of sweet potato protein to scavenge free radical and prevent lipid peroxidation were studied using the superoxide anion free radical system, hydroxyl radical system, DPPH radical system, and linoleic acid through oxidation system. These two capabilities reflect the antioxidant activity of sweet potato protein (Xue, 2006).

3.1.1 Superoxide Anion Radical Scavenging Activity of Sweet Potato Protein In 1968, McCord and Fridovich discovered superoxide dismutase and proposed the theory of toxic oxygen. They thought oxygen toxicity was due to the existence of superoxide anions, which have a long longevity and strong diffusibility. They can cause membrane damage, the

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Scavenging activity of O2 - (%)

12 10 8 6 4

2 0

0

1

2 3 4 Protein concentration (%)

5

6

7

Figure 2.16 Superoxide anion (O2 2) free radical scavenging activity of sweet potato protein.

inactivation of certain enzymes, and alterations of mitochondrial oxidative phosphorylation. The superoxide anion is the basis for the production of other oxygen free radicals, which can be transformed into other types of oxygen free radicals and cause great damage to the human body. Therefore, it is necessary to study the superoxide radical scavenging ability of sweet potato protein. Sweet potato protein has a certain scavenging activity on the superoxide anion free radicals produced by pyrogallol. Superoxide anion radical scavenging activity reached a maximum (B10%) when sweet potato protein concentration was 1%, and then remained unchanged as the protein concentration increased (Fig. 2.16).

3.1.2 Hydroxyl Radical Scavenging Activity of Sweet Potato Protein Hydroxyl radicals are a type of free radical with the most active chemical properties. They have a high reaction rate constant and cause the most harm among the free radicals. In a biological body, hydroxyl radicals attack the cell membrane, causing membrane damage and destroying sugar groups and DNA base sequences, inducing the disintegration of the double-helix structure, even causing cell death and mutations. Therefore, the scavenging activity of the hydroxyl radical is commonly used to evaluate the ability of scavenge free radicals of substance. Sweet potato protein has a strong hydroxyl radical scavenging activity. The scavenging rate of hydroxyl radicals amounted to 17.5% at a 2% protein concentration and continuously increased as sweet potato protein concentration increased. When sweet potato protein

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Scavenging activity of OH (%)

100 90

80 70 60 50 40 30 20 10 0 0

1

2

3 4 5 6 7 Protein concentration (%)

8

9

10

11

Figure 2.17 Hydroxyl radical (•OH) scavenging activity of sweet potato protein.

concentration was 6%, the scavenging rate reached 91.5% and was maintained at a similar level at higher concentrations (Fig. 2.17).

3.1.3 DPPH Radical Scavenging Activity of Sweet Potato Protein DPPH radicals are stable free radicals in organic solvents. The stability is mainly based on the resonance stabilization and the steric hindrance effects of three benzene rings. Thus, the unpaired electrons of the nitrogen atom cannot play a role in the pairing of electrons. The DPPH method can be applied to screening of free radical scavenging agents because DPPH radicals can provide information on the reaction between the specimen and the stable free radical. Sweet potato protein also has a strong DPPH radical scavenging activity. When the protein concentration was 0.1%, the DPPH radical scavenging rate reached to 38.5%, and then continuously increased as sweet potato protein concentration increased. When sweet potato protein concentration was 0.5%, the scavenging rate was 79.7% (Fig. 2.18).

3.1.4 Linoleic Acid Peroxidation-Inhibiting Effect of Sweet Potato Protein The destruction of proteins and membrane structures caused by lipid peroxides can result in damaged biological cells. Because of the great effects of lipid peroxidation on human health and life, it is important to study the preventive functions of sweet potato protein on lipid peroxidation. The linoleic acid
potassium thiocyanate detection method is mainly used to evaluate the abilities of antioxidants to prevent linoleic acid peroxidation and may partially reflect the ability of sweet

Scavenging activity of DPPH (%)

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90 80 70

60 50 40 30 20 10 0

0

0.5

1 1.5 Protein concentration (%)

2

2.5

Figure 2.18 DPPH radical (•DPPH) scavenging activity of sweet potato protein.

Antioxidant activity (%)

25 20 15 10 5 0

0

0.5

1

1.5

2

2.5

3

3.5

4

4.5

Protein concentration (%)

Figure 2.19 Antioxidant activity of sweet potato protein.

potato protein to prevent lipid peroxidation. Sweet potato protein has an inhibitory effect on linoleic acid oxidation systems. The antioxidant activity of sweet potato protein reached a maximum of B20%, when the concentration was 2% and maintained a similar level as the protein concentration increased (Fig. 2.19).

3.2 TRYPSIN INHIBITORY ACTIVITY OF SWEET POTATO PROTEIN Trypsin inhibitors are substances that inhibit the activity of trypsin and are usually divided into two categories, Browan-Birk-type

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inhibitors and Kunitz-type inhibitors, which both belong to the typical serine protease inhibitor family (Zhuang, 1999). The plant trypsin inhibitors are rich in nature. Generally, the molecular weights of plant trypsin inhibitors are small, and the active sites and action mechanisms are similar. In addition, they have a strong inhibitory effect on trypsin, plasma kallikrein, thrombin, fibrinolytic enzyme and coagulation factors Xa, and XIIa (Otlewski et al., 2000). Browan-Birk-type inhibitors have significant inhibitory effects on colorectal cancer, liver cancer, oral epithelial cancer, and lung cancer in the mammalian body (Kennedy et al., 1996, 2002). In addition, Kunitz-type inhibitor can greatly reduce the tumor burden and the formation of ascites in the peritoneal metastasis model of ovarian cancer. At present, trypsin inhibitory activity has been found and extracted from many kinds of plant tissues, such as those of soybean and squash trypsin inhibitors (Zhang et al., 1999; Mandal et al., 2002). Sweet potato protein extracted from sweet potato has trypsin inhibitory activity and can be used as a trypsin inhibitor. Yeh et al. (1997) compared the trypsin inhibitor isolated from the sweet potato root and the Sporamin expressed by Escherichia coli, and confirmed that the isolated inhibitor was Sporamin, which had a high trypsin inhibitor activity and the ability to induce injury. The genes encoding Sporamin proteins were highly homologous with the Kunitz-type soybean trypsin inhibitors and the injury-response genes of aspen (Zhang et al., 1999). Experimental studies proved that transferring the genes encoding Sporamin into tobacco and Brassica plants not only improved their trypsin inhibitory activities but also enhanced plant disease resistance and insect resistance (Morikami et al., 2005; Chen et al., 2006).

3.2.1 Determination of the Trypsin Inhibitor Activity of Sweet Potato There are two main methods for the determination of trypsin inhibitory activity, the Smith-recommended and the Kunitz methods (Mu and Li, 2010), which can both measure and evaluate the activity of Sporamin. The Smith-recommended method not only can detect Sporamin’s trypsin inhibitory activity, but it can also make a quantitative determination with good reproducibility. Because the Smithrecommended method makes determinations for blanks and samples, this method is more close to the true value. The Sporamin activity

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determined by the Kunitz method indicated that the inhibition of trypsin by Sporamin also showed good reproducibility.

3.2.2 Factors Influencing Trypsin Inhibitory Activity of Sweet Potato 3.2.2.1 Effect of Temperature on Trypsin Inhibitory Activity of Sweet Potato The treatment temperature has a great influence on the trypsin inhibition activity of sweet potato. The trypsin inhibition activities of roasted and cooked sweet potato are completely lost (Dickey et al., 1984). The trypsin inhibition activity of sweet potato still has thermal stability at 70 C, whereas maintaining 130 C for 30 min can completely destroy the trypsin inhibition activity of fresh sweet potato (Chein et al., 1980). Deng et al. (2009) studied the changes in trypsin inhibitor activity of sweet potato protein after heat treatment using the Smithrecommended method, and the trypsin inhibitory rate was significantly decreased as the treatment temperature increased. Sporamin has a high heat resistance and is not completely inactivated until the temperature reaches 127 C. 3.2.2.2 Effects of Growth Conditions on Trypsin Inhibition Activity of Sweet Potato The influence of growth conditions on the trypsin inhibitory activity of sweet potato should not be ignored. Studies have indicated that the growth conditions, such as season, cumulative precipitation, geographical location, and fertilization, of sweet potato could lead to differences in the trypsin inhibitory activity of sweet potato (Hou et al., 2004; Zhang et al., 1998; Bradbury et al., 1985). 3.2.2.3 Protein Content Related to Trypsin Inhibitory Activity of Sweet Potato The protein content of sweet potato varieties was directly related to the trypsin inhibitory activity. The trypsin inhibitory activity has a low positive correlation with the soluble protein level (Lin et al., 1980). 3.2.2.4 Effects of Structural Changes on Trypsin Inhibitory Activity The effects of structure showed that the trypsin inhibitory activity of Kunitz-I type and Bowman-Birk soybean trypsin inhibitors were related to differences in secondary structures. The inhibition of trypsin is a noncompetitive inhibition. The β-turn and random coil contents were decreased to 10.8% and 54%, respectively, after ultrasonic

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treatment with 65% amplitude for 20 min, whereas the β-sheet content was increased to 35.2%, and 71.5% of disulfide bonds were changed to thiol groups. These structural changes resulted in a 55% reduction in the trypsin inhibitory activity (Huang et al., 2004, 2005).

3.3 ANTICANCER ACTIVITY OF SWEET POTATO PROTEIN Cancer seriously endangers human life and health. There are B2 million new cases and 1.3 million people die of cancer each year in China. WHO experts warn that through the 2020s, the number of global cancer deaths per year may double, and 84 million people will die of cancer during the next decade. At present, cancer treatments have reached a certain level, but more effective adjuvant therapy is required to improve the efficacy of treatments. Invasive metastasis of cancer cells is a malignant feature of tumors and is the main cause of death in most patients having solid tumors. Trypsin inhibitors can inhibit the invasion and metastasis of malignant tumor cells. A urinary trypsin inhibitor has significant inhibitory effects on the metastasis of the extracellular matrix of breast cancer cells and Lewis lung cancers in mice (Liu et al., 2005). Moreover, bovine pancreatic trypsin inhibitor has strong inhibitory effects on experimental pulmonary metastasis of B16 melanoma tumors and Lewis lung cancers in mice. In addition, it has a greater inhibitory effect in combination with chemotherapy drugs (Du et al., 2004). In addition, soybean trypsin inhibitor inhibits the proliferation and metastasis of breast cancers. Sweet potato protein, like other kinds of trypsin inhibitors, also has a role in inhibiting tumor invasion and metastasis.

3.3.1 The Inhibitory Effects of Sweet Potato Protein on Cancer Cell Proliferation Sweet potato protein has a significant inhibitory effect on the proliferation of cancer cells. Deng (2009) found that sweet potato protein can inhibit the growth and proliferation of SW480 cells in a concentrationdependent manner. A significant inhibitory effect on the proliferation of SW480 cell can be observed clearly at lower protein concentrations, whereas it has a certain toxicity and effects on cell viability at a high concentration level. Table 2.4 shows that the survival rate of the cells is significantly reduced when sweet potato protein concentration is 2.5 mg/mL, and certain cytotoxicity on SW480 cells was produced.

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Table 2.4 Effects of Sweet Potato Protein Concentrations on the Cytotoxicity of SW480 Cells Concentration (mg/mL)

Survival Rate (%)

0.0

96.05 6 2.67

0.1

95.44 6 2.57

0.25

96.88 6 1.08

0.5

96.66 6 0.72

1.0

95.89 6 0.84

2.5

83.33 6 8.33a

P , 0.01.

a

3.3.2 Inhibitory Effects of Sweet Potato Protein on the In Vitro Migration and Invasion of Cancer Cells The inhibitory effects of sweet potato protein on the in vitro migration and invasion of cancer cells were studied mainly by two methods: One uses the Transwell method to determine the migration of cancer cells; the other adapted the ECM550 invasion assay kit to evaluate the antiinvasion properties of sweet potato protein. Deng (2009) investigated the inhibitory effects of sweet potato protein on the in vitro migration and invasion of cancer cells using both the Transwell method and the ECM550 invasion kit method. The migration of SW480 cells could be inhibited when sweet potato protein concentration was greater than 0.2 mg/mL. Furthermore, sweet potato protein significantly inhibited the invasion of SW480 cells when the concentration was 0.02 mg/mL, and the inhibitory effects were dose dependent. Thus, sweet potato protein can inhibit the migration of cancer cells and has a strong ability to resist the invasion of cancer cells.

3.3.3 Inhibitory Effects of Sweet Potato Protein on Tumor Metastasis Sweet potato protein has a strong inhibitory effect on tumor growth and metastasis in animals bearing tumors. Deng (2009) found that sweet potato protein can inhibit the metastasis of peritoneal diffuse transplanted tumors in nude mice. Compared with the model group, sweet potato protein significantly reduced the ascites and tumor weights of nude mice, as well as the intraperitoneal tumors. In addition, sweet potato protein also has a restraining effect on the metastasis of spontaneous Lewis lung cancer. After intraperitoneal injection and gavage protein treatment, the lung cancer tumor’s metastasis rate

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in mice significantly decreased compared with that of the control group. In particular, the weights of tumors in the mice receiving intraperitoneal injections of sweet potato protein were significantly lower than those of the control group.

3.4 OBESITY PREVENTION AND WEIGHT LOSS Previous studies (Xiong, 2009) hypothesized that sweet potato protein had the potential effect of preventing obesity, and assisting in weight and fat loss, which were then confirmed by cell and animal experiments. There are two ways to prevent obesity: reducing the number of fat cells by inhibiting the proliferation of preadipocytes and reducing the amount of fat in fat cells by preventing the preadipocyte transfer to mature adipocytes (Xiong, 2009).

3.4.1 Effects of Sweet Potato Protein on the Proliferation and Differentiation of 3T3-L1 Preadipocytes 3T3-L1 preadipocytes is the source of mouse embryos, having the morphology of fibroblast cells and the ability to differentiate into mature adipocytes under the stimulation of classic hormonal cocktails, including insulin, dexamethasone, and 3-isobutyl-1-methyl xanthine. They are commonly used in obesity studies and in the in vitro differentiation of adipocytes. The effects of sweet potato protein on the differentiation and proliferation of 3T3-L1 preadipocytes were studied through cell recovery, morphology, and cultures, as well as to induce differentiation of former 3T3-L1 fat cells. There was no significant change in the survival rate of the 3T3-L1 preadipocytes after different concentrations of sweet potato protein were added. Even as sweet potato protein concentration increased from 0.025 to 1.00 mg/mL, the cell survival rate remained above 92%, which indicated that sweet potato protein is not toxic to cells when the protein concentration is below 1.0 mg/mL (Table 2.5). After using different concentrations of sweet potato protein and differentiation culture medium to treat preadipocytes for 5 days, the contents of stained lipid droplets in the differentiated 3T3-L1 cells were significantly decreased as sweet potato protein concentration increased. The same significant inhibitory effect has also been seen with 0.020mg/mL berberine as the control (Fig. 2.20), which suggested that sweet

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Table 2.5 Effects of Different Sweet Potato Protein Concentrations on the Survival Rate of 3T3-L1 Preadipocytes (%) Sporamin (mg/mL)

24 h

48 h

0.000

93.3 6 1.30

97.4 6 0.12

0.025

94.9 6 1.52

97.8 6 1.15

0.125

92.2 6 1.16

95.0 6 3.07

0.250

93.3 6 1.33

99.3 6 0.10

0.500

93.7 6 1.02

98.5 6 0.95

1.000

93.1 6 1.04

97.5 6 1.61

3.5 a b

3

Absorbance (492 nm)

2.5 2 c 1.5 1 0.5 0

0

0.025

0.25

d

d

0.5

0.02 (berberine)

Protein concentration (mg/mL)

Figure 2.20 Inhibitory effect of sweet potato protein on the differentiation of 3T3-L1 preadipocytes.

potato protein also had a strong inhibitory effect on the differentiation of 3T3-L1 preadipocytes, like berberine (Xiong et al., 2009). Sporamin also has an effect on the proliferation of fat cells. This inhibitory effect gradually increased as the protein concentration and time increased and was shown to be time- and concentration-dependent (Xiong et al., 2009). The inhibition of 3T3-L1 preadipocyte proliferation at different Sporamin concentrations using the 3-(4,5-dimethyl-2thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide method (as MTT method) are shown in Fig. 2.21A. Compared with the blank, the OD value decreased after the 36-h treatment at a 0.025-mg/mL protein concentration, indicating that the concentration was able to effectively inhibit the proliferation of 3T3-L1 preadipocytes (P , 0.05). Moreover, the absorbance values decreased as the protein concentration increased, which indicated that the inhibitory effect gradually increased.

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Absorbance of elution by MTT method (492 nm)

(A) 2.5

adipocytes adding sporamin adding berberine

2 1.5 1 0.5 0

0

12

24 Time (h)

36

48

(B) Figure 2.21 Time- or dose-dependent inhibition of Sporamin on proliferation of 3T3-L1 preadipocytes: (A) Berberine (0.02 mg/mL) and different concentrations of 3T3-L1 preadipocytes treated with Sporamin for 36 h. (B) Sporamin (1.00 mg/mL) and berberine (0.02 mg/mL)-treated 3T3-L1 preadipocytes at different times.  Note: All of the data are means 6 standard deviations (n 5 6); P , 0.05.

Furthermore, as the processing time was extended from 12 to 48 h, the OD values of the treated groups had an increasing trend at a protein concentration of 1.00 mg/mL (Fig. 2.21B), which indicated that the number of cells increased. However, when compared with the blank, the OD values of Sporamin and the berberine group were significantly reduced after being treated for 24 h. This demonstrated that

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Sporamin proteins and berberine effectively inhibited the proliferation of preadipocytes (P , 0.05).

3.4.2 Experimental Animal Study of Obesity Prevention Obesity is a symptom caused by a long-term energy intake over consumption, leading to the storage of excessive fat that becomes damaging to the human health. Almost all health researchers and clinicians believe that prevention is the most important strategy to control the spread of obesity (Müller et al., 2001). Thus, there are important practical significances to the development of healthy foods to aid in the prevention of obesity. Xiong (2009) carried out some animal experiments on the ability of sweet potato protein to prevent obesity and, compared with normally fed mice, feeding with different dosages of sweet potato protein reduced the food intake levels and body weights of mice. However, no significant changes were observed in liver and kidney weight indices, indicating that the given irrigation gastric concentration was not toxic and had no effects on the accumulation of visceral adipose cells in mice. In addition, compared with the control group, the total cholesterol, triglyceride, and low-density lipoprotein cholesterol levels of mice in both model and sweet potato protein-fed groups were significantly increased. However, sweet potato protein can decrease the contents of low-density lipoprotein cholesterols and elevate the levels of highdensity lipoprotein cholesterols, ultimately promoting lipid transportation and excretion to prevent obesity.

3.4.3 Animal Experimental Study on Losing Weight and Reducing Lipid Levels Xiong (2009) carried out some animal experiments on the ability of sweet potato protein to reduce weight, and the body weights and lengths of mice in the obese model group were significantly greater than those of the blank control group. The diet intakes and weights of mice in the weight-loss group were decreased after feeding on sweet potato protein, and the greater sweet potato protein intake, the greater the weight loss, and feeding reduction. Compared with the obese mice model group, the weights of the total abdominal fat of mice in the reduced-weight model mice group were significantly decreased. Thus, sweet potato protein is likely to

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have a role in the inhibition of abdominal obesity. Sweet potato protein aids in weight loss in obese mice and did not cause toxicity but inhibited food intake. It can also reduce the blood lipid level by reducing the total lipid, total cholesterol and low-density lipoprotein cholesterol contents in the blood.

SECTION 4: FUNCTIONAL PROPERTIES OF SWEET POTATO PROTEIN Texture, flavor, color, appearance, and other sensory qualities affect people’s food choices. Protein has significant influences on food sensory qualities. The functional properties of protein usually include the physicochemical properties of protein that affect the performance of food systems during processing, storage, preparation, and consumption. The physicochemical properties that determine the functional protein properties include: size, shape, amino acid composition and sequence, distribution of the net charge, ratios of hydrophobicity and hydrophilicity, secondary, tertiary and quaternary structures, flexibility and rigidity of molecules, interactions between protein molecules, and the ability to interact with other groups. In addition, the external environment (such as temperature, pH value, salt concentration, and pressure) and storage conditions can also cause protein functional changes. Under nonreducing conditions, sweet potato protein has three isomers with different molecular weights, which are connected by disulfide bonds. Sweet potato protein is a kind of high-nutrient plant protein. To develop the applications of sweet potato protein in the food industry, it is necessary to understand its functional properties in food processing and storage. Accordingly, the solubility, emulsifying property, gel property, structural property, foaming ability, waterholding capacity, and oil-holding capacity of sweet potato protein are discussed below.

4.1 SOLUBILITY OF SWEET POTATO PROTEIN More than 80% of sweet potato storage protein is soluble, but the extraction and processing methods, such as pH value, ionic strength, temperature, and solvent type, can affect its solubility (Xue, 2006).

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120

Solubility (%)

100 80 60

0.1%

40

0.5% 1.0%

20 0

1

2

3

4

5 pH

6

7

8

9

10

Figure 2.22 pH
solubility curves of sweet potato protein solution at different concentrations.

120 Solubility (%)

100 80 60

Distilled water 0.1 mol/LNaCl 0.5 mol/LNaCl 1.0 mol/LNaCl

40 20 0 1

2

3

4

5

6

7

8

9

10

pH Figure 2.23 pH
solubility curves of 1% protein solution at different NaCl concentrations.

4.1.1 Effect of pH Value on the Solubility of Sweet Potato Protein The pH value of the aqueous solution has a great influence on the solubility of sweet potato protein. Regardless of the concentration of sweet potato protein, the lowest solubility values always appeared near pH 4 (Fig. 2.22). For most plant proteins, the lowest solubility is observed near the protein’s isoelectric point; however, sweet potato protein shows a high solubility level over a considerable pH range.

4.1.2 Effects of pH and NaCl on the Solubility of Sweet Potato Protein The solubility of sweet potato protein in different pH solutions decreased after the addition of NaCl (Fig. 2.23).

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The addition of NaCl significantly reduces the solubility of the protein at pH values less than 4 because of the interactions between proteins, and between protein and solvent. Under these conditions, the minimal solubility of sweet potato protein may be caused by the neutralization of positively charged protein and adsorption of chloride ions, which reduces the electrostatic repulsion to maintain protein separation and then enhances the interactions between proteins. Thus, the protein’s solubility is reduced accordingly. In the neutral pH range, with an increasing ion concentration, the electrostatic repulsion is slightly weakened by the neutralization between amino and carboxyl groups, leading to a slight decrease in protein solubility. Under alkaline conditions, the amino groups of the protein do not become charged; therefore, neutralization does not occur after addition of NaCl and a high solubility level is maintained.

4.1.3 Effects of Salt Species on the Solubility of Sweet Potato Protein Salt species have different effects on the solubility of sweet potato protein. The protein solubility after the addition of NaCl was lower than after the addition with CaCl2 or MgCl2 at pH 4, but higher than those of the other two salts at other pH levels. Under alkaline conditions, even though the addition of MgCl2 causes precipitation because of the reaction between Mg21 and OH2, the solubility of sweet potato protein after the addition of CaCl2 was still lower than that of MgCl2 (Fig. 2.24). This may be due to Ca21 combining with negatively charged proteins, forming a “bridge.” 120

Solubility (%)

100 80 60 Ca（1mol/L）

40

Mg（1mol/L） Na（1mol/L）

20 0

1

2

3

4

5

6

7

8

9

10

pH Figure 2.24 pH
solubility curves of 1% protein solution after the addition of different salt ions.

Sweet Potato Proteins

95

4.2 EMULSIFYING PROPERTIES OF SWEET POTATO PROTEIN Many foods are present as emulsions during processing. As an important component of the preparation of emulsified foods, the demand for emulsifiers is increasing year by year. Sweet potato protein is often used as an emulsifier. It has a medium-range molecular weight and has good emulsifying properties. It can be used as a natural emulsifier and as a kind of high-quality protein resource. In addition, it has low-energy requirements and health care
related functional characteristics. Therefore, sweet potato protein has good market prospects (Guo, 2010). In recent years, many scholars have studied the emulsifying properties of animal and plant proteins and explored the effects of various physical and chemical conditions on the emulsifying properties of proteins. The protein concentration, oil volume fraction, pH value, and salt concentration are important factors that affect the emulsifying properties of protein. In particular, the pH value and salt level can influence the formation and stability of an emulsion by changing the protein molecule’s structures, including surface hydrophobicity and surface charge distribution (Xue, 2006).

4.2.1 Effects of the Protein Concentration and Oil Volume Fraction on the Emulsifying Properties of Sweet Potato Protein 4.2.1.1 Effects of the Protein Concentration and Oil Volume Fraction on the Size of Emulsion Particles The mean size of the emulsion particles is a very important index for evaluating the properties of the emulsion. The smaller mean particle size of an emulsion, the stronger the emulsifying effects. As shown in Fig. 2.25, the volume-weighted mean diameter (D4,3) of sweet potato protein initially decreased rapidly as the protein concentration increased, but the rate of the mean particle sizes decreased slowly with further increases in the concentration (Fig. 2.25A). Moreover, the oil volume fraction is also an important factor that affects the size distribution. The D4,3 of the emulsion particles increased as the volume fraction of the oil phase increased in the emulsion (Fig. 2.25B) (Guo, 2010). 4.2.1.2 Effects of the Protein Concentration and Oil Volume Fraction on the Emulsifying Activity and Stability of Sweet Potato Protein The emulsifying activity index (EAI) indicates the efficiency of the emulsifier, which is an important parameter for characterizing the

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12 (A)

10

D 4,3 (µm)

8 6 4 2 0 0

0.5 1 1.5 Protein concentration (%, w/v)

2

12

(B)

D 4,3 (µm)

10 8 6 4 2 0 0

5

10

15

20

25

30

35

40

45

50

Oil volume fraction (%, v/v) Figure 2.25 Effects of protein concentration (A) and oil volume fraction (B) on the volume-weighted mean diameters (D4,3) of emulsion particles.

capacity of the emulsifier. As shown in Fig. 2.26A, the EAI value of sweet potato protein emulsion significantly decreased as the protein concentration (0.1%
2.0% w/v) increased but increased as the oil volume fraction increased (Fig. 2.26B). The emulsion stability index (ESI) value indicates the stability of sweet potato protein emulsion. As shown in Fig. 2.27A, the emulsion stability gradually increased as the protein concentration increased (0.1%
2.0% w/v) and increases rapidly at low protein concentrations (#0.5%). However, the stability decreased as the oil volume fraction increased (Fig. 2.27B), which indicates that protein concentration and oil volume fraction are two important factors that affect the stability of the emulsion. Moreover, they can improve the stability of the emulsion by increasing the protein concentration and reducing the oil volume fraction.

Sweet Potato Proteins

97

180 160

(A)

EAI (m2/g)

140 120 100

80 60 40 20 0

0

0.5

1

1.5

2

Protein concentration (%, w/v) 45

(B)

40 35 EAI (m2/g)

30 25

20 15 10 5 0

0

5

10

15

20

25

30

35

40

45

50

Oil volume fraction (%, v/v) Figure 2.26 Effects of protein concentration (A) and oil volume fraction (B) on the emulsifying activity of sweet potato protein.

4.2.1.3 The Concentration and Composition of Sweet Potato Protein Adsorbed on the Interface The stability of the emulsion is closely related to the composition and concentration of the adsorbed protein on the interface between oil and water. The protein composition of an SDS
PAGE pattern (Fig. 2.28) indicated that the main components of the interface adsorption are Sporamins A and B, compared with sweet potato protein, whereas the high molecular weight protein is not adsorbed. The interface-adsorbed protein concentration of sweet potato protein emulsion increased almost linearly as the protein concentration increased. When the protein concentration was greater than 1% (w/v), the rate of adsorbed protein decreased as the protein concentration increased. The adsorbed protein tended to saturation at protein

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120 (A) 100

ESI (min)

80 60 40 20 0

0

0.5

1

1.5

2

Protein concentration (%, w/v) 250

(B)

ESI (min)

200 150 100 50 0

0

5

10

15

20

25

30

35

40

45

50

Oil volume fraction (%, v/v) Figure 2.27 Effects of protein concentration (A) and oil volume fraction (B) on the emulsion stability of sweet potato protein.

concentrations of 1.5% and 2% (w/v), with no significant changes. However, the adsorbed protein concentration decreased rapidly as the oil volume fraction increased (Fig. 2.29).

4.2.2 Effects of pH on the Emulsifying Properties of Sweet Potato Protein 4.2.2.1 Effect of pH on the Particle Size of Emulsion Particles The solubility of sweet potato protein is dependent on the pH change of the aqueous solution. The protein has the lowest solubility at pH 4, at which the conformation of the protein molecule changes and has electrical neutrality. Moreover, the interactions between the protein molecules, including electrostatic and Van Edward forces, are enhanced and cause flocculation. Thus, the highest D4,3 value of the emulsion particles occurred at pH 4 (Fig. 2.30) (Guo, 2010).

Sweet Potato Proteins

99

Figure 2.28 SDS
PAGE pattern of the emulsion interface-adsorbed protein and nonadsorbed protein. Note: Lane 1, sweet potato protein; Lane 2, interface-adsorbed protein; Lane 3, nonadsorbed protein.

4.2.2.2 Effects of pH on the Emulsifying Activity of Sweet Potato Protein The pH value can affect the emulsifying properties of sweet potato protein by changing the molecular structure and surface charge distribution. Thus, the D4,3 of the emulsion particles was highest at pH 4. The EAI value of sweet potato protein emulsion increased as the molecular diffusion coefficient and repulsive forces increased. At a high pH (7 or 8), the EAI of the emulsion was higher and more stable (Figs. 2.31 and 2.32). As shown in Fig. 2.31, at pH 2, 3, 4, 5, 6, 7, and 8, the EAI values were 13.62, 26.56, 19.4, 27.01, 28.02, 31.6, and 30.83 m2/g, respectively. With increasing pH levels (pH 2
4), the EAI values initially increased and then decreased. As the pH level continued to increase (pH 5
8), so did the EAI values (Guo, 2010). 4.2.2.3 Effects of pH on the Stability of Sweet Potato Protein Emulsion The pH value significantly affected the stability of sweet potato protein emulsion (Fig. 2.32). At pH 2, 3, 4, 5, 6, 7, and 8, the ESI was 32.8, 54.1, 41.5, 24.2, 79.5, 85.7, and 125.3 min, respectively. The stability of sweet potato protein emulsion was the lowest at pH 5, while the stability was greater at pH 4, which may be because the interface-adsorbed protein concentration of sweet potato protein emulsion was higher,

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2.5 Interface adsorbed protein concentration (mg/m2)

(A) 2

1.5 1 0.5 0

0

0.5

1

1.5

2

Protein concentration (%, w/v)

Interface adsorbed protein concentration (mg/m2)

3.5 (B)

3 2.5 2

1.5 1 0.5 0

0

5

10

15

20

25

30

35

40

45

50

Oil volume fraction (%, v/v) Figure 2.29 Effect of protein concentration (A) and oil volume fraction (B) on interface-adsorbed protein concentration.

7

D 4,3 (µm)

6 5 4 3 2 1 2

3

4

5

6

7

8

pH Figure 2.30 Effects of pH on the volume-weighted mean diameters (D4,3) of sweet potato protein emulsion (1%, w/v).

Sweet Potato Proteins

101

35 30 EAI (m2/g)

25 20

15 10 5 0

2

3

4

5

6

7

8

pH Figure 2.31 Effects of pH on the emulsifying activity index (EAI) of sweet potato protein (1%, w/v).

140

ESI (min)

120 100 80 60 40 20 0

2

3

4

5 pH

6

7

8

Figure 2.32 Effects of pH on the emulsion stability index (ESI) of sweet potato protein emulsion (1%, w/v).

resulting in a lower level of aggregation among the emulsion particles. With the increase in pH (pH 6
8), the stability of sweet potato protein emulsion increased gradually, but it increased first and then decreased (pH 2
4), which may reflect the electrostatic repulsion among the emulsion particles (Guo, 2010).

4.2.3 Effects of NaCl on the Emulsifying Properties of Sweet Potato Protein 4.2.3.1 Effects of NaCl on the D4,3 of Sweet Potato Protein Emulsion NaCl is an important factor that affects the functional properties of proteins, especially at low concentrations. At low concentrations,

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8 7 D 4,3 (µm)

6 5 4 3 2 1 0

0

0.2

0.4

0.6

0.8

1

NaCl concentration (mol/L) Figure 2.33 Effects of the NaCl concentration on the volume-weighted mean diameters (D4,3) values of sweet potato protein emulsion.

NaCl reduces the electrostatic repulsion between emulsion particles, which increases the mutual attraction, promotes the aggregation of emulsified particles, and finally increases the particle size of the emulsion. The effect of the NaCl concentration on the D4,3 of sweet potato protein emulsion is shown in Fig. 2.33. At NaCl concentrations of 0.1, 0.2, 0.4, 0.6, 0.8, and 1.0 mol/L, the D4,3 values of the sweet potato emulsion were 4.1, 4.98, 4.35, 4.43, 4.5, and 4.45 μm, respectively. The D4,3 values significantly increased with the addition of different concentrations of NaCl, and they reached the maximum of 4.98 μm at 0.2mol/L NaCl (Guo, 2010). 4.2.3.2 Effects of NaCl on the Emulsifying Activity of Sweet Potato Protein At 0.2-mol/L NaCl, the mutual attraction between particles becomes frequent because of the strong electrostatic shielding, resulting in an increase in the D4,3 value of sweet potato protein emulsion, which leads to decreases in the EAI and ESI values (Figs. 2.34 and 2.35). Compared with the control condition (no addition of NaCl), the EAI of sweet potato protein emulsion significantly decreased from 30 to 18 mg/m2 when the NaCl concentration was 0.2 mol/L. With a further increase in the NaCl concentration (0.4
1.0 mol/L), the EAI of sweet potato protein emulsion was also decreased (28.9, 29.3, 28.8, and 27.1 mg/m2) but was only slightly lower than the control (Guo, 2010).

Sweet Potato Proteins

103

35 30 EAI (mg/m2)

25 20 15 10

5 0

0

0.2

0.4 0.6 NaCl concentration (mol/L)

0.8

1

ESI (min)

Figure 2.34 Effects of different NaCl concentrations on the emulsifying activity index (EAI) of sweet potato protein emulsion (1%, w/v).

90 80 70 60 50 40 30 20 10 0

0

0.2

0.4 0.6 NaCl concentration (mol/L)

0.8

1

Figure 2.35 Effect of different NaCl concentrations on the emulsifying activity index (EAI) of sweet potato protein emulsion (1%, w/v).

4.2.3.3 Effects of NaCl on the Stability of Sweet Potato Protein Emulsion The effects of NaCl on the stability of sweet potato protein emulsion are similar to those on the activity of sweet potato protein emulsion. As shown in Fig. 2.35, the stability of sweet potato protein emulsion at 0.2 mol/L NaCl was the lowest, but as the NaCl concentration increased, the stability of sweet potato protein emulsion increased (Guo, 2010).

4.2.4 Effects of High Hydrostatic Pressure Treatment on the Emulsifying Properties of Sweet Potato Protein High hydrostatic pressure treatments can significantly influence the emulsifying activity and emulsifying stability of sweet potato protein,

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as well as the particle size of the emulsion. However, for different concentrations of sweet potato protein, the effects of the high hydrostatic pressure treatment were different. Mehmood et al. (2013) studied the effects of high hydrostatic pressure treatments on the emulsifying properties of sweet potato protein at protein concentrations of 2%, 4%, and 6% (w/v). As shown in Fig. 2.36, the EAI value of sweet potato protein decreased after high hydrostatic pressure treatments (200
600 MPa), except the 400 MPa treatment on protein concentrations of 2% and 6%. The ESI values of sweet potato protein were also significantly reduced (P , 0.05) at 2% and 6% after high hydrostatic pressure treatment (200
600 MPa). However, the ESI value of sweet potato protein at a 4% (w/v) concentration increased rapidly after 400 and 600 MPa 100

SPP 2% SPP 4% SPP 6%

90 80 70

ab

(A)

ab

ab

a

ab

b

a

ab

ab

b

EAI (m2/g)

60

b

b

50 40 30 20 10 0

0.1

300

SPP 2% SPP 4% SPP 6%

250

ESI (min)

200

200 400 Pressure (MPa)

a

600

(B)

a b

c

b

d

150 a 100

b

c

d

d

50 0

c

0.1

200 400 Pressure (MPa)

600

Figure 2.36 Effects of pressure on the emulsifying properties of different concentrations of sweet potato protein.

Sweet Potato Proteins

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Table 2.6 Effects of High Hydrostatic Pressure Treatments on the Particle Size of Sweet Potato Protein Emulsion at Different Concentrations Sv (m2/mL)

Sample

Pressure (MPa)

D4,3 (μm)

2%

0.1

3.34 6 0.03b

2.60 6 0.01b

0.57 6 0.00b

200

3.70 6 0.04

a

2.74 6 0.00

a

0.54 6 0.00c

400

3.31 6 0.00

b

2.61 6 0.00

b

0.57 6 0.00b

600

3.28 6 0.03b

2.57 6 0.01c

0.58 6 0.00a

0.1

3.62 6 0.27

2.77 6 0.11

0.54 6 0.02b

200

3.65 6 0.71a

2.46 6 0.00b

0.56 6 0.05ab

400

3.26 6 0.00

a

2.66 6 0.29

ab

0.60 6 0.00a

600

3.06 6 0.06

a

2.46 6 0.02

b

0.60 6 0.00a

0.1

3.83 6 0.01b

2.76 6 0.01b

0.54 6 0.00a

200

4.02 6 0.58

a

2.89 6 0.09

a

0.51 6 0.01b

400

4.57 6 0.22

a

3.01 6 0.04

a

0.49 6 0.01b

600

4.48 6 0.24a

2.96 6 0.06a

0.50 6 0.01b

4%

6%

D3,2 (μm)

a

a

Different letters indicate significant differences (P , 0.05). D4,3 (μm) represents the volume-weighted means diameter; D3,2 (μm) represents the volume
surface mean diameter; Sv (m2/mL) represents the specific surface area.

treatment. This may have been due to the emulsion droplets having larger specific surface areas at the 4% (w/v) concentration, or a slight increase in the viscosity of the aqueous phase, weakening the collision rate of the droplets compared with the 200-MPa treatment. The 200-MPa treatment slightly increased the emulsion droplet size of sweet potato protein at a concentration of 2%, whereas the size decreased after the 400-MPa treatment. The emulsion droplet size of sweet potato protein was not significantly changed at a concentration of 4% after high hydrostatic pressure treatments (Table 2.6).

4.3 GELLING PROPERTIES OF SWEET POTATO PROTEIN Gelling is an important functional property of protein. Sweet potato protein can form a gel by heat treatment under neutral conditions, and the lowest gel concentration is 8% (w/v). The salt concentration has significant effects on the gel strength. With the addition of NaCl at a low concentration of 0.1 mol/L, the gel strength increased from 333.77 (in water) to 548.87 g but then decreased as the increase of NaCl concentration (0.5 and 1.0 mol/L), which were 394.38 and 348.78 g, respectively.

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Figure 2.37 Effects of NaCl concentration on the gel structure of sweet potato protein (10%, w/v): (A) native gel; (B
D) protein gels with 0.1, 0.5, and 1.0-mol/L NaCl, respectively.

As observed with a scanning electron microscope, the heated gel that was dissolved in distilled water showed a compact and smooth plate structure (Fig. 2.37A). After adding 0.1 mol/L NaCl, the gel showed a more closely packed granular structure (Fig. 2.37B). With increases in the NaCl concentration, the irregular particles became larger, and the interspace around them gradually increased (Fig. 2.37C and D), forming a relatively loose structure, which leads to a decrease in gel strength (Xue, 2006). Sweet potato proteins prepared by different methods show great differences in their gel properties. Arogundade et al. (2012) found that the hardness, elasticity, and recovery of sweet potato protein gels prepared using the isoelectric point precipitation method were stronger than those prepared using the ultrafiltration method. However, the structure of gels prepared using the ultrafiltration method was much more compact than those prepared using the isoelectric point precipitation method (Fig. 2.38).

Sweet Potato Proteins

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Figure 2.38 Scanning electron microscope of protein gels prepared using the (A) isoelectric point precipitation method and (B) ultrafiltration method.

Figure 2.39 SDS
PAGE of sweet potato protein at different pH values: (A) without β-mercaptoethanol, Coomassie brilliant blue staining; (B) with β-mercaptoethanol, Coomassie brilliant blue staining.

4.4 STRUCTURAL PROPERTIES OF SWEET POTATO PROTEIN Sweet potato protein, which is mainly composed of Sporamins A and B, having molecular weight of 31 and 22 kDa, respectively, is rich in β-sheet and β-turn structures. However, the structure and composition of sweet potato protein can be changed by different treatment methods.

4.4.1 Effects of Heat Treatment and pH on the Structure of Soluble Sweet Potato Protein The pH value has an effect on the structure of sweet potato protein. As shown in the SDS
PAGE (Fig. 2.39), strong acid conditions greatly influence sweet potato protein. Under nonreducing conditions

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(Fig. 2.39A), the 31-kDa-stained band was not present at pH 1 and 2 when compared with the other pH treatments. Under reducing conditions, all of the treatments showed the same protein-stained band of 25 kDa, indicating that it is difficult to form three kinds of isomers of sweet potato protein using disulfide bonds. The molecular structure of sweet potato protein changed under the 100 C heat treatment, forming high molecular weight polymers by disulfide bonds, and the alkaline condition promoted the reaction. Under nonreducing conditions (Fig. 2.40A), in addition to the polymer

Figure 2.40 SDS
PAGE of sweet potato protein after heating at 100 C for 10 min: (A) without β-mercaptoethanol, Coomassie brilliant blue staining; (B) with β-mercaptoethanol, Coomassie brilliant blue staining.

Sweet Potato Proteins

109

of B41 kDa, a higher molecular weight of polymer was also detected when the pH increased to 10. The protein staining bands of 31 kDa were not detected at all pH levels, indicating that the protein was unstable at 100 C. Under the reducing conditions, a single stained 25kDa band of sweet potato protein (Fig. 2.40B) was mainly detected, but the band intensity significantly decreased at pH 1
3, whereas a protein staining band of 14-kDa significantly increased, which suggested that a large amount of sweet potato protein degraded to the 14-kDa protein under acidic conditions at 100 C. Furthermore, high molecular weight polymers formed in a wide pH range, regardless of whether β-mercaptoethanol was added. Alkaline conditions promoted polymer formation (Xue, 2006).

4.4.2 Effects of High Hydrostatic Pressure Treatments on Structure of Sweet Potato Protein High hydrostatic pressure treatments can affect the secondary structures of proteins, promote, or damage the formation of chemical bonds in the protein and thus alter the functional properties of the protein. Sweet potato protein is rich in β-sheet and β-turn structures (Table 2.7). High hydrostatic pressure treatments promote the formation of secondary protein structures, and there was a significant difference in the composition of the secondary structures of sweet potato protein under different pH and pressure conditions. Table 2.7 Composition of Sweet Potato Protein at Different pH Levels Before and After High Hydrostatic Pressure Treatments (%) Sample

Composition

pH 3

α-Helix

15.50

3.90

10.70

9.60

β-Sheet

45.30

67.00

50.00

57.40

β-Turn

12.20

0.70

12.40

7.30

Random coil

27.00

28.40

26.90

25.80

α-Helix

14.40

8.80

5.00

30.40

β-Sheet

37.20

42.10

64.80

6.60

β-Turn

22.80

16.60

0.00

39.00

Random coil

25.70

32.50

30.20

24.10

pH 6

pH 9

0.1 MPa

200 MPa

400 MPa

600 MPa

α-Helix

8.90

5.50

16.50

6.00

β-Sheet

57.60

70.80

44.90

51.40

β-Turn

10.40

3.30

14.50

11.70

Random coil

23.10

20.40

24.10

30.90

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Sweet Potato Processing Technology

4.5 THE FOAMING PROPERTIES AND FOAM STABILITY OF SWEET POTATO PROTEIN Foaming occurs when liquid medium is combined with gas by physical means through agitation or aeration. Foaming is helpful in altering the texture, density, smell, and taste qualities of food, such as ice cream, cake, and bread. Foaming properties and foam stability are affected by protein concentration, protein type, degree of protein inactivation, other components, calcium ions, pH value, temperature, and agitation methods. The foam stability can be expressed as the water loss rate. The lower the water loss rate, the greater the foam stability. The soluble protein content in solution increased with increases in sweet potato protein concentration, as did the amount of bubbles forming. In addition, the foaming capacity improved, the water loss rate was reduced, and the foam stability was improved, but the extent of these changes depended on the sweet potato variety (Fig. 2.41). The foaming properties of sweet potato protein are dependent on pH, and the foaming ability is poor under the neutral conditions. The foam volume increased as the pH increased to over 6, and the largest amount of foam occurred at pH 10. The foam stability of sweet potato protein was best at pH 4 (Fig. 2.42). As shown in Fig. 2.42, the foaming curve of “Jing No. 6” purple sweet potato protein was slightly different from those of “Ya No. 1” and “No. 981,” which may be due to “Jing No. 6”s sweet potato protein powder containing some anthocyanins. Anthocyanins are water-soluble, and some of their properties are affected by pH conditions. Sweet potato protein varieties, such as “Yushu No. 7,” “Jing No. 6,” and “No. 95
837,” have good foaming capacities and foam stabilities (Table 2.8) (Liu, 2007).

4.6 WATER-HOLDING AND OIL-HOLDING CAPACITIES OF SWEET POTATO PROTEIN The water-holding capacity of protein is very important in meat processing, which can affect the juiciness, tenderness, and taste of the meat products. Protein with a good oil-holding capacity can be widely used in egg yolk products, meat products, dairy products, coffee mate, dough, and cake pastes. Previous results show that the water-holding capacity of sweet potato protein is relatively poor, mainly because of its good solubility, and most of the protein can be dissolved in water

Sweet Potato Proteins

Foaming capacity (Fc)

3.5

(A)

3 2.5 2

Jing No.6

1.5

Ya No.1

1

No.981

0.5 0

1

3

5 7 Protein concentration (%)

80

(B)

70 Water loss rate (%)

111

60

9

Jing No.6 Ya No.1 No.981

50 40 30 20 10 0

1

3 5 7 Protein concentration (%)

9

Figure 2.41 Effects of sweet potato protein concentration on the foaming properties (A) and foam water loss rate (B).

(Xue, 2006). This may be related to the large number of hydrophilic groups in the protein. In addition, sweet potato protein has a better oil-holding capacity than soybean protein, asparagus bean protein (Ragab et al., 2004), and sesame protein (Khalida et al., 2003). As shown in Table 2.8, sweet potato varieties, like “Jiangsu 55-1” (3.40 g/g) and “S-19” (3.00 g/g), have high water-holding capacities, whereas “Xushu No. 18” (3.52 g/g) and “Jiangsu 55-1” (2.88 g/g) have high oil-holding capacities.

SECTION 5: APPLICATIONS OF SWEET POTATO PROTEIN As basic material, protein is an indispensable part of the human dietary structure. Sweet potato protein is a vegetable protein of high quality, with a balanced amino acid ratio, that has many food-related functional

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Sweet Potato Processing Technology

(A) 4 Foaming capacity (Fc)

3.5 3 2.5 2

Jing No.6

1.5

Ya No.1

1

No.981

0.5 0 3

4

5

6

pH

7

8

9

10

(B) Water loss rate (%)

70

Jing No.6

60

Ya No.1

50

No.981

40 30 20

10 0

3

4

5

6

pH

7

8

9

10

Figure 2.42 Effects of pH on the foaming properties (A) and water loss rate (B) of sweet potato protein.

properties, such as solubility, emulsibility, gelation, water absorption, oil-holding capacity, and foamability. Different sweet potato protein contents could be widely used in different food processing areas.

5.1 EDIBLE PROTEIN POWDER Natural sweet potato protein is hard to digest because of its trypsin inhibitory activity; therefore, its admirable nutritional properties cannot be exhibited. However, the sweet potato powder made using thermal denaturation technology, whose trypsin inhibitory activity has been completely eliminated, is a good edible protein source, which can be digested and absorbed easily (Sun et al., 2012). In addition, heatdenatured protein can be used as a protein supplement and auxiliary material in various flour products, such as mixing sweet potato powder

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Table 2.8 Functional Assay Results of Sweet Potato Protein from Different Cultivars (%) Variety

Solubility

WHC

OHC

EAI

ESI

Foaming

Water-

Viscosity

(%)

(g/g)

(g/g)

(m2/g)

(h)

Capacity

Loss

(mPa s)

Rate Yz-2

88.98

2.43

1.50

73.20

42.89

2.04

46.83

1.84

S-19

85.63

3.00

1.56

69.67

44.53

2.41

40.06

3.03

Yushu No.7

80.30

2.92

2.29

68.81

43.03

2.96

32.84

1.65

YZ-4

99.89

2.64

2.23

64.21

45.58

1.44

69.68

1.72

Yz-1

96.14

2.58

2.15

73.11

46.76

2.36

42.61

3.97

Free sugar No.1

92.42

1.55

2.57

49.53

47.44

2.10

45.03

1.88

Jishu 98

91.94

1.62

2.36

63.62

43.35

2.30

40.00

2.05

Xushu 18

87.42

1.79

3.52

47.63

30.89

2.09

45.87

2.02

Yizi 138

83.66

1.83

1.99

62.86

67.3

1.51

65.45

3.92

Ya No.1

100.20

1.95

2.01

102.1

159.92

1.53

62.25

5.09

Jing No.1

90.97

2.16

2.29

52.54

251.93

1.55

64.75

2.24

Jing No.6

111.86

0.79

1.78

62.21

107.58

2.80

34.49

1.51

Black sweet potato

91.27

1.68

1.79

62.03

51.74

2.41

44.7

1.57

No. 981

85.66

2.58

1.60

53.52

106.44

1.72

56.41

2.34

Jiangsu 55-1

83.93

3.40

2.88

71.67

48.21

1.82

52.44

2.33

No. 95-837

85.2

2.76

2.09

43.76

62.3

2.76

35.09

3.09

Jing No. 2

67.03

2.02

2.64

62.12

90.41

2.28

40.92

4.82

No. 94-7-63

80.89

2.40

1.28

60.12

68.8

2.27

42.54

1.51

Hongxinwang

68.08

2.12

1.35

58.62

74.62

1.58

63.11

1.61

Note: All of the data are the mean values of three measurements, where WHC, OHC, EAI, and ESI are water-holding capacity, oil-holding capacity, emulsifying-activity index, and emulsion-stability index, respectively.

with flour to make steam breads, breads, biscuits, and cakes. Adding sweet potato powder to a recipe not only increases the protein content but also improves the color, taste, texture, and bulkiness of foods and prevents them from drying and hardening. Noodles, instant noodles, and macaroni will become more resistant to cooking, thereby maintaining higher nutritional values after adding sweet potato protein.

5.2 EMULSIFIER Protein is a surface active material and, as a typical amphiphilic compound (hydrophilic and hydrophilic), has the ability to decrease the interfacial tension. During the emulsifying process, the protein

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Sweet Potato Processing Technology

molecules absorb on the oil droplet quickly, forming a thin protein membrane, which deters the aggregation of droplets and increases the formation of homogeneous emulsifying particles, improving the emulsion stability. The molecular weight of sweet potato protein is about 25 kDa. Sweet potato protein has a good solubility and emulsifying activity in a wide range of pH values; moreover, it also possesses good functional and health care-related characteristics. Thus, sweet potato protein is a natural, nutritional, and functional emulsifier. In addition, this new protein emulsifier, with good performance, will meet the needs of multiple applications after protein modification.

5.3 HUMECTANT Sweet potato protein has good gelling characteristics and can retain much more water during food processing, avoiding water loss, which could act as a kind of humectant. Adding sweet potato protein to various meat products could improve the quality of the products and increase their nutritional values because of the supplementary protein. Sweet potato protein can also be used as binding agent and additive, instead of starch, in sausage and meat balls, which could improve taste, elasticity, and water absorption. In Chinese traditional foods, such as pie, steamed stuffed buns, and dumplings, adding sweet potato protein instead of meat will decrease the animal fat and cholesterol content, which will increase the protein content and improve the quality.

5.4 RAW MATERIAL OF ACTIVE PEPTIDES Active peptides for food resource can increase an organism’s immune response and prevent active oxygen from triggering various diseases. Thus, they have a vast potential market value. At present, various antioxidant peptides of food protein resources, such as soybean protein peptide, potato protein peptide, wheat germ protein peptide, casein peptide, fish protein peptide, can be produced after hydrolysis by different commercial proteases. The hydrolysate, after hydrolysis of sweet potato protein, has a greater free radical scavenging ability and angiotensin-converting enzyme inhibitory activity, as well as greater antioxidant and antihypertensive activities (Wang et al., 2011; Zhang et al., 2012a, 2012b). Sweet potato protein can be used as a good protein resource for producing active peptides.

Sweet Potato Proteins

115

5.5 BIOLOGICAL MEDICINE Research has shown that the trypsin inhibitors of animals and plants have obvious inhibitory effects on rectal cancer, liver cancer, oral epithelial cancer, and lung cancer, which can significantly decrease tumors. Natural sweet potato protein of a high purity, acting as a kind of trypsin inhibitor, may also have inhibitory effects on cancer proliferation and tumor metastasis. Thus, the extracted, natural high-purity sweet potato protein could be used clinically as a medicinal product.

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3), 134
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570. Zhang, D.P., Collins, W.W., Andrade, M., 1998. Genotype and fertilization effects on trypsin inhibitor activity in sweet potato. HortScience 33, 225
228. Zhang, M., Mu, T.H., Sun, M.J., 2012a. Sweet potato (Ipomoea batatas L.) protein hydrolysates: antioxidant activity and protective effects on oxidative DNA damage. Int. J. Food Sci. Technol. 47 (11), 2304
2310. Zhang, M., Mu, T.H., Wang, Y.B., et al., 2012b. Evaluation of free radical scavenging activities of sweet potato protein and its hydrolysates as affected by single and combination of enzyme systems. Int. J. Food Sci. Technol. 47 (4), 696
702. Zhang, Z., Wang, Z.H., Lin, R.F., 1999. Purification and some properties of the trypsin inhibitor from tartary buckwheat seeds. Chin. J. Biochem. Mol. Biol. 15 (2), 247
251 (in Chinese). Zhao, M., Ding, X.L., 2005. Study on extraction process of sweet potato glycoprotein. Food Res. Dev. 26 (1), 77
79 (in Chinese). Zhuang, B.C., 1999. Studies on the Biology of Wild Soybean in China. Beijing: Science Press, pp. 144
146 (in Chinese).

FURTHER READING Debora, V., Wanda, C., 1989. Ipomoein is the major soluble protein of sweet potato storage roots. HortScience 24 (5), 829
830. Fu, T.T., Mu, T.H., Chen, J.W., et al., 2012. Emulsifying properties of hydrolysates made by limit enzymatic hydrolysis of sweet potato heat-denatured protein. J. Nucl. Agric. Sci. 26 (07), 1018
1024 (in Chinese). Fu, T.T., Mu, T.H., 2013. The effect of high-pressure homogenization on emulsifying properties of enzymatic peptides obtained from sweet potato heat-denatured protein by alcalase. J. Nucl. Agric. Sci. 27 (1), 68
74 (in Chinese). Guo, Q., Mu, T.H., 2010. Effect of calcium chloride on emulsifying properties of sweet potato soluble protein. Sci. Agric. Sin. 43 (11), 2340
2346 (in Chinese). Kobayashi, H., Suzuki, M., Kanayama, N., et al., 2004. A soybean Kunitz trypsin inhibitor suppresses ovarian cancer cell invasion by blocking urokinase upregulation. Clin. Exp. Metastasis 21, 159
166. Kunitz, M., 1945. Crystallization of a trypsin inhibitor from soybean. Science 101 (2635), 668
669. Liu, L.L., Mu, T.H., Sun, Y.L., 2008. Nutritional composition and component correlation of sweet potato cultivars. J. Chin. Cereals Oils Assoc. 23 (1), 39
43 (in Chinese). Mignatti, P., Rifkin, D.B., 1993. Biology and biochemistry of proteinases in tumor invasion. Physiol. Rev 73 (1), 161
195.

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Smith, C., Van, M.W., Twaalfhoven, L., et al., 1980. The determination of trypsin inhibitor levels in foodstuffs. J. Sci. Food Agric. 31 (4), 341
350. Suzuki, M., Kobayashi, H., Tanaka, Y., et al., 2003. Suppression of invasion and peritoneal carcinomatosis of ovarian cancer cell line by overexpression of bikunin. Int. J. Cancer 104 (3), 289
302. Tan, X.W., Wu, Z.L., Jia, Y.S., Yu, G.H., 2005. Application of foam separation in multicomponent mixture of proteins. Chem. Ind. Eng. Prog. 24 (5), 510
513 (in Chinese). Xue, Y.L., Meng, X.J., Sun, Y.L., Mu, T.H., 2006. Quality comparison of four sweet potato protein powder. Food Res. Dev. 27 (2), 51
53 (in Chinese).

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CHAPTER

3

Sweet Potato Dietary Fiber

SECTION 1: INTRODUCTION OF DIETARY FIBER 1.1 The Definition of Dietary Fiber 1.2 The Composition of Dietary Fiber 1.3 The Classification of Dietary Fiber 1.4 Dietary Fiber Extraction Methods 1.4.1 Coarse Separation Method 1.4.2 Chemical Separation Method 1.4.3 Enzymatic Hydrolysis Method 1.4.4 Chemical Reagent in Combination with the Enzymatic Hydrolysis Method 1.5 The Physicochemical and Functional Properties of Dietary Fiber 1.6 The Mechanism of Dietary Fiber in Preventing Obesity 1.6.1 Physical Effects 1.6.2 Improving Intestinal Microbes 1.6.3 Regulating Short-Chain Fatty Acids in Intestine 1.6.4 Altering Metabolism SECTION 2: SWEET POTATO DIETARY FIBER EXTRACTION TECHNOLOGY 2.1 Sieve Method 2.1.1 Technological Processes 2.1.2 The Main Operation Steps 2.1.3 Results 2.2 Sieve Combined Enzymatic Hydrolysis Method 2.2.1 Technological Processes 2.2.2 The Main Operation Steps 2.2.3 Results Sweet Potato Processing Technology. DOI: http://dx.doi.org/10.1016/B978-0-12-812871-8.00003-9 Copyright © 2017 China Science Publishing & Media Ltd. Published by Elsevier Inc. All rights reserved.

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2.3 Biological Method 2.3.1 Single Enzyme Method 2.3.2 Fermentation Method 2.3.3 Mixed Enzymes Method 2.4 Chemical Separation Method 2.4.1 Water Extraction 2.4.2 Acid and Alkali Extraction 2.4.3 Chemical Reagent in Combination with the Enzymatic Method SECTION 3: THE PHYSIOLOGICAL PROPERTIES OF SWEET POTATO DIETARY FIBER 3.1 Effect of Sweet Potato Dietary Fiber on Preventing Obesity 3.1.1 Effects of Sweet Potato Dietary Fiber on the Body Weights of Rats 3.1.2 Glucose Tolerance Test 3.1.3 Blood Index Determination 3.1.4 Determination of Liver Indices 3.1.5 The Determination of the Organic Acid Content in Cecum 3.2 Effects of Sweet Potato Dietary Fiber on Treating Obesity 3.2.1 Effects of Sweet Potato Dietary Fiber on the Body Weights of Rats 3.2.2 Glucose Tolerance Test 3.2.3 Determination of Blood Indices 3.2.4 Determination of Liver Indices 3.2.5 The Determination of the Organic Acid Content in Cecum 3.3 Other Functional Properties of Sweet Potato Dietary Fiber 3.3.1 Protecting the Intestines From Cancer 3.3.2 The Absorption of Cholesterol 3.3.3 The Absorption of Na1, K1, Ca21, Fe31, and Pb21 3.3.4 Accelerating the Output Speed of Feces SECTION 4: THE PHYSICOCHEMICAL PROPERTIES OF SWEET POTATO DIETARY FIBER 4.1 Effects of Different Factors on the Water-Holding Capacity of Sweet Potato Dietary Fiber

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4.2 Effects of Different Factors on the Water-Swelling Capacity of Sweet Potato Dietary Fiber 4.3 Effects of Temperature on the Oil-Holding Capacity of Sweet Potato Dietary Fiber 4.4 Effects of Different Factors on the Viscosity of Sweet Potato Dietary Fiber 4.5 Water-Holding Capacity of Dietary Fibers From Different Sweet Potato Varieties 4.6 Water-Swelling Capacity of Dietary Fiber From Different Varieties of Sweet Potato 4.7 Oil-Holding Capacities of Dietary Fibers From Different Varieties of Sweet Potato SECTION 5: THE APPLICATIONS OF SWEET POTATO DIETARY FIBER 5.1 The Applications of Sweet Potato Dietary Fiber in Bread 5.1.1 The Sensory Qualities of Bread 5.1.2 The Physicochemical Properties of Bread 5.1.3 The Texture of Bread 5.2 The Applications of Sweet Potato Dietary Fiber in Beverages 5.3 The Applications of Sweet Potato Dietary Fiber in Meat Products 5.4 The Applications of Sweet Potato Dietary Fiber in Staple Foods 5.5 The Applications of Sweet Potato Dietary Fiber in Condiments 5.6 The Applications of Sweet Potato Dietary Fiber in Health Foods References Further Reading

Abstract In this chapter, dietary fiber extraction technology, such as sieve method, sieve combined enzymatic hydrolysis methods, biological methods, chemical separation method, and chemical reagent combined

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enzymatic hydrolysis method, are introduced. Physicochemical and functional properties of sweet potato dietary fiber, for example, viscosity, water-holding capacity, water-swelling capacity and oil-holding capacity, and physiological properties, are also evaluated. In addition, application of sweet potato dietary fiber in staple food, beverage, meat products, and functional foods are introduced. Results showed that sweet potato dietary fiber had high purity, good physicochemical and functional properties and exhibited better fat and glucose regulation. Therefore, dietary fiber extracted from sweet potato residue could be used as a fiber-rich ingredient in food and health product industries.

SECTION 1: INTRODUCTION OF DIETARY FIBER 1.1 THE DEFINITION OF DIETARY FIBER In 1953, dietary fiber was first defined by Hipsley, and it describes the components of plant cell wall materials that cannot be digested by the human intestinal tract, including cellulose, hemicellulose, and lignin (Devries et al., 1999). The definition and quantitative methods of determining dietary fiber were further improved and enriched in 1972 by Trowell, who indicated that dietary fiber was the plant cell residue that cannot be hydrolyzed by human digestive enzymes, including cellulose, hemicellulose, lignin, pectin, oligosaccharides, and gum (Rodriguez et al., 2006). In 1997, the Association of Official Analytical Chemists (AOAC) defined dietary fiber as plant polysaccharide substances that cannot be hydrolyzed by human digestive enzymes, including nonstarch polysaccharides, resistant starch, lignin, and other bioactive compounds (Cho et al., 1997). In 2001, the American Cereal Chemists Association redefined dietary fiber as plant substances or similar carbohydrates, including polysaccharides, oligosaccharides, lignin, and some plant materials, that cannot be digested and absorbed by the human small intestine but can be partially or fully fermented in the large intestine. This is a comprehensive and versatile definition of dietary fiber.

1.2 THE COMPOSITION OF DIETARY FIBER Dietary fiber is mainly composed of cellulose, hemicellulose, lignin, and pectin. Cellulose is the main component of the plant cell wall and

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is a kind of insoluble homogeneous polysaccharide, which is composed of glucose through the β-1,4-glucosidic bands (Wang and Li, 1999). Hemicellulose is mainly composed of xylose, glucose, mannose, galactose, arabinose, and rhamnose through the β-1,4-glucosidic bands, and it is also has galacturonic acid and glucuronic acid residues. Lignin is a nonpolysaccharide polymer that is formed by the connection of phenyl propane through ether and carbon
carbon bonds. Pectin is mainly composed of poly-galacturonic acid and rhamnogalacturonan. Polygalacturonic acid is composed of galacturonic acid residues through branched α-1,4-glucosidic bands, with methyl or acetyl groups as the substituent. Rhamnogalacturonan is composed of rhamnose through α-1,2-glucosidic bands, and of galacturonic acid residues through α-1,4-glucosidic bands. Rhamnose residues can be replaced by neutral galactose and arabinose.

1.3 THE CLASSIFICATION OF DIETARY FIBER Dietary fiber is a kind of carbohydrate and can be classified based on different contents. At present, the common classifications are as follows: Dietary fiber can be classified as soluble (SDF) and insoluble (IDF) based on the solubility. The former contains pectin, glucan, and gum. The latter contains cellulose, part of the hemicellulose, lignin, and wax  (Nawirska and Kwasniewska, 2005). Dietary fiber can be classified into partial fermentation and complete fermentation substances. The former contains lignin, cellulose, hemicellulose, vegetable wax, and cutin. The latter contains pectin, gum arabic, β-glucan, guar gum, seaweed glue, and inulin (Zheng, 2005). Dietary fiber can be classified as being from plant, animal, and microbial sources. Plant sources include lignin, cellulose, hemicellulose, galactomannan, and gum arabic. Animal sources include chitosan and collagen. Microbial sources include β-glucan, xanthan gum, and carboxymethylcellulose (Zheng, 2005).

1.4 DIETARY FIBER EXTRACTION METHODS At present, the methods for extracting dietary fiber mainly include coarse separation, chemical separation, enzymatic hydrolysis, and

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chemical reagent combined with enzymatic hydrolysis method (Zheng, 1995; Chen, 2002).

1.4.1 Coarse Separation Method The coarse separation method mainly includes the suspension and the air-flow classification methods. The content of dietary fiber can be increased by changing the relative content of each component in the raw material, such as decreasing the content of starch and phytic acid, thereby lowering the purity of the product obtained. The method is only suitable for the pretreatment of raw materials and does not apply to the preparation of high purity dietary fiber (Tang, 2002).

1.4.2 Chemical Separation Method In the chemical separation method, the crude product or raw material is soaked in acid or alkali solution after drying and then ground. The resulting solution is bleached and centrifuged after regulating the pH value to neutral. The obtained residue after centrifugation is the IDF, and then an ethanol solution is added to the supernatant to get precipitate, in which the precipitate obtained is the SDF (Cardoso et al., 2003).

1.4.3 Enzymatic Hydrolysis Method In the enzymatic hydrolysis method, amylase and protease are used, and some bioactive ingredients can be obtained by the introduction of other enzymes, including hemicellulose and arabanase (Anne et al., 2009; Wang et al., 2010). The advantage of enzymatic hydrolysis technology is that this method does not require high temperature and high pressure. This saves energy, decreases the processing equipment required, is convenient to operate, and is beneficial in protecting the environment (Min et al., 1998). Thus, it is particularly suitable for extracting dietary fiber from some materials that have high starch and protein contents.

1.4.4 Chemical Reagent in Combination with the Enzymatic Hydrolysis Method The detailed procedure of dietary fiber extraction using a chemical reagent in combination with the enzymatic hydrolysis method is as follows: First, raw materials are soaked in chemical reagents, such as phosphate buffer. Second, amylase is added, and protease, glucoamylase, and cellulase can be added under desirable pH conditions to degrade some ingredients. Third, an organic solvent, such as ethanol

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or acetone, is added. Then, after a period of reaction, the slurries are washed with water, filtered, and dried. This produces high purity dietary fiber (Baokun et al., 2011). In addition to the above methods, others, including membrane separation, fermentation, microwave assisted, ultrasonic and microwave combined ultrasonic, have also been used to extract dietary fiber (Margareta et al., 2002 Zhao, 2003; Lou et al., 2009; Wan et al., 2012).

1.5 THE PHYSICOCHEMICAL AND FUNCTIONAL PROPERTIES OF DIETARY FIBER The physicochemical properties of dietary fiber mainly include the water-holding capacity, oil-holding capacity, water-swelling capacity, and viscosity. The water and oil-holding capacities are the amounts of water and oil, respectively, combined with a certain amount of dietary fiber. Water-swelling is the volume of water that is absorbed by a certain mass of dietary fiber in the presence of an excess of water. Viscosity is considered to be related to the physiological properties. The viscosity of a liquid can be increased when dietary fiber is added. Viscous dietary fibers can change the concentrations of blood glucose and cholesterol, increase satiety, and shorten food retention times in the intestine. The functional properties of dietary fiber mainly include the glucose-absorption capacity, fat-absorption capacity, α-amylase activity inhibition ratio, and pancreatic lipase activity inhibition capacity (Chau et al., 2003). Dietary fiber can absorb some small molecules, such as glucose and fat, or delay their diffusion, because it has a larger specific surface area and exhibits a porous and loose structure (Chau et al., 2003). Chau et al. (2004) indicated that some characteristics are related to the α-amylase activity inhibition ratio because dietary fiber can pack starch and enzymes and prevent contact between enzymes and substrates, thus interfering with the enzymatic hydrolysis of starch substances by α-amylase. In addition, the structure of α-amylase can be affected by pectin and polyphenols in dietary fiber, thus inhibiting the activity of α-amylase. Chau et al. (2003) showed that the pancreatic

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lipase activity inhibition capacity of dietary fiber is related to the oilholding capacity because (1) dietary fiber can absorb fat, thereby preventing contact between pancreatic lipase and fat and (2) binding sites can be formed between hydrophobic groups of dietary fiber and pancreatic lipase, thereby affecting the activity and structure of pancreatic lipase. The above physicochemical and functional properties determine the applications of dietary fiber in the food industry (Sangnark and Noomhorm, 2003).

1.6 THE MECHANISM OF DIETARY FIBER IN PREVENTING OBESITY 1.6.1 Physical Effects Dietary fiber has a strong chewiness and high water-swelling capacity, and it can be used as a substitute for energy. Thus, it can increase satiety and reduce nutrient absorption in humans. Dietary fiber can absorb or bind cholesterol and bile acids, accelerate their excretion rates, reduce intestinal absorption of cholesterol and bile acids, and inhibit the intestinal-hepatic circulation of cholesterol and bile acids. In addition, dietary fiber can form gels in the intestine, thereby prolonging the time of gastric emptying and inhibiting the transport of glucose, triglycerides (TGs), and cholesterol in the intestines (Slavin, 2005). Some experiments (in vitro) confirmed that guar gum can reduce the influence of intestinal contractions, thus reducing glucose uptake (Edwards et al., 1988). Lairon et al. (2007) showed that viscous dietary fiber can form a gel, which reduces the contact between pancreatic lipase and the intestinal contents, preventing the formation of fat emulsion and micelles through the development of polymers between the dietary fiber and lipids.

1.6.2 Improving Intestinal Microbes Human intestinal microbes are mainly dominated by Bacillus and Firmicutes. Soluble dietary fiber can improve the intestinal microbes by supplying a substrate, such as fructo-oligosaccharide and galactooligosaccharide for microorganisms and can increase the contents of Bifidobacterium and Lactobacillus in infant feces (Boehm et al., 2002). Lactobacillus and Bifidobacterium can reduce blood cholesterol, enhance the early dissociation of bile, and remove extracellular

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cholesterol through absorption and precipitation (Turnbaugh et al., 2006). Some kinds of dietary fibers, such as inulin and oligosaccharides, can increase the content of Bifidobacterium and hinder the transformation of microorganisms related to obesity (Guigoz et al., 2002; Kolida et al., 2002).

1.6.3 Regulating Short-Chain Fatty Acids in Intestine Dietary fiber can be partially or completely fermented into short-chain fatty acids (SCFAs), such as acetic acid, propionate and butyrate, in the colons of humans and animals. Fermentation modes and the proportion of SCFAs are determined by the type of dietary fibers. Acetic acid is the main fermentation product of pectin, and propionic acid is the final fermentation product of gum arabic and cyclodextrine ́ ez-Escrig et al., 2008). Previous studies (Ushida et al., 2011; Jimen showed that the physiological properties of dietary fibers depend on the types and amounts of SCFAs, which can stimulate the intestinal absorption of water and sodium, and affect the intestinal pH and bile salt precipitation (Pylkas et al., 2005). SCFAs are associated with appetite, insulin sensitivity, and inflammation. Al-Lahham et al. (2010) showed that propionate can inhibit the stimulus
response of the inflammatory cytokine tumor necrosis factor-α (TNF-α). Studies (in vitro) indicated that SCFAs can inhibit the release of TNF-α and can also reduce the expression of the mRNA of interleukin-6 (IL-6) and protein (Tedelind et al., 2007). SCFAs can regulate physiological responses by activating different receptors, with the main receptors being free fatty acid (FFA)-receptors 2 and 3 (FFA2 and 3). Sleeth et al. (2010) showed that both FFA2 and 3 can regulate adipocyte differentiation, and FFA2 can reduce the storage of TG and the deposition of fat in adipose tissues. Vangaveti et al. (2010) confirmed that FFA2 could decrease the level of plasma SCFAs and increase the insulin sensitivity of fat cells. Corte et al. (2011) preformed experiments with mice that showed that the concentration of FFA2 in the colon increased by 31% when the mice were fed 5% cactus dietary fiber.

1.6.4 Altering Metabolism Dietary fiber can change the lipid and carbohydrate metabolism, mainly by regulating the expression levels of genes and hormones.

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1. Effects of dietary fiber on lipid metabolism Acetyl coenzyme A, which is controlled by a protein kinase, is the rate-limiting enzyme in the process of fat production. Experiments studying the effects of plantago dietary fiber on lean and obese mice for 10 weeks showed that the phosphorylation level of the acetyl coenzyme A-associated protein kinase increased when the obese mice were fed 5% plantago, thereby inhibiting the activity of acetyl CoA carboxylase. There were no significant differences between the control and the experimental group (Delzenne and Kok, 2001). Similarly, fructose-oligosaccharide (10 g/100 g) has also been shown to reduce the expression of acetyl coenzyme A carboxylase (Hu et al., 2010). Fatty acid synthase can catalyze the synthesis of fatty acids, and the main product is palmitic acid. The expression of fatty acid synthase can be reduced by adding resistant starch, fructose, inulin, β-glucan, plantato, and hydroxy propyl methyl cellulose to the diet of rodents (Demigné et al., 1995). Cani et al. (2008) indicated that the expression of TG lipase and the hormone sensitivity of adipose tissues in female mice adrenal glands increased after adding 1.0% gum arabic to drinking water for 180 days. These enzymes are involved in the fatty acid decomposition of adipocytes. TG lipase is the ratelimiting enzyme, which can hydrolyze TG s into diacylglycerol. 3-Hydroxy-3-methyl-glutaryl coenzyme A (HMG CoA) reductase, a restriction enzyme of cholesterol synthesis, can convert HMGCoA to mevalonate and can inhibit cholesterol synthesis by hindering HMG-CoA. Previous studies showed that hydroxy propyl methyl cellulose can increase the expression of HMG-CoA reductase (Zambell et al., 2003). Parnell and Reimer (2010) also determined that the combination of inulin and pectin can increase the expression of HMG-CoA reductase in mice. 2. Effects of dietary fiber on carbohydrates metabolism SDF and IDF may activate the insulin signaling pathway by participating in the regulation of hormones. For example, insulin polypeptide and glucagon-like peptide 1 (GLP-1), which are dependent on glucose, can stimulate insulin release, improve glucose tolerance, and delay gastric emptying. Glucagon is the precursor of GLP-1, and it is produced by L-cells at the end of the ileum and colon. Delzenne and Cani (2010) showed that dietary fiber can increase the amount of L-cells in the intestine, thus increasing the

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concentrations of glucagon and GLP-1. Reimer et al. (2011) also indicated that dietary fiber can increase the mRNA expression of glucagon in the ileum. Insulin is a protein hormone, which is secreted from pancreatic β cells after stimulation by endogenous or exogenous substances. The biological role of insulin is its ability to bind to specific receptors on the membranes of target cells and then activate them. The insulin receptor, which can form a complex with insulin or insulin-containing molecules, is a specific site on target cell membranes that has a high degree of specificity. After the binding of insulin to the insulin receptor, tyrosine kinase is activated and receptor substrates are phosphorylated, thereby regulating the intracellular enzyme system’s activity level and carbohydrate metabolism. Choi et al. (2010) showed that barley β-glucan can inhibit the serine phosphorylation of insulin receptor substrates, increase the tyrosine phosphorylation of insulin receptors, and activate the insulin signaling pathway by inhibiting transcription factors. In addition, SDF can regulate the metabolism of carbohydrates by altering gluconeogenesis. Choi et al. (2010) also showed that barley β-glucan can reduce the concentration of glucokinase, glucose-6phosphatase and pyruvate carboxykinase, which are involved in gluconeogenesis, in the liver.

SECTION 2: SWEET POTATO DIETARY FIBER EXTRACTION TECHNOLOGY Sweet potato residue, which is obtained from starch extraction, consists of more than 20% dietary fiber, and it is more valuable if dietary fiber can be extracted. Recently, there have been some reports on the extraction of sweet potato dietary fiber both in China and abroad, but the protocols cannot be used in industrial production because of their complexity. To solve this problem, the structures of sweet potato residues and starch particle sizes were studied, with the aim to develop a new and rapid production method for sweet potato dietary fiber. In this section, the sieve method, sieve combined enzymatic hydrolysis method, biological method, chemical separation method, and chemical reagent combined enzymatic hydrolysis method are introduced.

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2.1 SIEVE METHOD Preparation of sweet potato dietary fiber using the sieve method (SMSPDF) includes extracting the dietary fiber based on the different sizes of the raw materials. Compared with the chemical separation method, this method can reduce the loss of SDF and the cost. In addition, the color of the dietary fiber is light, and the purity is high. However, this method must be combined with enzymatic hydrolysis if nonstarch products are desired.

2.1.1 Technological Processes Sweet potato residue -wwashing -adrying -rgrinding -rslurry mixing (liquid:solid, pH value) -lstirring -tsieving (time and speed) - drying-grinding

2.1.2 The Main Operation Steps 1. Washing: Partial starch should be removed by water because large amounts of starch still exist in the sweet potato residue after starch extraction. 2. Drying: Sweet potato residue is drained after washing with water and dried at 60 C. During drying, the sample should be continuously rotated to evenly heat the raw material. 3. Grinding: Sweet potato residue is weighed accurately and is ground in a grinder. The best grinding time is selected by observing the effect of different milling times on the residue’s particle size distribution. 4. Slurry mixing: A certain amount of sweet potato residue is mixed with distilled water in a certain proportion, the pH value is adjusted, and then the sample is placed on a magnetic mixer to form evenly distributed slurry. 5. Sieving: The sweet potato residue slurry is sieved first using a screen (150 μm mesh) and then is sieved using another screen (35 μm mesh) to remove starch. The material between 35 and 150 μm in size is dietary fiber. 6. Drying: The material between 35 and 150 μm is dried for 24 h to produce dietary fiber.

2.1.3 Results 2.1.3.1 The Physicochemical Composition of Sweet Potato Dietary Fiber The physicochemical composition of the sweet potato residue and dietary fiber obtained by the sieve method are shown in Table 3.1.

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Table 3.1 The Main Components in Sweet Potato Residue and Dietary Fiber Obtained by the Sieve Method (%, Dry Base) Samples

Starch

TDF

SDF

IDF

SPR

44.74 6 3.45

27.40 6 0.51

2.66 6 0.57

26.63 6 2.79

SM-SPDF

11.84 6 0.76

81.25 6 0.28

15.47 6 1.19

60.65 6 1.74

Samples

Protein

Fat

Ash

Others

SPR

5.26 6 0.12

0.38 6 0.06

2.84 6 0.19

20.01 6 3.84

SM-SPDF

3.46 6 0.28

0.22 6 0.01

2.20 6 0.06

0.60 6 0.23

Note: SPR, sweet potato residue; SM-SPDF, sweet potato dietary fiber obtained by the sieve method; TDF, total dietary fiber; SDF, soluble dietary fiber; IDF, insoluble dietary fiber.

Table 3.2 The Component Analysis of the Sweet Potato Residue and Dietary Fiber (%, Dry Base) Samples SPR SM-SPDF

Pectin

Hemicellulose

Cellulose

Lignin

7.56 6 0.88

14.68 6 0.31

43.45 6 1.45

24.94 6 0.78

13.68 6 1.87

23.28 6 1.89

35.01 6 0.25

21.59 6 0.99

Note: SPR, sweet potato residue; SM-SPDF, sweet potato dietary fiber obtained by the sieve method.

The results main components of sweet potato residue are starch and dietary fiber. The total dietary fiber (TDF) content obtained by the sieve method was 2.96-fold of the sweet potato residue, and the SDF/ IDF increased from 1/10 to 1/4 in both sweet potato residue and sweet potato dietary fiber. The starch content of the dietary fiber decreased by 73.53% compared with that of the residue, indicating that the sieve method can remove starch effectively. In addition to starch, dietary fiber, protein, fat, and ash, along with 20% of other compounds, also exist in sweet potato residue. However, these compounds accounted for only 0.6% of the dietary fiber, indicating that these compounds may be soluble sugars and small molecules, which could be removed during the sieving process. 2.1.3.2 The Component Analysis of Sweet Potato Dietary Fiber The contents of pectin, hemicellulose, cellulose, and lignin in the sweet potato residue and dietary fiber obtained by the sieve method are shown in Table 3.2. The cellulose content in sweet potato residue is the highest, followed by lignin, hemicellulose, and pectin. However, the content of cellulose decreases in sweet potato dietary fiber, as the content of pectin and hemicellulose increases. The SDF content in sweet potato dietary fiber is high and has good physiological properties. In

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addition, the high content of lignin in the sweet potato residue and sweet potato dietary fiber may be related to the sweet potato peel not being removed during the starch extraction process.

2.2 SIEVE COMBINED ENZYMATIC HYDROLYSIS METHOD 2.2.1 Technological Processes Sweet potato residue - washing - drying - grinding - slurry mixing (liquid:solid, pH value) - sieving (time and speed) - slurry mixing (liquid:solid, pH value) - gelatinization - enzymatic hydrolysis - centrifugation-precipitate collecting - drying - grinding - dietary fiber

2.2.2 The Main Operation Steps 1. Washing: Partial starch should be removed by water because large amounts of starch exist in sweet potato residue after starch extraction. 2. Drying: Sweet potato residue is drained after washing with water and dried at 55 C. During drying, the sample should be continuously rotated to evenly heat the raw material. 3. Grinding: Sweet potato residue is weighed, ground in a grinder for 30 min, and then sieved using a screen (100 μm mesh). 4. Slurry mixing: Sweet potato residue is mixed with distilled water in a certain proportion (1:60), the pH value (5.0) is adjusted, and then the sample is placed on a magnetic mixer to make evenly distributed slurry. 5. Sieving: Sweet potato residue slurries are sieved first using a screen (150 μm mesh), and then are sieved using another screen (35 μm mesh) to remove starch. The material between 35 and 150 μm is collected. 6. Slurry mixing: The material between 35 and 150 μm is evenly mixed with distilled water (1:40). 7. Gelatinization: The above slurries are gelatinized under boiling water bath for 15 min. 8. Adjusting pH value: The pH of the gelatinized slurries was adjusted to 7.0 with 1 mol/L NaOH. 9. Enzymatic hydrolysis: An aliquot of 10 mL 5% α-amylase was added and enzymatically hydrolyzed at 60 C for 1 h.

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Table 3.3 The Composition of Sweet Potato Residues and Dietary Fibers Obtained Using the Sieve Combined Enzymatic Hydrolysis Method (%, Dry Base) Composition

SPR

SMEH-SPDF

Moisture

8.48 6 0.01

8.54 6 0.01

Ash

1.42 6 0.01

1.25 6 0.01

Protein

2.75 6 0.03

4.25 6 0.01

Starch

63.27 6 0.04

2.49 6 0.03

Fat Dietary fiber

0.31 6 0.04

1.38 6 0.02

19.04 6 0.18

81.31 6 0.23

Note: SPR, sweet potato residue; SMEH-SPDF, sweet potato dietary fiber obtained using the sieve combined with the enzymatic hydrolysis method.

10. Centrifugation: The enzymatic slurry is centrifuged at 4500 3 g for 20 min to remove soluble sugars that had been obtained from the starch during the enzymatic hydrolysis process. 11. Drying and grinding: The precipitate is collected and dried at 55 C, ground by a grinder, and sieved using a screen (100 μm mesh). The dietary fiber products are thus obtained.

2.2.3 Results The results in Table 3.3 show that the main components of sweet potato residue were starch and dietary fiber, and the other compounds, such as protein, ash, and fat, were limited. In addition, some other compounds, including soluble sugars, also exist in the sweet potato residue, with contents of B7%. After sieving and enzymatic hydrolysis by α-amylase, the content of dietary fiber was above 7%, and the starch content decreased to 2.49%. The purity of the dietary fiber is superior to that of the dietary fiber obtained by the sieve method (Table 3.1), perhaps because α-amylase could remove more starch.

2.3 BIOLOGICAL METHOD 2.3.1 Single Enzyme Method The single enzyme method uses only amylase to prepare dietary fiber from sweet potato residue (Wang and Li, 1999). The sweet potato dietary fiber obtained by the single enzyme method has a light color, high purity, no odor, and loose particles. In addition, the water-swelling and water-holding capacities are higher than those of dietary fibers that are obtained by the chemical separation method.

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2.3.1.1 Technological Process Sweet potato residue - slurry mixing (liquid:solid, pH value) - enzymatic hydrolysis - centrifugation - precipitate collection - drying - grinding -dietary fiber 2.3.1.2 The Main Operation Steps 1. Washing: Partial starch should be removed by water because a large amount of starch exists in the sweet potato residue after starch extraction. 2. Slurry mixing: Sweet potato residue is mixed with distilled water in a certain proportion, the pH value (5.0) is adjusted, and then the sample is placed on a magnetic mixer for 2 min to make evenly distributed slurry. 3. Enzymatic hydrolysis: Add α-amylase and enzymatically hydrolyze at 60 C for 1 h. 4. Centrifugation: The enzymatic slurry is centrifuged at 4500 3 g for 20 min to remove soluble sugars that were obtained from starch during the enzymatic hydrolysis process. 5. Drying and grinding: The precipitate is collected and dried at 55 C, ground using a grinder, and sieved using a screen (100 μm mesh). Thus, the dietary fiber products are obtained. 2.3.1.3 Results 1. Optimized technology for dietary fiber extraction Taking into account the cost and processing requirements, the optimum conditions for the production of sweet potato dietary fiber with high temperature resistance to α-amylase were determined as follows: liquid and solid ratio of 23 mL/g, enzyme and substrate ratio of 12.5 μL/g, enzymatic time of 25 min, and enzymatic temperature 82 C. Under these conditions, the starch content in the sweet potato’s dietary fiber was 1.97%. 2. The composition of sweet potato dietary fiber obtained using the single enzyme and AOAC methods. The chemical composition of sweet potato dietary fiber obtained by the single enzyme method (TαA-SPDF) and AOAC method (AOAC-SPDF) are shown in Table 3.4. The purity of the TαASPDF was greater than 85%, which was similar to that of AOACSPDF. This result indicated that the high purity dietary fiber could be obtained without protease and amyloglucosidase. The starch contents of TαA-SPDF and AOAC-SPDF, which increased 1.97% and 1.98%, respectively, were not significantly different. The protein

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Table 3.4 The Chemical Composition of the Sweet Potato Residue, TαA-SPDF, and AOAC-SPDF (%, Dry Base) Samples

Starch

Dietary Fiber

Protein

58.88 6 2.92

a

31.03 6 1.42

b

SPR TαA-SPDF

1.97 6 0.03

b

89.64 6 2.38

a

AOAC-SPDF

1.98 6 0.02

b

89.15 6 1.91

a

Fat

2.19 6 0.13

a

0.92 6 0.06

b

0.01 6 0.01

c

Ash

0.42 6 0.02

a

1.82 6 0.04c

0.39 6 0.01

a

2.2 6 0.11b

0.39 6 0.01

a

3.48 6 0.18a

Note: Values followed by the different letters in the same line are significantly different (P , 0.05). SPR, sweet potato residue; TαA-SPDF, sweet potato dietary fiber obtained using α-amylase; AOAC-SPDF, sweet potato dietary fiber obtained using the AOAC method.

Table 3.5 The Components of the Dietary Fiber in SPR, TαA-SPDF, and AOACSPDF (%, Dry Base) Samples

Pectin

Hemicellulose

Lignin

25.02 6 1.92

a

14.04 6 2.01

a

SPR TαA-SPDF

23.12 6 2.18

b

12.78 6 1.20

b

AOAC-SPDF

23.10 6 3.02b

12.12 6 0.96b

Cellulose

23.65 6 0.89

a

37.29 6 2.72b

24.01 6 1.14

a

40.08 6 3.25a

24.67 6 1.76a

40.11 6 1.08a

Note: Values followed by the different letters in the same line are significantly different (P , 0.05). SPR, sweet potato residue; TαA-SPDF, sweet potato dietary fiber obtained using α-amylase; AOAC-SPDF, sweet potato dietary fiber obtained using the AOAC method.

in AOAC-SPDF was lower than that of TαA-SPDF, perhaps because a large amount of protein was removed by the rotease using the AOAO method. However, the ash content in AOACSPDF was higher than that of TαA-SPDF, which may be related to the additions of MES-TRIS buffer, HCl, and NaOH. 3. The dietary fiber component in TαA-SPDF and AOAC-SPDF The results in Table 3.5 show that the cellulose content in sweet potato dietary fiber was the highest (B37%
40%), followed by lignin and pectin, with contents of 24% and 23%, respectively, with the hemicellulose content being the lowest (12%
14%). In theory, the contents of pectin and hemicellulose in AOAC-SPDF were higher than that of TαA-SPDF, which, because of the alcohol precipitation, could increase the SDF content. However, the results in Table 3.5 indicate that there is no significant difference in the pectin and hemicellulose contents between AOAC-SPDF and TαA-SPDF. The enzymatic temperature used with α-amylase is lower, and the enzymatic time is shorter, which may reduce the SDF loss. However, compared with the sweet potato residue, the contents of pectin and hemicellulose in the sweet potato residue were greater than those of AOAC-SPDF and TαA-SPDF, which indicated that the loss of SDF happens during the α-amylase extraction and AOAC991.43 method.

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2.3.2 Fermentation Method Microbial fermentation is a novel method to produce dietary fiber. Wu et al. (2005) studied the effect of medicinal fungi on the extraction of dietary fiber from sweet potato residue and found during the culture medium optimization experiment that the effect of the bran content on the IDF was the greatest, followed by sweet potato residue. The optimum fermentation culture medium was 9% sweet potato residue and 0.8% bran content, which produced 29.63 g/L of dietary fiber in fermentation liquor. In the 10-L fermentation tank, the dietary fiber content in fermentation liquor was 25.81 g/L using the same fermentation conditions for 4 days. Liu (2008) extracted dietary fiber from sweet potato residue using Schizophyllum commune for solid fermentation, and the optimum fermentation conditions were 29 C for 8 days, stirring once every 8 h. The TDF content was 72.70% under the above conditions. Moreover, they optimized the solid optimum fermentation conditions, which were 4% bran content, 70% moisture content, 4-dayold bacteria, 10% inoculation amount, and a fermentation time of 16 d, by using a shaking flask. The resulting TDF content was 70.50%.

2.3.3 Mixed Enzymes Method The mixed enzymes method is also widely used to extract dietary fiber from sweet potato residue in the food industry. Yin et al. (2008) extracted dietary fiber from sweet potato residue using α-amylase and protease. Zhang et al. (2009) studied the following process for extracting dietary fiber from sweet potato residue: sweet potato residue pretreatment - grinding and sieving - hydration - pH adjustment gelatinization - enzymatic hydrolysis - alkaline hydrolysis - cooling - washing -filtration - drying - dietary fiber. In the enzymatic hydrolysis process, starch was removed by α-amylase, protein was removed by papain, and fat was removed by NaOH. The TDF content was 62.45% of the final product. Li et al. (2007) studied the following process for extracting dietary fiber from sweet potato residue: sweet potato residue - drying - grinding - washing - filtration - gelatinization - cooling - enzymatic hydrolysis inactivating enzyme activity - washing - filtration - drying - dietary fiber. The TDF content was 76.12% of the final product. Sun et al. (2008) studied the following process for extracting dietary fiber from sweet potato residue using enzymatic hydrolysis: sweet potato residue - washing - drying - grinding - soaking in NaOH enzymatic hydrolysis by amylase - enzymatic hydrolysis by trypsin

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- enzymatic hydrolysis by glucoamylase - ethanol precipitation filtration - drying - grinding - dietary fiber. The resulting TDF content was 81.43% of the final product.

2.4 CHEMICAL SEPARATION METHOD Chemical reagents are used to extract dietary fiber after the raw material is dried and ground. The main methods are water extraction and acid and alkali extraction.

2.4.1 Water Extraction The water extraction method is the easiest method for extracting SDF from sweet potato residue. Zhang et al. (2009) extracted dietary fiber from sweet potato residue as follows: sweet potato residue pretreatment - grinding and sieving - hydration - heating - cooling washing - filtration - drying - dietary fiber.

2.4.2 Acid and Alkali Extraction Acid and alkali extractions are used to extract IDF from sweet potato residue. Yu (2005) extracted dietary fiber from sweet potato residue using an acid and alkali extraction as follows: sweet potato residue - soaking in 85 C hot water - filtration - residue - soaking 14 h in pH 1.5 acid water (solid:liquid 5 1:18) - extraction for 6 h at 85 C, pH 1.5 acid water - adjusting pH to neutral - soaking 12 h in 10% NaOH solution at room temperature (solid:liquid 5 1:10) - filtration - residue washing to neutral pH - adding 5% (v/v) sodium hypochlorite adjusting pH to 4.5
5.0 by 20% (v/v) NaOH -heating at 65 C for 3 h - filtration - residue - washing by water and alcohol - drying dietary fiber. The TDF content was 96.27% of the final product.

2.4.3 Chemical Reagent in Combination with the Enzymatic Method Small amounts of protein and starch exist in the dietary fiber obtained by chemical separation. Thus, the use of a chemical reagent in combination with the enzymatic method can be used to obtain high purity dietary fiber, which requires that some kinds of enzymes (such as α-amylase and protease) should be used to remove starch and protein during the chemical separation, Liu et al. (2005) extracted dietary fiber with NaOH and α-amylase. The α-amylase content was 6 U/g, at a pH 6.2. The enzymatic time was 30 min, the concentration of NaOH

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was 1%
2%, and the extraction time was longer than 60 min. The resulting TDF content was 80.70%.

SECTION 3: THE PHYSIOLOGICAL PROPERTIES OF SWEET POTATO DIETARY FIBER With the continuous improvement in people’s living standards in China, the intake of high calorie, high protein, and high fat foods increases, while DF intake decreases, leading to a series of healthrelated diseases. DF can regulate the body’s absorption of fat, glucose, and some other nutrients and can maintain the human body’s nutritional balance. This section mainly introduces the preventive and therapeutic effects, and other biological activities, of sweet potato dietary fiber on obese rats. This provides a preliminary understanding of the benefits of sweet potato dietary fiber.

3.1 EFFECT OF SWEET POTATO DIETARY FIBER ON PREVENTING OBESITY Obesity is a harmful worldwide epidemic, the main pathogenic factor of which is adipose cell metabolic disorders. These generate inflammatory substances, thereby reducing the sensitivity of insulin and leptin in skeletal muscle and liver, and affecting lipid and carbohydrate metabolism. The characteristics of obesity are abdominal obesity, dyslipidemia, insulin resistance, and chronic inflammation (Papathanasopoulos and Camilleri, 2010). Previous studies have indicated that dietary fiber can prevent obesity and improve related symptoms (Anderson et al., 1994; Salmeron et al., 1997).

3.1.1 Effects of Sweet Potato Dietary Fiber on the Body Weights of Rats The mean body weight changes of Wistar rats in different groups (prevention group) over the experimental period are shown in Table 3.6. The body weights of the control, high-fat diet group, low-dose dietary fiber group (2%), middle-dose dietary fiber group (6%), and high-dose dietary fiber group (10%) were not significantly different from 0 to 13 days. After day 14, the body weights of the middle- and high-dose dietary fiber groups were significantly lower than the weights of the other three groups. Thus, the weight gain of rats fed by sweet potato dietary

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Table 3.6 Effects of Sweet Potato Dietary Fiber (DF) on the Body Weights of Wistar Rats (g, Prevention Group) Time (day)

A

B

C

D

Control

High-Fat diet

Low-Dose DF

Middle-Dose DF

High-Dose DF

221.80 6 1.84

a

222.30 6 1.59

a

220.20 6 1.79a

248.80 6 2.48

a

250.10 6 2.76

a

247.90 6 2.77b

222.40 6 1.70

a

249.00 6 1.99

a

E

0

218.40 6 1.63

a

3.5

245.20 6 2.12

a

7

272.37 6 2.66a

271.19 6 2.24a

272.49 6 2.59a

271.44 6 3.19a

272.33 6 2.94a

10.5

286.68 6 3.06

a

283.83 6 2.65

a

284.37 6 3.17

a

285.65 6 3.89

a

283.01 6 3.06a

14

323.01 6 3.74

a

316.41 6 3.02

b

317.58 6 3.88

b

319.25 6 4.28

ab

316.82 6 3.62b

17.5

330.58 6 4.29a

324.56 6 4.27b

324.38 6 3.04b

331.89 6 4.42a

327.38 6 3.57b

21

357.63 6 4.98

359.34 6 4.69

358.17 6 4.22

356.61 6 5.71

351.55 6 4.21b

24.5

372.69 6 5.47a

374.86 6 5.38a

364.18 6 4.87b

369.89 6 6.08a,b

369.11 6 5.34a,b

28

397.10 6 5.77

b

405.35 6 6.04

a

403.88 6 5.96

a

398.37 6 7.02

b

395.26 6 5.91c

31.5

411.20 6 6.98

b

420.80 6 7.74

a

418.66 6 7.31

a,b

413.21 6 8.79

b

406.50 6 6.92c

35

422.44 6 7.44b

424.68 6 9.38b

416.99 6 7.69c

a

a

434.58 6 8.44a

a

429.66 6 8.02a,b

ab

Note: Values followed by the different letters in the same line are significantly different (P , 0.05).

fiber was significantly slower than that of the high-fat diet group, and their weights were lower than those of the control group, especially after day 28. The body weights of the rats in the high-dose dietary fiber group was significantly lower than that in the control group and the high-fat diet group, which showed that the sweet potato dietary fiber has the effect of preventing obesity.

3.1.2 Glucose Tolerance Test The effects of sweet potato dietary fiber on blood glucose level in Wistar rats are shown in Table 3.7. The area under the blood glucose curve (AUC) 5 0.25 3 (blood glucose value at 0 h 1 4 3 blood glucose value at 0.5 h 1 3 3 blood glucose value at 2 h). The smaller the area under the glucose curve, the lower the glucose tolerance, leading to greater insulin sensitivity. The AUC value of the control was 22.24, and the value increased to 26.27 after a high-fat diet intervention (P , 0.05), which indicated that the insulin sensitivity of the rats had decreased significantly. The AUC values of the three doses (low, middle, and high) of sweet potato dietary fiber were 25.17, 23.61, and 22.34, respectively, indicating that the insulin sensitivity increased with the increase of sweet potato dietary fiber. There were no significant differences between the AUC values of the middle and high-dose groups

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Table 3.7 Effects of Sweet Potato Dietary Fiber (DF) on the Blood Glucose and AUC Values of Wistar Rats (Prevention Group) Time (h)

Blood Glucose Value (mmol/L) Control 6.23 6 0.52

c

0.5

15.83 6 0.47

d

1

12.20 6 0.33c

2

0

AUC

6.47 6 0.84

d

22.24 6 1.00

b

High-Fat Diet

Low-Dose DF

7.27 6 0.69 18.6 6 0.93

a

a

14.8 6 1.22a

6.80 6 0.49

b

17.87 6 0.98

b

Middle-Dose DF

High-Dose DF

6.53 6 0.68

c

6.27 6 0.46d

16.67 6 1.27

c

15.80 6 0.82d

14.43 6 1.02a

13.40 6 0.74b

12.27 6 0.67c

7.80 6 0.57

a

7.47 6 0.37

b

7.07 6 0.69

6.63 6 0.24d

26.27 6 1.25

a

25.17 6 1.13

a

23.61 6 1.60

c

22.34 6 0.91b

ab

Note: Values followed by the different letters in the same line are significantly different (P , 0.05).

Table 3.8 Effects of Sweet Potato Dietary Fiber (DF) on Blood Indices of Wistar Rats (Prevention Group) Index

A

B

Control

C

D

High-Fat Diet

Low-Dose DF

10.27 6 0.49

9.80 6 0.37

E

Middle-Dose DF

High-Dose DF

b

9.53 6 0. 86

9.27 6 0.44d

Blood glucose (mmol/L)

9.23 6 0.32

TG (mg/mL)

53.53 6 1.07d

89.25 6 2.14a

84.42 6 2.12b

80.47 6 1.68c

80.63 6 2.02c

TC (mg/mL)

74.18 6 1.42d

122.29 6 2.06a

123.07 6 1.26a

114.86 6 1.48b

95.73 6 0.98c

HDL (mg/mL)

55.62 6 1.06

47.47 6 0.79

50.03 6 0.98

52.70 6 1.18

54.03 6 0.84b

d

a

a

e

d

c

c

Note: Values followed by the different letters in the same line are significantly different (P , 0.05). TG, triglyceride; TC, total cholesterol; HDL, high density lipoprotein cholesterol.

and the control rats, indicating that sweet potato dietary fiber had a significant effect on improving the insulin sensitivity of Wistar rats.

3.1.3 Blood Index Determination 3.1.3.1 The Blood Glucose and Blood Lipid Concentrations The blood indices of different Wistar rats are shown in Table 3.8. The concentrations of blood glucose (10.27 mmol/L) significantly increased after rats were fed on a high-fat diet compared with those of the control (9.23 mmol/L). In the glucose metabolism of rats fed a high-fat diet, the blood glucose sensitivity decreased, and, therefore, the body could not control the blood glucose concentration. However, after feeding rats different doses of sweet potato dietary fiber, the blood glucose levels decreased, and there was no significant difference between the blood glucose levels of the rats in the high-dose dietary fiber group and the control. Wang et al. (2007) found that a high-fat diet containing 10% plantain and 10% sugarcane dietary fiber could reduce the fasting

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143

blood glucose and fasting insulin levels in mice. Li and Wang (2010) also showed that wheat dietary fiber could decrease the fasting blood glucose concentration of mice and increase their glucose tolerance. The concentrations of TG and total cholesterol (TC) in the control group were 53.53 and 74.18 mg/mL, respectively, while the concentrations of TG and TC in the high-fat diet group were 89.25 and 122.29 mg/mL, respectively. A comprehensive comparison of TG, TC, and high density lipoprotein (HDL) in rats indicated that the TC and TG contents in blood increased significantly after rats were fed a high-fat diet, indicating that the obvious characteristics of obesity existed in these rats. HDLs are able to make the “bad” cholesterol return to the liver, thus avoiding obstruction in the blood vessels. Thus, the HDL concentration in rats fed a highfat diet was reduced, while the HDL concentration in rats was increased by adding sweet potato dietary fiber to the high-fat diet. With increases in the sweet potato dietary fiber doses, the HDL contents in the blood increased gradually, but the HDL contents in rats of the high-dose dietary fiber group were still less than those in the control. 3.1.3.2 The Plasma Leptin and Insulin Concentrations The concentrations of plasma leptin are shown in Fig. 3.1A. The concentrations of plasma leptin increased from 5.61 to 6.32 ng/mL after feeding rats a high-fat diet. Thus, leptin sensitivity occurred in Wistar rats after the high-fat diet intervention, and a lower concentration of plasma leptin cannot effectively regulate rat weight. The concentrations of plasma leptin decreased with the increase in the sweet potato dietary fiber dose, indicating that sweet potato dietary fiber can improve leptin sensitivity, and the sensitivity increased significantly with the dosage increase. The concentrations of plasma insulin are shown in Fig. 3.1B. The concentrations of insulin was lowest in the control group, and the insulin level increased significantly after a high-fat diet intervention, indicating that the insulin sensitivity decreased after feeding a high-fat diet. After treatment with 2% sweet potato dietary fiber, the plasma insulin concentration decreased, but not significantly. After intakes of 6% and 10% sweet potato dietary fiber, the plasma insulin concentrations were significantly lower than that of the high-fat diet group, showing that sweet potato dietary fiber can increase the insulin sensitivity of rats. The insulin sensitivity of the high-dose dietary fiber group was the highest, but was not significantly different from the control group. Ylonen et al. (2003)

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8

Plasma leptin˄ng/ml˅

7

a c

6

b

bc

C Low dose DF

D Middle dose DF

d

5 4 3

2 1 0

A Control

B High-fat diet group

E High dose DF

(A)

Plasma insulin˄uIU/ml˅

18 a

16 14

ab b

c

c

12 10 8 6 4 2 0

A Control

B High-fat diet group

C Low dose D Middle dose E High dose DF DF DF

(B) Figure 3.1 The effects of sweet potato dietary fiber (DF) on the plasma leptin (A) and insulin (B) levels in Wistar rats (prevention group). Note: Different letters indicate significant differences (P , 0.05).

determined that there was a negative correlation between the doses of TDF, SDF, and IDF and the insulin level. Choi et al. (2010) found that the addition of 2% or 4% β-glucan increases the glucose tolerance of mice and decreases their serum insulin level. 3.1.3.3 The Concentration of Plasma Adiponectin and GLP-1 Fig. 3.2A shows the levels of plasma adiponectin in Wistar rats. Adiponectin is an endogenous bioactive polypeptide or protein secreted by adipocytes. Adiponectin is a hormone that can improve insulin resistance and atherosclerosis in rats. In Fig. 3.2, the level of plasma adiponectin was highest in the control group, and the level of

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145

18

Adiponectin ˄mg/L˅

16

a b

14

c

b

D Middle dose DF

E High dose DF

d

12 10 8 6 4 2 0

A Control

B High-fat C Low dose diet group DF

Glucagon like peptide-1˄pmol/L˅

(A) 30

a

25

20

bc

b

c

C Low dose DF

D Middle dose DF

E High dose DF

d

15 10 5 0

A Control

B High-fat diet group

(B) Figure 3.2 The effects of sweet potato dietary fiber (DF) on the level of plasma adiponectin (A) and glucagonlike peptide-1 (B) in Wistar rats (prevention group). Note: Different letters indicate significant differences (P , 0.05).

plasma adiponectin was lowest after the intake of a high-fat diet. The plasma adiponectin levels increased slightly after the intake of lowand middle-doses of sweet potato dietary fiber, but there was no significant difference between the two groups. The plasma adiponectin level increased in the high-dose dietary fiber group, but there were still some differences compared with the control. Galisteo et al. (2005) found that psyllium peel, containing a high amount of SDF, could increase the circulating levels of adiponectin. Similarly, Galisteo et al. (2010) also found that rats fed psyllium peel and methyl cellulose had increased secretory levels of adiponectin.

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Fig. 3.2A shows the levels of plasma GLP-1 in Wistar rats. GLP-1 has a stimulating effect on the release of insulin after meals and can increase glucose tolerance, thus delaying gastric emptying. Fig. 3.2A shows that the level of GLP-1 was low in the control rats, but it increased after being fed a high-fat diet. The level of GLP-1 increased with the increase in the sweet potato dietary fiber dose. Delzenne and Cani (2010) showed that dietary fiber might increase the number of L-cells in the intestine, thus increasing the amount of GLP-1, which is consistent with the conclusion of this study. 3.1.3.4 The Plasma Inflammatory Factor Concentrations The concentration of plasma inflammatory factors is shown in Table 3.9. The FFA level was 8.09 mmol/mL in control group, while it increased to 11.31 mmol/mL after treatment with a high-fat diet, indicating that a certain chronic inflammation occurred in rats fed a highfat diet. The level of FFA was highest in the high-dose dietary fiber group, which was lower than that of the middle-dose dietary fiber group, indicating that sweet potato dietary fiber can reduce metabolic disorders associated with inflammation factors induced by obesity. TNF-α is an important marker of chronic inflammation. The level of TNF-α was 0.88 ng/mL in the control group, and it increased to 1.31 ng/mL after rats were fed a high-fat diet. After the intake of different doses of sweet potato dietary fiber, the level of TNF-α decreased. The level of C-reactive protein (CRP) decreased gradually with an increase in the sweet potato dietary fiber dose, but there was no significant difference between the high-fat diet group and the control. IL-6 is also an important inflammatory factor of the body. The level of IL-6 increased significantly after the intake of a high-fat diet, and there was a linear relationship between the IL-6 level and sweet Table 3.9 Effects of Sweet Potato Dietary Fiber (DF) on Inflammatory Factors of Wistar Rats (Prevention Group) Index

A

B

C

D

E

Control

High-Fat Diet

Low-Dose DF

Middle-Dose DF

High-Dose DF

FFA (mM/mL)

8.09 6 0.32d

11.31 6 0.14a

10.57 6 0.22a,b

10.15 6 0.16b

9.62 6 0.26c

TNF-α (ng/mL)

0.88 6 0.20

c

1.31 6 0.19

a

1.28 6 0.09

a

1.03 6 0.08

b

0.97 6 0.16d

CRP (mg/L)

2.63 6 0.07

c,d

2.88 6 0.13

b

3.04 6 0.18

a

2.70 6 0.07

c

2.55 6 0.12d

136.28 6 4.74b

130.66 6 3.67c

IL-6 (pg/mL)

110.38 6 3.33c

150.19 6 5.22a

141.69 6 2.02a,b

Note: Values followed by the different letters in the same line are significantly different (P , 0.05). FFA, free fatty acid; TNF-α, tumor necrosis factor-α; IL-6, interleukin-6.

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147

potato dietary fiber dose. The rats exhibited the lowest IL-6 level after treatment with a high-dose of sweet potato dietary fiber. Dietary fiber can reduce biomarkers associated with inflammation, including FFA, CRP, IL-6, and TNF-α, or reduce their biological activity (Galisteo et al., 2005).

3.1.4 Determination of Liver Indices 3.1.4.1 The Concentrations of TG and TC in Liver The TG concentrations in different Wistar rats are shown in Fig. 3.3A. The TG level of the control rats was 202.04 mg/mL, and after a

Liver triglycerides˄mg/mL˅

700

a

b

600 500 c

d

D Middle dose DF

F High dose DF

400 300

e 200 100 0

A Control

B High-fat C Low dose diet group DF

(A)

Liver total cholesterol˄mg/mL˅

250 a b 200 150 100 50

0

c

d

D Middle dose DF

F High dose DF

e

A Control

B High-fat C Low dose diet group DF

(B) Figure 3.3 The effect of sweet potato dietary fiber on the level of liver triglycerides (A) and total cholesterol (B) in Wistar rats (prevention group). Note: Different letters indicate significant differences (P , 0.05).

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high-fat diet treatment, the TG level increased to 617.15 mg/mL, which indicated that the liver lipid content increased after the intake of a high-fat diet, leading to the occurrence of fatty liver. Compared with the high-fat diet group, the level of TG in the low-dose dietary fibertreated rats showed no significant changes, indicating that a low-dose of sweet potato dietary fiber could not significantly decrease the TG content. However, after independent treatments with middle- and high-doses of sweet potato dietary fiber, the TG level decreased significantly, and the content of TG in the high-dose group was the lowest. This showed that sweet potato dietary fiber could significantly reduce the content of liver fat. The TC concentrations in different Wistar rats are shown in Fig. 3.3B. The TC level of the control rats was 36.00 mg/mL, and after a high-fat diet treatment, the TC level increased to 206.99 mg/mL. After independent treatments of low and middle doses of sweet potato dietary fiber, the TC level was 190.92 and 83.62 mg/mL, respectively, but there was no significant between the middle- and high-dose groups, indicating that 6% sweet potato dietary fiber could significantly reduce the TC content of liver.

3.1.4.2 Pathological Analysis of Liver Pathological rat liver sections from different experimental groups are shown in Fig. 3.4. Fig. 3.4A shows that the liver in the control group had normal tissue structures, and had no obvious pathological abnormalities. However, pathological sections of liver from the high-fat diet group showed that a large area of stem cell fatty degeneration appeared after the intake of a high-fat diet, and some white fat particles existed in the liver tissue, which had reached a stage of moderate cellular degeneration. After low-dose sweet potato dietary fiber treatment, the liver showed mild degeneration, and the amount of fat particles was significantly lower than that of rats in the high-fat diet group. After middle-dose sweet potato dietary fiber treatment, the degree of liver degeneration further decreased, and the amount of fat particles also further decreased. In the high-dose sweet potato dietary fiber group, little liver degeneration and few fat particles were observed. Thus, 10% sweet potato dietary fiber could prevent the fatty liver induced by a high-fat diet.

Figure 3.4 The liver pathology of Wistar rats (prevention group). DF, dietary fiber. (A) Control, (B) high-fat diet group, (C) low-dose DF, (D) middle-dose DF, and (E) high-dose DF.

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3.1.5 The Determination of the Organic Acid Content in Cecum Dietary fiber can be partially or totally fermented, and decomposed into SCFAs, in the cecum of humans and animals. The physiological properties of dietary fiber depend on the type and quantity of sweet potato dietary fiber, and SCFA can stimulate the absorption of water and sodium in the intestines, affect pH, and bile salt precipitation (Pylkas et al., 2005). The effects of sweet potato dietary fiber on the SCFA content of the cecum are shown in Table 3.10. Table 3.10 shows that the level of lactic acid decreased significantly after the intake of a high-fat diet, but there was no set trend between the high-fat diet group and the sweet potato dietary fiber groups. However, we can still conclude that the lactic acid content in obese rats was lower than that of control rats. The acetic acid and propionic acid contents were 2671.88 and 891.17 μg/mL in the control group, respectively; these contents increased to 2922.25 and 1356.43 μg/mL after the high-fat diet treatment, and the contents of acetic acid and propionic acid decreased in a linear trend with the increase in the sweet potato dietary fiber dose. In addition, the content of butyric acid was 829.90 μg/mL in the control group. The butyric acid content decreased significantly after the intake of a high-fat diet, and the content decreased in a linear trend with the increase in the sweet potato dietary fiber. These results indicated that the SCFA content could be used to evaluate obesity.

Table 3.10 Effect of Sweet Potato Dietary Fiber (DF) on the SCFA Content of the Cecum (Prevention Group) Index

A

(μg/mL)

Control

B

C

D

E

High-Fat Diet

Low-Dose DF

Middle-Dose DF

High-Dose DF

Group Lactic acid

407.83 6 16.28a

129.58 6 10.37c

196.09 6 18.68b

105.30 6 6.42c

Acetic acid

2671.88 6 56.38c

2922.25 6 26.84a

2828.18 6 18.28b

2800.53 6 15.83b

2670.87 6 17.67c

Propionic acid

891.17 6 17.19d

1356.43 6 35.16a

1158.48 6 33.14b

1133.76 6 18.84b

936.62 6 22.32c

Butyric acid

829.90 6 14.35a

315.35 6 19.44e

425.87 6 15.90d

475.83 6 25.35c

594.15 6 15.87b

160.07 6 16.24bc

Note: Values followed by the different letters in the same line are significantly different (P , 0.05).

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3.2 EFFECTS OF SWEET POTATO DIETARY FIBER ON TREATING OBESITY 3.2.1 Effects of Sweet Potato Dietary Fiber on the Body Weights of Rats Male Wistar rats were treated with a high-fat diet from day 1 to 45, and an obesity model was established after 45 days, and the body weight, plasma TC, TG, and HDL cholesterol (HDL-C) of every rat were determined. The rats were divided randomly into five groups. The average initial weight of the rats was 455.87 g, which increased to 606.7 g after the intake of a high-fat diet for 45 days. The weight gain was lower in the low-dose sweet potato dietary fiber group than in the high-fat diet group. The weight gain of the middle- and high-dose sweet potato dietary fiber groups was reduced significantly compared to that of the high-fat diet group, and the decrement was positively correlated to the amount of sweet potato dietary fiber. This result indicated that sweet potato dietary fiber can effectively treat obesity caused by a high-fat diet. Simvastatin has obvious effects on lowering body weights and blood lipid levels, and the results showed that the body weight of rats treated with simvastatin was significantly lower than those of the high-fat diet group, and middle- and low-dose dietary fiber groups, but there was no significant difference between the simvastatin group and the high-dose sweet potato dietary fiber group. These results indicated that 10% sweet potato dietary fiber could effectively decrease body weight.

3.2.2 Glucose Tolerance Test The average AUC value of Wistar rats in the high-fat diet group was 26.27, and the value increased to 32.91, 30.70, and 28.59 after treatment of 2%, 6%, and 10% sweet potato dietary fiber, respectively, indicating that sweet potato dietary fiber can improve insulin sensitivity and that the increase of insulin sensitivity depended linearly on the amount of sweet potato dietary fiber. The average AUC values were not significantly different between the rats treated with middle or high doses of sweet potato dietary fiber and the rats treated with simvastatin (29.26), indicating that the effect of sweet potato dietary fiber on improving the insulin sensitivity of rats was equivalent to the effect of the western medicine simvastatin (Table 3.11).

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Table 3.11 Effects of Sweet Potato Dietary Fiber (DF) on the Blood Glucose and AUC Values of Wistar Rats (Treatment Group) Time (h)

Blood Glucose Value (mmol/L) High-Fat Diet

Low-Dose DF

Group 10.44 6 1.24

Middle-Dose

High-Dose

DF

DF

Group

10.20 6 1.92

b

9.76 6 0.68

c

21.6 6 0.93

a

20.07 6 0.98

b

18.67 6 1.27

c

18.80 6 0.82c

15.8 6 1.22

a

14.43 6 1.02

b

14.40 6 0.74

b

14.27 6 0.67b

9.97 6 0.69c

10.63 6 0.24b

0

10.83 6 1.39

a

0.5

21.83 6 0.47

a

1

16.20 6 0.33

a

2

12.47 6 0.84a

11.60 6 0.57a,b

10.77 6 0.37b

AUC

33.89 6 1.18

32.91 6 1.36

30.70 6 1.42

a

Simvastatin

a,b

a

28.59 6 1.60

bc

9.93 6 0.46b,c

29.26 6 0.91c

c

Note: Values followed by the different letters in the same line are significantly different (P , 0.05).

Table 3.12 Effects of Sweet Potato Dietary Fiber (DF) on the Blood Indices of Wistar Rats (Treatment Group) A

B

C

D

E

High-Fat Diet

Low-Dose DF

Middle-Dose

High-Dose

Simvastatin

DF

DF

Group 10.44 6 1.24

10.20 6 1.92

9.76 6 0.68

Group 9.93 6 0.46b,c

Blood glucose (mmol/L)

10.83 6 1.39

TG (mg/mL)

88.74 6 9.36a

84.22 6 2.48a

70.06 6 6.40b

60.36 6 4.22c

60.69 6 3.68c

TC (mg/mL)

107.88 6 6.74a

102.67 6 9.27a

92.09 6 6.09b

86.09 6 8.39c

90.38 6 6.28b

45.34 6 3.26

46.86 6 4.97

47.29 6 3.14

48.17 6 2.98

52.97 6 3.45a

HDL(mg/mL)

a

c

a,b

b

b

b

c

b

Note: Values followed by the different letters in the same line are significantly different (P , 0.05). TG, triglyceride; TC, total cholesterol; HDL, high density lipoprotein cholesterol.

3.2.3 Determination of Blood Indices 3.2.3.1 The Blood Glucose and Blood Lipid Concentrations The blood indices of different Wistar rats are shown in Table 3.12. The blood glucose level of the high-fat diet group was 10.83 mmol/L, and there was no significant difference between the high-fat diet group and low-dose dietary fiber group. The average blood glucose levels in middle- and high-dose rats were 10.20 and 9.76 mmol/L, respectively, and these values were lower to some degree than the high-fat group, indicating that 6% and 10% sweet potato dietary fiber could alleviate the abnormal glucose metabolism of obese rats. The glucose sensitivity was improved, and the body could regulate fasting blood glucose values through a more efficient glycemic regulation system. Similarly, after a simvastatin treatment, the blood glucose value decreased to

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some degree, indicating that simvastatin also had a certain role in the regulation of fasting blood glucose. The TG and TC concentrations in the high-fat diet group were 88.74 and 107.88 mg/mL, respectively, and there were no significant differences with the low-dose sweet potato dietary fiber group, indicating that the low dose of sweet potato dietary fiber was not enough to lower the TG and TC levels in the blood. However, the TG and TC levels in the middle- and high-dose sweet potato dietary fiber groups decreased significantly, and the decrement was positively related to the amount of sweet potato dietary fiber, indicating that 6% and 10% sweet potato dietary fiber could significantly reduce the TG and TC levels. The HDL-C level in the high-fat diet group was the lowest at 45.34 mg/mL. The HDL-C level increased with the increase in the sweet potato dietary fiber dose, but there was a significant difference between the high-dose dietary fiber group and the simvastatin group. 3.2.3.2 Plasma Leptin and Insulin Concentrations The concentrations of plasma leptin are shown in Fig. 3.5A. The plasma leptin concentration in the high-fat diet group was the highest, indicating that the leptin sensitivity was the lowest. However, higher concentrations of leptin could regulate the body’s metabolism. The concentration of plasma leptin was higher in the low-dose sweet potato dietary fiber group, but it was not significantly different from the highfat diet group, indicating that 2% sweet potato dietary fiber was not enough to regulate leptin sensitivity. However, the leptin level decreased significantly after middle- and high-dose sweet potato dietary fiber treatments, which showed that higher amounts of sweet potato dietary fiber could regulate leptin sensitivity. The rats that were fed a high-dose of sweet potato dietary fiber exhibited the greatest leptin sensitivity. Galisteo et al. (2005) found that psyllium peel containing SDF could increase the leptin sensitivity in obese Zucker rats. The concentrations of plasma insulin are shown in Fig. 3.5B. The plasma insulin level was highest in the high-fat diet and low-dose sweet potato dietary fiber groups, indicating that the insulin sensitivity of these two groups was the lowest, However, after independent treatments of 6% and 10% sweet potato dietary fiber, the insulin levels decreased significantly, and there was no significant difference between the high-dose dietary fiber and simvastatin groups, which indicated

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8

a

a

b

Leptin (ng/mL)

7

d

c

6 5 4

3 2 1

0

A High-fat B Low dose diet group DF

C Middle dose DF

(A) 18

a

16

b

c d

14 Insulin (µIU/mL)

D High dose E DF Simvastatin group

e

12 10 8 6

4 2 0

A High-fat diet group

B Low dose DF

C Middle dose DF

D High dose E Simvastatin DF group

(B) Figure 3.5 The effects of sweet potato dietary fiber (DF) on the plasma leptin (A) and insulin (B) level in Wistar rats (treatment group). Note: Different letters indicate significant differences (P , 0.05).

that both sweet potato dietary fiber and simvastatin could improve the insulin sensitivity of rats. 3.2.3.3 The Plasma Adiponectin and GLP-1 Concentrations Fig. 3.6A shows the level of plasma adiponectin in Wistar rats. The level of adiponectin in the high-fat diet group was the lowest, while, after independent treatments of 6% and 10% sweet potato dietary fiber, this value increased, and the increment was positively related to the amount of sweet potato dietary fiber. The level of adiponectin in the simvastatin group was equivalent to that of the middle-dose dietary fiber group, which indicated that the promoting effect of simvastatin

Sweet Potato Dietary Fiber

a

Adiponectin˄mg/L˅

16

b

c

e

155

d

12

8

4

0

A High-fat B Low dose diet group DF

C Middle dose DF

D High dose E DF Simvastatin group

Glucagon-like peptide-1˄pmol/L˅

(A) 35 a 30 25

b

c

d e

20 15

10 5 0

A High-fat B Low dose diet group DF

C Middle dose DF

D High dose E DF Simvastatin group

(B) Figure 3.6 The effects of sweet potato dietary fiber (DF) on the plasma adiponectin (A) and glucagon like peptide-1 (B) levels in Wistar rats (treatment group). Note: Different letters indicate significant differences (P , 0.05).

on the secretion of adiponectin in adipose tissue is not as great as that of the high-dose of sweet potato dietary fiber. Fig. 3.6B shows the level of plasma GLP-1 in Wistar rats. The level of GLP-1 in the high-fat diet group was the lowest, while, after independent treatments of 6% and 10% sweet potato dietary fiber, this value increased, which showed that higher amounts of sweet potato dietary fiber could stimulate the release of GLP-1. The level of GLP-1 in the simvastatin group increased, and the effect was equivalent to that of 10% sweet potato dietary fiber.

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Table 3.13 Effects of Sweet Potato Dietary Fiber (DF) on Inflammatory Factors of Wistar Rats (Treatment Group) Index

A

B

C

D

E

High-Fat Diet

Low-Dose DF

Middle-Dose

High-Dose

Simvastatin

Group

DF

DF

Group

FFA (mM/mL)

14.09 6 0.24

a

14.31 6 0.16

a

13.57 6 0.22

b

12.45 6 0.14

c

TNF-α (ng/mL)

1.38 6 0.20

a

1.19 6 0.17

b

1.09 6 0.08

c

0.92 6 0.04

c

0.92 6 0.16c

CRP (mg/L)

2.26 6 0.17

b

2.31 6 0.12

ab

2.38 6 0.16

a

2.21 6 0.17

b

2.43 6 0.14a

122.74 6 4.47c

123.30 6 3.62c

IL-6 (pg/mL)

142.94 6 2.33a

141.92 6 5.52a

131.86 6 2.20b

12.62 6 0.24c

Note: Values followed by the different letters in the same line are significantly different (P , 0.05). FFA, free fatty acid; TNF-α, tumor necrosis factor-α; IL-6, interleukin-6.

3.2.3.4 The Plasma Inflammatory Factor Concentrations The concentrations of plasma inflammatory factors are shown in Table 3.13. The FFA level was 14.09 mmol/mL in the high-fat diet group, while this value decreased after independent treatments of middle- and high doses of sweet potato dietary fiber, indicating that a higher amount of sweet potato dietary fiber could reduce the inflammatory factor secretion disorder caused by obesity, and the effect of the high dose of sweet potato dietary fiber was equivalent to the effect of simvastatin. TNF-α is an important marker of chronic inflammation. Sweet potato dietary fiber could decrease the level of plasma TNF-α, and this effect was positively related to the amount of sweet potato dietary fiber. The same conclusion, that sweet potato dietary fiber could reduce the metabolic disorder caused by obesity, could be drawn from the analysis of the IL-6 levels in the plasma,

3.2.4 Determination of Liver Indices 3.2.4.1 The TG and TC Concentrations of Liver The TG concentrations in different Wistar rats are shown in Fig. 3.7A. The liver TG level was the highest (786.66 mg/mL) in rats fed a highfat diet, indicating that they exhibit metabolic abnormalities in their liver lipids. The TG level in the low-dose sweet potato dietary fiber group was less than the high-fat diet intervention, and the decrement was positively related to the amount of sweet potato dietary fiber. The TG levels in the low-, middle- and high-dose dietary fiber groups were 702.83, 612.81, and 506.31 mg/mL, respectively. Thus, sweet potato dietary fiber had a significant effect on reducing liver TG levels, and 10% sweet potato dietary fiber had a better effect than simvastatin.

Sweet Potato Dietary Fiber

157

900

a

Liver triglyceride˄mg/mL˅

800

b

700

c

d

600

e

500

400 300 200 100 0 A High-fat B Low dose diet group DF

C Middle dose DF

D High dose E DF Simvastatin group

(A) Liver total cholesterol˄mg/mL˅

250 a

b

200

c

150

d e

100 50 0

A High-fat diet group

B Low dose DF

C Middle dose DF

D High dose E DF Simvastatin group

(B) Figure 3.7 The Effects of Sweet Potato Dietary Fiber (DF) on the Liver Triglycerides (A) and Total Cholesterol (B) Level in Wistar Rats (Treatment Group). Note: Different letters indicate significant differences (P , 0.05).

Similarly, the liver TC content in the high-fat diet group was the highest (202.73 mg/mL) (Fig. 3.7B), which showed that the liver’s cholesterol metabolism in the high-fat diet group was abnormal. After treatment with different doses (low, middle, and high) of sweet potato dietary fiber, the liver TC of rats decreased to 178.16, 128.78, and 92.92 mg/mL, respectively, indicating that sweet potato dietary fiber could effectively reduce the TC content of the liver. 3.2.4.2 Pathological Analysis of Liver Pathological rat liver sections from different experimental groups are shown in Fig. 3.8. The pathological sections of livers from the high-fat

Figure 3.8 The pathology of livers from Wistar rats (treatment group); DF, dietary fiber. (A) High-fat diet group, (B) low-dose DF, (C) middle-dose DF, (D) high-dose DF, and (E) simvastatin group.

Sweet Potato Dietary Fiber

159

Table 3.14 Effects of Sweet Potato Dietary Fiber (DF) on the SCFA Content in the Cecum (Treatment Group) Index

A

B

C

D

(μg/mL)

High-Fat Diet

Low-Dose DF

Middle-Dose DF

High-Dose DF

Group

E Simvastatin Group

Lactic acid

405.26 6 16.42b

448.40 6 38.34a

446.50 6 24.42a

414.83 6 8.79a

406.90 6 18.64a

Acetic acid

2931.83 6 42.11a

2861.54 6 28.69b

2767.56 6 36.28c

2643.58 6 34.48d

2637.66 6 20.08d

Propionic acid

1130.07 6 17.64a

1111.17 6 22.88a

1070.01 6 12.69b

986.82 6 14.57c

873.54 6 18.99d

Butyric acid

1191.47 6 29.17a

870.15 6 9.46b

823.34 6 17.54c

817.28 6 26.37c

897.95 6 16.21a,b

Note: Values followed by the different letters in the same line are significantly different (P , 0.05).

diet group showed that a large area of stem cell fatty degeneration appeared after the intake of a high-fat diet, and some white fat particles existed in the liver tissue, which had reached a stage of severe cellular degeneration (Fig. 3.8A). After treatment with low-dose sweet potato dietary fiber, the liver showed mild liver degeneration, and the amount of fat particles was significantly lower than in rats from the high-fat diet group (Fig. 3.8B). After treatment with middle-dose sweet potato dietary fiber, the degree of liver degeneration further decreased, and the amount of fat particles also further decreased (Fig. 3.8C). In the high-dose sweet potato dietary fiber group, little liver degeneration and few fat particles were observed (Fig. 3.8D). These results indicated that sweet potato dietary fiber could alleviate fatty liver induced by a high-fat diet. Fig. 3.8C showed that rat livers still exhibited a mild cellular degeneration and that the amount of fat particles was higher than that of the high-dose sweet potato dietary fiber group after treatment with simvastatin. Based on liver pathology, the diet containing 10% sweet potato dietary fiber had better fatty liver therapeutic effects than simvastatin.

3.2.5 The Determination of the Organic Acid Content in Cecum The effects of sweet potato dietary fiber on the SCFA content of the cecum are shown in Table 3.14. The table shows that the lactic acid levels were not significantly different among the groups. Therefore, we could not determine obesity symptoms using the lactic acid content. However, the acetic acid level was highest in the high-fat diet group at 2931.83 μg/mL, and the value decreased with an increase in sweet potato dietary fiber. In addition, the acetic acid value was not

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significantly different between the high-dose dietary fiber group (10%) and the simvastatin group. Therefore, we concluded that sweet potato dietary fiber can reduce the acetic acid content in obese rats. We also found that the rats in the high-fat diet and low-dose dietary fiber groups exhibited the highest propionic acid contents, with values of 1130.07 and 1111.17 μg/mL, respectively, and there was a negative correlation between the propionic acid content and dietary fiber dose. Similarly, we also found that the rats in the high-fat diet group exhibited the highest butyric acid level. The value decreased gradually with increases in the sweet potato dietary fiber dose, and the butyric acid content in high-dose sweet potato dietary fiber group was equivalent to that of the simvastatin group.

3.3 OTHER FUNCTIONAL PROPERTIES OF SWEET POTATO DIETARY FIBER 3.3.1 Protecting the Intestines From Cancer A main cause of intestinal cancer is irritating or toxic substances that remain in the intestine for a long period of time. Sweet potato dietary fiber, which exhibits a high water-holding capacity, can increase feces volume and increase the bowel movement rates in the human body, being beneficial to prevent intestinal cancer.

3.3.2 The Absorption of Cholesterol Sweet potato dietary fiber can reduce the cholesterol concentration in the body because sweet potato dietary fiber has a high molecular weight. Thus, it can form a network structure in some solutions, allowing it to absorb bile acids and salt materials. These substances are combined with dietary fiber in the small intestine, and then discharged with the feces. Thus, extra cholesterol in the body is converted into bile acid to compensate for the discharged amount. This reduces the cholesterol concentration.

3.3.3 The Absorption of Na1, K1, Ca21, Fe31, and Pb21 In 1981, Sentenac proposed that there are two interaction mechanisms between dietary fiber and cations. The first mechanism is an electrostatic interaction, which is constrained by Boltzmann’s law and is influenced by ion valence. The second mechanism is an intrinsic affinity that exists between anionic groups in the dietary fiber system and cation groups in the solution system. Thus, anions and cations can

Sweet Potato Dietary Fiber

161

form ion pairs and share the same molecular orbital. This chemical behavior follows the law of mass action, and, therefore, it affects the metabolism of some minerals.

3.3.4 Accelerating the Output Speed of Feces Constipation is also known as dry stool. In general, constipation refers to a stool interval of more than 48 h, resulting in dry feces. Chymus in the cecum is converted into feces and then arrives at the end of the colon, which may take 12
14 h. Crude fiber and some edible dietary fiber can expand the colon’s volume, and absorb and retain moisture, softening the feces and stimulating the secretion of digestive enzymes. This accelerates the feces’ output speed.

SECTION 4: THE PHYSICOCHEMICAL PROPERTIES OF SWEET POTATO DIETARY FIBER The physicochemical properties of sweet potato dietary fiber include the water-holding capacity, water-swelling capacity, oil-holding capacity and viscosity. In general, the physicochemical properties of dietary fiber are not only closely related to its source, composition ratio and particle size, but are also affected by the temperature, pH, ionic strength and other factors (Grigelmo-Miguel and MartínBelloso, 1998). The effects of different factors (including temperature, pH value, and salt concentration) on the water-holding capacity, water-swelling capacity, oil-holding capacity, and viscosity of sweet potato dietary fiber are analyzed and compared in this section.

4.1 EFFECTS OF DIFFERENT FACTORS ON THE WATERHOLDING CAPACITY OF SWEET POTATO DIETARY FIBER Temperature, pH value, and salt concentration have effects on the waterholding capacity. Fig. 3.9 shows that the water-holding capacity of sweet potato dietary fiber was 8.23 g/g when the temperature was 20 C, and its value increased with the rising temperature. The water-holding capacity reached its highest level (1.106 g/g) when the temperature was 70 C. The water-holding capacities of the other three materials, sweet potato residue, sweet potato powder, and soybean dietary fiber, increased with the increase in temperature. However, a large amount of starch exists in sweet

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Water holding capacity˄ g/g ˅

14

Sweet potato residue

Sweet potato powder

Soybean dietary fiber

Sweet potato dietary fiber

12 10 8 6 4 2 0

20

30

40

50

60

70

80

90

100

Temperature˄°C˅ Figure 3.9 Effects of temperature on the water-holding capacity (g/g) of sweet potato dietary fiber.

potato residue and sweet potato powder, and starch can absorb water and swell, leading to higher water-holding capacities. Water-holding capacities further increased with the temperature. The water-holding capacity decreased from 8.13 to 7.14 g/g when the pH value changed from 3 to 11. The water-holding capacity changed little when the NaCl concentration increased from 0% to 10%, and this value was lower when the NaCl concentration was 1% (Figs. 3.10 and 3.11).

4.2 EFFECTS OF DIFFERENT FACTORS ON THE WATERSWELLING CAPACITY OF SWEET POTATO DIETARY FIBER Dietary fiber can absorb water, increasing its weight and volume. The effects of the external temperature, pH value, and salt concentration on the water-swelling capacity of sweet potato dietary fiber were studied, and the results showed that the water-swelling capacity was influenced by temperature and pH value. Fig. 3.12 shows that temperature had no significant effect on the water-swelling capacity of sweet potato dietary fiber, while it had a significant effect on soybean dietary fiber. The water-swelling capacities of sweet potato and soybean dietary fibers were 10.58 and 10.18 mL/g, respectively, when the temperature was 100 C. Sweet potato residue and sweet potato powder also exhibited higher water-swelling capacities because they contain large amounts of starch.

Sweet Potato Dietary Fiber

10

Sweet potato residue

Sweet potato powder

Soybean dietary fiber

Sweet potato dietary fiber

163

Water holding capacity˄g/g)

8

6

4

2

0 3

4

5

6

7 8 pH Value

9

10

11

Figure 3.10 Effects of the pH value on the water-holding capacity (g/g) of sweet potato dietary fiber.

Water holding capacity˄g /g)

9

Sweet potato residue

Sweet potato powder

Soybean dietary fiber

Sweet potato dietary fiber

8 7 6 5 4 3 2 1 0

0%

1%

3%

5%

7%

10%

Salt concentration

Figure 3.11 Effects of the salt concentration on the water-holding capacity (g/g) of sweet potato dietary fiber.

Fig. 3.13 shows that the water-swelling capacity of dietary fibers was the lowest when at pH 3. This value increased as the pH increased. The water-swelling capacity of sweet potato powder was the highest (5.30 mL/g) at pH 6, and then it decreased. However, the waterswelling capacity of sweet potato dietary fiber, soybean dietary fiber, and sweet potato residue further increased, reaching 6.63, 5.75, and

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Water swelling capacity˄mL/g)

14

Sweet potato residue

Sweet potato powder

Soybean dietary fiber

Sweet potato dietary fiber

12

10 8 6 4 2 0 20

30

40

50 60 Temperature (°C)

70

80

90

100

Figure 3.12 Effects of temperature on the water-swelling capacity (mL/g) of sweet potato dietary fiber.

Sweet potato residue Soybean dietary fiber

Water swellinging capacity (mL/g)

8

Sweet potato powder Sweet potato dietary fiber

7 6 5 4 3 2

1 0

3

4

5

6

7

8

9

10

11

pH Value

Figure 3.13 Effects of pH value on the water-swelling capacity (mL/g) of sweet potato dietary fiber.

7.50 mL/g, respectively, at pH 11. Fig. 3.14 shows that the salt concentration has no significant effect on the water-swelling capacities of these four materials.

4.3 EFFECTS OF TEMPERATURE ON THE OIL-HOLDING CAPACITY OF SWEET POTATO DIETARY FIBER The oil-holding capacity of dietary fiber, as well as the water-holding capacity, is affected by temperature. Fig. 3.15 indicates that the oil-

Water swelling capacity (mL/g)

Sweet Potato Dietary Fiber

Sweet potato residue Soybean dietary fiber

10 9 8 7 6 5 4 3 2 1 0 0%

1%

165

Sweet potato powder Sweet potato dietary fiber

3% 5% Salt concentration

7%

10%

Figure 3.14 Effects of salt concentration on the water-swelling capacity (mL/g) of sweet potato dietary fiber.

Oil holding capacity (g/g)

4

Sweet potato residue

Sweet potato powder

Soybean dietary fiber

Sweet potato dietary fiber

3

2

1

0 20

30

40

50

60 70 Temperature (°C)

80

90

100

Figure 3.15 Effects of temperature on the oil-holding capacity (g/g) of sweet potato dietary fiber.

holding capacities of the four samples increased when the temperature was 50
60 C. When the temperature reached 60 C, the oil-holding capacities were greatest at 3.10, 2.33, 1.45, and 1.58 g/g. The oilholding capacity decreased sharply when the temperature was between 60 and 70 C. However, decrement was greatest when the temperature was 90 C. Therefore, it is beneficial to improve the oil-holding capacity of dietary fiber at temperatures between 60 and 90 C, which occur during food processing.

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4.4 EFFECTS OF DIFFERENT FACTORS ON THE VISCOSITY OF SWEET POTATO DIETARY FIBER Fig. 3.16 shows that the viscosity of sweet potato dietary fiber, soybean dietary fiber, and sweet potato powder changed little at temperatures from 20 to 100 C, while the viscosity of the sweet potato residue increased significantly at temperatures from 60 to 100 C. Fig. 3.17 shows that the pH level had no significant effects on the viscosities of dietary fibers. Fig. 3.18 shows that the viscosity of sweet potato dietary

Sweet potato residue Soybean dietary fiber

Viscosity (cP· rpm)

5

Sweet potato powder Sweet potato dietary fiber

4 3

2 1 0 20

40

60 Temperature (°C)

80

100

Figure 3.16 Effects of temperature on the viscosity (cP rpm) of sweet potato dietary fiber.

10

Sweet potato residue

Sweet potato powder

9

Soybean dietary fiber

Sweet potato dietary fiber

Viscosity (cP.rpm)

8 7 6 5 4 3 2 1 0 3

4

5

6

7

8

9

10

pH Value

Figure 3.17 Effects of pH value on the viscosity (cP rpm) of sweet potato dietary fiber.

11

Sweet Potato Dietary Fiber

Viscosity (cP· rpm)

4

Sweet potato residue

Sweet potato powder

Soybean dietary fiber

Sweet potato dietary fiber

167

3

2

1

0

0%

0.50%

1.00%

1.50%

2.00%

3.00%

Salt concentration Figure 3.18 Effects of salt concentration on the viscosity (cP rpm) of sweet potato dietary fiber.

fiber increased with increases in the NaCl concentration. The viscosity was 3.44 cP rpm at a 3% NaCl concentration. The viscosities of the other three materials increased with increasing salt concentrations, and the viscosities of the sweet potato residue, sweet potato powder, and soybean dietary fiber were not different at the same NaCl concentrations.

4.5 WATER-HOLDING CAPACITY OF DIETARY FIBERS FROM DIFFERENT SWEET POTATO VARIETIES Fig. 3.19 shows that the water-holding capacities of dietary fibers from different sweet potato varieties were significantly different (P , 0.05). The water-holding capacities of dietary fibers from 10 sweet potato varieties were as follows: “Xu 55-2” (6.15 g/g) . “Xinong 431” (5.56 g/g) . “Jishu 82” (5.09 g/g) . “Weiduol” (4.96 g/g) . “Lvya 18” (4.82 g/g) . “Jishu 99” (4.78 g/g) . “Jishu 7-1” (4.58 g/g) . “Jishu 21” (4.55 g/g) . “Beijing 553” (4.21 g/g) . “Jishu 98” (3.54 g/ g). The average water-holding capacity was 4.82 g/g, which was lower than that of guar gum (63.07 g/g), citrus pectin (28.07 g/g), and apple pectin (16.51 g/g), but higher than that of cellulose (0.71 g/g). It was similar to that of cocoa dietary fiber (4.76 g/g) and Canavalia dietary fiber (5.53 g/g). Raghavendra et al. (2004) showed that the waterholding capacity of dietary fibers from different sources had obvious

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Water holding capacity (g/g)

7

a b

6

5

i

h

g

f

c

e

d

j

4 3

2 1 0

1

2

3

4 5 6 Variety number

7

8

9

10

Figure 3.19 Water-holding capacities (g/g) of dietary fibers from different varieties of sweet potato: (1) “Beijing 553”, (2) “Jishu 21”, (3) “Jishu 7-1”, (4) “Jishu 82”, (5) “Jishu 98”, (6) “Jishu 99”, (7) “Lvya 18”, (8) “Weiduoli”, (9) “Xinong 431”, (10) “Xu 55-2”.

differences, with citrus and cocoa dietary fibers having the highest water-holding capacity (7.0 g/g), while wheat bran exhibited the lowest water-holding capacity (1.9 g/g).

4.6 WATER-SWELLING CAPACITY OF DIETARY FIBER FROM DIFFERENT VARIETIES OF SWEET POTATO Fig. 3.20 shows that the water-swelling capacity of dietary fibers from different sweet potato varieties were significantly different (P , 0.05). The water-swelling capacities of dietary fibers from 10 sweet potato varieties were as follows: “Xinong 431” (12.56 mL/g) . “Weiduoli” (12.26 mL/g) . “Jishu 99” (10.92 mL/g) . “Xu 55-2” (10.51 mL/g) . “Jishu 82” (10.51 mL/g) . “Beijing 553” (10.12 mL/g) . “Jishu 7-1” (9.81 mL/g) . “Jishu 98” (9.34 mL/g) . “Jishu 21” (9.09 mL/g) . “Lvya 18” (8.11 mL/g). The average water-swelling capacity was 10.32 mL/g, which was higher than that of cocoa dietary fiber (6.51 mL/g) and apple pectin (7.42 mL/g), but it was similar to that of citrus pectin (10.45 mL/g) (Lecumberri et al., 2007). Figuerola et al. (2005) also found that the water-swelling capacities of dietary fibers from different apple, lemon, and grape varieties were significantly different. In addition, dietary fibers extracted from oat bran, wheat bran, peas, peas shell, and coconut also exhibited different swelling capacities, with values from 5.3 to 20 mL/g (Raghavendra et al., 2004).

Sweet Potato Dietary Fiber

14 Water swelling capacity (mL/g)

b 12 e 10

h

f

a

c

d

169

d

g i

8 6 4

2 0

1

2

3

4

5

6

7

8

9

10

Variety number

Figure 3.20 Water-swelling capacities (mL/g) of dietary fibers from different varieties of sweet potato: (1) “Beijing 553”, (2) “Jishu 21”, (3) “Jishu 7-1”, (4) “Jishu 82”, (5) “Jishu 98”, (6) “Jishu 99”, (7) “Lvya 18”, (8) “Weiduoli”, (9) “Xinong 431”, (10) “Xu 55-2”.

Oil holding capacity (g/g)

3 b

2.5

c

e

2 g

a

d f

h j

1.5

i

1 0.5 0

1

2

3

4

5

6

7

8

9

10

Variety number Figure 3.21 Oil-holding capacities (g/g) of dietary fibers from different varieties of sweet potato: (1) “Beijing 553”, (2) “Jishu 21”, (3) “Jishu 7-1”, (4) “Jishu 82”, (5) “Jishu 98”, (6) “Jishu 99”, (7) “Lvya 18”, (8) “Weiduoli”, (9) “Xinong 431”, (10) “Xu 55-2”.

4.7 OIL-HOLDING CAPACITIES OF DIETARY FIBERS FROM DIFFERENT VARIETIES OF SWEET POTATO Fig. 3.21 shows that the oil-holding capacities of dietary fibers from different sweet potato varieties were significantly different (P , 0.05). The oil-holding capacities of dietary fibers from 10 different sweet potato varieties were as follows: “Xu 55-2” (2.48 g/g) . “Jishu 99” (2.38 g/g) . “Xinong 431” (2.35 g/g) . “Weiduoli” (2.19 g/g) . “Jishu 21” (2.06 g/g) . “Lvya 18” (1.76 g/g) . “Beijng 553” (1.67 g/g) . “Jishu 7-1” (1.65 g/g) . “Jishu 98” (1.56 g/g) . “Jishu 82” (1.43 g/g).

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The average oil-holding capacity was 1.95 g/g, which was lower than that of date dietary fiber (9.6 g/g) (Elleuch et al., 2008), and it was similar to that of dietary fibers from apple (1.3 g/g), pea (0.9 g/g), wheat (1.3 g/g), beet (1.5 g/g), and carrot (1.2 g/g) (Raghavendra et al., 2004). Figuerola et al. (2005) also found that the oil-holding capacities of dietary fibers from different varieties of grape, lemon, and apple showed different oilholding capacities, ranging from 0.60 to 1.81 g/g.

SECTION 5: THE APPLICATIONS OF SWEET POTATO DIETARY FIBER Sweet potato dietary fiber can be used for baked foods, beverages, meat products, and other foods. Zhang et al. (2009) found that sweet potato dietary fiber could improve the quality of bread and help to prolong its preservation period. Yu (2005) showed that cake and steamed bread tasted good, had no rough sense, and no acerbity when sweet potato dietary fiber was added. In addition, the stability and taste of orange beverages was improved when sweet potato dietary fiber was added. This section introduces the applications of sweet potato dietary fiber to bread, beverages, meat products, staple food, seasoning and health foods, to provide a theoretical basis for the application of sweet potato dietary fiber in the food and health product industries.

5.1 THE APPLICATIONS OF SWEET POTATO DIETARY FIBER IN BREAD Bread is becoming a necessary food for breakfast. Different amounts of sweet potato dietary fiber and sweet potato residue were added in bread by Guo (2010), and the sensory, textural, and physicochemical properties were determined to evaluate the effects of the addition of different amounts of sweet potato dietary fiber on the quality of bread, thus providing a theoretical basis for the applications of sweet potato dietary fiber in flour products.

5.1.1 The Sensory Qualities of Bread The effects of different amounts of sweet potato dietary fiber on the sensory qualities of bread are shown in Tables 3.15
3.17. The

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appearance and intrinsic qualities of bread were improved when the added amounts of sweet potato dietary fiber and residue were 1%
3% and 3%, respectively, while these two qualities decreased when the amounts of sweet potato dietary fiber and residue were 5% and 6%, respectively. When the amounts of sweet potato dietary fiber and residue were 1%
3% and 3%, respectively, the bread’s quality level was greater than that of the control group. When the amounts of sweet potato dietary fiber and residue were above 4% and 6%, respectively, the quality level of the bread decreased.

5.1.2 The Physicochemical Properties of Bread With increases in the added amounts of sweet potato dietary fiber and sweet potato residues, the volume and specific volume of bread decreased, but the weight increased. However, the specific volume of bread changed little when the added amounts of sweet potato dietary fiber and sweet potato residue were 1%
3% and 3%, respectively. When the added amounts were higher than 4% and 6%, respectively, the specific volume of bread decreased. Table 3.18 also shows that the moisture content of the bread increased with increases in the added amounts of sweet potato dietary fiber and sweet potato residue. This was because the gluten content decreased and the interactions between gluten and fiber materials decreased. Thus, the water retention capacity of bread increased (Zhang et al., 2009).

5.1.3 The Texture of Bread Table 3.19 shows that the hardness of bread decreased as the added amounts of sweet potato dietary fiber and sweet potato residue Table 3.15 Phenotypic Evaluation of Bread Addition Amount

Volume

Color

Texture

Appearance

Tactility

Score

Control

9.42 6 0.76

8.66 6 0.50

7.43 6 0.55

3.13 6 0.43

3.74 6 0.73

32.38 6 0.34

1% SPDF

8.75 6 0.45

8.72 6 0.39

8.45 6 0.86

3.15 6 0.48

3.83 6 0.67

32.90 6 0.27

2% SPDF

8.13 6 0.78

8.95 6 0.42

8.91 6 0.73

3.17 6 0.51

4.25 6 0.59

33.41 6 0.35

3% SPDF

7.82 6 0.62

9.07 6 0.34

8.96 6 0.67

3.26 6 0.72

4.79 6 0.48

33.90 6 0.22

4% SPDF

7.09 6 0.54

8.74 6 0.38

7.93 6 0.53

2.44 6 0.65

4.50 6 0.54

30.70 6 0.38

5% SPDF

6.83 6 0.81

7.45 6 0.42

6.86 6 0.44

2.63 6 0.57

3.80 6 0.46

27.57 6 0.40

3% SPR

8.56 6 0.39

8.73 6 0.28

7.56 6 0.37

3.26 6 0.45

3.84 6 0.56

33.15 6 0.27

6% SPR

7.50 6 0.62

5.87 6 0.66

5.02 6 0.52

2.45 6 0.55

3.72 6 0.55

24.56 6 0.24

Note: SPDF, sweet potato dietary fiber; SPR, sweet potato residue.

Table 3.16 Quality Evaluation of Bread Addition Amount

Internal Tissue

Flesh Color

Tactility

Texture

Flavor

Odor

Score

Control

8.93 6 0.65

9.00 6 0.55

5.67 6 0.78

9.21 6 0.72

7.93 6 0.46

2.98 6 0.65

43.72 6 0.25

1% SPDF

7.64 6 0.53

8.93 6 0.62

9.15 6 0.82

13.54 6 0.85

8.57 6 0.38

4.25 6 0.73

52.08 6 0.27

2% SPDF

8.87 6 0.37

8.78 6 0.57

9.13 6 0.68

12.89 6 0.78

8.64 6 0.29

4.33 6 0.82

52.64 6 0.31

3% SPDF

9.13 6 0.42

8.75 6 0.59

8.97 6 0.59

12.75 6 0.66

8.91 6 0.22

3.92 6 0.67

52.43 6 0.22

4% SPDF

8.65 6 0.34

8.07 6 0.44

8.76 6 0.83

11.86 6 0.58

8.75 6 0.31

4.03 6 0.59

50.12 6 0.17

5% SPDF

7.52 6 0.67

8.65 6 0.38

7.92 6 0.75

10.34 6 0.76

8.42 6 0.36

3.87 6 0.63

46.72 6 0.35

3% SPR

9.06 6 0.33

9.18 6 0.42

9.21 6 0.56

12.44 6 0.65

8.59 6 0.45

3.88 6 0.56

52.36 6 0.23

6% SPR

8.55 6 0.72

8.64 6 0.76

7.93 6 0.81

10.57 6 0.77

8.87 6 0.28

3.54 6 0.62

48.10 6 0.16

Note: SPDF, sweet potato dietary fiber; SPR, sweet potato residue.

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Table 3.17 Sensory Evaluation of Bread Addition Amount

Appearance Score

Internal Score

Total Score

Quality Level

Control

32.38 6 0.34

43.72 6 0.25

76.10 6 0.34

Better

1% SPDF

32.90 6 0.27

52.08 6 0.27

84.98 6 0.25

Good

2% SPDF

33.41 6 0.35

52.64 6 0.31

86.05 6 0.33

Good

3% SPDF

33.90 6 0.22

52.43 6 0.22

86.33 6 0.27

Good

4% SPDF

30.70 6 0.38

50.12 6 0.17

80.82 6 0.18

Good

5% SPDF

27.57 6 0.40

46.72 6 0.35

74.29 6 0.41

Better

3% SPR

29.74 6 0.33

52.36 6 0.23

82.10 6 0.37

Good

6%SPR

24.56 6 0.24

48.10 6 0.16

72.66 6 0.24

Better

Note: SPDF, sweet potato dietary fiber; SPR, sweet potato residue.

Table 3.18 The Physicochemical Properties of Bread Addition Amount

Volume (cm3)

Weight (g)

Specific Volume (cm2/g)

Moisture Content (%)

Control

494.07

85.48

5.78

19.16

1% SPDF

478.32

88.25

5.42

23.57

2% SPDF

429.97

89.39

4.81

24.12

3% SPDF

427.72

89.67

4.77

24.98

4% SPDF

365.14

92.44

3.95

25.74

5% SPDF

263.16

93.65

2.81

26.86

3% SPR

471.28

88.92

5.30

23.65

6% SPR

416.86

90.23

4.62

25.03

Note: SPDF, sweet potato dietary fiber; SPR, sweet potato residue.

Table 3.19 The Texture of Bread Addition Amount

Hardness

Cohesiveness

Viscosity

Elasticity

Chewiness

Control

1314.83 6 0.56

0.407 6 0.44

535.13 6 0.58

0.781 6 0.72

157.48 6 0.43

1% SPDF

541.35 6 0.35

0.448 6 0.38

242.52 6 0.48

0.815 6 0.66

167.06 6 0.54

2% SPDF

583.14 6 0.63

0.452 6 0.45

263.57 6 0.53

0.822 6 0.80

174.52 6 0.66

3% SPDF

624.37 6 0.44

0.453 6 0.56

282.83 6 0.46

0.874 6 0.75

188.23 6 0.57

4% SPDF

736.64 6 0.92

0.479 6 0.59

352.85 6 0.65

0.849 6 0.67

135.22 6 0.49

5% SPDF

878.99 6 0.47

0.581 6 0.47

510.69 6 0.32

0.745 6 0.59

116.34 6 0.52

3% SPR

745.92 6 0.85

0.439 6 0.63

327.45 6 0.89

0.863 6 0.62

175.35 6 0.64

6% SPR

931.82 6 0.53

0.418 6 0.58

389.50 6 0.68

0.755 6 0.55

160.59 6 0.72

Note: SPDF, sweet potato dietary fiber; SPR, sweet potato residue.

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increased. With increasing added amounts, a compact network structure between the fibrous materials and the gluten protein was formed, increasing the water interception of bread, thereby reducing the bread’s hardness (Zhang et al., 2009). In addition, with increasing added amounts, the cohesiveness, chewiness, and elasticity of bread increased, while the viscosity decreased. This may be due to the decrease in gluten protein content, making the viscosity of the bread decrease. In addition, the dense structure, which was formed between the fiber material and gluten protein, further increased the cohesiveness, chewiness, and elasticity of the bread (Zhang et al., 2009). The data in Table 3.19 indicate that when the sweet potato dietary fiber content was 2%
3% and the sweet potato residue content was 3%, the bread had better textural characteristics.

5.2 THE APPLICATIONS OF SWEET POTATO DIETARY FIBER IN BEVERAGES Beverages containing dietary fiber are popular in Europe, the United States, Japan, and other developed countries. This kind of beverage is also popular in China. Here, the dietary fiber is mainly used for liquids, solids, and carbonated drinks. The dietary fiber is also fermented using lactic acid bacteria and made into whey beverages.

5.3 THE APPLICATIONS OF SWEET POTATO DIETARY FIBER IN MEAT PRODUCTS Adding dietary fiber to meat products can retain the moisture and reduce the calories. The added amounts of dietary fiber in meat products are generally 1%
5% (Jin, 2004).

5.4 THE APPLICATIONS OF SWEET POTATO DIETARY FIBER IN STAPLE FOODS Dietary fiber can be used in noodles and steamed bread, and the additional amount of dietary fiber is generally 5%
6%. The strength of raw noodles having added dietary fiber decreased, while the strength increased after being cooked. Dietary fiber can also be added to cereal raw materials to enhance breakfast foods. In addition, dietary fiber can also be added to snack foods, such as pudding, chocolate, candy,

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175

and chewing gum. The added amounts differ depending on the type of food (Yue, 2004).

5.5 THE APPLICATIONS OF SWEET POTATO DIETARY FIBER IN CONDIMENTS Dietary fiber can also be added to some food additives, such as caramel pigment, animal and vegetable oils and fats, sorbic acid, trace elements and other nutrients, and xylitol, to make fillings that are used for beef pies, burgers, and stuffing.

5.6 THE APPLICATIONS OF SWEET POTATO DIETARY FIBER IN HEALTH FOODS Dietary fiber has been widely used in health foods, in addition to being used in food additives. Currently, there are many capsules and tablets, as well as other health products, based on dietary fiber, which have good economic benefits.

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1642. Wu, J.G., Zhou, S., Zhang, X.Y., 2005. Study on fermentation technology of dietary fiber from sweet potato residue by liquid fermentation of medicinal fungi. Food Ferment. Ind. 7, 42
44 (in Chinese). Yin, C.C., 2008. Research on the enzymatic preparation of sweet potato dietary fiber and its pressure modification. HuaNan Agricultural University, GuangZhou, China. Ylonen, K.K., Alfthan, G.G., Group, L.L., Saloranta, C.C., Aro, A.A., Virtanen, S.M.S.M., 2003. Dietary intakes and plasma concentrations of carotenoids and tocopherols in relation to glucose metabolism in subjects at high risk of type 2 diabetes: the Botnia Dietary Study. Am. J. Clin. Nutr 77 (6), 1434
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671. Yu, Y.J., 2005. Extraction, Function and Application of Dietary Fiber from Sweet Potato Residue. Shanghai University, Shanghai (in Chinese). Zhang, J.Q., Guan, X.S., Wan, J., 2009. Preparation of dietary fiber from sweet potato residues and its effect on the quality of bread. J. Jiangxi Food Ind. 11 (1), 32
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Zhao, Q.L., 2003. Study on the Extraction of Active Components from Plant Cells by Microwave Pretreatment. Guangxi University, Guangxi (in Chinese). Zheng, J.X., 2005. Functional Dietary Fiber. Chemical Industry Press, Beijing (in Chinese). Zheng, J.J., 1995. Some questions should be considered in the research of dietary fiber. Chin. Foreign Technical Intell 2, 5
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FURTHER READING Alfredo, V.O., Gabriel, R.R., Luis, C.G., David, B.A., 2009. Physicochemical properties of a fibrous fraction from chia (Salvia hispanica L.). LWT—Food Sci. Technol. 42 (1), 168
173. AOAC Official Method 991.43 Total, Soluble, and Insoluble Dietary Fiber in Foods EnzymaticGravimetric Method, MES-TRIS Buffer First Action 1991 Final Action, 1994. Bligh, E.G., Dyer, W.J., 1959. A rapid method of total lipid extraction and purification. Can. J. Biochem. Physiol. 37 (8), 911
917. Boehm, G., Lidestri, M., Casetta, P., Jelinek, J., Negretti, F., Stahl, B., et al., 2002. Supplementation of a bovine milk formula with an oligosaccharide mixture increases counts of faecal bifidobacteria in preterm infants. Arch. Dis. Childhood-Fetal Neonatal Ed. 86 (3), F178
F181. Bovell-Benjamin, A.C., 2007. Sweet potato: a review of its past, present, and future role in human nutrition. Adv. Food. Nutr. Res. 52, 1
59. Cao, Y.Y., 2007a. Study on the extraction technology of sweet potato dietary fiber from sweet potato by sieve method. Food Sci. Technol. 28 (7), 131
133 (in Chinese). Cao, Y.Y., 2007b. Study on Preparation and Physicochemical Properties of Sweet Potato Dietary Fiber. Xinjiang Agricultural University, Xinjing, (in Chinese). Caprez, A., Arrigoni, E., Amadò, R., Neukom, H., 1986. Influence of different types of thermal treatment on the chemical composition and physical properties of wheat bran. J. Cereal Sci. 4 (3), 233
239. Chuang, S.C., Vermeulen, R., Sharabiani, M.T., Sacerdote, C., Saberi Hosnijeh, F., Berrino, F., et al., 2011. The intake of grain fibers modulates cytokine levels in blood. Biomarkers. 16 (6), 504
510. Corte, O.L., Martínez, F.H., Ortiz, A.R., 2010. Effect of dietary fiber in the quantitative expression of butyrate receptor GPR43 in rats colon. Nutricion Hospitalaria 26 (5), 1052
1058. Cummings, J.H., Macfarlane, G.T., Englyst, H.N., 2001. Prebiotic digestion and fermentation. Am. J. Clin. Nutr. 73 (2), 415s
420s. de Delahaye, E.P., Jiménez, P., Pérez, E., 2005. Effect of enrichment with high content dietary fiber stabilized rice bran flour on chemical and functional properties of storage frozen pizzas. J. Food Eng. 68 (1), 1
7. de Escalada Pla, M.F., Ponce, N.M., Stortz, C.A., Gerschenson, L.N., Rojas, A.M., 2007. Composition and functional properties of enriched fiber products obtained from pumpkin (Cucurbita moschata Duchesne ex Poiret). LWT—Food Sci. Technol. 40 (7), 1176
1185. Elleuch, Mohamed, Bedigian, Dorothea, Roiseux, Olivier, 2011. Dietary fiber and fiber-rich byproducts of food processing: characterisation, technological functionality and commercial applications: A review. Food. Chem. 124, 411
421.

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Femenia, A., Lefebvre, A.C., Thebaudin, J.Y., Robertson, J.A., Bourgeoious, C.M., 1997. Physical and sensory properties of model foods supplemented with cauliflower fiber. J. Food. Sci. 62 (4), 635
639. Fernandez, M.L., Lin, E.C., Trejo, A., McNamara, D.J., 1994. Prickly pear (Opuntia sp.) pectin alters hepatic cholesterol metabolism without affecting cholesterol absorption in guinea pigs fed a hypercholesterolemic diet. J. Nutr. 124 (6), 817
824. Fernandez-Gines, J.M., Fernandez-Lopez, J., Sayas-Barbera, E., Sendra, E., Perez-Alvarez, J.A., 2004. Lemon albedo as a new source of dietary fiber: application to bologna sausages. Meat Sci. 67 (1), 7
13. Haarman, M., Knol, J., 2006. Quantitative real-time PCR analysis of fecal Lactobacillus species in infants receiving a prebiotic infant formula. Appl. Environ. Microbiol. 72 (4), 2359
2365. Han, J.J., 2009. Analysis of the main components of sweet potato dietary fiber from 10 kinds of sweet potato residues and its screening materials. J. Chin. Cereals Oils Assoc. 24 (1), 40
43 (in Chinese). Kaczmarczyk, M.M., Miller, M.J., Freund, G.G., 2012. The health benefits of dietary fiber: beyond the usual suspects of type 2 diabetes mellitus, cardiovascular disease and colon cancer. Metabolism. 61 (8), 1058
1066. Kennedy, R.L., Vangaveti, V., Jarrod, G., Shashidhar, V., Baune, B.T., 2010. Review: free fatty acid receptors: emerging targets for treatment of diabetes and its complications. Ther. Adv. Endocrinol. Metabol. 1 (4), 165
175. Liu, C.M., Li, Z.L., Liang, R.H., Tu, Z.C., Liu, W., 2006. Physiological properties and application status of dietary fiber. Food Res. Dev. 27 (1), 122
125 (in Chinese). Ma, Y., Griffith, J.A., Chasan-Taber, L., Olendzki, B.C., Jackson, E., Stanek, E.J., et al., 2006. Association between dietary fiber and serum C-reactive protein. Am. J. Clin. Nutr. 83 (4), 760
766. Ma, Z.W., Zhang, X.Z., 2004. Long term effect of dietary fiber on lipid metabolism in rats and its mechanism. Chin. J. Clin. Rehab. 8 (9), 1677
1679 (in Chinese). Mei, X., Mu, T.H., Han, J.J., 2010. Composition and physicochemical properties of dietary fiber extracted from residues of 10 varieties of sweet potato by a sieving method. J. Agric. Food. Chem. 58 (12), 7305
7310. Meyer, A.S., Dam, B.P., Lærke, H.N., 2009. Enzymatic solubilization of a pectinaceous dietary fiber fraction from potato pulp: optimization of the fiber extraction process. Biochem. Eng. J. 43 (1), 106
112. Mu, T.H., Tan, S.S., Xue, Y.L., 2009. The amino acid composition, solubility and emulsifying properties of sweet potato protein. Food. Chem. 112 (4), 1002
1005. Noda, T., Takahata, Y., Nagata, T., Shibuya, N., 1994. Chemical composition of cell wall material from sweet potato starch residue. Starch
Stärke 46 (6), 232
236. Nyman, E.M.G.L., Svanberg, S.M., 2002. Modification of physicochemical properties of dietary fibre in carrots by mono- and divalent cations. Food. Chem. 76 (3), 273
280. Qi, B., Jiang, L., Li, Y., Chen, S., Sui, X., 2011. Extract dietary fiber from the soy pods by chemistry-enzymatic methods. Procedia Engineering 15, 4862
4873. Raninen, K., Lappi, J., Mykkänen, H., Poutanen, K., 2011. Dietary fiber type reflects physiological functionality: comparison of grain fiber, inulin, and polydextrose. Nutr. Rev. 69 (1), 9
21. Redondo-Cuenca, A., Villanueva-Suárez, M.J., Rodríguez-Sevilla, M.D., Mateos-Aparicio, I., 2007. Chemical composition and dietary fibre of yellow and green commercial soybeans (Glycine max). Food. Chem. 101 (3), 1216
1222. Roberfroid, M., Gibson, G.R., Hoyles, L., McCartney, A.L., Rastall, R., Rowland, I., et al., 2010. Prebiotic effects: metabolic and health benefits. Br. J. Nutr. 104 (S2), S1
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Sa’ad, H., Peppelenbosch, M.P., Roelofsen, H., Vonk, R.J., Venema, K., 2010. Biological effects of propionic acid in humans; metabolism, potential applications and underlying mechanisms. Biochim. Biophys. Acta (BBA)-Mol. Cell Biol. Lipids 1801 (11), 1175
1183. Sato, Y., Kawabuchi, S., Irimoto, Y., Miyawaki, O., 2004. Effect of water activity and solventordering on intermolecular interaction of high-methoxyl pectins in various sugar solutions. Food Hydrocoll. 18 (4), 527
534. Sentenac, H., Grignon, C., 1981. A model for predicting ionic equilibrium concentrations in cell walls. Plant. Physiol. 68 (2), 415
419. Sowbhagya, H.B., Suma, P.F., Mahadevamma, S., Tharanathan, R.N., 2007. Spent residue from cumin—a potential source of dietary fiber. Food. Chem. 104 (3), 1220
1225. Staffolo, M.D., Bertola, N., Martino, M., 2004. Influence of dietary fiber addition on sensory and rheological properties of yogurt. Int. Dairy J. 14 (3), 263
268. Takamine, K., Abe, J., Iwaya, A., 2000. A new manufacturing process for dietary fiber from sweetpotato residue and its physical characteristics. Jpn. Soc. Appl. Glycosci. 47, 67
72. Takamine, K., Hotta, H., Degawa, Y., 2005. Effect of dietary fiber prepared from sweet potato pulp on cecal fermentation products and microflora in rats. Jpn. Soc. Appl. Glycosci. 52, 1
5. Tappy, L., Gügolz, E., Würsch, P., 1996. Effects of breakfast cereals containing various amounts of β-glucan fibers on plasma glucose and insulin responses in NIDDM subjects. Diabetes. Care 19 (8), 831
834. Tedelind, S., Westberg, F., Kjerrulf, M., Tebelind, S., Westberg, F., Kjerrulf, M., et al., 2006. Anti-Inflammatory properties of the short-chain fatty acid acetate and propionate: a study with relevance to inflammatory bowel disease. World J. Gastroenterol. 13, 2826
2832, World Journal of Gastroenterology, 2007, 13(20): 2826
2832. Thomassen, L.V., Meyer, A.S., 2010. Statistically designed optimisation of enzyme catalysed starch removal from potato pulp. Enzyme. Microb. Technol. 46 (3), 297
303. Tighe, P., Duthie, G., Vaughan, N., Brittenden, J., Simpson, W.G., Duthie, S., et al., 2010. Effect of increased consumption of whole-grain foods on blood pressure and other cardiovascular risk markers in healthy middle-aged persons: a randomized controlled trial. Am. J. Clin. Nutr. 92 (4), 733
740. Wang, S., Liu, F., 2000. Preparation, properties and application of high activity corn dietary fiber. Food Sci. 21 (7), 22
24 (in Chinese). Wang, W.X., 2010. Study on preparation of high activity soybean dietary fiber by cellulase. Food Mach. 118
122 (in Chinese). Wang, X.M., 2013. Study on the extraction technology of sweet potato dietary fiber and the effect of preventing and treating obese Wistar rats. Chin. Acad. Agric. Sci. (in Chinese). Wu, S.F., Hou, C.Y., 2011. Study on preparation technology of purple sweet potato dietary fiber by enzymatic method. Chin. J. Food Sci. 11 (1), 60
68 (in Chinese). Xie, S.L., Shi, Y.G., 2006. Effects of soybean oligosaccharides and soybean peptides on blood lipid metabolism in rats. J. Third Mil. Med. Univ. 28 (9), 945
948 (in Chinese). Yang, J.J., Bao, J.G., 1993. Influence of apple fiber on water holding capacity and quality of cake. J. Wuxi Light Ind. Sch. 12 (4), 263
268 (in Chinese). Zhang, L.L., Wu, X.K., Chen, L.L., 2010. The content of total dietary fiber in different sweet potato varieties determined by enzymatic gravimetric method. Mod. Prev. Med. 37 (2), 238
240 (in Chinese).

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CHAPTER

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SECTION 1: AN OVERVIEW OF PECTIN 1.1 The Distribution of Pectin 1.2 The Structure of Pectin 1.2.1 Homogalacturonan 1.2.2 RG-I 1.2.3 RG-II 1.3 The Research Methods Determining Pectin Structure 1.3.1 Monosaccharide Composition Analysis 1.3.2 Estimation of the Degree of Esterification 1.3.3 Relative Molecular Weight Determination of Pectin 1.4 Extraction of Pectin 1.4.1 Pretreatment of Raw Materials 1.4.2 Pectin Hydrolysis and Extraction 1.4.3 Pectin Purification 1.4.4 Drying Pectin 1.5 Functional Properties of Pectin 1.5.1 Physiological Functions of Pectin 1.5.2 Physicochemical Functions of Pectin SECTION 2: PRODUCTION TECHNOLOGY OF SWEET POTATO PECTIN 2.1 Extraction Process of Pectin from Sweet Potato 2.2 The Determination of the Galacturonic Acid Content in the Pectin Solution 2.3 Factors Affecting the Yield and Galacturonic Acid Content of Sweet Potato Pectin 2.3.1 Effects of Temperature on the Extraction Yield and Galacturonic Acid Content of Sweet Potato Pectin Sweet Potato Processing Technology. DOI: http://dx.doi.org/10.1016/B978-0-12-812871-8.00004-0 Copyright © 2017 China Science Publishing & Media Ltd. Published by Elsevier Inc. All rights reserved.

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2.3.2 Effects of pH Value on the Extraction Yield and Galacturonic Acid Content of Sweet Potato Pectin 2.3.3 Effects of Extraction Time on the Yield and the Galacturonic Acid Content of Sweet Potato Pectin 2.3.4 Effects of the Solid/Liquid Ratio on the Pectin Yield and the Galacturonic Acid Content of Sweet Potato Pectin 2.4 Optimization of Pectin Extraction from Sweet Potato Pulp SECTION 3: BIOLOGICAL ACTIVITIES OF SWEET POTATO PECTIN 3.1 The Preparation of pH-Modified Pectin 3.2 The Preparation of Thermal-Modified Pectin 3.3 The Cell Cultures 3.4 Effects of Sweet Potato Pectin on Cancer Cell Survival Rates 3.5 Effects of Sweet Potato Pectin on Cancer Cell Proliferation 3.5.1 Effects of Sweet Potato Pectin on the Proliferation of HT-29 Colon Cancer Cells 3.5.2 Effects of Sweet Potato Pectin on Bcap-37 Breast Cancer Cell Proliferation 3.5.3 Effects of Sweet Potato Pectin on SMMC-7721 Liver Cancer Cell Proliferation 3.6 Effects of Sweet Potato Pectin on Cancer Cell Metastasis 3.6.1 Effects of Sweet Potato Pectin on Cancer Cell Adhesion 3.6.2 The Effects of Sweet Potato Pectin on Cancer Cell Migration 3.6.3 Effects of Sweet Potato Pectin on the Urokinase-Type Plasminogen Activator (uPA) Content in Cancer Cells SECTION 4: PHYSICOCHEMICAL CHARACTERISTICS OF SWEET POTATO PECTIN 4.1 A Viscosity Analysis of Sweet Potato Pectin 4.1.1 Effects of Pectin Concentration and pH Value on the Viscosity of a Pectin Solution 4.1.2 Effects of Sucrose, NaCl, and Ca21 Concentrations on the Viscosity of a Pectin Solution 4.1.3 Effects of Temperature and Shear Rate on the Viscosity of a Pectin Solution

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4.2 The Gelation Properties of Sweet Potato Pectin 4.2.1 Determination of the Sweet Potato Pectin Gel’s Linear Viscoelastic Region 4.2.2 The Effects of Pectin Concentration on the Moduli and the Complex Viscosity of Pectin Gelation 4.2.3 The Effects of the Sucrose Concentration on the Moduli and the Complex Viscosity of Pectin During Gelation 4.2.4 The Effect of Ca21 Concentration on the Moduli and the Complex Viscosity of Pectin Gelation 4.2.5 The Effects of pH Value on the Moduli and the Complex Viscosity of Pectin Gelation 4.2.6 Gel Texture Analysis of Sweet Potato Pectin 4.2.7 The Microstructure of Sweet Potato Pectin Gel 4.3 The Emulsifying Properties of Sweet Potato Pectin 4.3.1 Effects of the Pectin Concentration and Oil-Phase Volume Fraction on the Particle Sizes of Pectin Emulsions 4.3.2 Effects of pH Value, and NaCl and Ca21 Concentrations on the Particle Sizes of Pectin Emulsions 4.3.3 Effects of Emulsion Particles on the Adsorption of Pectin 4.3.4 Emulsifying Activity 4.3.5 Emulsion Viscosity 4.3.6 Emulsion Stability 4.3.7 Emulsion Micro Imaging SECTION 5: APPLICATIONS OF SWEET POTATO PECTIN 5.1 Candied Fruit 5.2 Bread 5.3 Frozen Food 5.4 Yogurt Products 5.5 Beverages 5.6 Others References Further Reading

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Abstract This chapter gives a relatively comprehensive introduction about sweet potato pectin. It starts by presenting a well-rounded overview on pectin. Then, the production technologies of sweet potato pectin are introduced. Response surface design is adopted to determine the best production conditions for pectin: the temperature is 93 C, extraction time is 2.2 h, pH value is 1.7, and the solid/liquid ratio is 1:30. The effects of pH- and thermal-modified pectin on the proliferation, inhibition, and migration of HT-29 colon cancer cells, Bcap-37breast cancer cells and SMMC-7721 liver cancer cells are discussed. It then explores factors that influence viscosity, gelation properties, and emulsifying properties of pectin. By the end of this chapter, the applications of sweet potato pectin in different products are presented.

SECTION 1: AN OVERVIEW OF PECTIN In most parts of China, large amounts of sweet potato pulp are available as by-products of sweet potato starch, vermicelli noodle and silk noodle processing. The water content of fresh sweet potato pulp can be as high as 80%, making it difficult to store and transport. In addition, the spoilage odor of pulp can lead to serious environmental pollution. Because of this, sweet potato processing factories invest a good deal of manpower and material resources to sweet potato pulp and waste-water treatment every year. Sweet potato pulp contains at least 20%
30% pectin. At present, in China’s food industry, pectin is mainly extracted from orange peel and apple pomace, and there are no reports on using sweet potato as a raw material for the isolation of pectin during production. If sweet potato pulp can be developed into a new material to extract pectin, then we can increase the added value to the sweet potato processing industry and develop new raw resources for pectin. In addition, we can also protect the environment.

1.1 THE DISTRIBUTION OF PECTIN Pectin is the most structurally complex polysaccharide in nature. It is a component of the cell walls of higher plants, gymnosperms, pteridophytes, mosses, and algae and makes up 35% of the primary walls of nongraminaceae dicotyledons and monocotyledons, 2%
10% of graminaceous monocot primary walls and 5% of the cell walls in woody

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tissues. Pectin is abundant in growing and dividing cell walls and the cell walls of plants’ soft parts (Mohnen, 2008). The highest concentration of pectin is in the middle lamellal cell wall, with a gradual decrease from the primary cell wall to the plasma membrane. Pectin is not only related to the adhesion of plant cells but also plays a role in stabilizing plant tissue structures. The existence of pectin guarantees the extension and sliding of the fiber-polysaccharide structures, and it participates in controlling cell wall porosity. The pectin network is also involved in cell wall growth, development, differentiation, and a variety of other physiological reactions (Thakur et al., 1997). A schematic diagram of the cell wall structure is shown in Fig. 4.1. Pectin is generally produced at the beginning of the growth stage in primary cells, accounting for about 1/3 dry matter in cell walls. The highest content of pectin is in the middle lamella, with a gradual decrease from primary cell wall to the plasma membrane. The pectin in the middle lamella is mostly connected with each other by Ca21 bridge (McCann and Roberts,1994). When immature, most of the pectins exist in the protopectin form, which is a water-insoluble pectic substance. The combinations between pectin substances or pectin, and hemicellulose or calcium salt, coexist in a mechanical or chemical way. Under the enzyme actions, protopectin gradually becomes pectinic acid, which

Figure 4.1 The schematic diagram of the cell wall structure.

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Figure 4.2 A schematic diagram of the three major polysaccharide domains of pectin (Willats et al., 2001).

include galacturonic acids that have different degrees of methyl esterification. When no carbomethoxylation of the pectinic acids occurs, pectic acids were formed.

1.2 THE STRUCTURE OF PECTIN Pectin is a kind of complex polysaccharide substance that mainly consists of α-(1,4)-linked D-galacturonic acid units. The relative molar mass of pectin is 400,000, and the relative degree of polymerization is more than 1000 units. Three kinds of pectin materials, including homogalacturonan (HGA), rhamnogalacturonan-I (RG-I), and RG-II, can be found in the primary cell walls of the plants. The structure of pectin is shown in Fig. 4.2.

1.2.1 Homogalacturonan HGA is an essentially linear polymer containing B100
200 (1,4)linked α-galacturonic acids (Willats et al., 2001), in which parts of the carboxyl groups could be methyl esterified. According to the different plant sources, HG may also be partially O-acetylated at C-3 or C-2. The free carboxyl groups were present in the form of free acids and in the forms of potassium, sodium, and calcium salts.

1.2.2 RG-I RG-I contains the repeating disaccharide [(1,2)-α-L-rhap-(1,4)-α-DGalpA] backbone (Fig. 4.3) in which parts of the galacturonic acid resides may be O-acetylated at C-3 or C-2, but at present, no studies showed that they could be methyl esterified. According to different plant sources and extraction methods, roughly 2%
80% of the rhamnoses of RG-I are substituted by neutral sugars and acid oligosaccharide side chains. The main side chains include linear and branched

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Figure 4.3 A major structural model of RG-I (Lerogue et al., 1993).

α-L-arabinose residues and/or β-D-galactose residues that differ in relative proportions and side-chain lengths because of plant sources. In addition, fructose residues, including β-D-glucuronic acid residues, 4-Omethyl-β-D-glucuronic acid residues, ferulic acids, and coumaric acids, exist in RG-I (Sun and Tang, 2004).

1.2.3 RG-II Unlike the structure of RG-I, the backbone of RG-II consists of 1,4linked α-D-galacturonic acids with four side branches of the polymer structure. The RG-II domain, which has a lower molecular weight, contains 11 different types of sugar residues, and the structure is very complex.

1.3 THE RESEARCH METHODS DETERMINING PECTIN STRUCTURE 1.3.1 Monosaccharide Composition Analysis Pectin is mainly composed of galacturonic acids, which account for B70% of total sugars in pectin, and the remaining neutral sugars, such as rhamnose, arabinose, galactose, glucose, xylose, and mannose, mostly exist in the pectin side chains. The monosaccharide composition of pectin is an important index that helps describe pectin’s chemical composition. The pectin monosaccharide compositional analysis is mainly based on determining the sugar component residues after chemical or enzymatic hydrolysis. Several chemical methods have been used for the hydrolysis of neutral sugar side chains, which vary among

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themselves because of the differences in the acid types, the acid concentrations, and the time and temperature of hydrolysis. Sulfuric acid, trifluoroacetic acid, hydrochloric acid, methanol-sulfuric acid solution, methanol-hydrochloric acid solution, and hydrofluoric acid are all commonly used as hydrolysis solvents. The common acid concentration, temperature, and hydrolysis time are 1
2 mol/L, 100
121 C, and 2
3 h, respectively. At present, the methods to determine the monosaccharide composition mainly include colorimetry, gas
liquid chromatography, and high-performance ion-exchange liquid chromatography. The colorimetric method mainly adopts an m-phenylenediamine sulfate-cresol method but the accuracy is low. Uronic and nonuronic acids exhibit the same color, and each pectin monosaccharide that is present at the same concentration shows a different color intensity. Gas
liquid chromatography and high-performance ion-exchange liquid chromatography have high accuracy levels, but compared with gas
liquid chromatography, the high-performance ionexchange liquid chromatography, equipped with an ion-exchange resin and a pulse amperometric detector, use a simple sample treatment and have a high accuracy and good repeatability. Thus, it is the most commonly used method to determine the pectin monosaccharide composition (Garna et al., 2007).

1.3.2 Estimation of the Degree of Esterification The degree of esterification of pectin refers to the percentage of esterified galacturonic acids out of the total galacturonic acid residues, and it usually represents the degree of methoxylation. Methods for determining the esterification are titration, colorimetry, gas chromatography, infrared spectroscopy, high-performance liquid chromatography, and nuclear magnetic resonance spectroscopy. The operation of the titration method is simple. In infrared spectroscopy, the degree of esterification is a ratio of the amount of esterified carboxyl groups’ absorption peaks in the absorption peaks of all of the carboxyl groups. High-performance liquid chromatography is more complicated. Before measuring, pectin needs to be saponified, and then, the amounts of methanol and acetic acid are used to divide the contents of the galacturonic acids to determine the degree of esterification value. Nuclear magnetic resonance spectroscopy has certain requirements for the molecular weights of samples, and the requirements of the test instruments are also greater.

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1.3.3 Relative Molecular Weight Determination of Pectin Pectin molecular weight not only affects the dissolution, absorption, and distribution of pectin, but also has a significant effect on the interactions between the target molecules. Therefore, it is necessary to determine the molecular weight of pectin. At present, the methods reported to determine the molecular weight of pectin include gel chromatography, viscosity, high-performance molecular exclusion chromatography, laser scattering, and ultrafiltration centrifugation methods. Among these methods, the operation of high-performance molecular exclusion chromatography is the easiest and has the lowest cost. High-performance molecular exclusion chromatography has been widely used to determine the molecular weight of pectin and its derivatives, and it can measure the weight-average molecular weight, number-average molecular weight, z-average molecular weight, and polymer dispersion. In addition, the rotational radius of the pectin molecules in solution can also be determined by high-performance molecular exclusion chromatography.

1.4 EXTRACTION OF PECTIN The extraction of pectin generally includes the following four parts: The pretreatment of the raw materials (to remove the small molecular weight carbohydrates and pigments), pectin hydrolysis and extraction, pectin purification, and pectin drying. The extraction and purification of pectin are the key steps because they can directly affect the quality of pectin.

1.4.1 Pretreatment of Raw Materials At present, the raw materials for pectin extraction are mainly byproducts in food processing, such as apple pomace, citrus peel, sugar beet pulp, peach residue, mango peel, passion fruit peel, and sunflower heads. Some raw materials contain large amounts of pigments and, therefore, must be decolorized, by using acetone, ether, and ethanol, before pectin extraction. Some plants, such as potatoes and soybean, have high levels of starch in their fruit and other tissues need to be treated with enzymes to break down the starch (Turquois et al., 1999). Thus, pretreatments are necessary to remove as much of the pigments and low molecular weight substances as possible, while keeping the loss of pectin (especially water-soluble pectin) to a minimum.

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1.4.2 Pectin Hydrolysis and Extraction Protopectin hydrolysis and extraction are the key steps in pectin preparation, and, normally, the same solvent is used in both steps. At present, the pectin extraction methods include acid, alkali, and salt extractions, as well as nontraditional methods. In addition, combinations of extraction methods are also widely used. Acid extraction is the most common way to extract pectin, and it is usually used under high temperature and low pH value conditions. Yapo and Koffi (2008) used citric, nitric, and sulfuric acids as solvents to extract pectin from yellow passion fruit peels, and they found that the different acids significantly influenced the recovered pectin molecules and the gel properties. The pectin extracted by citric acid had a higher molecular weight, higher degree of esterification, and better gel performance. Canteri-Schemin et al. (2005) used nitric, citric, hydrochloric, sulfuric, tartaric, and phosphoric acids to extract pectin from apple pomace and found that the highest pectin yield was obtained using citric acid solvent (13.75%) and the lowest yield was obtained using malic and phosphoric acids.

1.4.3 Pectin Purification During the pectin extraction process, some small molecules, as well as a certain amount of mineral salts, can easily be simultaneously extracted. Pectin purification removes the impurities from crude pectin, and this step influences the pectin’s quality and color. The alcohol precipitation method is often used to purify pectin, and the suitable ratio of alcohol to pectin crude extract is 1:3. Therefore, the alcohol precipitation method can consume a large amount of alcohol, especially at the industrial level. To reduce the consumption of alcohol, the pectin extract should be concentrated in advance. The vacuum concentration method is a traditional method that can remove volatile materials and moisture but cannot remove the protein and ash. Thus, this method is not selective. In recent years, several separation membranes (reverse osmosis, microfiltration, and ultrafiltration membranes) have been applied to pectin purification. The ultrafiltration method can be carried out at room temperature to avoid heating the samples, and the membrane stress is relative low. Precipitation with metal ions is another pectin purification method that can cause serious pollution problems, and the attained pectin has a higher ash content as well as poor solubility. In addition, the ion-exchange resin, which can remove all kinds of metal ions and does not react with high molecular weight pectin, has been used to purify the pectin. Wang et al. (2007) used an

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ion-exchange resin to purify pectin, and the main performance index and the pectin yield were superior to those achieved by traditional methods. The purification methods had effects on the yield, chemical composition and physicochemical properties of pectin (Hwang et al., 1992; Yapo and Koffi, 2008). Yapo and Koffi (2008) used three precipitation methods, ethanol, dialysis, and metal ion, to purify the yellow passion fruit pectin extracted by the acid method and found that the pectin precipitated by the metal ion method produced the highest galacturonic acid content [78.9% (w/w)], the lowest neutral sugar [1.4% (w/w)], ash [0.9% (w/w)], and protein [1.4% (w/w)] contents and a high gel strength.

1.4.4 Drying Pectin The drying method has an important influence on pectin quality, and several methods, including low temperature, freeze, vacuum, and spray drying, are often used. At present, spray drying is commonly used in foreign countries, and it does not require pectin precipitation. The attained pectin has a high solubility and good gel performance. The drying method most suitable to the raw materials and production capacity should be chosen.

1.5 FUNCTIONAL PROPERTIES OF PECTIN 1.5.1 Physiological Functions of Pectin Pectin is strongly hydrophilic and, therefore, it cannot be enzymatically digested in the gut. However, it can be fermented to produce short chain fatty acids by colonic microbes. Pectin influences the metabolism of nutrients, such as lipid and protein, and it regulates the growth of intestinal flora, promotes the apoptosis of cancer cells, and inhibits the growth and metastasis of tumors. In addition, it has a strong metal ion-binding capability. 1.5.1.1 Regulation of the Blood Glucose and Blood Lipid Levels With changing human dietary structures, the morbidities associated with diabetes, obesity, and hyperlipidemia, which are characterized by high blood glucose and high blood lipid levels, have increased dramatically. At present, drugs are often used to regulate blood glucose and blood lipid in clinical practice. These have a certain positive effect but also have undesirable side effects (Vaccitolo et al., 2001). Pectin can reduce the levels of blood glucose and blood lipid in patients with type

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II diabetes, possibly because the pectin delayed the absorption of sugar and fat in the digestive tract (Dongowski and Lorenz,2004) and affected the metabolism of bile acid, therefore, indirectly changing the activities of lipid metabolism-related enzymes (Ide et a1., 1990). Tests using Guinea pigs showed that pectin could regulate liver cholesterol homeostasis and lipoprotein metabolism, increase the expression of liver lipoprotein receptor B/E, promote the metabolism of low density lipoprotein, significantly reduce the cholesterol content of low density lipoprotein, and prevent the uptake and recycling of bile acid (Fernandez et al., 1994). Amidated pectin could also significantly increase the excretion of neutral sterols and bile acids from rats and reduce the level of cholesterol in the plasma and liver. There was no significant effect on cholestyramine (Marounek et al., 2010). Experiments using rats also found that cucumber pectin could promote glycogen synthesis, decrease the breakdown of glycogens, and reduce the level of blood glucose. These results may be related to pectin enhancing protein kinase C activities in the pancreas and the brain, while reducing the protein kinase C activity in the liver and increasing insulin secretion (Sudheesh and Vijayalakshmi, 2007). 1.5.1.2 Regulation of Lipid Peroxidation Pectin decreases the level of plasma cholesterol and triglyceride of hyperlipidemia rats, which were induced by a high fat diet, restrains the accumulation of body fat (Li et al., 2010) and weight loss, and also improves the activities of superoxide dismutase and glutathione peroxidase (Zhang et al., 2013), removes free radicals effectively to protect cells against oxidative damage, inhibits the synthesis and accumulation of malondialdehyde in the plasma (Li et al., 2010), maintains the metabolic homeostasis, and protects the body from damage. 1.5.1.3 Effects on Heavy Metals Heavy metals, such as arsenic, cadmium, lead, and mercury, can be enriched in human organs, which can cause dysfunctions of the kidney, liver, brain, reproductive system, immune system, and central nervous system (Carpenter, 2001). In vitro research found that sugar beet pectin could chelate copper, lead and chromium (Mata et al., 2009). Khotimchenko et al. (2004) found that pectin could promote the excretion of lead from rats and reduce thyroid damage caused by lead. In addition, the ability of pectin to absorb lead may be connected with its degree of methyl esterification (Khotimchernko et al.,

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2007). Eliaz et al. (2006) applied modified citrus pectin in clinical practice, and the average reduction rate in pectin-treated cases was 74% (five cases were used for the detoxification effect test) with no side effects. It was hypothesized that RG-II may be related to the ability to chelate heavy metals. Thus, pectin has the potential to be developed as a safe chelating agent for removing heavy metals from the body. 1.5.1.4 Inhibiting Effects on Cancer Metastasis Cancer metastasis, which is how cancer spreads from the positions of initial tumor growth to distant organs and tissues, increases cancer morbidity levels and death rates. Inhibiting this process is an enormous challenge facing clinical medicine (Glinsky and Raz, 2009), and there is an urgent need to develop drugs that can effectively inhibit cancer metastasis with limited adverse side effects. In vitro, pectin could impede the same types of tumor cells from aggregating (Platt and Raz, 1992), interfere with the prostate MAT-LyLu cells and MDA-MB-435 cells adhesion to human endothelial cells (Nangia-Makker et al., 2002), bond B16-F1 melanoma tumor cells (Platt and Raz, 1992), and inhibit the metastasis of the B16-F1 melanoma cells in mice and the MAT-LyLu prostate cancer cells in rats (Platt and Raz, 1992). At the same time, in vivo studies also found that pectin could inhibit the metastasis of breast cancer cells and prostate cancer cells to the lungs and bones, with a 90% inhibition rate (Glinskii et al., 2005). Inohara and Raz (1994) found that when inhibiting tumor metastasis, pectin had no side effects on cells, which may be because it bonds to galectins on the surface of cancer cells, interfering with the adhesion between cells or between cells and the matrix. Pectin contains galactose side chains, so it could be used as a galectin ligand. Thus, although pectin could not directly affect the expression and secretion of galectin in tumor cells, it could cause competitive inhibition by competing with natural ligands to bind the galectins (Platt and Raz, 1992). Therefore, it is possible to inhibit metastasis by impeding or interfering with the galectin-mediated cancers. 1.5.1.5 Effects on the Proliferation and Apoptosis of Cancer Cells In vitro studies found that apple pectin could activate caspase-3 and induced the apoptosis of HT-29 human colon cancer cells (OlanoMartin et al., 2003). In addition, apple pectin acids could restrain the proliferation of rat GH3/B6 pituitary tumor cells when the

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concentrations were at 100 μg/mL
1 mg/mL, and this effect was dosedependent. Concentrations of 2.5
5 mg/mL could induce cancer cell gangrene (Attaril et al., 2009). Caspase could directly lead to the disintegration of cells, and it was at the core of cancer-cell-mediated death and the regulatory mechanism of cell apoptosis (Mese et al., 2000). Pectin may activate caspase and induce the apoptosis of cancer cells. To some extent, pectin could activate the receptor δ gene, and the expression of prostaglandin E2, by inhibiting the peroxidase proliferation in rats injected with azoxymethane, enhance the production of prostaglandin E3 and induce the apoptosis of colon cancer cells (Vanamala et al., 2008). Vayssade et al. (2010) used the okra pectin RG-I region to treat B16F10 melanoma cells in vitro and found that treated cells could stay in the cell cycle (G2/M), while the cancer cells could not enter mitosis. In addition, the expression levels of cadherin and the α5 integrin group were inhibited. Thus, the signal pathway mediated by the previous two substances was hindered. This may reduce the aggregation and recognition among cells and may induce apoptosis. However, the signal paths of key molecules need to be further discussed. 1.5.1.6 pH-Modified Pectin and its Anticancer Activities At present, high pH value modifications are commonly used to modify pectin. Pectin extracted by acid is treated by an alkali (generally pH 10) and incubated at a relatively mild temperature (50
60 C) for a period of time (generally 60 min). Then, the pH is adjusted to be acidic (generally pH 3.0). Pectin prepared by this method is often called pHmodified pectin (Pienta et al., 1995). Citrus pectin is often chosen as a raw material to make pH-modified pectin, and the modifications of other raw materials are limited. However, there has also been research on the adsorptive ability of pH-modified durian pectin for heavy metals. It found that the adsorptive effects of pH-modified pectin were improved (Wei et al., 2011). Most of the pH-modified citrus pectin research concentrated on the prevention and inhibition of cancer. pHmodified pectin has a good inhibitory effect on cancer cell’s metastasis and proliferation, and it promotes the apoptosis of cancer cells. Platt and Raz (1992) injected mice with cancer cells and found that commercial pectin with a relatively high molecular weight could promote the aggregation of the same types of cancer cells and the metastasis to the lungs, while pH-modified pectin of a relatively low molecular weight

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could inhibit metastasis to the lungs. Pienta et al. (1995) found that treating rats with oral pH-modified citrus pectin could significantly reduce the metastasis of MAT-LyLu prostate cancer cells to the lungs. Chauhan et al. (2005) found that GCS-100 modified citrus pectin could induce damage to the DNA in myeloma cells and activate caspase-8, caspase-3, and poly ADP-ribose polymerase. This induction could treat people who had demonstrated resistance to conventional and bortezomib therapies, and GCS-100 had no side effects on the normal viability of lymphocytes. GCS-100 basically induced apoptosis through the mitochondrial pathway, and it could also inhibit the apoptosis of refined multiple myeloma tumor cells from patients. Zhang et al. (2010) showed that the pH-modified citrus pectin had a strong inhibitory effect on H22 liver cancer cells, and the inhibition rate reached 47.8% at a high dose. 1.5.1.7 Thermal-Modified Pectin and its Anticancer Activities Jackson et al. (2007) found that commercial grade fractionated pectin powder could strongly promote prostate cancer cell apoptosis in male hormone-sensitive and male hormone-insensitive people, and produced thermal-modified pectin used in fractionated pectin powder production, which had similar effects on prostate cancer cell apoptosis. The heating temperatures were 100
132 C, and the heating time ranged from 20 min to 5.5 h. Jackson et al. (2007) used 30
60 min as the heating time and found no significant difference in the induction of apoptotic activity. They selected a heating time of 30 min and named the pectin produced under these conditions as thermal-modified pectin. The mechanisms that thermal-modified citrus pectin uses to induce the apoptosis of prostate cancer cells are not known, but exposing thermal-modified pectin to an alkali treatment significantly decreased the prostate cancer cells apoptosis-inducing activity, which indicates that the alkali-sensitive structures of thermal-modified pectin are required to induce the apoptosis of prostate cancer cells. This may be related to the thermal-modified citrus pectin’s ability to activate caspase-3 and poly ADP-ribose polymerase, promoting the apoptosis of prostate cancer cells. Determining whether there are other active fragments in thermal-modified pectin requires further study. Moreover, citrus pectin without thermal modifications could not induce the apoptosis of prostate cancer cells, and it has been hypothesized that the thermal modification can expose and enrich the pectin active fragments, which can induce apoptosis.

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The inhibition mechanisms of modified pectins on cancer cells are not known. The low molecular weight of modified pectin and its simple structure may make it more easily used by the body (Wai et al., 2010; Kidd, 1996; Jackson et al., 2007), or the inhibition mechanism may relate to galectin-3 in cancer cell surfaces. Modified pectin is rich in galactose, it has shorter chains, and it easily interacts with galectin-3. Thus, modified pectin can inhibit its antiadhesion and antiapoptotic properties.

1.5.2 Physicochemical Functions of Pectin 1.5.2.1 Viscosity-Associated Properties of Pectin Viscosity is the deformation of fluid under the action of external forces. The viscosity of pectin is related to the molecular weight of pectin, the extension of molecules in solution, the number of side chains, and the degree of methylation. In a pectin solution, pectin molecules can be connected by hydrogen bonds or hydrophobic interactions, resulting in different configurations. Pectin molecules with a low degree of esterification, because of the ionization, have negatively charged galactose acid residues, which can result in an electrostatic repulsion between pectin molecules. The electrostatic repulsion also has an effect on the viscosity of the pectin solution. Pectin solution viscosity can also be affected by the sugar and ion concentrations. Because of the presence of sugar, the hydration capability of pectin molecules may change, or pectin may interact with sugar molecules by hydrogen bonds to influence the pectin viscosity. The presence of ions that produce electrostatic shielding effects, to a certain extent, can eliminate the mutual exclusions between charged pectin molecules. 1.5.2.2 Gel Properties of Pectin Pectin can be used as a gelling agent in the food processing industry, especially in preserved fruits, jellies, and jams. Natural pectins with different degrees of esterification can be divided into high methoxyl pectin ($50%) and a low-methoxyl pectin (#50%). High methoxyl pectin can form a gel under acidic conditions (pH 2.0
3.5) when sucrose is added to a concentration greater than 55% in which hydrogen bonds and hydrophobic interactions play important roles in gel formation. The pH value range in which low methoxyl pectin forms a gel is wide (2.0
6.0) and metal ions, such as calcium, are essential for gel formation. This is due to the low degree of esterification of

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low methoxyl pectin and the great amount of free carboxyls in the pectin molecule. The inter- or intrachain carboxyl groups are connected to each other by calcium ions to form the gel. Internal factors, such as the contents of galacturonic acid and neutral sugars, the degree of esterification, the degree of acetylation, and molecular weight (Yapo and Koffi, 2008; O’Donoghue and Somerfield, 2008), and also various external factors, such as the ash content and ionic strength in a pectin solution, also affect the gelation of pectin. Among the factors, contrary to the influences of the galacturonic acid content, degree of esterification and molecular weight, and increasing the amounts of other factors had a negative influence on gel formation (Miyamoto and Chang, 1992; Pippen and Mccready, 1950; Sosulski et al., 1978). Pectin extracted from chicory, pumpkin, and sugar beet by-products had a high degree of acetylation, high neutral sugar content, and low molecular weight (Robert et al., 2006; Shkodina et al., 1998; Yapo et al., 2007) and showed poor gelling properties and needed structural modifications. In addition, pectin gel properties were also influenced by the pH value, temperature, pectin concentration, coexisting solutes, such as sucrose, and the concentration of ions, such as calcium (Thakur et al., 1997). 1.5.2.3 Emulsifying Properties of Pectin In addition to soybean soluble polysaccharide, gum arabic, and guar gum, pectin is also a nature polysaccharide which has emulsifying abilities (Leroux et al., 2003; Nakauma et al., 2008). The emulsifying properties of pectin are mainly described by the particle size of the emulsion, emulsifying activity, emulsion stability, and emulsion viscosity. In previous studies, the emulsifying properties of citrus and sugar beet have been reported. Akhtar et al. (2002) found that with an increase in the citrus pectin concentration, the emulsion stability gradually increased. Bueno et al. (2008) used light microscopy to observe fresh citrus pectin and soybean protein emulsions at pH 6.2 and found that, compared with soybean protein emulsion, the emulsion particles of the citrus pectin were highly uniform with a smaller particle size. Yapo et al. (2007) showed that when the pectin concentration and oilphase volume fractions increasing, changes in the sugar beet pectin emulsion particle diameter showed an “inverted trapezoid” trend. When the pectin concentrations were between 1.0% and 3.0% and the

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oil phase volume fractions were between 20% and 50%, the particle size did not change significantly. Nakauma et al. (2008) compared the emulsion particle diameters and viscosity of sugar beet pectin, soybean soluble polysaccharides, and arabic gum and found that the emulsifying ability of sugar beet pectin was the strongest, and the emulsion ability of 1% sugar beet pectin was equal to the emulsion abilities of 5.0% soybean soluble polysaccharide and 10% arabic gum. Leroux et al. (2003) showed that at the same pectin concentration in the same oil phase volume fraction, the emulsion particle size of sugar beet pectin was less than that of citrus pectin, and the proportion of the emulsion particles absorbed on the sugar beet pectin was greater than that of the orange pectin. Sharma et al. (1998) discussed the effects of the oil phase volume fraction on tomato pectin emulsion’s stability. The results showed that the emulsions in the high oil phase volume fraction were more stable than the emulsions in the low oil phase volume fraction. In addition to the pectin concentration and oil phase volume fraction, the emulsifying properties of pectin were influenced by pH value, NaCl concentration, and calcium concentration (Nakauma et al., 2008). In addition, pectin can be added to protein solutions to enhance the stability of the emulsion and reduce its particle size. Pectin and protein can be connected by hydrogen bonds or covalent bonds, and the contents of hydrophobic groups in proteins were high, which can fully wrap the surfaces of the oil droplets. While the hydrophilicity of pectin is strong, it can extend into the aqueous solution. The emulsion particles formed in this way not only have strong hydrophobic proteins but also have strong hydrophilic pectin at the surface, which greatly enhances the stability of the emulsion (Mishra et al., 2001; Neirynck et al., 2004). Thus, pectin has been widely used as an emulsifier, gelling agent, and thickening agent in the food processing industry to increase the viscosity, produce colloidal structures, control crystallization, prevent dehydration, improve textures, prevent the loss of flavor, and prolong the stability of food systems. In the pharmaceutical industry, pectin can be used alone or mixed with other excipients to produce ointments, film agents, suppositories, and microcapsules. In addition, the adsorptive capability of pectin for metal ions can be used to remove heavy metal ions during waste-water treatment.

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SECTION 2: PRODUCTION TECHNOLOGY OF SWEET POTATO PECTIN At present, acid extraction is commonly used to extract pectin during industrial production. Pectin is often extracted from industrial byproducts, such as orange peel, apple pomace, and banana peel. Extracting pectin from sweet potato residue as a high added-value product can increase the utilization rates of raw materials and reduce the waste of resources and production of environmental pollutants, which has important practical significance. Our laboratory has carried out related research. We used hydrochloric acid as solvent to extract pectin from sweet potato pulp and used the pectin extraction ratio as an indicator to optimize the process. The maximum extraction ratio was 65.86% (Wei et al., 2008). However, the pectin extraction ratio did not accurately reflect the impact of pectin extraction conditions on the yield or its galacturonic acid content. FAO regulates that pectin with a galacturonic acid content greater than 65% can be used as an additive. Therefore, we used hydrochloric acid to extract pectin from sweet potato pulp. The pectin yield and galacturonic acid content were used as evaluation indices for response surface analysis. We will introduce the conditions that produced a high yield of pectin with a high galacturonic acid content to provide a theoretical basis for sweet potato pectin processing.

2.1 EXTRACTION PROCESS OF PECTIN FROM SWEET POTATO In total, 10 g of sweet potato pulp (W0) was mixed with distilled water at a certain solid to liquid ratio in a conical flack to produce a suspension liquid. Then, 1 mol/L hydrochloric acid was used to adjust the suspension to the required pH and the mixture was placed in a thermostat oscillator, which was set to a suitable temperature for a suitable period of time. After that, the mixture was centrifuged at 6148 3 g for 30 min to obtain a supernatant. The supernatant was concentrated in a rotary evaporator and treated with 1 mL 0.1% α-amylase at 60 C for 1.5 h to remove the starch from the samples. The supernatant was boiled at 100 C for 10 min to inactivate the enzymes. The enzymatic hydrolysis was precipitated by a 33 volume

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of the supernatant overnight and then centrifuged at 6148 3 g for 30 min to obtain the precipitate. The precipitate was then washed using 70%, 80%, and 90% (v/v) ethanol. After washing, the pectin was dissolved in deionized water, filtered using membranes with 10,000 Da cut-off, and then the resulting solution was freeze dried to obtain the pectin. Y1 5

W1 3 100 W0

(4.1)

in which Y1 represents pectin yield, %; W0 represents the mass of sweet potato pulp, g; and W1 represents the mass of sweet potato pectin, g.

2.2 THE DETERMINATION OF THE GALACTURONIC ACID CONTENT IN THE PECTIN SOLUTION The purity of pectin is often expressed by the content of galacturonic acid, and the carbazole sulfuric acid method is commonly used to determine the content of galacturonic acid in pectin. 1. The principle of the galacturonic acid content determination by the carbazole sulfuric acid method Under acidic conditions, pectin can hydrolyze to generate galacturonic acids. The galacturonic acids will combine with carbazole to form a purple compound that has a strong absorption at 530 nm. The standard curve is produced by measuring the absorbance values of galacturonic acid solutions of different known concentrations. Thus, the concentration of galacturonic acids can be determined by comparing the absorbance value of the solution to the standard. 2. The production of a standard curve of known galacturonic acid solutions and their absorbance values Aliquots of 1.0 mL galacturonic acid solutions, with concentrations ranging from 10 to 90 μg/mL, and 6.0 mL of concentrated sulfuric acid were placed in an ice bath for thoroughly mixing, and then the mixture was placed in a boiling water bath for 15 min, followed by cooling to room temperature. After adding 0.5 mL of 0.15% carbazole, the mixtures were mixed well at room temperature for 30 min while the color developed. A standard curve was produced, and the regression equation was y 5 0.0056x 1 0.0061, with R2 5 0.9971. 3. Determination of the galacturonic acid content

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Samples of 0.1 g freeze-dried pectin were dissolved in distilled water to a final solution volume of 250 mL. The solution was diluted 100 times with distilled water before the test. Then, 1 mL of diluted pectin solution and 6 mL concentrated sulfuric acid were mixed together in an ice bath, after which the mixture was placed in a boiling water bath for 15 min. The mixture was allowed to cool to room temperature. Then, 0.5 mL of 0.15% carbazole solution was added to the mixture and the mixture was kept for 30 min at room temperature to allow the color to develop. The absorbance value was determined at 530 nm. Lastly, based on the regression equation, the galacturonic acid content of the diluted solution was calculated, and then, the total content of galacturonic acid in solution was obtained by multiplying the diluted factor.

2.3 FACTORS AFFECTING THE YIELD AND GALACTURONIC ACID CONTENT OF SWEET POTATO PECTIN 2.3.1 Effects of Temperature on the Extraction Yield and Galacturonic Acid Content of Sweet Potato Pectin

As shown in Fig. 4.4, as the temperature increased in the 60
90 C range, the pectin yield increased significantly (P , 0.05) and reached a the maximum value at 90 C. At higher temperatures, the pectin yield

Figure 4.4 Effects of temperatures on the yield and the galacturonic acid content of sweet potato pectin. Values followed by the same letter on the same curve are not significantly different (P . 0.05).

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showed no significant increase (P . 0.05). As the temperature increased in the 60
90 C range, the galacturonic acid content also increased significantly (P , 0.05), reaching a maximum value of 69% at 90 C. At higher temperatures, the galacturonic acid content decreased significantly (P , 0.05). At 90 C, both the galacturonic acid content and the pectin yield reached their maximum values.

2.3.2 Effects of pH Value on the Extraction Yield and Galacturonic Acid Content of Sweet Potato Pectin As shown in Fig. 4.5, as pH values increased, but remained less than 1.5, the pectin yield significantly increased. When the pH values were at 1.5
2.0, the pectin yield showed no significant difference (P . 0.05). When the pH value was higher than 2.0, the pectin yield decreased significantly (P , 0.05). There were no significant differences in the galacturonic acid contents when the pH values were in the 1.0
2.0 range (P . 0.05). With an increase in the pH values (2
4), the galacturonic acid content decreased significantly (P , 0.05). This suggested that pectin was suitable to be extracted at a low pH value and that higher pH values were not beneficial to dissolving and extracting pectin.

Figure 4.5 Effects of pH values on the yield and the galacturonic acid content of sweet potato pectin. Values followed by the same letter on the same curve are not significantly different (P . 0.05).

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Figure 4.6 Effects of extraction time on the pectin yield and the galacturonic acid content of sweet potato pectin. Values followed by the same letter on the same curve are not significantly different (P . 0.05).

2.3.3 Effects of Extraction Time on the Yield and the Galacturonic Acid Content of Sweet Potato Pectin As shown in Fig. 4.6, in a period of time (1
2 h), the pectin yield increased with the prolonged time, reaching a maximum value of 5.01% at 2 h. The pectin yield decreased significantly (P , 0.05) when the extraction time was prolonged. With an increase in the extraction time (1
2.5 h), the galacturonic acid content significantly increased (P , 0.05). When the time was extended to 3 h, the galacturonic acid content of showed no difference (P . 0.05). As the time was further prolonged (3
4 h), the galacturonic acid content decreased significantly (P , 0.05). This indicated that a long extraction time increased free pectin hydrolysis, which resulted in a decrease in the pectin yield and galacturonic acid content.

2.3.4 Effects of the Solid/Liquid Ratio on the Pectin Yield and the Galacturonic Acid Content of Sweet Potato Pectin With an increase in the solid/liquid ratio from 15:1 to 30:1, the pectin yield significantly increased (P , 0.05) and reached a maximum value of 4.97% at 30:1. A further increase of the solid/liquid ratio to more than 30:1 resulted in no significant difference in the pectin yield (P.0.05) (Fig. 4.7). When increasing the solid/liquid ratio from 15:1

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Figure 4.7 Effects of the solid/liquid ratio on the pectin yield and the galacturonic acid content of sweet potato pectin. Values followed by the same letter on the same curve are not significantly different (P . 0.05).

to 30:1, the galacturonic acid content significantly increased (P,0.05) and reached a maximum value at the 30:1 solid/liquid ratio. The galacturonic acid content showed no significant change when the solid/liquid ratio continued to be increased (P . 0.05). When the solid/liquid ratio was at 30:1, the pectin yield and the galacturonic acid content reached their maximum values.

2.4 OPTIMIZATION OF PECTIN EXTRACTION FROM SWEET POTATO PULP An analysis of the experimental results was carried out using Design Expert software. The variations of the responses, the sweet potato pectin yield (%) and the galacturonic acid content (%), were determined as functions of the independent variables, including extraction temperature, extraction time, pH value, and solid/liquid ratio. Four factors with three levels of response surface analysis tests were designed. Based on the results of the single factor experiments, the range and the levels of the four independent variables are shown in Table 4.1. Response surface design and test results are shown in Table 4.2. Design Expert software (Version 7.0) was used for the multiple regression fitting. After considering the requirements of practical operations, the best conditions were determined as follows: the temperature

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Table 4.1 Independent Variables and Their Levels for Box
Behnken Design X1

Factor Level

X2

X3

X4

Temperature ( C)

Time (h)

pH Value

Solid/Liquid Ratio (mL/g)



21

80

1

1

20

0

90

2

2

25

1

100

3

3

30

Table 4.2 Response Surface Design and Experimental Results Run

Factors

Yield (%)

Galacturonic Acid Content (%)

X1

X2

1

1

0

0

1

4.94

67.90

2

0

0

21

1

4.70

67.35

3

0

0

0

0

5.00

68.02

4

1

0

1

0

4.21

59.88

5

0

0

1

21

4.19

60.30

6

21

1

0

0

4.40

64.87

7

0

21

1

0

4.11

58.00

8

0

1

1

0

4.05

59.05

9

21

0

1

0

3.75

56.20

10

0

0

21

21

4.67

67.33

11

21

0

21

0

4.34

64.24

12

21

0

0

1

4.55

65.12

13

0

1

0

1

4.87

67.56

14

0

21

0

21

4.80

65.90

15

0

0

0

0

5.05

68.18

16

1

1

0

0

4.78

66.28

X3

X4

17

0

1

21

0

4.68

66.73

18

1

21

0

0

4.65

65.92

19

1

0

0

21

4.93

67.55

20

0

0

0

0

5.02

68.00

21

0

1

0

21

4.75

67.58

22

0

21

21

0

4.31

64.54

23

0

0

0

0

5.00

67.95

24

0

21

0

1

4.79

66.50

25

21

0

0

21

4.50

64.72

26

0

0

1

1

4.23

60.90

27

0

0

0

0

5.07

68.15

28

21

21

0

0

4.30

61.51

29

1

0

21

0

4.60

66.42

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was 93 C, the extraction time was 2.2 h, the pH value was 1.7, and the solid/liquid ratio was 30:1. Under the optimized conditions, the pectin yield and the galacturonic acid content were 5.09% and 70.03%, respectively, which met the FAO regulations for pectin additives (galacturonic acid content $ 65%). This technology can meet the requirements of industrial production with low-cost raw materials and an easy production process. There are broad market prospects for this technology.

SECTION 3: BIOLOGICAL ACTIVITIES OF SWEET POTATO PECTIN We mentioned in the first section that pectin could inhibit the growth and metastasis of cancer cells and promote cancer cell apoptosis; however, high molecular weights or poor water solubility levels may hinder or limit the anticancer effects. Modifications can produce pectin with shorter chains and fewer sugar branches, reduce the molecular weight and the degree of the esterification, increase the galacturonic acid content, and increase the water solubility of the pectin. In this section, we will describe the effects of pH- and thermal-modified pectins on the proliferation, inhibition, and migration of HT-29 colon cancer cells, Bcap-37breast cancer cells, and SMMC-7721liver cancer cells.

3.1 THE PREPARATION OF pH-MODIFIED PECTIN The experiment was carried out using the Platt and Raz (1992) method with slight modifications. The specific steps were as follows: a sample of 1.5 g unmodified sweet potato pectin was used as the raw material and mixed with 100 mL water to obtain a 1.5% solution. Then, 3 mol/L NaOH solution was slowly added to adjust the pH to 10.0. The mixture was heated at 55 C for 30 min and then cooled to room temperature. The mixture was allowed to stand overnight at room temperature after adjusting the pH to 3.0 using 3 mol/L hydrochloric acid. After adding an equal volume of 95% ethanol, the mixture was placed in the refrigerator (220 C) for 2 h, then the mixture was filtered, and the entrapped solids were collected to be dissolved in deionized water. The solution was dialyzed against water using membranes with a 3.5-kDa cut-off to remove the salts, and then it was freeze dried. This produced pH-modified pectin.

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3.2 THE PREPARATION OF THERMAL-MODIFIED PECTIN The preparation of thermal-modified pectin was performed according to the method of Jackson et al (2007). Briefly, 1 g unmodified sweet potato pectin was dissolved in deionized water to produce a 0.1% solution. The mixture was heated under the conditions of 123.2 C at 17.2
21.7 psi for 60 min and was then cooled to room temperature. The mixture was allowed to stand overnight at 4 C. The supernatant was obtained and freeze dried to produce thermal-modified pectin.

3.3 THE CELL CULTURES The HT-29 human colon, Bcap-37 breast, and SMMC-7721 liver cancer cells were placed in McCoy’s 5 A culture medium supplemented with 10% fetal bovine serum and 1% each penicillin and streptomycin. The medium was cultured in an incubator under the conditions of 5% CO2 and 37 C. The culture medium was replaced every 2 days and passaged once every 4 days. In general, one culture should be passaged into three new cultures.

3.4 EFFECTS OF SWEET POTATO PECTIN ON CANCER CELL SURVIVAL RATES As shown in Table 4.3, the medium was treated with 1.00 mg/mL sweet potato pectin for 48 h. The survival rates of the HT-29, SMMC7721, and Bcap-37 cancer cells were not different than those of the control group (without sweet potato pectin) (P , 0.05). Cancer cells treated with unmodified pectin, pH-modified pectin and thermalmodified pectin all had survival rates of greater than 90%. This suggested that pectin had no influence on the survival rates of cancer cells. When the concentration of pectin was less than 1.00 mg/mL and Table 4.3 Effects of Sweet Potato Pectin on Survival Rates of Cancer Cells (%) Pectin Types

Pectin Concentration (mg/mL)

Cancer Cells Types HT-29

Bcap-37

SMMC-7721

Survival Rates of Cells (%) Control group

0.00

95.60 6 1.21

98.71 6 2.31

96.52 6 1.68

Unmodified pectin

1.00

97.21 6 2.05

98.89 6 1.87

93.98 6 2.68

pH-modified pectin

1.00

93.84 6 0.53

99.13 6 2.76

92.74 6 1.14

Thermal-modified pectin

1.00

97.49 6 2.02

94.87 6 3.12

93.68 6 1.81

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the treatment time was 48 h, pectin showed no toxicity towards cancer cells. This was consistent with the results of Inohara et al. (1994). Thus, pectin may inhibit the growth of cancer cells by enhancing the immune functions of the body.

3.5 EFFECTS OF SWEET POTATO PECTIN ON CANCER CELL PROLIFERATION 3.5.1 Effects of Sweet Potato Pectin on the Proliferation of HT-29 Colon Cancer Cells The inhibitory effects of three types of sweet potato pectins on the proliferation of HT-29 cells all occurred in concentration and timedependent manners. When HT-29 cells were treated with different concentrations of pectins (0.01
1.00 mg/mL), compared with unmodified pectin, the inhibitory effects of the thermal- and pH-modified pectins increased significantly (P , 0.05). When the pectin concentration was 1.00 mg/mL and the treated time was 24 h, the inhibition rates of the unmodified pectin, thermal-modified pectin, and pH-modified pectin were 46.64%, 67.77%, and 75.10%, respectively (Fig. 4.8). Compared with the unmodified pectin, the inhibition rate of thermal-modified pectin increased by 45.32% and that of pH-modified pectin increased by 61.03%. As seen in Fig. 4.9, under different treatment times (12
48 h), the effects of thermal- and pH-modified pectins on the inhibitory effects of HT-29 cells were significantly greater than that of

Figure 4.8 Effects of pectin concentrations on HT-29 colon cancer cells (24 h). Values followed by the same letter for the same kind of pectin are not significantly different (P . 0.05).

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Figure 4.9 Effects of pectin (1.00 mg/mL) treatment time on HT-29 colon cancer cells. Values followed by the same letter for the same kind of pectin are not significantly different (P . 0.05).

the unmodified pectin (P , 0.05). For thermal-modified pectin, with the extension of the treatment time, the inhibitory effect increased, but when the treatment time was longer than 24 h, the inhibitory effects had no significant differences (P . 0.05). For pH-modified pectin, with the extension of the treatment time, the inhibitory effect increased, and at 36 h, the inhibitory effect was the greatest. These two kinds of modification methods significantly increased the inhibitory effects of sweet potato pectin on HT-29 human colon cancer cells, and at a high concentration, the inhibitory effect of pH-modified sweet potato pectin was significantly higher than that of thermal-modified sweet potato pectin (P , 0.05).

3.5.2 Effects of Sweet Potato Pectin on Bcap-37 Breast Cancer Cell Proliferation The inhibitory effects of three types of sweet potato pectins on Bcap37 cell proliferation occurred in concentration and time-dependent manners. When treating Bcap-37 cells with different concentrations of pectin (0.01
1.00 mg/mL), compared with the unmodified pectin, the inhibitory effects of thermal- and pH-modified pectins on Bcap-37 cells significantly increased (P , 0.05). The inhibition rates of unmodified, thermal-modified, and pH-modified pectins on Bcap-37 proliferation were 42.64%, 61.10%, and 60.77%, respectively, when the pectin

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Figure 4.10 Effects of pectin concentrations on Bcap-37 breast cancer cells (24 h). Values followed by the same letter for the same kind of pectin are not significantly different (P . 0.05).

Figure 4.11 Effects of pectin (1.00 mg/mL) treatment times on Bcap-37 breast cancer cells. Values followed by the same letter for the same kind of pectin are not significantly different (P . 0.05).

concentration was 1.00 mg/mL, and the treatment time was 24 h (Fig. 4.10). Compared with the unmodified pectin, the inhibition rate of thermal-modified pectin increased by 43.31%, while the rate of pHmodified pectin increased by 42.54%. As shown in Fig. 4.11, at different treatment times, the inhibitory effects of thermal-modified and pH-modified pectins on Bcap-37 cell proliferation were significantly greater than that of unmodified pectin (P , 0.05). With the extension

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of time, the inhibitory effects of thermal-modified pectin increased and reached a maximum value at 24 h. The inhibitory effects of pHmodified pectin increased as the treatment time increased. The effects showed no significant differences when the treatment time was greater than 24 h (P . 0.05). Thus, both modification methods enhanced the inhibitory effects of pectin on Bcap-37 cells and under a high concentration, the effects of pH-modified and thermal-modified pectins were not significantly different (P . 0.05).

3.5.3 Effects of Sweet Potato Pectin on SMMC-7721 Liver Cancer Cell Proliferation As shown in Figs. 4.12 and 4.13, the inhibitory effects of the three types of pectin on SMMC-7721 cell proliferation occurred in dose and time-dependent manners. Compared with unmodified pectin, the inhibitory effects of thermal-modified and pH-modified pectins on SMMC7721 cell proliferation significantly increased (P , 0.05). When the pectin concentration was 1.00 mg/mL and the treatment time was 24 h, the inhibitory rates of unmodified, thermal-modified, and pH-modified pectins were 21.57%, 24.62%, and 29.68%, respectively (Fig. 4.12). After thermal modifications, the inhibitory rate increased by 14.16%, while the inhibitory rate of the pH-modified pectin increased by 37.59%. Fig. 4.13 shows that during different treatment times

Figure 4.12 Effects of pectin concentrations on SMMC-7721 liver cancer cells (24 h). Values followed by the same letter for the same kind of pectin are not significantly different (P . 0.05).

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Figure 4.13 Effects of pectin (1.00 mg/mL) treatment times on SMMC-7721 liver cancer cells. Values followed by the same letter for the same kind of pectin are not significantly different (P . 0.05).

(12
48 h), the inhibitory functions of thermal-modified and pHmodified pectins were greater than that of the unmodified pectin (P , 0.05). With the increase in treatment time, the inhibitory effects of thermal-modified pectin improved greatly and the best effect occurred at 24 h. With the increase in treatment time, the inhibitory effects of pH-modified pectin increased. The inhibitory effect was enhanced when the treatment time was less than 24 h, and the inhibition rate decreased significantly because of shortening treatment time (P,0.05). When the time was longer than 24 h, the inhibition rates were not significantly different at longer treatment times (P . 0.05). Thus, these two kinds of modification methods significantly increased the inhibitory effects of sweet potato pectin on SMMC-7721 liver cancer cells, and the inhibitory effect of pH-modified pectin was significantly greater than that of the thermal-modified pectin (P , 0.05).

3.6 EFFECTS OF SWEET POTATO PECTIN ON CANCER CELL METASTASIS 3.6.1 Effects of Sweet Potato Pectin on Cancer Cell Adhesion After the digestion of cultured HT-29 and Bcap-37 cells, they were repeatedly pipetted to distribute them. After counting cells with an inverted microscope, cell concentrations were adjusted. The cell suspension with the density of 5 3 105 (for every hole) was injected into the Tissue Culture Plate 12 (1.0 mL solution in a hole). The plate was placed in an incubator under conditions of 5% CO2 and 37 C for 24 h.

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215

Then, the culture medium was abandoned and the plate was cleaned with PBS solution. To each plate, 1.0 mL of fresh samples (0.1, 0.25, 0.5 and 1 mg/mL) and culture medium were added, except for the control group, and all of the groups contained 100 ng/mL of PMA (a cancer promoting agent that can strongly promote the metastasis of cancer cells). The plates were cultured under the same conditions for another 24 h. Then, the culture medium was abandoned, and the plates were washed twice with PBS solution, and 600 μL of pancreatin was needed to digest the samples in each well. Finally, the tissue culture plates were shaken at 100 rpm at room temperature and the time (min) to completely digest cancer cells was calculated. 3.6.1.1 Effects of Unmodified Sweet Potato Pectin on Cancer Cell Adhesion In Fig. 4.14, the effects of unmodified pectin on the adhesion of HT-29 and Bcap-37 cells after a 24 h treatment are seen. The complete digestion time of cancer cells treated only with PMA was significantly longer than those of the control groups and the treatment groups that were treated with pectin. In the pectin-treated groups, as the pectin concentration (0.1, 0.25, 0.5, and 1 mg/mL) increased, the adhesion effects of cancer cells significantly decreased (P , 0.05). For HT-29, the complete digestion time of the group treated with PMA (without pectin) was 12.4 min, while the time of the pectin-treated group (1 mg/mL) was

Figure 4.14 The effects of unmodified pectin on the adhesive capabilities of Bcap-37and HT-29 cancer cells. Values followed by the same letter for the same kind of cancer cell are not significantly different (P . 0.05).

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9.1 min, which suggested that the full digestion time of cells treated with 1 mg/mL pectin solution could be reduced by 26.6%. For Bcap-37 cells, the complete digestion time of the PMA only group was 10.2 min; however, the time of the pectin-treated group (1 mg/mL) was 8.2 min, which indicated that the time of complete digestion with a pectin treatment could be reduced by 19.6%. Thus, unmodified sweet potato pectin could reduce the adhesion effects of HT-29 and Bcap-37 cancer cells and that its antiadhesion effect on HT-29 was greater than the effect on Bcap-37.

3.6.1.2 Effects of Thermal-Modified Sweet Potato Pectin on Cancer Cell Adhesion Fig. 4.15 shows the effects of thermal-modified sweet potato pectin on the adhesive capabilities of HT-29 and Bcap-37 cells after a 24 h treatment. The complete digestion time for cancer cells only treated with PMA was significantly longer than those of the control and the pectintreated groups (P , 0.05). When the cancer cells were treated with increasing pectin concentrations (0.1, 0.25, 0.5, and 1 mg/mL), the adhesive capabilities of cancer cells were significantly decreased (P , 0.05). When the pectin concentration was less than 0.5 mg/mL, compared with the control group (without pectin and PMA), the complete cell digestion time of was significantly longer than that of the control group (P , 0.05). In addition, when the concentration of pectin

Figure 4.15 The effects of thermal-modified pectin on the adhesive capabilities of Bcap-37 and HT-29 cancer cells. Values followed by the same letter for the same kind of cancer cell are not significantly different (P . 0.05).

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increased to 1 mg/mL, the complete digestion times of the pectintreated and control groups showed no significant differences (P.0.05). For HT-29, in the PMA-treated group, the complete cell digestion time was 12.4 min; however, the complete digestion time of cells treated with 1 mg/mL of thermal-modified pectin was 8.5 min. Thus, the complete digestion time decreased by 31.45%. For Bcap-37, in the PMA-treated group, the complete cell digestion time was 10.2 min, while the complete digestion time of cells treated with 1 mg/mL of thermal-modified pectin was 6.5 min, which meant that the complete digestion time was reduced by 36.23%. Compared with unmodified pectin, the digestion times of HT-29 and Bcap-37 cells treated with thermal-modified pectin were reduced by 6.6% and 20.73%, respectively. Thus, sweet potato pectin with thermal modifications could reduce the adhesion capabilities of cancer cells (especially for Bcap-37 cells), and this effect was superior to the effect of unmodified pectin. 3.6.1.3 Effects of pH-Modified Sweet Potato Pectin on Cancer Cell Adhesion In Fig. 4.16, the effect of pH-modified pectin on the adhesive capabilities of HT-29 and Bcap-37 cells after 24 h treatments is seen. The time required for the complete digestion of cancer cells only treated with PMA was significantly longer than that of the control and the pectin treatment groups (P,0.05). Among the pectin treatment groups, an

Figure 4.16 The effects of pH-modified pectins on the adhesive capabilities of Bcap-37and HT-29 cancer cells. Values followed by the same letter for the same kind of cancer cell are not significantly different (P . 0.05).

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increase in the pectin concentration (0.1, 0.25, 0.5, 1 mg/ml), significantly reduced the adhesive capabilities of cancer cells (P , 0.05). When the pectin concentrations were less than 0.5 mg/mL, compared with the control group, the complete cell digestion time was longer than that of the control group (P , 0.05), while as the concentration increased to 1 mg/mL, the digestion time between pH-modified pectin and control groups showed no significant difference (P . 0.05). For HT-29 cells, in the PMA-treated group, the complete cell digestion time was 12.4 min; however, the complete digestion time of cells treated with 1 mg/mL of pH-modified pectin was 8.4 min. Thus, the complete digestion time was reduced by 32.25%. For Bcap-37 cells, in the PMA-treated group, the complete cell digestion time was 10.2 min, while the complete digestion time for cells treated with 1 mg/mL of pH-modified pectin was 6.4 min, which meant that the complete digestion time was reduced by 37.25%. When compared with unmodified pectin, the pH-modified pectin decreased the digestion times of HT-29 cells and Bcap-37 cells by 7.69% and 21.95%, respectively. Thus, the pH-modified pectin could reduce the adhesive capabilities of HT-29 and Bcap-37 cancer cells, and the effect was particularly significant for Bcap-37 (P , 0.05).

3.6.2 The Effects of Sweet Potato Pectin on Cancer Cell Migration A cell suspension of 5 3 105 cells per well was injected into the tissue culture plates, and 1 ml solution was added to every well. The plates were incubated under 5% CO2 and 37 C conditions for 24 h. After that, the culture medium was abandoned, and the cell monolayer of cells was removed by scraping with yellow micropipette tips. The castoff cells were washed twice with PBS and blotted. Then, 1.0 mL of freshly prepared samples (0.1, 0.25, 0.5, and 1 mg/mL) and culture medium were added, except for the control group. All of the groups were treated with 100 ng/ml of PMA. The cells were placed under an electron microscope and images at 1003 magnification were taken. The cells were incubated in the incubator (37 C and 5% CO2) for another 24 h, washed with PBS twice and fixed in 4% paraformaldehyde. The cells were placed under the electron microscope and images at 1003 magnification were taken. NIH ImageJ software was used to analyze the images of cells after 0 h and 24 h incubation times and measure the widths as L0 and L24, respectively.

Sweet Potato Pectin

The migration rate ð%Þ 5

L0 2 L24 L0

219

(4.2)

3.6.2.1 Effects of Unmodified Sweet Potato Pectin on Cancer Cell Migration The migration rate of cells treated only with PMA was significantly higher than those of the control and pectin-treated groups. In the groups receiving pectin treatments, as the pectin concentration increased, the cancer cell migration rates significantly decreased (P , 0.05). When compared with the PMA-treatment group, the cancer cell migration rate significantly decreased (P , 0.05), but was higher than that of the control group (P , 0.05). For the HT-29 cells (Fig. 4.17), the PMA group’s migration rate was 40.76%, while that of the pectin-treated group (1 mg/mL) was 27.98%. Thus, the migration rate was reduced by 31.35%. For the Bcap-37 cells (Fig. 4.18), the migration rate of the PMA-treated group was 52.35%, and the migration rate of cells treated with 1 mg/mL pectin was 39.64%, which meant that the migration rate decreased by 24.28%. Thus, it can be concluded that a treatment of unmodified pectin could reduce the migration rates of cancer cells. 3.6.2.2 Effects of Thermal-Modified Sweet Potato Pectin on Cancer Cell Migration In the PMA group, in which the cells were only treated with PMA, the migration rate was much higher than that of the control and the

Figure 4.17 Inhibitory effects of unmodified pectin on HT-29 cells after a 24 h-treatment. Values followed by the same letter are not significantly different (P . 0.05).

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Figure 4.18 Inhibitory effects of unmodified pectin on Bcap-37 cells after a 24 h-treatment. Values followed by the same letter are not significantly different (P . 0.05).

HT-29 cells migraton rates˄%˅

45

a

40

b

35

c

30 d

25

e

20 15

f

10 5

0 Pectin(mg/ml) 1– PMA –

–2 +

3 +0.1 +

4 +0.25 +

5 +0.5 +

6 +1.0 +

Figure 4.19 Inhibitory effects of thermal-modified pectin on HT-29 cells after a 24-h treatment. Values followed by the same letter are not significantly different (P . 0.05).

thermal-modified pectin groups. In the thermal-modified pectin-treated group, as the pectin concentration increased, the cell migration rate significantly decreased (P , 0.05), but the rate was still faster than that of the control group (without pectin and PMA; P , 0.05). For HT-29 cells (Fig. 4.19), the migration rates of PMA only treated group and the 1 mg/mL thermal-modified pectin treatment group were 40.76% and 18.66%, respectively. Thus, thermal-modified pectin reduce the

Sweet Potato Pectin

221

Bcap-37 cells migration rates˄%˅

60 a b

50

c d

40 30 20

e f

10

0 Pectin(mg/ml)1– PMA –

2– +

3 +0.1 +

4 +0.25 +

5 +0.5 +

6 +1.0 +

Figure 4.20 Inhibitory effects of thermal-modified pectin on Bcap-37 cells after a 24 h-treatment. Values followed by the same letter are not significantly different (P . 0.05).

rate by 54.22%. For Bcap-37 cells (Fig. 4.20), the migration rate of the PMA group was 52.35%, while the rate of the pectin-treated group (1 mg/mL) was 25.98%. Thus, the migration rate was reduced by 50.37%. When the thermal-modified pectin concentration was 1 mg/mL, compared with the unmodified pectin, the migration rates of HT-29 and Bcap-37 cells decreased by 33.31% and 34.46%, respectively. This suggested that the thermal-modified sweet potato pectin could inhibit the migration of cancer cells and that the effects were better than that of the unmodified pectin. 3.6.2.3 Effects of pH-Modified Sweet Potato Pectin on Cancer Cell Migration After treating only with PMA, the migration rates of cancer cells were significantly faster than those of the control and pH-modified pectin treatment groups. In the groups treated with pH-modified pectin, the cell migration rates decreased while the pectin concentration increased (P , 0.05). When the pectin concentration was less than 0.5 mg/mL, the migration rate was still faster than that of the control group (P , 0.05). When the pectin concentration was increased to 1 mg/mL, the migration rate was not significantly different from the control group (P . 0.05). For HT-29 cells (Fig. 4.21), the migration rate of the PMA group was 40.76%, however, the migration rate of the pHmodified pectin-treated group (1 mg/mL) was 14.45%. Thus, the migration rate was reduced by 64.55%. For Bcap-37 cells (Fig. 4.22),

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HT-29 cells migration rates˄%˅

45

a

40

b

35 c

30 25

d

20 15

e

e

10 5

0 Pectin(mg/ml)0.44 – PMA –

0.5 – +

0.5508 +0.1 +

0.442 +0.25 +

0.4 +0.5 +

0.45 +1.0 +

Figure 4.21 Inhibitory effects of pH-modified pectin on HT-29 cells after a 24 h-treatment. Values followed by the same letter are not significantly different (P . 0.05).

60 Bcap-37 cells migration rates˄%˅

a 50

b c

40

d

30 20

e

e

10

0 Pectin(mg/ml) 1– PMA –

–2 +

3 +0.1 +

4 +0.25 +

5 +0.5 +

6 +1.0 +

Figure 4.22 Inhibitory effects of pH-modified pectin on Bcap-37 cells after a 24 h-treatment. Values followed by the same letter are not significantly different (P . 0.05).

the migration rates of the PMA and pH-modified pectin-treated groups (1 mg/mL) were 52.35% and 19.64%, respectively, and the migration rate was decreased by 62.48% by the pH-modified pectin. Compared with the migration rates of unmodified pectin, after pH modification, the migration rates of HT-29 and Bcap-37 cells were reduced by 48.36% and 50.45%, respectively. pH-modified pectin could greatly decrease the migration rates.

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3.6.3 Effects of Sweet Potato Pectin on the Urokinase-Type Plasminogen Activator (uPA) Content in Cancer Cells The HT-29 and Bcap-37 cell suspensions at densities of 1 3 106 cells/ mL were inoculated into the culture dishes. The inoculation volumes were 5.0 mL. The culture dishes were incubated for 24 h under 5% of CO2 at 37 C. After abandoning the culture medium, different concentrations (0.1, 0.25, 0.5, and 1 mg/mL) of pectin samples and culture medium were added, and 100 ng/mL PMA was also added, except for to the control group. The culture dishes were incubated again for another 24 h under the same conditions. After abandoning the cultures, the cells were washed with 1 mL cold PBS solution, and the cells were scraped with a scraper. After centrifuging at 2,000 rpm for 30 s at 4 C, the supernatant was discarded and 0.2 mL RIPA’s buffer was added to the samples. Cells were disrupted by ultrasound treatments and were dissolved on ice for 2 h. The samples were then centrifuged in 12,000 rpm for 10 min at 4 C to obtain the supernatant. The supernatants were transferred to new tubes, and the protein concentrations were measured using a reagent kit. After adjusting the protein concentrations to the same levels, the uPA content was measured using an uPA kit. The specific steps were as follows: 1. One well was established as the blank control, the rest of the wells contained 100 μL of samples or standards and the wells were covered with tape. The mixtures were incubated at 37 C for 30 min. 2. The liquid in the every well was removed and 200 μL washing solution was added three times to each well. After washing, the microplates were placed on blotting paper to allow the residual liquid to dry. 3. Then, 100 μL horseradish peroxidase-antibodies were added to every well and plates were incubated for 30 min at 37 C. If the horseradish peroxidase-antibodies appeared turbid, then the plates were allowed to reach room temperature and the solution was gently mixed until uniform. 4. The solution in the wells was discarded, and each well was washed five times. 5. Then, 90 μL 3,30 ,5,50 -tetramethylbenzidine substrate was added to each well and incubated at 37 C for 15 min. 6. Termination solution (50 μL) was added to each well. If the change of color was not uniform, then the microwell plate was gently shaken to mix well. 7. The absorbance of each well was measured by at 450 nm.

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3.6.3.1 Effects of Unmodified Sweet Potato Pectin on the uPA Content of Cancer Cells As shown in Fig. 4.23, after treating only with PMA, the uPA content was significantly greater than those of the control and pectin-treated groups. In the groups receiving pectin treatments, an increasing pectin concentration resulted in significantly decreasing the uPA contents in cancer cells (P , 0.05). The uPA content of HT-29 cancer cells treated with 1 mg/mL unmodified pectin decreased by 18.46% compared with the uPA groups. For Bcap-37 cells, the uPA content of the pectintreated group was reduced by 18.80%. Thus, unmodified pectin may decrease the uPA contents of Bcap-37 and HT-29 cells. 3.6.3.2 Effects of Thermal-Modified Sweet Potato Pectin on the uPA Contents of Cancer Cells As shown in Fig. 4.24, after being treated only with PMA, the uPA content was greater than those of the control and pectin-treated groups. When treating with thermal-modified pectin, as the pectin concentration increased, the uPA content in cancer cells significantly decreased (P , 0.05). When the concentration of thermal-modified pectin reached 1 mg/mL, the uPA content level in HT-29 cells was not significantly different with that of the control group (P . 0.05); however, the uPA level in Bcap-37 cells was significantly greater than that of the control group. This indicated that the thermal-modified

Figure 4.23 Effects of unmodified sweet potato pectin on the uPA contents of cancer cells. Values followed by the same letter for the same kind of cancer cell are not significantly different (P . 0.05).

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Figure 4.24 Effects of thermal-modified sweet potato pectin on the uPA contents of cancer cells. Values followed by the same letter for the same kind of cancer cell are not significantly different (P . 0.05).

pectin had a greater inhibitory effect on the uPA content in HT-29 cells than in Bcap-37 cells. For HT-29 cells, the uPA content of the cancer cells treated with 1 mg/mL thermal-modified pectin was reduced by 33.76% compared with the control groups. For Bcap-37 cells, the uPA content of the cancer cells in the pectin-treated group was decreased by 31.52%. Compared with the unmodified pectin, after thermal pectin modification, the uPA content of HT-29 cells was reduced by 18.76%, while the uPA content of Bcap-37 cells was reduced by 15.67%. This indicated that the thermal modification of pectin had significantly increased the inhibitory effects on the uPA content in cancer cells. 3.6.3.3 Effects of pH-Modified Sweet Potato Pectin on the uPA Contents of Cancer Cells As shown in Fig. 4.25, the uPA content in cells treated only with PMA was greater than those of the control and the pH-modified pectin-treated group. With the increasing pH-modified pectin concentration, the uPA content in cancer cells significantly decreased (P , 0.05). When the concentration reached 1 mg/mL, the uPA contents in both HT-29 and Bcap-37 cells were not different from that of the control group (P . 0.05). When treating HT-29 cells with 1 mg/mL pH-modified pectin, the uPA content decreased by 36.11%, while in Bcap-37 cells the content decreased by 35.33%. After the pH modification of pectin, the uPA contents of HT-29 and Bcap-37 cells were reduced by 21.65%

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Figure 4.25 Effects of pH-modified sweet potato pectin on the uPA contents of cancer cells. Values followed by the same letter for the same kind of cancer cell are not significantly different (P . 0.05).

and 20.36% when compared with the respective unmodified pectintreated cells. Thus, the pH-modified pectin had obviously increased the inhibitory effect on the uPA contents in cancer cells. In summary, pectin had no toxic effects on HT-29 colon cancer cells, Bcap-37 breast cancer cells or SMMC-7721 liver cancer cells. The inhibitory effects of modified sweet potato pectin on cancer cell proliferation were enhanced significantly and the inhibitory effects on HT-29 and Bcap-37 were particularly significant. This may be because the anticancer effects of modified pectin were selective. The extent of adhesion, the migration rate and the uPA content all decreased significantly when cells were treated with modified sweet potato pectin. When the pectin concentration reached 1 mg/mL and the treatment time reached 24 h, the pH-modified pectin could decrease the complete digestion time of HT-29 and Bcap-37 cells by 7.69% and 21.95%, respectively, decrease the migration rates by 48.36% and 50.45%, respectively, and decrease the uPA content by 21.65% and 20.36%, respectively, compared with unmodified pectin. In addition, the thermal-modified pectin could decrease the complete digestion time of HT-29 and Bcap-37 cells by 6.60% and 20.73%, respectively, reduce the migration rates by 33.31% and 34.46%, respectively, and decrease the uPA content by 18.76% and 15.67%, respectively. This indicated that pectin modification could significantly improve the inhibitory effects on cancer cell migration.

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SECTION 4: PHYSICOCHEMICAL CHARACTERISTICS OF SWEET POTATO PECTIN In this section, the effects of temperature, shear rate, pectin concentration, ionic strength (NaCl concentration), Ca21 concentration, and the coexistence of solute (sucrose concentration) on the viscosity of a sweet potato pectin solution were studied using the acid method. In addition, the storage and loss moduli variation trends, phase angle tangent changes under different shear rates, are discussed under different conditions. Thus, we explored the optimal conditions for pectin gel formation, to provide a theoretical basis for the application of acid method-based sweet potato pectin in food industry.

4.1 A VISCOSITY ANALYSIS OF SWEET POTATO PECTIN Single factor experiments were used to explore the effects of pectin concentration [0.1%
2.0% (w/v)], pH value (2.0
10.0), sucrose concentration [10%
60% (w/v)], the concentration of NaCl (0.05
1.0 M) and Ca21 concentration (1
50 mM), measuring temperature (10
90 C), and measuring shear rate (1
1000 s21) on the viscosity of a pectin solution. In the single factor experiments, when discussing one factor, the other factors were maintained as: 1.0% pectin concentration, pH 5.0, no sucrose, NaCl and Ca21, 25 C measuring temperature, and 50 s21 measuring shear rate.

4.1.1 Effects of Pectin Concentration and pH Value on the Viscosity of a Pectin Solution With the pectin concentration increased (Fig. 4.26), the viscosity of the pectin solution significantly increased. When the pectin concentration was 0.1%, the viscosity of the solution was 1.18 mP s. When the pectin concentration was increased to 2.0%, the viscosity of the solution reached 7.06 mP s, because as the number of the pectin molecules in solution increases, the interactions (hydrogen bonds and hydrophobic interactions) between molecules are enhanced, as are the entanglements between molecules. With an increasing pH (Fig. 4.26), the viscosity of the pectin solution was changed to “M” type, with the three inflection points. pH 5.0 and 9.0 were at the top, having high viscosity levels of 3.82 and 3.65 mP s, respectively, while pH 7.0 was at the bottom with a low viscosity of 3.17 mP s. At pH 5.0 and 9.0, the net charges of many pectin molecules might result in stronger intermolecular

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Figure 4.26 Effects of pectin concentration and pH value on the viscosity of a pectin solution.

electrostatic interactions, with the pectin molecules in solution becoming stretched and highly viscous; however, at pH 7.0, there were limited ions in solution, the electrostatic interaction was weak and the viscosity decreased.

4.1.2 Effects of Sucrose, NaCl, and Ca21 Concentrations on the Viscosity of a Pectin Solution As the sucrose concentration increased (Fig. 4.27), the viscosity of the pectin solution increased. As the sucrose concentration increased from 10% to 40%, the viscosity of the pectin solution rose relatively slowly, but when the concentration increased from 40% to 60%, the viscosity increased significantly (P , 0.05). Sucrose in the pectin solution interacted with pectin molecules through hydrogen bonding. In addition, the strong hydrating ability of sucrose reduced the level of pectin hydration greatly, and the interactions between pectin molecules were intensified as the sucrose concentration increased. As the interactions became stronger, the viscosity increased more quickly. When the sucrose concentration was 10%, 40%, and 60%, the viscosity levels of the pectin solution were 3.77, 13.04, and 49.6 mP s, respectively. As the NaCl concentration increased, but was less than 0.4 M, the viscosity of the pectin solution changed little, but when the NaCl concentration was greater than 0.4 M, the viscosity of the pectin solution increased significantly. When the NaCl concentrations were 0.05, 0.4, and 1.0 M, the viscosity levels of the solution were 2.79, 3.26, and 6.94 mP s, respectively. In the presence of NaCl, the electrostatic effect of pectin was shielded. The interactions between pectin molecules

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Figure 4.27 Effects of sucrose, NaCl and Ca21 concentrations on the viscosity of a pectin solution.

increased as the NaCl concentration increased, as did the viscosity (Fig. 4.27). Ca21 significantly affected the viscosity of pectin solution (Fig. 4.27). As the Ca21 concentration increased, the viscosity of the solution increased. When the Ca21 concentration ranged from 0 to 50 mmol/L, the viscosity of the pectin solution showed a certain

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correlation, and R250.88. Under the condition of no Ca21, the viscosity of the pectin solution was 3.31 mP s, while at a 50 mmol/L Ca21 concentration, the viscosity of the pectin solution reached 1067.64 mP s. Sweet potato pectin has a low degree of esterification (31.71%), and there is still part of the galacturonic acid in the pectin chain that is not esterified. The nonesterified galacturonic acid forms negatively charged carboxyl groups due to ionization, and it can form ionic bonds between Ca21 ions. One Ca21 can be combined with two galacturonic acid residues, thereby causing cross-linking between pectin molecules. Thus, because of the presence of Ca21, the viscosity of the solution increased.

4.1.3 Effects of Temperature and Shear Rate on the Viscosity of a Pectin Solution With increasing temperature (Fig. 4.28), the viscosity of the pectin solution decreased, and when the temperature was over 50 C, the rate of decline slowed down. When the temperature was 10 C, the pectin solution’s viscosity increased to 13.0 mP s, and when the temperature increased to 90 C, the viscosity dropped to 1.66 mP s. With an increase in temperature, the molecular thermal motion of pectin intensified and the interaction forces between molecules decreased. The viscosity decreased at a constant shear rate (Fig. 4.28). As the shear rate increased in the range of 1
1000 s21, a shear thinning-associated phenomenon appeared in the pectin solution, which showed the characteristics of a non-Newtonian fluid. When the shear rate was 1 s21, the viscosity of the pectin solution was 15.0 mP s, and when the shear rate rose to 1000 s21, the viscosity was only 3.72 mP s. When shear

Figure 4.28 Effects of temperature and shear rate on the viscosity of a pectin solution.

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rate was greater than 35 s21, it had little influence on the viscosity of the pectin solution, and the viscosity of the solution tended to be stable, which may be because the shear force formed at a rate of more than 35 s21, which is far more than the interaction force between pectin molecules in solution.

4.2 THE GELATION PROPERTIES OF SWEET POTATO PECTIN Single factor experiments were employed to explore the effects of pectin concentration (0.5%
3.0%), sucrose concentration (10%
60%), Ca21 concentrations (10
50 mM), and pH value (2.0
7.0) on the gelling properties of pectin, when discussing one of the factors, the values of the other factors were maintained as: pectin concentration 1.5%, sucrose concentration 30%, Ca21 concentration 30 mM, and pH value 5.0.

4.2.1 Determination of the Sweet Potato Pectin Gel’s Linear Viscoelastic Region Under the pectin gel-forming conditions of 1.5% pectin concentration, 30% sucrose concentration, 30 mM Ca21 concentration, and pH value of 5.0, strain scanning was performed (Fig. 4.29). Strain was applied to the gel to produce deformation forces, which were used to determine the stress at which the transformation of the shear frequency will not damage the gel’s structure. According to the pectin gel storage and loss moduli’s changes under different strains, the sweet potato pectin gel’s

Figure 4.29 Determination of the pectin gel’s linear viscoelastic region. G0 indicates the storage modulus, and Gv indicates the loss modulus.

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Figure 4.30 The effects of pectin concentration on moduli and the complex viscosity of pectin gelation. G0 indicates the storage modulus, and Gv indicates the loss modulus.

linear viscoelastic strain range was determined to be between 0.3% and 3%. The rheometer’s recommended strain is 1%.

4.2.2 The Effects of Pectin Concentration on the Moduli and the Complex Viscosity of Pectin Gelation The effects of pectin concentration on the storage modulus, loss modulus, composite viscosity, and phase angle tangent during the sweet potato pectin gelation process were determined (Fig. 4.30). At different concentrations, the gel storage modulus, loss modulus, and complex viscosity increased as the temperature decreased, and the energy of the storage modulus was always greater than that of the loss modulus, indicating that pectin is in a gelatenous state. With an increasing pectin concentration, the storage and loss moduli increased, and the tan δ (phase angle tangent) value decreased, indicating that as the storage modulus increased more quickly, the gel strength also increased.

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Figure 4.31 Effects of sucrose concentration on the moduli and the complex viscosity of pectin gelation.

At low pectin concentrations, the shear frequency had a greater impact on the pectin gel modulus. When the pectin concentration was 0.5%, the shear frequency was over 10 Hz, the tan δ was greater than 1, and the loss modulus value was greater than that of the storage modulus, which indicated that, under low pectin concentration conditions, the gel-forming strength was weak. When the shear frequency was high, the gel transformed from a solid to liquid state, and when the pectin concentration was high (greater than 2.0%), the gel strength increased and was less affected by the shear rate.

4.2.3 The Effects of the Sucrose Concentration on the Moduli and the Complex Viscosity of Pectin During Gelation In Fig. 4.31, the effects of sucrose concentration on the storage modulus, loss modulus, composite viscosity, and tan δ of sweet potato pectin during gelation are shown. They indicate that the sweet potato pectin

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gel storage modulus, loss modulus and complex viscosity gradually increased as the temperature decreased. In this process, the energy of the loss modulus is greater than that of the storage modulus, indicating that the pectin is in a gelatinous state. As the sucrose concentration increased, the storage and loss moduli also increased, and under different shear rates, the tan δ trend was roughly the same as the sucrose concentration change, decreasing first, then increasing, and finally decreasing again. When the sucrose concentration was 20%, the tan δ value was significantly (P , 0.05) lower than the other tan δ values at other sucrose concentrations, which indicated that the gel strength was its greatest. However, in general, the tan δ value varied less with the sucrose concentration’s change, which indicated that the sucrose concentration was not the main factor affecting the sweet potato pectin’s gel strength.

4.2.4 The Effect of Ca21 Concentration on the Moduli and the Complex Viscosity of Pectin Gelation The effects of Ca21 concentration on sweet potato pectin gelation are shown in Fig. 4.32. As the temperature decreased and the Ca21 concentration increased, the gel storage modulus, loss modulus, and composite viscosity increased, indicating that a low temperature can accelerate the sweet potato pectin gel network formation and enhance sweet potato pectin gel strength. At low Ca21 concentrations, the sweet potato pectin gel modulus and complex viscosity declined, and as the Ca21 concentration increased, the pectin modulus and complex viscosity increased until the concentration was greater than 20 mM, at which point the modulus and complex viscosity varied less. At different Ca21 concentrations less than 20 mM, the tan δ value and the shear frequency decreased as the Ca21 concentration increased, and the gel strength increased. When the Ca21 concentrations were greater than 20 mM, tan δ values increased as the Ca21 increased, and gel strength decreased. This may be because of the relatively high Ca21 concentration, which promoted the formation of calcium pectin acid precipitation, destroying the gel structure. Thus, during sweet potato pectin gel formation, too high of a Ca21 concentration will decrease the gel strength, which is not conducive to the formation of the gel structure. Pectin can also form a gel without the presence of Ca21, but the strength of the gel structure is weak, and at a high shear frequency, the gel shows fluid characteristics (tan δ . 1).

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Figure 4.32 Effects of Ca21 concentration on the moduli and the complex viscosity of pectin gelation.

4.2.5 The Effects of pH Value on the Moduli and the Complex Viscosity of Pectin Gelation In Fig. 4.33, the effects of pH value on sweet potato pectin gelation are revealed. As the temperature decreases, the storage modulus, loss modulus, and composite viscosity of sweet potato pectin gel significantly increased, the structure of the gel formed continuously, and the gel strength increased gradually. When the pH value increased from 2 to 5, the gel modulus and complex viscosity increased significantly; however, when the pH was increased from 5 to 7, the gel modulus and compound viscosity decreased slightly. The tan δ value at different shear frequencies showed that as the pH value increased to 5.0, the tan δ value decreased and the gel strength increased. However, when the pH increased above 5.0, the tan δ value increased, indicating that the energy of the storage modulus was lower than that of the loss

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Figure 4.33 Effects of pH value on the moduli and the complex viscosity of pectin gelation.

modulus, weakening the gel strength. In addition, under low pH conditions, sweet potato pectin can also form a solid gel, but the gel strength is weak, while under a high shear rate, the loss modulus of the gel is greater than the storage modulus, which is a fluid characteristic.

4.2.6 Gel Texture Analysis of Sweet Potato Pectin 4.2.6.1 Effects of Pectin Concentration on the Texture Parameters of Sweet Potato Pectin Gel A texture analysis of sweet potato pectin gels containing different sweet potato pectin concentrations is shown in Table 4.4. As the pectin concentration increased, the gel hardness significantly increased (P , 0.05). When the pectin concentration was 0.5%, the gel hardness was 7.70 g, and when the pectin concentration was 3.0%, the hardness reached to 84.53 g. Gel adhesiveness and chewiness increased along

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Table 4.4 Effects of Pectin Concentration on Gel Texture Concentration (% w/v)

Hardness (g)

Adhesiveness

Chewiness

0.5

7.70 6 0.88

2.69 6 0.71

0.79 6 0.28d

1.0

21.84 6 2.25

1.5

45.47 6 3.41d

28.66 6 2.37b

25.02 6 4.66b

2.0

58.48 6 3.29

c

36.66 6 3.12

a,b

33.34 6 3.55a

2.5

75.28 6 2.79

b

42.44 6 4.01

a

40.13 6 3.78a

3.0

84.53 6 3.37a

43.88 6 3.96a

41.19 6 4.06a

f e

d

12.91 6 1.06

c

9.83 6 0.77c

Concentration (% w/v)

Tackiness

Elasticity

Recovery rate

0.5

0.35 6 0.08b

0.29 6 0.02b

0.49 6 0.05a

1.0

0.59 6 0.11a,b

0.76 6 0.11a

0.34 6 0.03b

1.5

0.63 6 0.05

0.87 6 0.08

a

0.023 6 0.005c

2.0

0.63 6 0.09a,b

0.91 6 0.04a

0.017 6 0.007c

2.5

0.56 6 0.04

a

0.95 6 0.02

a

0.020 6 0.004c

3.0

0.52 6 0.04

a

0.94 6 0.02

a

0.015 6 0.001c

a,b

Values followed by the same letter in the same column are not significantly different (P . 0.05).

with the pectin concentration. At concentrations was greater than 2.0%, changes did not have significant effects on the two indicators. With an increase in the pectin concentration, the tackiness and elasticity of the gel first increased and then decreased, and the recovery rate decreased gradually. When the concentration was greater than 1.5%, further increases had no significant effects on the three indices. 4.2.6.2 Effects of Sucrose Concentration on the Texture Parameters of Pectin Gel Table 4.5 lists the texture indices of hardness, gumminess, chewiness, cohesiveness, elasticity, and recovery rate of sweet potato pectin gel under different sucrose concentrations. As the sucrose concentration increased, the other texture parameters of sweet potato pectin gel showed no significant (P , 0.05) changes, except for gumminess and chewiness. When the sucrose concentration was 10%, the gumminess and chewiness of sweet potato pectin gel were significantly lower than their counterparts under other concentrations, while the gumminess and chewiness under other concentrations showed no significant differences. The hardness, gumminess, and chewiness of the sweet potato pectin gel were highest, at values of 45.47, 28.66 and 25.02 g, respectively, when the sucrose concentration were 30%, but the elasticity was the lowest, at only 0.87.

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Table 4.5 Effects of Sucrose Concentration on Gel Texture Concentration (% w/v)

Hardness (g)

Gumminess

10

42.17 6 1.88

a

20

43.25 6 2.25

a

30

Chewiness

17.37 6 1.01

b

15.55 6 0.28b

22.95 6 1.36

a

21.01 6 0.77a

45.47 6 1.41a

28.66 6 2.37a

25.02 6 1.66a

40

44.23 6 1.29

a

25.01 6 1.12

a

23.52 6 1.55a

50

44.41 6 2.79

a

23.76 6 1.01

a

22.09 6 0.78a

60

43.93 6 1.37a

27.51 6 1.96a

24.70 6 1.06a

Concentration (% w/v)

Cohesiveness

Elasticity

Recovery rate

10

0.44 6 0.08a

0.90 6 0.02a

0.019 6 0.005a

20

0.53 6 0.11a

0.92 6 0.07a

0.021 6 0.003a

30

0.61 6 0.05

0.87 6 0.05

a

0.023 6 0.005a

40

0.57 6 0.09a

0.94 6 0.04a

0.022 6 0.007a

50

0.52 6 0.04

a

0.93 6 0.02

a

0.017 6 0.004a

60

0.63 6 0.04

a

0.90 6 0.02

a

0.021 6 0.001a

a

Values followed by the same letter in the same column are not significantly different (P . 0.05).

Table 4.6 Effects of Ca21 Concentration on Gel Texture Concentration (% w/v)

Hardness (g)

Gumminess

10

119.56 6 2.25a

58.97 6 2.06a

50.85 6 1.77a

20

48.05 6 2.41

d

21.89 6 1.31

d

19.27 6 1.06d

30

45.76 6 2.01

d

26.47 6 2.12

c,d

22.53 6 2.15d

40

59.92 6 1.79c

32.82 6 1.99c

31.00 6 1.78c

50

79.12 6 1.37

45.68 6 1.76

42.15 6 2.66b

Concentration (% w/v)

Cohesiveness

Elasticity

10

0.49 6 0.05

a

20

0.46 6 0.04

a

30

b

Chewiness

b

Recovery Rate

0.86 6 0.02

a

0.068 6 0.007a

0.88 6 0.05

a

0.027 6 0.005b

0.58 6 0.05a

0.85 6 0.03a

0.023 6 0.003b

40

0.55 6 0.04

a

0.94 6 0.04

a

0.024 6 0.004b

50

0.58 6 0.02

a

0.92 6 0.03

a

0.020 6 0.003b

Values followed by the same letter in the same column are not significantly different (P . 0.05).

4.2.6.3 Effects of Ca21 Concentration on the Texture Parameters of Pectin Gel Table 4.6 lists the effects of different concentrations of Ca21 on sweet potato pectin gel texture. As the Ca21 concentration increased, the sweet potato pectin gel’s hardness, cohesiveness, and chewiness decreased and then increased. At a Ca21 concentration of 10 mM, sweet potato pectin gel’s hardness, gelling, and chewiness reached their

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maximum values of 119.56, 58.97, and 50.85 g, respectively. When the Ca21 concentration was 30 mM, the hardness value of sweet potato pectin gel was its lowest, at only 45.76 g. The cohesiveness and chewiness values were lowest, only 21.89 and 19.27, respectively, at a 20 mM Ca21 concentration. The increased Ca21 concentration had no significant (P , 0.05) effects on the sweet potato pectin gel’s bonding strength and elasticity. The gel recovery rate was greatest when the Ca21 concentration was 10 mM, and there were significant differences between other gel recovery rates at other Ca21 concentrations. When the concentration of Ca21 was 10 mM, the sweet potato pectin gel structure was stable. 4.2.6.4 Effects of pH Value on the Texture Parameters of Pectin Gel Table 4.7 lists the effects of the pH value of the system on the sweet potato pectin gel texture. As the pH value increased, the hardness, gumminess, and chewiness of sweet potato pectin gel first increased and then decreased. At pH 6.0, the sweet potato pectin gel’s hardness, gumminess, and chewiness reached their maximum values of 45.91, 24.91, and 22.01 g, respectively, and these values did not differ significantly (P , 0.05) from those at pH 5.0. The pectin gel’s cohesiveness and elasticity first increased and then decreased as the pH value increased. When the pH was 3.0, the cohesiveness and elasticity values Table 4.7 Effects of pH Value on Gel Texture pH Value

Hardness (g)

Gumminess

Chewiness

2

9.01 6 0.88d

3.70 6 0.31d

2.17 6 0.28d

3

7.98 6 1.25

3.35 6 1.06

4

41.39 6 1.41

5

d

1.85 6 0.77d

d c

15.20 6 1.06c

45.20 6 1.29a

24.44 6 1.12a

21.84 6 1.15a

6

45.91 6 1.79

a

24.91 6 1.01

a

22.01 6 0.78a

7

34.74 6 1.37

c

21.67 6 0.96

b

18.09 6 0.66b

pH Value

Cohesiveness

Elasticity

2

0.41 6 0.05

c

3

0.33 6 0.04

d

4

0.42 6 0.05c

0.88 6 0.03a

0.031 6 0.005b

5

0.54 6 0.04

0.89 6 0.04

a

0.019 6 0.003b

6

0.56 6 0.02a,b

0.88 6 0.02a

0.018 6 0.008b

7

0.62 6 0.04

0.83 6 0.02

0.018 6 0.007b

b

b

a

17.36 6 1.37

Recovery Rate

0.59 6 0.02

b

0.70 6 0.05a

0.42 6 0.05

c

0.81 6 0.07a

a

Values followed by the same letter in the same column are not significantly different (P . 0.05).

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Figure 4.34 Microstructure of sweet potato pectin gel.

were lowest. In addition, the recovery rate of the gel first increased and then decreased as the pH increased, reaching a maximum at pH 3.0. When the pH was greater than three, it had no significant effects on the elasticity or recovery rate of the pectin gel.

4.2.7 The Microstructure of Sweet Potato Pectin Gel Fig. 4.34 shows the microstructure of a sweet potato pectin gel. Scanning electron micrographs of the gel magnified 100 and 300 times are shown in Fig. 4.34A and B, respectively. The skeleton of the pectin gel is a porous network structure and water molecules are wrapped in these pores, connecting pectin molecules through hydrogen bonds or hydration forces.

4.3 THE EMULSIFYING PROPERTIES OF SWEET POTATO PECTIN Single factor tests were used to investigate the effects of pectin concentrations (0.5%
4.0%), oil-phase volume fractions (5%
40%), pH values (2.0
7.0), NaCl concentrations (0.05
1.0 M) and Ca21 concentrations (0.5
8 mM) on its emulsifying properties. In the single factor experiments, when one of the factors was discussed, the remaining factors were maintained as: 2% pectin concentration, 20% oil-phase volume fraction, pH 5, and neither NaCl nor Ca21.

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Based on the desired pectin concentration, a certain amount of pectin was measured separately and dispersed into distilled water containing 0.02% (w/v) sodium azide. The pH was adjusted, the pectin solution was stirred at room temperature for 24 h. The pectin solution was mixed with soybean oil at a certain (v/v) mixing ratio. After homogenizing at 24,000 rpm for 1 min, the properties of the characteristic indices, including emulsion particle size, viscosity, emulsifying activity index (EAI), emulsion stability index (ESI), and the absorption of emulsified particles to pectin, of sweet potato pectin emulsions prepared under different conditions were explored. In addition, emulsions under different conditions were microscopically observed.

4.3.1 Effects of the Pectin Concentration and Oil-Phase Volume Fraction on the Particle Sizes of Pectin Emulsions The particle size is an important index for evaluating the merits of the emulsion (Tesch and Schubert, 2002). Fig. 4.35 shows that the concentration of pectin and the oil-phase volume fraction significantly influenced the particle size of the emulsion. As the pectin concentration increased, the particle size of the emulsion gradually decreased. At a pectin concentration of less than 2.0%, the emulsion particle diameter increased significantly (P , 0.05) after being maintained at room temperature for 1 and 7 days, when compared with the fresh emulsion, and when the pectin concentration was greater than 2.0%, the resting time had no significant effect on the emulsion particle size. As the oilphase volume fraction increased, the emulsion particle size increased.

Figure 4.35 Effects of the pectin concentration and oil-phase volume fraction on the particle diameter of the pectin emulsion.

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Compared with the fresh emulsion’s particle size, when the oil-phase volume fraction was greater than 25%, the emulsion particle diameter increased significantly after being placed at room temperature for 1 and 7 days, and when the oil phase volume fraction was less than 25%, the resting time had no significant effect on the emulsion particle size.

4.3.2 Effects of pH Value, and NaCl and Ca21 Concentrations on the Particle Sizes of Pectin Emulsions The pH value also had significant effects (Fig. 4.36) on the sweet potato pectin emulsion’s particle diameter. When the pH increased from 2.0 to 7.0, the pectin emulsion particle size first decreased and then increased. At pH 5.0, the emulsion particle diameter was significantly less than the other emulsion particle diameters (P , 0.05) under other pH values. After placing the emulsion at room temperature for 1 and 7 days, the emulsion particle size increased compared with that of the fresh emulsion, especially when the pH value was less than 5, and the 7 days size was significantly different from that of the fresh emulsion.

Figure 4.36 Effects of pH value, and NaCl and Ca21 concentrations on the particle sizes of pectin emulsions.

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NaCl and Ca21 concentrations had significant effects on the emulsion particle size of sweet potato pectin (Fig. 4.36). As the NaCl concentration increased, the emulsion particle size first decreased and then increased. When the NaCl concentration was 0.2 M, the fresh emulsion particle size was the smallest, only 11.11 μm after 1 day. When the NaCl concentration ranged from 0.05 to 0.2 M and 0.4 to 1.0 M, the pectin emulsion particle size significantly increased (P , 0.05); however, at NaCl concentrations between 0.2 and 0.4 M, the emulsion particle diameter did not obviously change. After the emulsion was maintained for 7 day, the emulsion particle size did not change significantly compared with the size at 1 day. As the Ca21 concentration increased from 0.5 to 7 mM, the emulsion particle size increased also. The particle size increased greatly when the Ca21 concentration increased from 7 to 8 mM, and there was a significant difference between the sizes at these concentrations. Compared with the fresh emulsion, the emulsions containing different Ca21 concentrations had significantly increasing particle sizes after being placed at room temperature for 1 and 7 days (Fig. 4.36).

4.3.3 Effects of Emulsion Particles on the Adsorption of Pectin 4.3.3.1 Effects of the Pectin Concentration and Oil-Phase Volume Fraction on the Adsorption of Pectin to Emulsion Particles The pectin concentration and oil-phase volume fraction had significant effects on the adsorption of pectin to emulsion particles (Fig. 4.37). As the pectin concentration increased, the amount of pectin adsorbed on the surface of emulsified particles (Γ ) increased significantly (P , 0.05), and the increase was positively correlated

Figure 4.37 Effects of the pectin concentration and oil-phase volume fraction on pectin adsorption by emulsion particles.

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(R2 5 0.96) with the pectin concentration. As the concentration increased, the proportion of pectin adsorbed by the emulsified particles significantly increased, and when the concentration was less than 3.5%, there were significant differences among the pectin concentrations. As the oil-phase volume fraction increased, the Γ values first increased and then decreased. Different oil-phase volume fractions showed obvious dissimilarities, and the 15% oil-phase volume fraction, the Γ value was highest. When the oil-phase volume fraction was less than 10%, the emulsified particles’ pectin adsorption ratio increased slightly, but the difference was not significant. When the oil phase volume fraction was greater than 10%, the emulsified particle’s pectin adsorption ratio decreased significantly. 4.3.3.2 Effects of pH Value, and the NaCl and Ca21 Concentrations on the Adsorption of Pectin by Emulsion Particles The pH effects on the emulsified particles’ adsorption of pectin is shown in Fig. 4.38. At different pH values, the Γ value was not

Figure 4.38 Effects of pH, and the NaCl and Ca21 concentrations on the adsorption of pectin by emulsion particles.

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significantly affected. The pectin adsorption ratio first increased and then decreased as the pH value increased, and the emulsion particle size change showed the same trend. At pH 5.0, the Γ value was the highest because the emulsion particle size was smaller at pH 5 and the relative total surface area was larger. Thus, when the Γ values were the same, the proportion of adsorbed pectin was greater. The concentrations of NaCl and Ca21 had significant effects on the adsorption of pectin by the emulsion. When the NaCl concentration was less than 0.3 M (Fig. 4.38), the Γ values were not significantly different (P , 0.05) at different NaCl concentrations. At NaCl concentration greater than 0.3 M, the Γ value decreased significantly. At a 0.05 M NaCl concentration, the Γ value increased to 1.90 mg/m2, while at a 1.0 M NaCl concentration, the Γ value was only 0.73 mg/m2. As the NaCl concentration increased, the proportion of pectin significantly decreased. The Γ value increased slightly under different Ca21 concentrations less than 2.0 mm, but the differences were not significant. When the Ca21 concentration was greater than 2.0 mM, the Ca21 concentration increased (Fig. 4.38), and the Γ value decreased significantly. As the Ca21 concentration increased, the adsorbed pectin ratio in the emulsion showed the same trend as the Γ value. When the Ca21 concentration was less than 2.0 mM, the emulsion’s adsorbed pectin ratio was not significantly different, and as the Ca21 concentration increased above 2.0 mM, the adsorbed pectin ratio decreased significantly.

4.3.4 Emulsifying Activity 4.3.4.1 Effects of the Pectin Concentration and Oil-Phase Volume Fraction on the Emulsifying Activity of Pectin The pectin concentration and oil-phase volume fraction had significant effects on the EAI of the sweet potato pectin emulsion. As the pectin concentration and oil-phase volume fraction increased, the EAI increased significantly (P , 0.05) and was positively correlated with the pectin concentration and oil-phase volume fraction (Fig. 4.39), with R2 5 0.91 and 0.99, respectively. When the pectin concentration was 0.5%, the EAI was 9.27%, and when the pectin concentration was 4.0%, the EAI reached 100%. When the oil-phase volume fraction was 5%, the EAI was relatively small, only 8.59%, while as the oilphase volume fraction increased to 40%, the EAI increased rapidly to 51.42%.

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Figure 4.39 Effects of the pectin concentration and oil-phase volume fraction on the EAI of the pectin emulsion.

4.3.4.2 Effects of pH, and NaCl and Ca21 Concentrations on Emulsifying Activity of Pectin The pH value had a great impact (Fig. 4.40) on EAI of the emulsion. As the pH value increased, the EAI first increased and then decreased, At pH 5.0, the EAI was its highest, at 28.96%, but was not significantly different (P , 0.05) from the value at pH 4.0. However, it was significantly higher than other EAI values at different pH levels. When the pH value was 3.0, the EAI was lowest, only 24.12%, but was not significantly different from the EAI value at pH 2.0. The EAI values of the sweet potato pectin emulsion were significantly affected by the NaCl and Ca21 concentrations (Fig. 4.40). As the NaCl concentration increased, the EAI first decreased slightly and then increased. When the NaCl concentration increased from 0.05 to 0.1 M, the EAI decreased, and when the NaCl concentration was greater than 0.1 M, the EAI increased. At NaCl concentrations from 0.2 to 0.5 M, no significant (P , 0.05) changes in the EAI occurred. The EAI was highest, at 55.36%, when the NaCl concentration was 1.0 M. As the Ca21 concentration increased, the EAI increased, and at Ca21 concentrations between 0.5 and 6 mM, the EAI increased within a smaller range but there were no significant differences within this concentration range. When the Ca21 concentration was greater than 6.0 mM, the EAI values increased significantly, and the EAI reached 55.39% at a Ca21 concentration of 8.0 mM.

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Figure 4.40 Effects of pH value, and the NaCl and Ca21 concentrations on the EAI of the pectin emulsion.

4.3.5 Emulsion Viscosity 4.3.5.1 Effects of the Pectin Concentration and Oil-Phase Volume Fraction on the Viscosity of the Pectin Emulsion The viscosity of the emulsion has a significant effect on its stability. A high viscosity can prevent the flocculation of emulsified particles (Mishra et al., 2001), and maintain the stability of the emulsion. The emulsion’s viscosity was significantly (P , 0.05) affected by the pectin concentration and oil-phase volume fraction (Fig. 4.41). As the pectin concentration increased, the viscosity of the emulsion increased. When the pectin concentration was at the relatively low value of 0.5%, the emulsion viscosity was 6.85 mPa s, and when the pectin concentration increased to 4%, the viscosity of the emulsion was 133.33 mPa s. As the oil-phase volume fraction increased, the emulsion viscosity also increased significantly. When the oil-phase volume fraction was 5%,

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Figure 4.41 Effects of the pectin concentration and oil-phase volume fraction on the viscosity of the emulsion.

the viscosity of the emulsion was 12.23 mPa s; however, when the oilphase volume fraction increased to 40%, the viscosity of the emulsion reached 63.90 mPa s. 4.3.5.2 Effects of pH, and the NaCl and Ca21 Concentrations on the Viscosity of the Pectin Emulsion As the pH value increased, the viscosity of the sweet potato pectin emulsion first decreased and then increased (Fig. 4.42). The emulsion viscosity was lowest at pH 3, at only 21.14 mP s, but it was not significantly different than that at pH 2. When the pH value was 5, the viscosity of the emulsion was highest, reaching 25.73 mP s, and the viscosity of the emulsion decreased when the pH value continued to rise. The concentrations of NaCl and Ca21 greatly influenced the viscosity of the emulsion (Fig. 4.42). With the NaCl concentration increased, the viscosity of the emulsion increased, and a significant emulsion viscosity difference (P , 0.05) existed between different NaCl concentrations. When the NaCl concentration was 0.05 M, the viscosity of the emulsion was 27.54 mP s, and viscosity of the emulsion was 101.34 mP s when the NaCl concentration was 1.0 M. As the Ca21 concentration increased, there was a significant increase in the viscosity of the emulsion, and the rate of the viscosity increase, under Ca21 concentrations between 6.0 and 8.0 mM, was significantly greater than that of the Ca21 concentration between 0.5 and 5.0 mM. When the Ca21 concentration was 0.5 mM, the emulsion viscosity was 30.37 mP s, but when the Ca21 concentration was 8.0 mM, the emulsion viscosity was 208.78 mP s.

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Figure 4.42 Effects of pH value, and the NaCl and Ca21 concentrations on the viscosity of the pectin emulsion.

4.3.6 Emulsion Stability 4.3.6.1 Effects of the Pectin Concentration and Oil-Phase Volume Fraction on the Emulsion Stability The pectin concentration and oil-phase volume fraction greatly influenced the ESI. As the pectin concentration increased (Fig. 4.43), the ESI change occurred in two stages. When the pectin concentration was between 0.5% and 1.5%, the ESI increased significantly (P , 0.05), but when the concentration was between 1.5% and 4.0%, the ESI increased gradually, a eventually reaching 100%, and no significant differences existed between the different concentrations. When the oil-phase volume fraction ranged from 5% to 30%, the ESI gradually increased; however, the 5% oil-phase volume fraction of ESI was significantly different from other counterparts. When the oil-phase volume fraction was greater than 30%, the ESI significantly decreased, but there was

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Figure 4.43 Effect of pectin concentration and oil-phase volume fraction on ESI of pectin emulsion.

Figure 4.44 Effects of the pH, and the NaCl and Ca21 concentrations on the pectin ESI.

no significant difference between ESI values with 35% and 40% oilphase volume fraction (Fig. 4.43). 4.3.6.2 Effects of pH Value, and the NaCl and Ca21 Concentrations on the Pectin Emulsion Stability The effects of the NaCl and Ca21 concentrations on pectin ESI values are shown in Fig. 4.44. As the pH increased, the ESI first decreased,

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then increased, and then decreased again. At pH 2.0, the ESI value was lowest, only 74.07%, and the value was not significantly different (P , 0.05) than that at pH 3.0. The ESI value was the highest at pH 5.0, reaching 89.96%. It was not significantly different than the ESI value at pH 6.0, but it was significantly higher than other ESI values at other pH levels. As the NaCl concentration increased, the ESI increased. The ESI values at NaCl concentrations of 0.05 and 1.0 M were significantly different (P , 0.05) than their counterparts under other NaCl concentrations. The ESI values were 90.15% and 100% for NaCl concentrations of 0.1 and 0.5 M, respectively. They were not significantly different. As the Ca21 concentration increased, the ESI increased. The increase was significant when the Ca21 concentration was between 0.5 and 3.0 mM. However, there were no significant differences at Ca21 concentrations between 3.0 and 5.0 mM. When the Ca21 concentration increased from 5.0 to 6.0 mM, the ESI significantly increased, reaching the maximum of 100%. If the Ca21 concentration increased, then the ESI did not change. At a Ca21 concentration of 0.5 mM, the ESI was 83.94%, and the ESI was 100% when the Ca21 concentration was 8.0 mM.

4.3.7 Emulsion Micro Imaging As shown in Fig. 4.45, sweet potato pectin emulsion particles appear regular and round. After microscopic imaging, and emulsion particle size analysis and calculations, using the Mivnt image analysis system, 0.5% of fresh sweet potato pectin emulsion particle size was calculated as 18.35 μm (Fig. 4.45A). After being placed at room temperature for 1 day (Fig. 4.45B), the emulsion particles significantly increased to 29.76 μm (P , 0.05). This may due to the low 0.5% pectin concentration, at which the emulsion stability is poor. When the emulsion was placed at room temperature for 1 day, the adjacent emulsion particles flocculated and then formed larger emulsion particles. The emulsion particle size was 5.08 μm (Fig. 4.45C) in the 5% oil-phase volume fraction, and the emulsion particle size increased to 20.11 μm when the oilphase volume fraction was 40% (Fig. 4.45D), which indicated that under higher oil-phase volume fraction conditions, the particle emulsion’s mutual coalescence was intensified, leading to the formation of larger emulsion particles. When the emulsion’s NaCl concentration was 0.05 M (Fig. 4.45E), the average particle diameter was 12.25 μm, and the particle size reached 23.57 μm when the NaCl concentration

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Figure 4.45 Pectin emulsion micro images taken under different conditions.

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Figure 4.45 (Continued).

increased to 1.0 M (Fig. 4.45F). At higher NaCl concentrations, because of the high ionic strength of the emulsion, the mutual aggregation between pectin molecules and emulsified particles increased, and the amount of pectin covering emulsion particles’ surfaces decreased, resulting in increasing granularity. At a Ca21 concentration of 0.5 mM (Fig. 4.45G), the emulsion particle size was 10.12 μm, and when the Ca21 concentration was 8.0 mM (Fig. 4.45H), the emulsion particle size increased, but only slightly to 12.83 μm. As the Ca21concentration increased, the pectin formed a gel network, and the emulsion particles were confined to a network structure. The decreased fluidity prevented the emulsion particles from aggregating. The pH value had more obvious effects on particle size, and at pH 2 (Fig. 4.45I), 3 (Fig. 4.45J), 5 (Fig. 4.45K), and 7 (Fig. 4.45L), fresh emulsion particle sizes were 10.27, 9.89, 9.36, and 10.01 μm, respectively. pH value changes led to differently charged pectin molecules, affecting the emulsion particles’ charge number, and different electrostatic interactions existed between emulsion particles, resulting in emulsion particle size differences at various pH values.

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In summary, the water solubility of sweet potato pectin is relatively high, which makes it a good emulsifying stabilizer. The EAI increased as the pectin concentration increased, indicating that a high pectin concentration could reduce the interfacial tension more effectively and promote the formation of emulsion particles. The oil-phase volume fraction may have affected the EAI because, along with the increase in the oil-phase volume fraction, a certain amount of sweet potato pectin can produce a greater number of emulsion particles, leading to an increase in the EAI. In addition, because the oil-phase volume fraction increased, the viscosity increased, which prevented emulsion particles from mutually coalescing, to a certain extent. The particle size and ESI measurements showed that the emulsion of higher pectin concentrations [3.5%
4% (w/v)] and the oil-phase volume fraction [10%
30% (v/v)] led to greater ESI values. Lower pectin concentrations or higher oil-phase volume fractions of the emulsion led to relatively poor ESI values, which may be because, under these conditions, the Γ value was lower. Thus, the pectin coverage rate of the emulsion particle surface was low. Emulsified particles share pectin molecules with adjacent emulsified particles to reduce the surface tension, resulting in a larger floc particle size. Then stability of the emulsion was then reduced. This also explains the apparent increase in the size of the particle after placing the emulsion at room temperature 1 or 7 days under a lower pectin concentration and high oil-phase volume fraction conditions. The optical microscopy observations confirmed the above results. After being placed at room temperature for 1 day, the particle diameter of the fresh 0.5% (w/v) low concentration pectin emulsion increased significantly. As the NaCl concentration increased, the ionic strength of the emulsion increased, the pectin mutual aggregation increased, and the hydration capacity decreased. Thus, the number of pectin molecules attached to the emulsified particles decreased. During emulsion formation, the aggregation between adjacent emulsion particles increased, forming emulsion particles of a large size. However, the high viscosity of the emulsion prevented aggregation and flocculation during maintenance and centrifugation; therefore, the EAI and ESI values were greater. The effects of Ca21 on pectin emulsifying properties were related to its ability to change the pectin molecular structure. Ca21 can form network structures with pectin molecules

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through ionic bonds, and the effects of pectin molecular configurations on the stability of the emulsion are far greater than those of the ionic strength and electrostatic interactions (Nakauma et al., 2008). The network structure consequentially enhanced the hydration capability and the distribution of water molecules around the emulsion particles, as well as preventing the aggregation and flocculation of emulsion particles. The effects of the pH on the emulsion of sweet potato pectin showed that at pH 5, sweet potato pectin had a strong emulsifying ability, and the emulsion stability was high. Decreasing or increasing pH values contributed to the decline in the emulsifying ability and stability of pectin, which may be related to the electric charge associated with pectin in solution.

SECTION 5: APPLICATIONS OF SWEET POTATO PECTIN 5.1 CANDIED FRUIT Candied fruit products do not contain sweeteners in addition to fruit juice or concentrated fruit juice, so their solid contents are slightly lower than the contents of products that contain sweeteners. Because of the lack of additional sweeteners, consumers consider candied fruit products to be of high quality. The solid content of candied fruit is 55%
62%. In the upper solid content range, high amounts of methoxyl pectin are used for gel formation; while in the lower range, small amounts of methoxyl pectin are added to provide the ideal taste and texture.

5.2 BREAD Water can absorb a high level of methoxyl pectin, which increases the amount of dough, and significantly improves the dough’s freshness, hardness and stability. Therefore, adding pectin to dough can improve the dough and increase the baking volume of the bread. For example, adding pectin can reduce the flour amount by 30%, while maintain the existing Hamburg bread volume. Pectin can effectively extend the shelf life of bread, and during the normal shelf life, bread containing pectin maintains a good hardness.

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5.3 FROZEN FOOD Pectin can slow down the growth rate of crystals, reduce the loss of syrup, and improve the quality of frozen products. Ca21 and pectin have stabilize freeze
thaw effects on fruit. A low level of methoxyl pectin can improve the fruit quality in ice cream products. Pectin can improve the quality of frozen foods by controlling the size of ice crystals. In ice cream, pectin can prevent the reduction of flavor and pigment. When pectin is used for preparing gel pudding desserts, no freezing is needed to produce a sweet taste with a pudding consistency.

5.4 YOGURT PRODUCTS As a stabilizer in fruit yogurt, compared with starch and other vegetable gums, pectin provides an excellent flavor and texture. During the preparation of yogurt, low levels of methoxyl pectin and the other gums used together can prevent whey separation. The stable yogurt products can be obtained by using a high ester pectin, but if the pectin addition is limited, then the electrical charge will be neutralized. Because the repulsive force elimination system is not stable, continuing to add pectin will create a new repulsive force, which makes the acidified milk products stable. Hydrophobic and electrostatic reactions have very important roles in the stability of pectin
casein systems. In a whey protein emulsion, modified pectin can stabilize high concentrations of whey protein (Einhorn-Stoll et al., 1996). Thus, adding pectin can extend the shelf life of yogurt.

5.5 BEVERAGES At present, low sugar soft drinks possess a huge proportion of the soft drink market, but reducing the sweetening agent can affect the traditional beverage’s taste and sensory quality. This can be compensated by adding 0.05%
0.10% of high methoxyl pectin. High methoxyl pectin, as a suspension agent, when added to functional juice drinks with pulp, can gelatinize with calcium ions, reducing pulp precipitate from forming a material that is difficult to disperse, achieving a uniform fruit grain suspension. In addition, it provides an excellent flavor, enhancing the taste, and overcoming the alginate’s shortcomings, such as poor pseudoplasticity, rubber-like odor and high turbidity. Adding

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high-methoxy pectin gel to a beverage also has positive health effects, such as stimulating the stomach and relieving lead poisoning.

5.6 OTHERS Pectin can prevent gallstone formation, lower cholesterol, and remove intestinal cholic acid. It plays a role in regulating blood glucose and lowering the lipid content. It promotes intestinal peristalsis, prevents colitis, and it also protects the gastrointestinal mucosa, preventing damage. Pectin also can effectively prevent gastric ulcers. Thus, it can be developed into therapeutic and health care-related products. Pectin can be used as a food additive, with its excellent performance under low sugar and low heat conditions, making it the best-selling additive on the foreign market. The pectin production scale in China is not large, the scope of use is narrow, and sources are limited. With the increasing demand for low calorie foods, the amount of pectin used as a substitute for fat and sugar will increase dramatically. Therefore, we should make full use of waste materials, such as sweet potato residue, to extract pectin and develop large-scale energy-saving pectin industrialized production technology. Research on more applicable and feasible modification methods to improve pectin quality, giving it a market competitiveness, is necessary. In addition, further studies on sweet potato pectin structure and to establish the relationship between the structural composition and functional properties should be performed, along with studies on the physiological health functions of sweet potato pectin and clarifying its functional factors and mechanisms. These will promote the development and application of sweet potato pectin and its derivatives in the food, drug, and cosmetic industries.

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312. Thakur, B.R., Singh, R.K., Handa, A.K., 1997. Chemistry and uses of pectin- a review. Crit. Rev. Food Sci. Nutr. 37, 47
73. Turquois, T., Rinaudob, M., Taravelb, F.R., et al., 1999. Extraction of highly gelling pectic substances from sugar beet pulp and potato pulp: influence of extrinsic parameters on their gelling properties. Food Hydrocolloids 13, 255
262. Vaccitolo, J.A., Connolly, H.M., Rubenson, D.S., et al., 2001. Operation for anorexigenassociated valvular heart disease. Thorac. Cardiovasc. Surg. 122 (4), 656. Vanamala, J., Glagolenko, A., Yang, P., et al., 2008. Dietary fish oil and pectin enhance colonocyte apoptosis in part through suppression of PPARδ/PGE2 and elevation of PGE3. Carcinogenesis 29 (4), 790
796. Vayssade, M., Sengkhamparn, N., Verhoef, R., et al., 2010. Antiproliferative and proapoptotic actions of okra pectin on B16F10 melanoma cells. Phytother. Res. 24, 982
989. Wai, W.W., AlKarkhi, A.F.M., Easa, A.M., 2010. Comparing biosorbent ability of modified citrus and durian rind pectin. Carbohydr. Polym. 79, 584
589. Wang, S.J., Chen, F., Wu, J.H., et al., 2007. Optiimization of pectin extraction assisted by microwave from apple pomace using response surface methodology. J. Food Eng. 78, 693
700. Wei, H.X., Liang, B.D., Mu, T.H., 2008. Study on sweet potato pectin extraction technology. J. Food Ind. 4, 26
29, in Chinese. Wei, S., Guo, X.H., 2011. Optimum Extraction Process of Pectin from Jackfruit Filum. Food Res. Dev 6, 020, in Chinese. Willats, W.G., McCartney, L., Mackie, W., Knox, J.P., 2001. Pectin: cell biology and prospects for functional analysis. Plant Cell Walls. Springer, Netherlands. Yapo, B.M., Koffi, K.L., 2008. The polysaccharide composition of yellow passion fruit rind cell wall: chemical and macromolecular features of extracted pectins and hemicellulosic polysaccharides. J. Sci. Food Agric. 88 (12), 2125
2133. Yapo, B.M., Robert, C., Etienne, I., et al., 2007. Effect of extraction conditions on the yield, purity and surface properties of sugar beet pulp pectin extracts. Food Chem. 100, 1356
1364. Zhang, W.B., Liu, C.Z., Gao, L., 2010. Modified citrus pectin: preparation, characterization and anti-cancer activities. Chem. J. Chin. Univ. 31 (5), 964
969 (in Chinese). Zhang, Y.Y., Mu, T.H., Zhang, M., 2013. Optimisation of acid extraction of pectin from sweet potato residues by response surface met hodology and its antiproliferation effect on cancer cells. Int. J. Food Sci. Technol. 48 (4), 778
785.

FURTHER READING Arslan, N., Kar, F., 1998. Filtration of sugar-beet pulp pectin extract and flow properties of pectin solutions. J. Food Eng. 36, 113
122. Axelos, M.A., Thibault, J.F., 1991. The Chemistry of Low-Methoxyl Pectin Gelation. Academic Press, New York, pp. 109
118. Bergman, M., Djaldetti, M., Salman, H., et al., 2009. Effect of citrus pectin on malignant cell proliferation. Biomed. Pharmacother. 2824, 4. Kelco, C.P., 2001. GENU Pectin Book. CP Kelco ApS, Denmark.

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Fishman, M.L., Chau, H.K., Hoagland, P.D., et al., 2006. Microwave assisted extraction of lime pectin. Food Hydrocolloids 20, 1170
1177. Funami, T., Zhang, G.Y., Hiroe, M., et al., 2007. Effects of the proteinaceous moiety on the emulsifying properties of sugar beet pectin. Food Hydrocolloids 21, 1319
1329. Hayashi, A., Gillen, A.C., Lott, J.R., 2000. Effects of daily oral administration of quercetin chalcone and modified citrus pectin on implanted colon-25 tumor growth in balb-c mice. Altern. Med. Rev. 5 (6), 546
548. Hillman, L.C., Peters, S.G., Fisher, C.A., 1985. The effects of the fiber components pectin, cellulose and lignin on serum cholesterol levels. Am. Soc. Nutr. 42 (2), 207
213. Hu, D.L., Liao, J.K., Wu, X.L., et al., 2002. Free radicals and the oxidative damage of DNA. J. Environ. Hyg. 29 (5), 261
263 (in Chinese). Islam, A.M., Phillips, G.O., Sljivo, A., et al., 1997. A review of recent developments on the regulatory, structural and functional aspects of gum Arabic. Food Hydrocolloids 11, 493
505. Kasapis, S., Norton, L.T., Ubbink, J.B., 2009. Modern biopolymer science: bridging the divide between fundamental treatise and industrial application//McCclements. D. J. Biopolymers Food Emulsions. Academic Press, London, pp. 129
166. Liu, H.Y., Li, H.L., Dun, B.Y., 2008. Extraction of pectin from shaddock peel by microwave heating. Trans. CSAE 24 (8), 302
304 (in Chinese). Raz, A., Lotan, R., 1987. Endogenous galactoside-binding lectins: a new class of functional tumor cell surface molecules related to metastasis. Cancer Metastasis Rev. 6, 433
452. Sakamoto, T., Hours, R.A., Sakai, T., 1995. Enzymic pectin extraction from protopectins using microbial protopectinases. Process Biochem. 30 (5), 403
409. Salvador, L.D., Suganuma, T., Kitahara, K., et al., 2000. Monosaccharide composition of sweetpotato fiber and cell wall polysaccharides from sweetpotato, cassava, and potato analyzed by the high-performance anion exchange chromatography with pulsed amperometric detection method. J. Agric. Food Chem. 48, 3448
3454. Tamaki, Y., Konishi, T., Fukuta, M., et al., 2008. Isolation and structural characterization of pectin from endocarp of Citrus depressa. Food Chem. 107, 352
361. Terpstra, A.H.M., Lapre, J.A., de Vries, H.T., et al., 1998. Dietary pectin with high viscosity lowers plasma and liver cholesterol concentration and plasma cholesteryl ester transfer protein activity in hamsters. Nutr. Metab. 128 (11), 1944
1949. Yoo, S.H., Fishman, M.L., Hotchkiss, J.A.T., et al., 2006. Viscometric behavior of high-methoxy and low-methoxy pectin solutions. Food Hydrocolloids 20 (1), 62
67.

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CHAPTER

5

Sweet Potato Granules

SECTION 1: BACKGROUND FOR DEVELOPING SWEET POTATO GRANULES SECTION 2: TECHNOLOGIES AND KEY POINTS IN MANUFACTURING SWEET POTATO GRANULES 2.1 Manufacturing Technologies 2.2 Key Points 2.2.1 Selection of Special Cultivars 2.2.2 Storage of Raw Sweet Potatoes 2.2.3 Selection of Materials 2.2.4 Cleaning 2.2.5 Peeling 2.2.6 Slicing 2.2.7 Color Preservation 2.2.8 Calcium Soaking 2.2.9 Steaming 2.2.10 Mashing 2.2.11 Addition of Emulsifier 2.2.12 Drying SECTION 3: APPLICATIONS OF SWEET POTATO GRANULES 3.1 Applications of Sweet Potato Granules in Bread 3.1.1 Procedure of Making Bread with Sweet Potato Granules 3.1.2 Skills for Making Bread with Sweet Potato Granules 3.1.3 Effects of Sweet Potato Granules on Bread Production 3.2 Applications of Sweet Potato Granules in Biscuits 3.2.1 Procedure of Making Biscuits with Sweet Potato Granules

Sweet Potato Processing Technology. DOI: http://dx.doi.org/10.1016/B978-0-12-812871-8.00005-2 Copyright © 2017 China Science Publishing & Media Ltd. Published by Elsevier Inc. All rights reserved.

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3.2.2 Key Points in the Procedure for Making Biscuits with Sweet Potato Granules 3.2.3 Effects of Sweet Potato Granules on Biscuit Production 3.3 Applications of Sweet Potato Granules in Noodles 3.3.1 Manufacturing Technologies 3.3.2 Key Points in the Procedure for Making Noodles with Sweet Potato Granules 3.3.3 Effects of Sweet Potato Granules on Noodle Production 3.4 Applications of Sweet Potato Granules in Thick Slurries References

Abstract As a new product, sweet potato granules have been concerned by more and more people because of the characteristics, such as high cell integrity, wide reprocessing prospects, excellent storage properties, and so on. In this chapter, the background for developing, technologies and key points in manufacturing, methods of preparing bread, biscuits, noodles, and thick slurries with sweet potato granules were described.

SECTION 1: BACKGROUND FOR DEVELOPING SWEET POTATO GRANULES China is a top sweet potato production country in terms of the planting area and annual output. However, the sweet potato processing industry in China has developed slowly because of various problems, such as a low processing rate, limited finished products, and few processing components. Sweet potatoes in China are utilized as follows: 50% for feed, 14% for human consumption, 6% for cultivars, and only 15% for industrial processing. The other 15% decay due to improper storage (Lei et al., 2001). The sweet potato processing rate in China is significantly lower than those in the developed countries, such as South Korea and Japan. At present, Chinese sweet potatoes are mainly made into elementary commodities, such as starch, noodles, vermicelli, and chips, and a further expansion of the product range is required. In addition, the main component of sweet potatoes involved in processing is starch, and other nutritious components, including dietary fibers, proteins, and pectin, are rarely used during sweet potato processing.

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The preceding issues make it necessary to study and develop new products and to increase the comprehensive sweet potato processing rate. This is also expected by the Chinese government to promote key agricultural products that will improve regional agricultural economies. Sweet potato granules are produced from raw sweet potatoes. After a series procedures, including the selection, washing, peeling, slicing, color preservation, and boiling, the sweet potato cells are decomposed into several whole cells under certain conditions. After that, granules are obtained by drying with a low shear strength and a low extrusion pressure. Unlike simple dehydrated sweet potato products, sweet potato granules are a type of powder having a relatively high cell integrity, which contributes to a large number of processing advantages, including a high nutrition retention rate, wide reprocessing prospects, and excellent storage properties. Thus, sweet potato granules gradually became an ideal new sweet potato product in China. This chapter provides new perspectives on the research and development of sweet potato granules, and is aimed at exchanging information regarding, and promoting the development of, the sweet potato granule industry in China.

SECTION 2: TECHNOLOGIES AND KEY POINTS IN MANUFACTURING SWEET POTATO GRANULES 2.1 MANUFACTURING TECHNOLOGIES After years of painstaking research, our laboratory proposed an entire high-quality sweet potato granule manufacturing process involving several important technologies. Compared with the manufacturing technologies in other reports, those presented in our study were simple, conserved energy, and required only a low-cost equipment investment. The process is as follows:

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2.2 KEY POINTS 2.2.1 Selection of Special Cultivars The deviation in the quality of sweet potato granules (including free starch contents) produced from different sweet potato cultivars can reach 26.41%. Therefore, appropriate sweet potato cultivars need to be selected before the manufacturing of sweet potato granules. In a series of experiments, we found that the cultivars “Xu 25-2,” “Shang 0110-3,” “Suyu 303,” were suitable for making sweet potato granules.

2.2.2 Storage of Raw Sweet Potatoes Similar to other agro-products, sweet potatoes can be stored before granule production to prolong their lifecycle and increase the comprehensive use of equipment. In addition, sweet potatoes have thin skins and high water concentrations, making them prone to decay not stored correctly. The decay of raw materials causes nutritional losses, which further damages the economic benefits of sweet potato granule manufacturing. Thus, appropriate storage methods for raw sweet potatoes must be used to optimize the manufacturing of sweet potato granules. The process of storing raw sweet potatoes is as follows:

Raw sweet potatoes that are not damaged, without scabs, and with intact skins should be selected for storage. To sterilize raw sweet potatoes, hot steam and disinfectant can be used. The steam sterilization conditions are: Application mode: Spraying; spraying pressure: 1.2 times the standard atmospheric pressure; spray time: 3 s; steam temperature at the potato surface: 90 C. Disinfectants are used as follows: Application mode: Soaking; disinfectant type: Iprodione or carbendazim; disinfectant concentration: 0.1%; soaking time: 5 s (Afek and Orenstein, 2003). For wound healing of raw sweet potatoes, the following parameters are used: Temperature: 30 C; humidity: 92%; duration: 4
7 days (Van Oirschot et al., 2006). The appropriate sweet potato storage temperature ranges from 13 to 16 C. Because of changes in the color, flavor, and taste during sweet potato storage, the recommended storage temperature is 16 C.

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2.2.3 Selection of Materials The first food priority is safety. As a new type of sweet potato product, considerable attention to safety during their production is required. When sweet potatoes encounter the black spot pathogen, a toxin is produced, which consists of ipomeamarone and sweet potato ketoalcohol. The two toxins cause nausea, or even vomiting, dizziness, fatigue, and numbness in serious scenarios, with a fatality rate of 16%. After sweet potatoes are infected by black spot pathogen, its threats to human bodies cannot be eliminated or destroyed by boiling, steaming, or roasting. Therefore, it is important to filter out raw sweet potatoes impacted by the black spot pathogen during the manufacturing of sweet potato granules. After being infected by black spot pathogen, sweet potatoes have black spots, became hardened, and have a bitter taste. Thus, before manufacturing of sweet potato granules, sweet potatoes damaged by worms, with black spots, and showing decay need to be manually removed.

2.2.4 Cleaning Sweet potatoes are a rhizomatous crop, which grow underground and have impurities, like mud, on their surfaces when they are harvested. To keep the impurity concentration at a relatively low level in sweet potato granules, raw sweet potatoes need to be washed before manufacturing. A traditional potato cleaning method is based on a blowing cleaner. After potatoes are immersed and washed, they are sent to a blowing machine. By air-stirring and the rubbing of the roller, as well as the spray of high-pressure water, the potatoes are cleaned (Liu et al., 1999). Alternatively, the vibration of a rotating screen is employed to remove large-sized mud particles and sand, and then high-pressure water is used to rinse the potatoes (Kang et al., 2002). In addition, a cleaning method with both vertical and horizontal washing machines has been used for potato cleaning (He et al., 2005a,b).

2.2.5 Peeling Differences in the skin and flesh of sweet potato varieties generally exist. The taste of sweet potato skin is unpleasant; therefore, unpeeled

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sweet potatoes have a negative influence on the appearance and taste of sweet potato granules. At present, the main peeling methods include alkaline, steam, and abrasive peeling. Alkaline peeling treats materials with an alkaline solution at a certain concentration, and then washes them to remove their skins (Liu et al., 1999). Steam peeling places potatoes into a steam tank, ripens the potato skins with high-pressure steam, and abruptly reduces the pressure to peel their skins (Fang et al., 2002). Abrasive peeling uses a device with multiple rotation shafts. When potatoes and brushes on the rotation shafts rotate in different directions, abrasive force is produced to peel potato skins. Sweet potatoes have deep eyes, and therefore, most manufacturers use the abrasive or manual method to remove sweet potato skins.

2.2.6 Slicing During the production of sweet potato granules, the cell tissues of sweet potatoes are generally softened and loosened by heat. Sweet potatoes usually have irregular shapes, and thus, the direct heating of sweet potatoes results in excessive or insufficient softening and loosening of some parts, while other parts are appropriately softened and loosened. In this case, sweet potatoes need to be sliced to pieces of the same thickness to achieve even heating during the softening and loosening of the materials. The most appropriate slice thickness ranges from 8 to 15 mm for the manufacturing of sweet potato granules.

2.2.7 Color Preservation Color is an important factor influencing food quality. In food processing, enzyme-triggered and nonenzymatic browning introduce inevitable changes in the finished products’ color. Therefore, protective coloration is a crucial procedure in the manufacturing of sweet potato granules. Our study demonstrates that 0.1% ascorbic acid is an ideal color-retention agent during the production of sweet potato granules.

2.2.8 Calcium Soaking Our study shows that the calcium concentration is a key factor influencing the quality of sweet potato granules. After sweet potatoes are soaked in calcium solutions at different concentrations, the produced granules contain different free starch contents, with a variance of 6%.

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The most appropriate calcium concentration ranges from 56.04 to 63.96 ppm for the calcium soaking of sweet potato granules.

2.2.9 Steaming Steaming is a core procedure during the production of sweet potato granules. The purpose of steaming is to loosen the sweet potato cell tissue to cause it to decompose into whole single-cells in later procedures. Insufficient or excessive steaming is unfavorable for the complete separation of sweet potato cell tissues. The most appropriate steaming duration is 11.21
12.80 min for the manufacturing of sweet potato granules.

2.2.10 Mashing Unlike traditional sweet potato products, sweet potato granules are powdered products that are dried after compacted sweet potato cells are separated. Mashing is the main procedure for breaking the compacted cells. At present, sweet potato mashing is usually performed by squeezing and stirring. The latter is more fitted to the production of sweet potato granules because it retains a higher cell integrity level.

2.2.11 Addition of Emulsifier According to earlier reports, after a certain amount of emulsifier is added to sweet potatoes during mashing, the stickiness of the products is decreased, improving their dispersion (Ooraikul et al., 1978). Thus, the free starch content of sweet potato granules is significantly decreased. During the manufacturing of sweet potato granules, the most suitable emulsifier added to the products is 0.1% glycerin monostearate.

2.2.12 Drying The following methods can be used to dry sweet potato granules: Drying by air flow, freeze
drying, spray
drying, and drying using a rake dryer (Zhang, 2004). The procedure for drying sweet potato granules varies according to the drying method. If sweet potato granules are dried by air flow, then dried granules must added to mashed sweet potatoes to ensure the smooth loading of materials and to prevent sticking. In this way, the water content in mashed sweet potatoes is decreased to B30%, which satisfies the requirement of the air flow dryer. The freeze
drying and spray
drying procedures are simpler, requiring only the loading of sample materials.

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SECTION 3: APPLICATIONS OF SWEET POTATO GRANULES In recent years, research in China reported on the methods of utilizing sweet potato granules. According to the reports, sweet potato granules can be applied in bread (Lin, 2009); biscuits (Liang, 2009); noodles (Zhang et al., 2012); and thick slurries (Yuan, 2009). The following sections described the preparations of bread, biscuits, noodles, and thick slurry-associated food with sweet potato granules.

3.1 APPLICATIONS OF SWEET POTATO GRANULES IN BREAD Bread is important in human diets because it is convenient for consumption, easy to carry, and rich in nutrition. However, the dietary fiber content in bread is low due to its raw materials, which do not meet the modern requirements of a balanced human diet. Sweet potato granules represent a special raw material for making bread. Their addition during bread production helps to increase the nutrient value and improve the bread quality because of their thickening, water absorption, and water holding capabilities. They also provide supplemental dietary fiber.

3.1.1 Procedure of Making Bread with Sweet Potato Granules The making of bread with sweet potato granules involves flour mixing and production. Flour mixing skills:

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Production skills:

3.1.2 Skills for Making Bread with Sweet Potato Granules 3.1.2.1 Activation of Dry Yeast Dry yeast was dissolved in 30 C water and then left for 30 min before use. When dry yeast was used, mixing with white sugar was prohibited. 3.1.2.2 Flour Mixing White sugar, margarine, sweet potato granules, and bread improver were adequately stirred before they were mixed with flour. The flour was agitated for 8 min at a low speed and then agitated for 4 min at a high speed. 3.1.2.3 Static Fermentation Fermentation parameters are important to achieve a static fermentation effect. The fermentation temperature was under 27 or 28 C, the relative humidity was kept between 75% and 80%, and the fermentation time was 2 h. 3.1.2.4 Bread Formation Fermented dough was ready for division. Generally, dough was cut into B100 g pieces. The gas in dough must be eliminated by squeezing. Then, the dough can be made into a bread base of the required shape. 3.1.2.5 Proofing The suitable proofing conditions were as follows: Temperature: 35
40 C; relative humidity: 90%
95%; and duration: 45 min. 3.1.2.6 Baking To properly bake bread with sweet potato granules, the suitable temperature was determined to be 210
220 C, and the baking duration was 15 min.

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3.1.2.7 Cooling and Packing Bread with sweet potato granules was naturally cooled. When the central temperature becomes 35 C, the bread should be immediately packaged. Delayed packing will result in a great water loss, accelerated aging, flavor changes, and an increase in the risk of fungal infection. 3.1.2.8 Optimization of the Recipe for Making Bread with Sweet Potato Granules An optimized recipe for bread with sweet potato granules includes the following additives: 8% sweet potato granules; 12% white sugar; 0.8% bread improver; and 1.5% yeast.

3.1.3 Effects of Sweet Potato Granules on Bread Production Bread with sweet potato granules made using the preceding method is soft, tasty, not sticky, and elastic. The bread aroma is concordant with sweet potatoes. In addition, the storage stability of bread containing sweet potato granules is significantly improved when compared with bread without sweet potato granules.

3.2 APPLICATIONS OF SWEET POTATO GRANULES IN BISCUITS People are paying increasing attention to nutrition and health issues, resulting in greater food requirements. Because of the high nutrient content and the protective health-related functions of sweet potatoes and related products, such as sweet potato granules, the applications of sweet potato granules in biscuit manufacturing not only extend the sweet potato industry but also meet the consumers growing requirements. Biscuits containing sweet potato granules have a market great potential. The following section describes the method of producing biscuits using sweet potato granules.

3.2.1 Procedure of Making Biscuits with Sweet Potato Granules Biscuits with sweet potato granules are manufactured as follows:

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3.2.2 Key Points in the Procedure for Making Biscuits with Sweet Potato Granules 3.2.2.1 Supplemental Ingredient Treatment Supplemental ingredients used in biscuits with sweet potato granules included vegetable oil, white sugar, sodium bicarbonate, ammonium bicarbonate, gluconolactone, salt, eggs, vitamin C, water, and flavors. These ingredients required treatment before use. For example, white sugar must be ground to powder, and ammonium bicarbonate (leavening agent), sodium bicarbonate, and salt must be completely dissolved. The leavening agent and salt can be dissolved in eggs. Incompletely dissolved ammonium bicarbonate (leavening agent) and sodium bicarbonate will exist in the biscuit base as granules. When the base is baked, decomposed leavening agents will aggregate, which easily causes a foam on the biscuit, leading to black spots on the surface and blank pockets inside the biscuit. As a result, the biscuit quality is degraded. 3.2.2.2 Recommended Ratio of Granules to Supplemental Ingredients in Biscuits with Sweet Potato Granules The ratio of sweet potato granules to supplemental ingredients affects the quality of biscuits containing sweet potato granules. A low proportion of sweet potato granules indicates a high proportion of supplement ingredients, leading to an inadequate sweet potato flavor. A high proportion of sweet potato granules results in a thick sweet potato flavor, but a deteriorated dough performance, which makes it difficult to form biscuits, which are also hard to chew. The amounts of grease and white sugar added to biscuits containing sweet potato granules greatly influence the biscuit quality. A low

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grease volume makes biscuits hard to chew and causes a dry nonsmooth surface. A high grease volume leads to crisp biscuits, but they have an oily taste and are easy-to-break. A low white sugar content leads to inadequate caramelization during biscuit baking, unsatisfactory colorization, and haad to rd#to#chew biscuits. A high white sugar content leads to crisp biscuits, but they are dark in color and excessively sweet tasting. Our study showed that the proper proportions of sweet potato granules and other ingredients in the biscuit were as follows: 100 g high-gluten flour; 30 g sweet potato granules; 15 g grease; 20 g white sugar; 1 g sodium bicarbonate; 0.8 g ammonium bicarbonate; 1.6 g gluconolactone; 0.5 g salt; 4 g egg; 8 mg vitamin-C; an appropriate volume of flavoring; and water, as determined by the dough mixing conditions. 3.2.2.3 Mixing After sweet potato granules were evenly mixed with flour, the supplemental ingredients were added and mixed evenly. 3.2.2.4 Dough Concoction The temperature of the dough concoction should be kept between 22 and 28 C. 3.2.2.5 Rolling Dough was rolled flat and then dried before biscuits took shape. 3.2.2.6 Baking The most proper parameters for baking biscuits with sweet potato granules included the surface temperature of 220 C, bottom temperature of 190 C, and baking time of 9 min. 3.2.2.7 Cooling The baked biscuits containing sweet potato granules were quickly taken out and cooled to 38
40 C under room temperature, which allowed the water in the biscuits to continue evaporating, preventing biscuit deformation. 3.2.2.8 Arrangement and Packing Broken and irregular biscuits were eliminated, and then, the other biscuits were packed and sealed.

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3.2.3 Effects of Sweet Potato Granules on Biscuit Production Biscuits with sweet potato granules made using the preceding method that have a homogeneous color of gold or light purple. The taste is mildly sweet and crisp, with a loose structure. There is a sweet potato flavor, and high levels of dietary fiber and protein. Its taste and texture are like those of crisp biscuits, but the added grease volume does not meet the requirement of crisp biscuits. It is a new type of biscuit, between semihard and crisp.

3.3 APPLICATIONS OF SWEET POTATO GRANULES IN NOODLES Noodles are an important flour product consumed daily by people. With the elevation of living standards, people have increased their requirements for noodles. A large number of nutritious materials, including oat, buckwheat, mushroom, and egg, have been added to noodles to enhance their healthy properties. Sweet potato granules are nutritious and have many health-related functions and, therefore, can be added to noodles. The following section describes the method of producing noodles containing sweet potato granules.

3.3.1 Manufacturing Technologies The technologies for manufacturing noodles with sweet potato granules are as follows:

3.3.2 Key Points in the Procedure for Making Noodles with Sweet Potato Granules 3.3.2.1 Suitable Sweet Potato Granules for Making Noodles The quality of noodles containing sweet potato granules is significantly affected by the sweet potato granules. The sweet potato granule’s size is negatively correlated to the rate of broken noodles during its cooking. In contrast, the cooking loss rate of noodles increases

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concordantly with the size of sweet potato granules. The bend-to-break rate becomes lower and then higher as the size increases. Taking into account the rate of broken noodles during cooking, the cooking loss rate, and the bend-to-break rate, Zhang et al. (2012) concluded that the proper sweet potato granule size is 100 mesh for making noodles. 3.3.2.2 Proper Amount of Sweet Potato Granules for Noodle Production If excessive sweet potato granules are added, the gluten content of noodles is insufficient, making the noodles less elastic, easy-to-break, and less resistant to cooking. Taking into account the rate of broken noodles during cooking, the cooking loss rate, and the bend-to-break rate, Zhang et al. (2012) found that 10% sweet potato granules was the optimum amount for making noodles. 3.3.2.3 Mixing with Water and Flour and Static Aging With the increase in the dough’s water content, the breaking rate during the cooking of noodles containing sweet potato granules rose, and the cooking loss rate and bend-to-break rate first decreased and then increased. Therefore, the water content in dough is a parameter requiring special attention. During the preparation of noodles with sweet potato granules, the following parameters were recommended: Water content in dough: 24%; dough kneading time: 18 min; dough kneading temperature: 24
28 C; static aging time: 30 min (Zhang et al., 2012). 3.3.2.4 Rolling The thickness requires special attention during the rolling procedure. Thinly rolled noodles have a high cooking loss rate, while thickly rolled noodles have a high bend-to-break rate. The appropriate thickness for rolled noodles with sweet potato granules was 1.3 mm (Zhang et al., 2012).

3.3.3 Effects of Sweet Potato Granules on Noodle Production Noodles with sweet potato granules made in the preceding method have good chewiness, satisfactory hardness, excellent stickiness, and extraordinary performance.

3.4 APPLICATIONS OF SWEET POTATO GRANULES IN THICK SLURRIES Sweet potato granules can be used as materials for producing bread, biscuits, and noodles. They can also be put into concerted applications

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containing milk powder to make nutritionally balanced thick-slurry foods with sweet potato granules. Yuan (2009) conducted measurements on the amino acid content in sweet potato granules and whole milk powder, calculated the amino acid modes of sweet potato granules and whole milk powder, and evaluated the thick-slurry foods made with different proportions of sweet potato granules and milk powder based on the amino acid ratio coefficient and chemical scores. Then, based on approximations among the amino acid mode, standard World Health Organization (WHO)/Food and Agriculture Organization (FAO) mode, and all-egg mode of the prepared products, the best ratio of sweet potato granules to milk powder in thick-slurry foods was found to be 8:2. In addition, when considering stability and stickiness, the best stabilizer amounts of the thick-slurry foods containing sweet potato granules, as determined using the univariate analysis of variance (ANOVA) and orthogonal tests, were as follows: 0.65% sodium carboxymethylcellulose; 0.6% xanthan gum; 0.2% monoglyceride; and 12% saccharose.

REFERENCES Afek, U., Orenstein, J., 2003. Decreased sweet potato decay during storage by steam treatments. Crop Prot. 22, 321
324. Fang, X.F., Wu, G., 2002. A study on the production technologies of potato granules. Grain Oil Process. Food Mach 1, 14
16 (in Chinese). He, X.Y., Yang, S., 2005a. Production control of potato granules. Grain Oil Food Sci. Technol. 2 (13), 39
40 (in Chinese). He, X.Y., Yang, S., 2005b. Quality and production control of potato granules. Grain Oil 2, 15
17 (in Chinese). Kang, W.Y., Zhang, T., 2002. Processing technologies and product quality standards of potato granules. New Technol. New Prod. 2, 16
18 (in Chinese). Lei, M., Lu, X.L., Mao, L.J., 2001. Recent sweet potato food development and trend in China. Grain Oil Process. Food Mach. 11, 12
14 (in Chinese). Liang, M.F., 2009. A Study on the Processing Characteristics of Sweet Potato Granules and the Technologies of Applying Them to Biscuit Production. Changsha: Hunan Agricultural University (in Chinese). Lin, H., 2009. A Study on the Processing Characteristics of Sweet Potato Granules and Their Application in Bread Making. Changsha: Hunan Agricultural University (in Chinese). Liu, J.G., Chen, X.W., 1999. A brief introduction to the technologies for processing potato granules. Potato Mag 13 (1), 58
60 (in Chinese). Ooraikul, B., 1978. Production of Potato Granules. U.S. Patent: 4110478, 1978. Van Oirschot, Q.E.A., Rees, D., Aked, J., et al., 2006. Sweetpotato cultivars differ in efficiency of wound healing. Postharvest Biol. Technol. 42, 65
74.

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Yuan, L.N., 2009 Research on the Anti-breaking of Sweet Potato Granule Cells and Their Thickslurry Food. Wuhan: Central China Agricultural University (in Chinese). Zhang, T., 2004. New Technologies for Producing Whole-cell Sweet Potato Granules. People’s Republic of China: 03100273.0 (in Chinese). Zhang, Y., Xiao, Y.L., Chen, G., et al., 2012. A study on the influence of sweet potato granules on noodle quality. Food Ferment. Technol. 48 (1), 44
48 (in Chinese).

CHAPTER

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Sweet Potato Anthocyanins

SECTION 1: REVIEW OF SWEET POTATO ANTHOCYANIN 1.1 Introduction to Anthocyanins 1.1.1 Anthocyanin Structure 1.1.2 Anthocyanin Degradation 1.1.3 Copigmentation Effect 1.2 Status of Sweet Potato Anthocyanins 1.2.1 The Extraction Method of Purple Sweet Potato 1.2.2 Purification Methods for Sweet Potato Anthocyanin 1.2.3 The Structures of Sweet Potato Anthocyanins 1.2.4 Bioactivity of Sweet Potato Anthocyanins SECTION 2: THE PREPARATION OF SWEET POTATO ANTHOCYANINS 2.1 The Extraction of Sweet Potato Anthocyanins 2.1.1 Chemical Analysis of Purple Sweet Potato Powders 2.1.2 The Extraction of ATPE 2.1.3 Experimental Design for the Selection of Extraction Parameters 2.1.4 Plackett
Burman Experiment 2.1.5 Experimental Design for Optimizing Extraction Parameters 2.1.6 Comparing ATPS to the Other Methods 2.2 Purification of Sweet Potato Anthocyanins 2.2.1 The Adsorption and the Desorption Ratios of AB-8 Resin 2.2.2 Dynamic Adsorption and Desorption SECTION 3: THE STABILITIES OF ANTHOCYANINS FROM SWEET POTATO 3.1 The Effects of Temperature on the Thermal Stabilities of Anthocyanins

Sweet Potato Processing Technology. DOI: http://dx.doi.org/10.1016/B978-0-12-812871-8.00006-4 Copyright © 2017 China Science Publishing & Media Ltd. Published by Elsevier Inc. All rights reserved.

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3.2 The Effects of High Pressure on the Thermal Stability of Anthocyanins 3.3 The Effects of Different pH Levels on the Thermal Stability of Anthocyanins 3.4 Effects of Different Solvent Treatments on the Thermal Degradation of Sweet Potato Anthocyanins 3.4.1 Thermal Degradation Experiment 3.4.2 Thermal Degradation Model 3.4.3 Degradation Kinetic Parameters of Anthocyanins in Deionized Water 3.4.4 Degradation Kinetic Parameters of Anthocyanins in 10% Ethanol 3.4.5 Degradation Kinetic Parameters of Anthocyanins in 20% Ethanol 3.4.6 Degradation Kinetic Parameters of Anthocyanins in 50% Ethanol SECTION 4: BIOLOGICAL ACTIVITY OF SWEET POTATO ANTHOCYANIN 4.1 The Effects of Sweet Potato Anthocyanin on Acute Alcoholic Liver Damage and Dealcoholic Effects 4.1.1 Experimental Model for Investigating Acute ALD and Dealcoholic Effects 4.1.2 Effect of Sweet Potato Anthocyanins on Alcoholic Intoxication 4.1.3 Effect of Sweet Potato Anthocyanins on Growth and the Liver 4.1.4 Effects of Sweet Potato Anthocyanins on Serum ALT, AST, and LDH Activities 4.1.5 Effects of Sweet Potato Anthocyanins on Serum TG, TCH, and LDL-C Activity Levels 4.1.6 Effects of Sweet Potato Anthocyanins on the Hepatic MDA Content 4.1.7 Effects of Sweet Potato Anthocyanins on Hepatic Superoxide Dismutase (SOD) and Glutathione S-transferases (GST) Activity Levels 4.1.8 Effects of Sweet Potato Anthocyanins on the Hepatic ADH Activity Level 4.1.9 Effects of Sweet Potato Anthocyanins on the Histopathological Analysis

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4.2 The Effects of Sweet Potato Anthocyanin on Subacute Alcoholic Liver Damage 4.2.1 Subacute ALD Mice Models 4.2.2 Effects of Sweet Potato Anthocyanins on Growth and Liver Index 4.2.3 Effects of Sweet Potato Anthocyanins on Serum ALT, AST, and LDH Activity Levels 4.2.4 Effects of Sweet Potato Anthocyanins on Serum TG, and LDL-C Levels and TCH Activity 4.2.5 Effects of Sweet Potato Anthocyanins on the Hepatic MDA Activity Level 4.2.6 Effects of Sweet Potato Anthocyanins on Hepatic GST and SOD Activity Levels 4.2.7 Effects of Sweet Potato Anthocyanins on the Hepatic ADH Activity Level 4.2.8 Effects of Sweet Potato Anthocyanins on the Histopathological Analysis SECTION 5: APPLICATIONS OF SWEET POTATO ANTHOCYANINS 5.1 Pharmaceutical Industry 5.2 Food Industry 5.3 Cosmetics Industry References Further Reading

Abstract In this chapter, purple sweet potato anthocyanins (PSPAs) were extracted using aqueous two-phase system, and the conditions were optimized using response surface methodology; at the optimum conditions, the degradation kinetics and the effects on the acute and subacute alcohol liver disease were studied. The results showed that the optimal technological parameters for the aqueous two-phase extraction of PSPAs were as follows: liquid
solid ratio of 45:1 (mL/g), ethanol concentration of 27% (w/w), ammonium sulphate concentration of 20% (w/w), and pH 3.4. PSPA is a good antioxidant and exerts a protective effect in serum and in liver of mice intoxicated with ethanol. In conclusion, this study provides theoretic support for the development of functional foods with antioxidant and hepatoprotective activities.

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SECTION 1: REVIEW OF SWEET POTATO ANTHOCYANIN 1.1 INTRODUCTION TO ANTHOCYANINS 1.1.1 Anthocyanin Structure In recent years, natural food-coloring pigments have been increasingly favored because synthetic pigments have mutagenic, teratogenic, carcinogenic, and other harmful effects on human health (Yang et al., 2006). It is generally accepted that anthocyanins are the most important water-soluble pigments in many fruits and vegetables, such as grape, blueberry, cabbage, and purple corn. In addition, anthocyanins are widely used as natural pigment around the world, with the majority of countries (including the United States of America, China, Japan, and the European Community) allowing their use as food colorings, in place of synthetic pigments, because of their safety. In addition, the Food and Drug Administration (United States of America) included anthocyanin as a colorant without a license (Yao, 2009). Anthocyanins have the typical structure of flavonoids, with C6
C3
C6 as the basic skeleton, and they are derivatives of 2-phenylbenzopyrilium. Depending on the number and position of the hydroxyl and methoxyl substituents, a dozen different anthocyanidins have been described in publications, of which six are commonly found in fruits and vegetables (Fig. 6.1; Table 6.1).

Figure 6.1 Anthocyanin structure.

Table 6.1 Structures of the Anthocyanin Most Commonly Found in Foods Name

R1

R2

Pelargonidin

H

H

Cyanidin

OH

H

Delphinidin

OH

OH

Peonidin

OMe

H

Petunidin

OMe

OH

Malvidin

OMe

OMe

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Anthocyanin in fruits and vegetables is linked to one or more glycosidic unit. Sugars may be linked as mono-, di-, or triglycosides and may, in addition, be acylated with different organic acids. The glycosidic units may be linked to the anthocyanin by α or β linkage, and always occurs in position 3 of the aglycon. When additional sugars are present in the anthocyanin molecule, they are linked to positions 5 and 7. Glucose and galactose as hexoses, and rhamnose, arabinose and xylose as pentoses, are the sugars that most commonly form part of the anthocyanidin. The most common acylating agents are cinnamic acids, frequently p-cinnamic or caffeic acid, but also ferulic and sinapic acids (Clifford, 2000). They coexist as four main equilibrium species: the flavylium cation, the quinoidal base, the carbinol or pseudobase, and the chalcone C in aqueous solution (Fig. 6.2) (Sonia, 2008).

1.1.2 Anthocyanin Degradation In the plant, anthocyanins have a specific structure that can protect them from the pro-nuclear role, but once they have been removed from the environment, anthocyanins are unstable in food processing and cooking. The degradation occurs in various conversions, resulting in phenolic acids, aldehydes and diketones (Fig. 6.3) (Seeram et al., 2001).

Figure 6.2 Chemical transformations of anthocyanins.

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Figure 6.3 Anthocyanin degradation.

1.1.3 Copigmentation Effect Copigmentation is a phenomenon in which the pigments and other colorless organic compounds, or metallic ions, form molecular or complex associations, generating a change or an increment in the color intensity (Boulton, 2001). In food science, this phenomenon is considered a very important interaction because color is one of the main quality factors crucial to a product’s acceptance (Eiro et al., 2002). The anthocyanin
copigment interaction can be carried out in different ways depending on the interacting species: Intramolecular copigmentation, intermolecular copigmentation, self-association, and interaction with a metal (Fig. 6.4) (Castaneda-Ovando et al., 2009). Intramolecular and intermolecular copigmentation are the two main forms. Intramolecular copigmentation occurs when the copigment is part of the anthocyanin. For example, the acylation of anthocyanins stabilizes the pigment mainly because of the enhanced interaction.

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Figure 6.4 Anthocyanin interactions. (A) self-association, (B) intramolecular copigmentation, (C) metal complex formation, (D) intermolecular copigmentation (Castaneda-Ovando et al., 2009).

Intermolecular copigmentation occurs when the colorless copigment and colored anthocyanin are combined by a noncovalent (hydrophobic) force. “Anthocyanin-copigments” form horizontal or vertical laminated composites, reinforce anthocyanin coloration, and the copigments mainly include flavonoids and polyphenols. The interactions of different copigments with the same anthocyanins can result in different colors, and the effects of the same copigment with different threedimensional structures vary. The B rings of delphinidin and petunidin have at least one free hydroxyl group that can interact with metallic iron, aluminum, copper, magnesium, and potassium chelate.

1.2 STATUS OF SWEET POTATO ANTHOCYANINS Sweet potato (Ipomoea batatas) is an important plant, with mainly yellow or white flesh. However, Japan developed, by crossbreeding, purple sweet potato that has high yields and a high content of anthocyanins (Suda, 2003). China began to introduce purple sweet potato

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varieties in 1980, and then bred new varieties suitable for domestic cultivation, such as the purple sweet potato “135,” “Yan 337,” “Yan 176,” “Xushu 4,” “Jingshu 16,” “Yuzi 1,” and “Qunzi 1.” At present, purple sweet potato are widely planted in northeastern China, including Hebei, Shandong, Jiangsu, Guangxi, and Guangdong Provinces (Peng et al., 2010). Compared with anthocyanins from grapes, basil, purple rice, and black beans, the thermal stability of purple sweet potato is similar to purple corn, but greater than the other sources, and the light stability is the highest (Pu et al., 2010). Purple sweet potato is a nutrient-rich source, with a high yield and a low price. In addition, animal and clinical experimental results showed that sweet potato anthocyanins can be ingested in their complete molecular forms and increase the serum’s antioxidant capacity (Tomoyuki et al., 2006). These in vivo experiments showed that sweet potato anthocyanins can be used in their entirety by the body, leading to an increase in related research.

1.2.1 The Extraction Method of Purple Sweet Potato Since April 16, 2012, the Ministry of Health of China has approved the use of purple sweet potato pigments as food additives. The extraction methods for sweet potato anthocyanins are as follows: 1.2.1.1 Solvent Extraction Solvent extraction involves the removal of active ingredients with an appropriate solvent. It is divided into hot and cold extraction methods, which are performed at room or high temperature, respectively. Cold extraction extracts ingredients that are easily destroyed by heat and substances containing starches, gums, pectin, and mucus. Heat extraction is preferred over cold extraction because it can increase the solubility of the extracted component. Because anthocyanins are relatively stable under acidic conditions, acidified ethanol, acetic acid, hydrochloric acid, formic acid, and citric acid are used as extraction solvents (Wu et al., (2011). For the extraction of sweet potato anthocyanins, hydrochloride methanol and 0.5% citric acid, and ethanol or acidified ethanol (85:15 volume ratio) have been used as solvents (Yin et al., 2002). However, because methanol is highly volatile and has a certain toxicity, it should not be used as a natural pigment extraction agent but is

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mainly used for anthocyanin analyses and structural identification. Yao (2009) optimized an anthocyanin extraction method, using an extraction temperature of 60 C, the 5% formic acid solvent was used for 60 min, at a solid
liquid ratio of 1:10. Qiu et al. (2009) optimized the acidified ethanol method. The optimal extraction conditions for sweet potato anthocyanins were as follows: Extraction temperature of 80 C, extraction time of 60 min, and solid
liquid ratio of 1:32. Zhai et al. (2012) investigated the effects of extraction agent type, solid
liquid ratio, extraction time, and temperature on the extraction of pigments from purple sweet potato, and determined the optimum conditions of purple sweet potato pigment were, 40% ethanol as the extraction solvent at a solid
liquid ratio of 1/20 (g/mL) for 60 min at 30 C. 1.2.1.2 Microwave Extraction The microwave extraction has a heating effect on the solvent and material. The microwave’s heating mechanism is different from conventional heating, because it can penetrate the material and the extraction solvent, resulting in a uniformly heated system. This saves the time required for the outside to inside heating of conventional methods. Thus, the extraction efficiency is significantly greater compared with conventional heating methods. At present, the application of microwave heating to extract natural pigments is being promoted because of the time and thermal efficiencies, high product quality, high raw material utilization and its simplicity. Liu et al. (2012) extracted sweet potato anthocyanins using an enzyme-microwave assisted solvent extraction method, and compared with the conventional hydrochloric acid
ethanol method, the anthocyanin yield was increased 1.86-fold and the extraction time was shortened by 82%. 1.2.1.3 Supercritical Fluid Extraction Supercritical fluid extraction is a new extraction and separation technology, which has been widely used in the chemical, pharmaceutical, food, and other industries for the past two decades. He and Liang (1999) studied the artemisinin extraction method using supercritical fluid extraction technology, which greatly improved the solubility and color value of the pigments, and resulted in a good refining effect. However, the application requires costly equipment.

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1.2.1.4 Ultrasonic Extraction Ultrasonic extraction can accelerate exposing the target component to the solvent through its intense cavitation, mechanical vibration, emulsification, diffusion, crushing and stirring actions, and other multilevel effects, which can increase the frequency and speed of the material elements’ movements. Ultrasonic extraction is a powerful tool for extracting natural products and biologically active ingredients. Gu et al. (2004) extracted sweet potato anthocyanins using the ultrasonic extraction and compared the extraction effect with the freeze
thawing and solvent extraction methods. The ultrasonic method and freeze
thawing extraction were more effective than the solvent extraction, and the ultrasonic method was the most effective. Lien et al. (2012) showed that sonication could improve the anthocyanin extraction rate at 25 C for 22 min. 1.2.1.5 Aqueous Two-phase Extraction Different polymers of different solutions, or a polymer and a watersoluble inorganic salt mixture, separate into a high water concentration phase and an immiscible phase at a certain concentration. This is the basis of the Aqueous two-phase extraction (ATPE) method (Planas et al., 1996). Until now, traditional ATPE was based either on a polyethylene glycol (PEG)/salt or a polymer/polymer (e.g., PEG/dextran) system. However, because of the high cost of the polymers and difficulty in isolating the extracted molecules from the polymer phase, these systems cannot be used for large-scale production (Aydoggan et al., 2011). Recently, a short-chain alcohol/inorganic salt system has been used as a novel aqueous two-phase system (ATPS) to purify natural compounds. This ATPS has many advantages, such as low cost, low interfacial tension, good resolution, high yield, high capacity, and simple scale-up. ATPE is based on the impact of charges, hydrogen and ionic bonding, and other forces of the different distribution coefficients in the upper and lower phases (Kwon, 1996). Because of the unique advantages of the aqueous two-phase method, it has been studied by researchers (Lu and Deng, 2000). Relevant technical literature on the ATPE of natural products, demonstrating its value, has been reported, although in limited amounts. Ethanol
ammonium sulfate is a common and an economic ATPE system that has been applied to separate 1,3-propanediol and

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2,3-butanediol, with recovery rates of 98.1% and 91.7%, respectively, and the coefficients were 38.3 and 7.1, respectively (Li et al., 2010, 2011). Zhang et al. (2009) applied this method to separate bilberry anthocyanins, capsaicin, and red pigment. Wu et al. (2011) used this method to extract anthocyanins from Mulberry, which resulted in a high purity, removing almost 90% of the free sugar. 1.2.1.6 Other Extraction Methods To improve the yield of anthocyanins, mechanical crushing, heating, refrigeration, enzymes (such as pectinase, cellulase, and protease), pulsed-electric field, and pressing-assisted extraction have been used to shorten the extraction time and improve pigment yields. Pectinase was widely used in the production of fruit juice and wine. When extracting grape pigments and phenolic compounds, pectinase can break down the cell wall, promote the release of pigments and phenols, and shorten the soaking time, making it easier to clarify and filter (Capounova et al., 2002). Guo and Xu (2012) extracted sweet potato anthocyanins using α-amylase hydrolysis, with extraction conditions of 60 C, pH 5.5, 70 min extraction time, the 400 U/mL enzyme concentration 400 U/mL. In addition, pressure extraction technology has been developed in recent years. In this method, the inside of the cell breaks to accelerate the diffusion of the solvent. It can also improve the solubility and selectivity, thereby increasing the rate of leaching (Li et al., 2004). Truong et al. (2012) studied the pressurized liquid extraction process of sweet potato anthocyanins, and showed that the best extraction conditions were a pressure of 1500 psi and a acetic acid:methanol: water solvent [7/75/18 (v/v/v)]. The sweet potato anthocyanins from 335 cultivars were measured, and the cyanidin-3-glucoside content was 0
663 mg/100 g dry weight and 0
200 mg /100 g fresh weight. Pulsed electric-assisted extraction is a method that applies a potential voltage pulse to breakdown the cell membrane, thereby, increasing the membrane permeability and promoting intracellular substance elution. Pulsed electric-assisted extraction is a rapid new nonthermal processing technology with a low extraction temperature, resulting in a high extraction rate and good product quality (Mustafa et al., 2004). Eduardo et al. (2013) studied the pulsed electric-assisted extraction process of sweet potato anthocyanins, and found that the best

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conditions included an electric pulse of 3.4 kV/cm and 105 μs. The results of this method were similar to that using 96% ethanol at 40 C for 8 h. Traditional solvent extraction processes have widespread applications, but there is still the potential to improve. Ultrasonic, microwaveassisted extraction, supercritical dioxide extraction, enzyme-assisted extraction, and ATPE methods are at present not widely used in production. Combinations of these extraction methods will be a trend in the development of more applicable technology.

1.2.2 Purification Methods for Sweet Potato Anthocyanin The anthocyanin product obtained using traditional extraction processes is a crude natural pigment, and the color value is low and impurity content is high, which directly affects the stability of natural pigments and dyes, thus limiting its applications. Thus, the natural pigment production industry has introduced advanced chemical technologies for refining crude natural pigments. Thus, the purification process not only can improve the performance of the natural pigment, it can also expand the scope of application. Sweet potato anthocyanin contains many types of impurities, such as high contents of sugar, starch, and protein. In addition, the molecular weights of the sugars were similar to those of the anthocyanins, which made it difficult to remove the carbohydrates. Thus, to improve the color value of the product, the crude extracts must be purified. At present, there are five purification methods (Table 6.2).

1.2.3 The Structures of Sweet Potato Anthocyanins The structures of sweet potato anthocyanins have been studied by many researchers. Terahara et al. (1999) isolated eight acylated anthocyanins from the storage roots of the purple sweet potato, I. batatas cv Yamagawamurasaki, which is the source of the food colorant “purple sweet potato color.” Of these, six pigments were identified mainly by NMR analyses as diacylated anthocyanins, cyanidin, and peonidin 3-O-(6-O-(E)-caffeyl-2-O-(6-O-acyl-β-D-glucopyranosyl)-β-D-glucopyranoside)-5-O-β-D-gluco-pyranosides, in which each acyl substituent was a p-hydroxybenzoyl, (E)-caffeyl or (E)ferulyl residue. Furthermore, it was possible to isolate the monoacylated cyanidin-3-(6-caffeoylsophoroside)-5-glucoside, as well as three diacylated

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Table 6.2 Purification Methods of Sweet Potato Anthocyanin Method

Application

Feature

Resin

Ou et al. (2010) and Li and Gao (2003) studied purification processes that used different kinds of resin

The process is based on physical separation. There are limited chemical and biological changes, resulting in the maintenance of the stability and physiological activity of the pigment

Membrane separation

Li (2007) purified a sweet potato anthocyanin solution using 100 μm ultrafiltration. The impurity content was decreased, with a 38% removal rate

This process requires low energy and chemical inputs, leading to little environmental pollution. Moreover, it is easy to operate and industrialize

Grade alcohol precipitation

The method is based on the different solubility levels of anthocyanins, polysaccharides, starch, and protein in ethanol, with the ethanol concentration being adjusted repeatedly to precipitate the macromolecules

The grade alcohol precipitation method is simple; however, it is relatively complex, large amount of alcohol is consumed, and is accompanied by adsorption and double-teaming, resulting in different degrees of pigment loss

Lead acetate precipitation

Anthocyanin crude extract was precipitated by adding lead acetate, and the precipitate was collected by filtration

The method is not suitable for food or pharmaceutical grade anthocyanin extractions because of the toxicity of lead acetate

High-speed countercurrent chromatography

Qiu et al. (2009) purified four potato anthocyanins using high-speed countercurrent chromatography

The operation is simple and easy, and it results in a high separation efficiency and recovery rate. However, the equipment is costly

major pigments, cyanidin-3-(6, dicaffeoylsophoroside)-5-glucoside, cyanidin-3-(6-caffeoyl-6-p-hydroxy-benzoyl-sophoroside)-5-glucoside, and peonidin-3-(6-caffeoyl-6-p-hydroxybenzoyl-sophoroside)-5-glucoside using high-speed countercurrent chromatography (Montilla et al., 2010). With the increasing amount of purple sweet potato acreage in China, many researchers have studied the sweet potato anthocyanins structures because of their high photothermal stability. Yao (2009) found that the main anthocyanin of purple sweet potato (“Yan 176”) was 3-O-(6-O-trans-caffeyl-2-O-β-glucopyranosyl-β-glucopyranoside)-5O-β-glucoside peonidin. Qiu et al. (2009) found four anthocyanins, peonidin 3-O-(6-O-(E)-caffeoyl-2-O-β-D-glucopyranosyl-β-D-glucopyranoside)5-O-β-D-glucoside, cyanidin 3-O-(6-O-p-coumaroyl)-β-D-glucopyranoside, peonidin 3-O-(2-O-(6-O-(E)-caffeoyl-β-D-glucopyranosyl)-6-O-(E)-caffeoylβ-D-glucopyranoside)-5-O-β-D-glucopyranoside, and peonidin 3-O-(2-O(6-O-(E)-feruloyl-β-D-glucopyranosyl)-6-O-(E)-caffeoyl-β-D-glucopyranoside)5-O-β-D-glucopyranoside, from purple sweet potato using high-speed counter-current chromatography.

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Figure 6.5 The structures of sweet potato anthocyanins. Note: Me, methyl; Cy, cyanidin; Pn, peonidin; Caf, caffeic acid; So, sophoroside; PHB, hydroxybenzoic acid; Fer, ferulic acid; Glc, glucoside.

The structures of sweet potato anthocyanins are shown in Fig. 6.5. The main structures are A, B, C, D, E, F, G, and H because of different substituents, Cy-CafSop-Glc, Cy-diCafSop-Glc, CyCafPHBSop-Glc, Pn-CafSop-Glc, Cy-CafFerSop-Glc, Pn-diCafSopGlc, Pn-CafPHBSop-Glc, and Pn-CafFerSop-Glc, respectively.

1.2.4 Bioactivity of Sweet Potato Anthocyanins 1.2.4.1 Antioxidant Human metabolic processes produce a variety of free radicals, which can cause damage to proteins and fats, and through nucleic acid oxidation. This damage is an important cause of cancer, cardiovascular disease, and neurological disorders. The antioxidant activities of anthocyanins was 20-fold that of Vitamin C and 50-fold that of Vitamin E (Castaneda-Ovando et al., 2009). Teow et al. (2007) investigated the antioxidant activities of 19 sweet potato genotypes with distinctive flesh colors (white, cream, yellow, orange, and purple) through the oxygen radical absorbance capacity, 2,2-diphenyl-1-picrylhydrazyl (DPPH), and 2,2-azinobis (3-ethylbenzothiazoline-6-sulfonic acid), and found that the antioxidant

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activity of the purple flesh colored sweet potato was the highest. Moreover, the anthocyanins from purple sweet potato showed a stronger DPPH radical-scavenging activity than anthocyanins from red cabbage, grape skin, elderberry, or purple corn, and eight major components of the anthocyanins from purple sweet potato showed higher activity levels than ascorbic acid (Kano et al., 2005). In addition, the clear hydroxyl radical (  OH), superoxide anion radical (O2  ) and DPPH scavenging abilities of the anthocyanins reached 85%, 75%, and 99%, respectively, at a 5 mg/mL concentration (Yao, 2009). Han et al. (2006) suggested that purple potato flakes had antioxidant functions, such as radical scavenging activity and the inhibition of linoleic acid oxidation, and that they improved the antioxidant potential in rats by enhancing hepatic Mn-superoxide dismutase (SOD), Cu/Zn-SOD, and glutathione peroxidase mRNA expression. Zhang et al. (2009) found that sweet potato anthocyanins could effectively suppress D-gal-induced histology changes, including structural damage and leucocyte infiltration in mouse liver. In addition, anthocyanins could largely attenuate the D-gal-induced malondialdehyde (MDA) increase and could markedly renew the activities of copper, Zn-SOD, catalase, and Glutathione peroxidase in the livers of D-gal-treated mice. The western blot analysis showed that anthocyanins could inhibit the upregulation of the expression of nuclear factor-κB p65, Cyclooxygenase 2, and inducible nitric oxide synthase caused by D-gal. Han et al. (2011) studied the influence of sweet potato anthocyanins on thymocytes and found that sweet potato anthocyanins removed reactive oxygen species (ROS) by regulating the expression of Bcl-2 antagonist X-related p53 inhibition. 1.2.4.2 Antihyperglycemia and Hypertension An α-glucosidase inhibitor is used in the treatment of hyperglycemia as an effective drug that may decrease the postprandial blood glucose levels. Purple sweet potato anthocyanins (PSPAs) caused high oxidation resistance levels in 28 plant species, and a further study showed that α-glucosidase inhibition was high correlated with antioxidant activity (R2 5 0.82) (Mai et al., 2007). Fujise et al. (2008) evaluated the duration of the antihyperglycemic effects of 6-O-caffeoylsophorose (CS), a newly identified natural α-glucosidase inhibitor from fermented purple sweet potato, and found that the simultaneous or

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proadministration of CS with maltose eliminated the antihyperglycemic effect compared with the control. Angiotensin converting enzyme (ACE) can cause the catalytically inert activation of angiotensin Ito angiotensin II, and inhibit the vasodilator and bradykinin. Thus, inhibiting ACE activity is important in the prevention and treatment of hypertension. Yamakawa et al. (1998) studied the ACE inhibitory effects of different cultivated sweet potato anthocyanins, and found that the effect of purple sweet potato was the greatest compared with the other cultivars. When 1% sweet potato anthocyanin was added to food, it can significantly improve the blood pressures and heart rates of hypertensive rats (Shindo et al., 2007). The blood pressure was significantly lowered after consuming 400 mg/kg body weight of sweet potato anthocyanins. After an intake time of eight weeks, the blood pressure levels were quite similar to normal mice. 1.2.4.3 Antiatherosclerosis Purple sweet potato alcohol extract could enhance the Fe31-reducing ability, reduce the MDA content, and improve the low-density lipoprotein (LDL) content. The effects are similar to that of Vc (Park et al., 2010). Miyazaki et al. (2008) studied animal models using foods rich in cholesterol and fat, and found that sweet potato anthocyanins reduced arterial plaque areas, and lipid peroxidation (a serum oxidative stress marker) and sulfur thiobarbituric acid reactive substances (a liver and kidney oxidative stress marker) levels. Chen et al. (2011) studied the influence of sweet potato anthocyanins on lipid metabolism and oxidative stress in hyperlipidemia rats, compared with the control group. The total cholesterol potato anthocyanin (TCH), triglyceride (TG), and low-density lipoprotein cholesterol (LDL-C) levels, as well as the atherosclerosis index (AI), were significantly decreased, and the highdensity lipoprotein cholesterol level was increased significantly by the addition of sweet potato anthocyanins. 1.2.4.4 Liver Protection During liver fibrosis, hepatic stellate cells play critical roles in the increased formation and reduced degradation of the extracellular matrix in the liver. The anthocyanins suppressed hepatic stellate cell activation, including platelet-derived growth factor (PDGF)induced proliferation and α-smooth muscle actin expression. In addition, anthocyanins inhibited PDGF-induced Akt and ERK1/2

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phosphorylation. Anthocyanins also inhibited the phosphorylation of PDGF receptor-β (PDGFR-β) following PDGF-BB stimulation, providing a mechanism for the inhibition of aggregation factor -mediated kinase (Choi et al., 2011). Zhang et al. (2010a,b) indicated that PSPAs could protect mouse liver against D-gal-induced hepatocyte apoptosis by attenuating oxidative stress, inhibiting the activation of caspase-3, and enhancing cell survival signaling, which involves enhancing the level of the antiapoptotic protein Bcl-2 and the activation of the PI3K/Akt pathway. Hwang et al. (2011a,b) and Choi et al. (2010) studied the liver damage model of dimethyl nitrosamine (DMA), t-butyl hydroperoxide (T-BHP), and amino-ethyl phenol (APAP), and the results showed that PSPAs decreased the liver damage level lowered the alanine aminotransferase (ALT), aspartate aminotransferase (AST), and DMA contents, thereby, reducing the incidence of liver disease. Suda et al. (2008) examined the effects of purple sweet potato beverages rich in acylated anthocyanins on serum hepatic biomarkers in healthy Japanese men. The intake of the purple sweet potato beverage significantly decreased the serum levels of hepatic biomarkers, particularly the gamma glutamyltransferase level, in healthy men with borderline hepatitis. 1.2.4.5 Enhanced Memory Anthocyanins have antioxidant activities, which can cross the blood
brain
barrier into the central nervous system (Milbury and Kalt, 2010). D-Galactose is produced by body metabolites, and if its content is too high, then its degradation produces synapsins, cerebral cortical neurons that cause injury, learning and memory loss. Shan et al. (2009) evaluated the effect of anthocyanins on the brain aging in mice induced by D-galactose, and sweet potato anthocyanin intake significantly enhanced mouse passive avoidance and reduced D-galactoseinduced cognitive impairment. Lu et al. (2011) evaluated the effects of purple sweet potato color on cognitive deficits induced by hippocampal mitochondrial dysfunction in domoic acid-treated mice and explored the potential mechanisms underlying this effect. The oral administration of purple sweet potato color to domoic acid-treated mice significantly improved their behavioral performance in a step-through

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passive avoidance task and a Morris water maze task. Furthermore, purple sweet potato color significantly suppressed endoplasmic reticulum stress-induced apoptosis, which prevented neuronal loss and restored the expression of memory-related proteins. 1.2.4.6 Antibacterial Anthocyanins contain phenolic hydroxyl groups in their molecular structures, which can be incorporated into proteins or enzymes by hydrogen, damaging, or inactivating the protein structures, resulting in cytoplasmic condensation and disintegration. Thus, they have bacteriostatic effects. Wang et al. (2005) found that the inhibitory effects of anthocyanin from sweet potato on the growth of Escherichia coli, Pseudomonas aeruginosa, and Staphylococcus aureus were discovered and showed a positive correlation with the anthocyanin concentration of sweet potato. The results of a bacteriostatic experiment with anthocyanins from purple sweet potato by Han et al. (2008) showed that S. aureus and E. coli growth was inhibited in a dose-dependent manner, but Aspergillus niger and Saccharomyces cerevisiae were not affected by anthocyanin addition. 1.2.4.7 Other Bioactivities Potato anthocyanins has antiobesity properties and inhibits lipogenic factors, such as fatty acid synthase, lipoprotein lipase, and acetyl synthetase (Guo et al., 2011). Moreover, Ju et al. (2011) found that anthocyanins increased the phosphorylation of protein kinases and downstream targets of acetyl coenzyme A, thereby, activating the expression of carnitine acyl transferase, which increased fatty acid metabolism. Gout is a clinical syndrome in which tissue damage is induced by a chronic metabolic disorder associated with increased concentrations of uric acid in the blood. The study of Hwa et al. (2011) investigated the hypouricemic effects of anthocyanin extracts from purple sweet potato, and allopurinol, on serum uric acid levels in hyperuricemic mice. The administration of a single oral dose of 100 mg/kg anthocyanin of purple sweet potato on animals reduced the serum uric acid concentration to 4.10 6 0.04 mg/dL, compared with a concentration of 10.25 6 0.63 mg/dL in the hyperuricemic control group.

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In conclusion, sweet potato anthocyanins are safe, nontoxic, odorless natural pigments, and they have some nutritional and pharmacological effects, which have great potential in the food, cosmetic, and medical industries.

SECTION 2: THE PREPARATION OF SWEET POTATO ANTHOCYANINS 2.1 THE EXTRACTION OF SWEET POTATO ANTHOCYANINS ATPE has been widely applied to the separation of biomacromolecules, such as proteins and antibiotics, because of its mild conditions and high capacity. Until now, most ATPE purification was based either on PEG/salt or polymer/polymer (e.g., PEG/dextran) systems. However, because of the high cost of the polymers and difficulty in isolating the extracted molecules from the polymer phase, these systems cannot be used for large-scale production. Recently, short-chain alcohol/inorganic salt systems have been used as novel ATPE methods to purify natural compounds. The ATPE system has many advantages such as low cost, low interfacial tension, good resolution, high yield, high capacity, and simple scale-up (Rito-Palomares, 2004). Moreover, because of its structure, these are suitable for hydrophilic compounds. Short-chain alcohols, such as ethanol, methanol, and 2-propanol, can form stable and adjustable ATPE systems with inorganic salts, such as phosphate and sulfate. Ethanol
ammonium sulfate is a common and economic ATPE system, which has been applied to the extraction of anthocyanins from Mulberry (Wu et al., 2011), as well as 1,3-propanediol (Li et al., 2009), piceid, emodin, and resveratrol (Wang et al., 2008). In the present study, the applicability of ATPE formed by ethanol
ammonium sulfate to the extraction of PSPAs was investigated using the Box
Behnken design combined with the response surface methodology, and the structures were measured using HPLC
ESI
MS.

2.1.1 Chemical Analysis of Purple Sweet Potato Powders An entire batch of purple sweet potatoes was washed, cut, freeze dried, and then smashed to obtain purple sweet potato powder. The starch, crude fiber, ash, protein, and fat contents of the purple sweet potato powder were 63.35, 4.87, 3.98, 8.21, and 0.7 g/100 g dry matter,

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respectively, which indicated that starch was the predominant component of the powder followed by crude fiber, ash, protein, and fat. The anthocyanin content of the purple sweet potato powder was 311 mg/ 100 g, which was slightly lower than the anthocyanin content of grape skins (435.8 mg/100 g) reported by Margarita et al. (2009). However, this value was obviously greater than that of Korean black raspberries (37.2 mg/100 g) and purple corn (185.1 mg/100 g) (Ku and Mun, 2008; Yang and Zhai, 2010; Jing et al., 2012). Thus, it is a suitable raw material for the extraction of anthocyanins.

2.1.2 The Extraction of ATPE An ATPE system was prepared according to Li et al. (2010). A predetermined quantity of ammonium sulfate was dissolved in water, and then, certain volumes of ethanol and purple sweet potato powders were added to the solution and mixed well to form two phases. The mixture was mingled thoroughly and held until the two phases were completely separated. The PSPA concentrations in both the top and bottom phases were analyzed and the residues that accumulated at the interface of the two phases were discarded. The yield (Y1) was the ratio of the PSPAs partitioned in the top phase to the total amount of PSPAs. The partition coefficient (Y2) was the ratio of equilibrium concentrations of PSPAs in the top phase and bottom phase, which were calculated using the following equations: Y1 5 Ct 3 Y2 5

Vt ; Mtotal

Ct : Cb

(6.1) (6.2)

where Ct and Cb indicate the equilibrium concentrations of PSPAs in the top phase and bottom phase, respectively, Vt indicate the equilibrium volume of PSPAs in the top phase, and Mtotal represents the total amount of PSPAs.

2.1.3 Experimental Design for the Selection of Extraction Parameters The purple sweet potato powder was treated by various temperatures (20
60 C), concentrations of ethanol (20%
30% w/w) and concentrations of ammonium sulfate (17%
22% w/w), with a ratio of

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299

liquid-solid (mL/g) (ranging from 10:1 to 100:1 v/w), for a given time (from 20 to 120 min), while the pH value of deionized water ranged from 1 to 5. The anthocyanin yield (Y1) and the partition coefficient (Y2) were indices for evaluating the effects of different parameters.

2.1.4 Plackett
Burman Experiment The results of the Plackett
Burman experiment are shown in Table 6.3. The extraction and time had no significant effect on the anthocyanin yield (Y1) or the partition coefficient (Y2). Considering the cost of production and the economic efficiency, room temperature, and 20 min were chosen as extraction conditions.

2.1.5 Experimental Design for Optimizing Extraction Parameters Response surface methodology was applied to determine the working conditions for anthocyanin extraction from purple sweet potato based on the single-factor experiment results. A Box
Behnken design with four independent factors (X1, liquid
solid ratio; X2, ethanol concentration; X3, ammonium sulfate concentration; and X4, pH) set at three levels was carried out (Table 6.4). Table 6.3 Results of the Plackett
Burman Experiment Factors

Degrees of Freedom

F Value

Pr . F

pH

1

29.36

0.0066

Temperature

1

0.9

0.4532

Time

1

13.99

0.4926

Ratio of liquid
solid

1

60.90

0.0182

Ammonium sulfate

1

13.95

0.0153

Ethanol

1

20.38

0.0449

Block error

1

5.36

0.2349

Table 6.4 Design Approach and Experimental Results of the Response Surface Methodology X1 (mL/g)

X2 (%)

X3 (%)

X4

21

40

23

20

3

0

50

25

21

3.5

1

60

27

22

4

Note: X1, liquid
solid ratio; X2, ethanol concentration; X3, ammonium sulfate concentration, and X4, pH.

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The experimental results were fitted to the second-order regression: Y 5 β 0 1 β 1 X1 1 β 2 X2 1 β 3 X3 1 β 4 X4 1 β 11 X11 2 1 β 22 X22 2 1 β 33 X33 2 1 β 44 X44 2 1 β 12 X1 X2 1 β 13 X1 X3 1 β 14 X1 X4 1 β 23 X2 X3 1 β 24 X2 X4 1 β 34 X3 X4 ;

(6.3)

where Y represents the predicted response (the yield and the partition coefficient), β 0 represents the intercept; β 1, β 2, β 3, and β 4 represent linear coefficients; β 11, β 22, β 33, and β 44 represent squared coefficients; and β 12, β 13, β 14, β 23, β 24, and β 34 represent interaction coefficients. The goodness-of-fit of the regression model and the significance of parameter estimates were determined by the analysis of variance (Table 6.5). 2.1.5.1 Statistical Analysis and Model Fitting Response surface methodology was used to optimize the extraction conditions of anthocyanins. Experimental values obtained for Y1 and Y2 at the designed points are shown in Table 6.5. After application of the response surface regression procedure, the predicted model can be described by the following equations: Y1 585:9621:66X1 21:81X2 11:95X3 28:76X4 20:5X1 X2 27:31X1 X3 20:33X1 X4 20:19X2 X3 20:99X2 X4 20:93X3 X4 22:87X1 2 12:25X2 2 24:11X3 2 28:07X4 2 and (6.4) Y2 518:3110:041X1 20:66X2 12:81X3 12:35X4 21:99X1 X2 10:011X1 X3 20:88X1 X4 11:95X2 X3 21:35X2 X4 22:04X3 X4 22:99X1 2 22:04X2 2 20:63X3 2 23:14X4 2 : (6.5) An analysis of variance is required to test the significance and adequacy of the quadratic model. The results for Y1 and Y2 are shown in Tables 6.6 and 6.7, respectively. 2.1.5.1.1 Analysis of Variance for the Extraction Yield of PSPAs An analysis of variance for the model Y1 is shown in Table 6.6. The regression model was highly significant (P , 0.01), while the lack of fit was not significant (P 5 0.9673 . 0.05). The determination coefficient (R2) of the predicted model was 0.9977, indicating that only 0.23% of the total variation was not explained by the model. Meanwhile, a very low value of 0.37 for the coefficient of variation clearly indicated that

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Table 6.5 Experiment Results of the Response Surface Methodology Run

Y1 (%)

Parameters

Y2

X1

X2

X3

X4

1

50

23

21

3

89.56

9.78

2

60

27

21

3.5

81.46

10.82

3

50

23

20

3.5

83.80

15.66

4

50

27

21

3

88.13

11.41

5

50

25

21

3.5

85.44

18.82

6

60

23

21

3.5

85.99

16

7

40

25

20

3.5

71.4

11.67

8

60

25

21

4

64.26

13.49

9

50

25

21

3.5

86.46

18.36

10

60

25

20

3.5

82.78

11.89

11

40

23

21

3.5

88.49

12.02

12

40

25

21

4

68.14

15.25

13

50

27

20

3.5

80.26

10.03

14

50

25

22

3

83.78

17.08

15

40

25

22

3.5

89.65

17.18

16

50

25

20

4

62.19

16.35

17

50

25

20

4

68.617

13.49

18

50

23

22

3.5

88.16

17.38

19

50

27

21

4

68.62

13.49

20

40

25

21

3

84.98

9.13

21

50

23

21

4

73.99

17.25

22

50

25

22

4

68.12

17.98

23

60

25

22

3.5

71.81

17.45

24

60

25

21

3

82.42

10.9

25

40

27

21

3.5

85.97

14.82

26

50

27

22

3.5

83.86

18.55

27

50

25

21

3.5

85.97

17.75

Note: X1, liquid
solid ratio; X2, ethanol concentration; X3, ammonium sulfate concentration, and X4, pH, Y1 represents the yield, Y2 represents the partition coefficient.

the experimental values were associated with a very high degree of precision and a good deal of reliability. Thus, the model explained the response adequately. The result of the regression coefficient analysis showed that the linear effects of all of the variables were very significant (P , 0.01). Based on the significance of the regression coefficients of the quadratic polynomial model, pH value was found to be the

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Table 6.6 Analysis of Variance of the Anthocyanin Yield Test Source

Sum of Squares

Degrees of Freedom

Mean Square

F-Value

P-Value

Model

1805.5

14

128.93

1472.97

,0.0001

X1

33.02

1

33.02

77.19

,0.0001

X2

39.23

1

39.23

448.08

,0.0001

X3

45.58

1

45.58

520.71

,0.0001

X4

920.71

1

920.71

10518.49

,0.0001

X1X2

1.02

1

1.02

11.64

0.0052

X1X3

213.47

1

213.47

2438.076

,0.0001

X1X4

0.43

1

0.43

4.93

0.0465

X2X3

0.14

1

0.14

1.6

0.2304

X2X4

3.88

1

3.88

44.38

,0.0001

X3X4

3.44

1

3.44

39.32

,0.0001

X12

43.84

1

43.84

500.88

,0.0001

X22

27.03

1

27.03

308.84

,0.0001

X32

90.27

1

90.27

1031.33

,0.0001

X42

347.20

1

347.20

3966.56

,0.0001

Total error

1.05

12

1.088

Lack of fit

0.53

10

0.053

0.20

0.9673

Pure error

0.52

2

0.26

Total SS

1806.10

26 Predicted R2 5 0.9977

most significant factor affecting the anthocyanin yield, followed by ammonium sulfate concentration, ethanol concentration and the liquid-solid ratio. The quadratic terms (X12, X22, X32, and X42) were very significant (P , 0.01). Among the interaction terms, the interactions between X1X3, X2X4, and X3X4 (P , 0.01) were very significant with regard to the anthocyanin yield, and X1X2 and X1X4 (P , 0.05) were also significant. The other terms were not significant (P . 0.05). In addition, the standard errors of all of the regression coefficients ranged from 0.085 to 0.17. 2.1.5.1.2 Analysis of Variance (ANOVA) for the Partition Coefficient of PSPAs Table 6.7 showed the analysis of partition coefficient variance for the regression equation. It revealed that the most relevant variable (P , 0.0001) concerning the partition coefficient was pH value, followed by ammonium sulfate and ethanol concentrations, whereas the

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Table 6.7 Analysis of Variance of Anthocyanin Partition Coefficient Test Source

Sum of Squares

Degree of Freedom

Mean Square

F-Value

P-Value

Model

308.52

14

22.04

187.28

,0.0001

X1

0.020

1

0.020

0.17

0.6890

X2

5.3

1

5.3

45.02

,0.0001

X3

94.88

1

94.88

806.34

,0.0001

X4

66.45

1

66.45

564.68

,0.0001

X1X2

15.89

1

15.89

135.07

,0.0001

X1X3

4.731E-004

1

4.731E-004

4.020E-003

0.9505

X1X4

3.11

1

3.11

26.47

0.0002

X2X3

15.20

1

15.20

129.21

,0.0001

X2X4

7.26

1

7.26

61.73

,0.0001

X3X4

16.72

1

16.72

142.07

,0.0001

X12

47.62

1

47.62

404.70

,0.0001

X22

22.22

1

22.22

188.82

,0.0001

X32

2.12

1

2.12

18.04

0.0011

448.13

,0.0001

0.29

0.9803

X42

52.73

1

52.73

Total error

1.41

12

0.12

Lack of fit

0.83

10

0.083

Pure error

0.58

2

0.29

Total SS

309.93

26 Predicted R2 5 0.9803

liquid
solid ratio was not significant (P . 0.05); The quadratic terms (X12, X22, X32, and X42) were very significant (P , 0.01). In addition to the interaction term X1X3 (P . 0.05), the other interaction terms were also highly relevant, affecting the partition coefficient (P , 0.01). The determination coefficient (R2) of the model was 0.9803, which indicated that the model had adequately represented the real relationship between the parameters chosen. The regression model was highly significant (P , 0.01), and the lack of fit was not significant (P 5 0.9298 . 0.05). The coefficient of variation was 2.38 and the standard errors of the regression coefficients ranged from 0.099 to 0.20, which showed that the model can be used with accuracy. 2.1.5.2 Analysis of the Response Surface Response surfaces were plotted between two independent variables, while keeping the other independent variables at the zero coded level.

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Figure 6.6 Response surface plots of the extraction parameters on anthocyanin yield. (A) At varying liquid
solid ratios and ammonium sulfate concentrations, (B) at varying liquid
solid ratios and pH values, (C) at varying ethanol concentrations and pH values, (D) at varying ammonium sulfate concentrations and pH values.

Effects on the response functions Y1 and Y2 are shown in Figs. 6.6 and 6.7, respectively. 2.1.5.2.1 The Interaction Between the Variables on Y1 The effects of different X1 and X3 on the Y1 when X2 and X4 were fixed at zero levels are illustrated in Fig. 6.6A. With the liquid
solid ratio and ammonium sulfate concentration rising, the anthocyanin yield increased, which was probably because the anthocyanins mostly combined with proteins and polysaccharides. With the liquid
solid ratio increasing, more solvent could enter cells and more anthocyanins could permeate into the solvent (Prasad et al., 2009). When the ammonium sulfate concentration increased from 20% to 21.5%, the anthocyanin yield increased from 70% to 85%. This was probably because of the enhanced salting-out effect. Our results were in good agreement with those of Li et al. (2010), where the 2,3-butanediol content increased as the ammonium sulfate concentration increased.

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Figure 6.7 Response surface plots of the extraction parameters on the anthocyanin partition coefficient. (A) At varying liquid
solid ratios and ethanol concentrations, (B) at varying ethanol concentrations and ammonium sulfate concentrations, (C) at varying ethanol concentrations and pH values, (D) at varying ammonium sulfate concentrations and pH values.

Fig. 6.6B shows a response surface plot of the effects of pH and the liquid
solid ratio, which were very significant (P , 0.01) on the anthocyanin yield (Table 6.6). However, the interaction between them was mainly the effect of the pH value because pH can usually change the electrical properties of assigned materials, and then influence the electric potential of the ATPS. With the pH level increasing, a decline in the anthocyanin yield was observed, which may be because the anthocyanins were stable under acidic conditions, and a higher pH can lead to the degradation of anthocyanins, which results in the destruction of the structure (Fan et al., 2008). While lower pH values result in the hydrolysis of anthocyanins (Yang et al., 2008a,b), the appropriate pH value is needed in the extraction process. Fig. 6.6C showed the response surface plot of various ethanol concentrations and pH values at an ammonium sulfate concentration of

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20% and liquid
solid ratio of 50:1 (mL/g). The interaction effect was similar to that seen in Fig. 6.6B. The influence of the ethanol concentration was not significant, which was probably because the ethanol concentration increased from 23% to 27%, and the ATPS was not changed significantly. Acidified ethanol is a traditional method of extracting anthocyanins. However, our results were obviously different than those of Jayaprakasha et al. (2008), where the total polyphenol extraction yield of grapes increased when the ethanol concentration increased. This may have been because the ethanol concentration was in the range of 23% to 27%. When the ethanol is redistributed in the top and bottom phases, the ethanol concentration in top phase can reach to 60%. Ghafoor et al. (2009) reported that an ethanol concentration of 60% was sufficient for extracting high quantities of anthocyanins from grape seeds. The influence of the interaction between ammonium sulfate concentration and pH value are illustrated in Fig. 6.6D. The interaction effect was similar to those in Fig. 6.6B and C, which were influenced by pH, and was significant compared with that of the ammonium sulfate concentration. Therefore, the pH value should be strictly controlled in the extraction process. 2.1.5.2.2 The Interaction Between the Variables on the Anthocyanin Partition Coefficient (Y2) The effects of the ethanol concentration and liquid
solid ratio shown in Fig. 6.7A demonstrated that the anthocyanin partition coefficient increased rapidly with the increase in the ethanol concentration and liquid
solid ratio. This may be because increasing the ethanol concentration can lead to an increase in the concentration in the top phase, which favors the distribution of anthocyanins to the top phase. Guo et al. (2013) have reported that when the ethanol concentration increased from 15% to 19%, the partition coefficient of lignans increased from 17.05 to 74.93. Wang et al. (2008) obtained higher partition coefficients of piceid, resveratrol, and emodin from Polygonum cuspidatum, when an ethanol concentration of 25% was used. Our results were in good agreement with these reports. The influence of the liquid
solid ratio on the anthocyanin partition coefficient was not significant (P . 0.05), but the interaction with ethanol concentration was significant, which may be because the extraction process was more complex and affected by many factors. Although the liquid
solid ratio

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307

did not change in the ATPS, if the liquid
solid ratio increased, then the rate of penetrating and spreading to purple sweet potato cell increased. Thus, increasing the rate of damage to the hydrophobic bonds and diffusing to the top phase, thereby, affecting the anthocyanin partition coefficient. The effects of the ammonium sulfate and ethanol concentrations on the anthocyanin partition coefficient are shown in Fig. 6.7B. The anthocyanin partition coefficient increased from 13.29 to 19.05 with an increase in the ammonium sulfate concentration from 20% to 22%. This is probably because of the “ion dipole” between salt ions and water molecules, which can reduce the quantity of free water in the bottom phase, thus crowding out ethanol and target objects assigned to the top phase (Wang et al., 2008). Our results were consentience with those of Li et al. (2010), in which the increase of the ammonium sulfate concentration from 20% to 28% resulted in the partition coefficient of 2,3-butanediol increasing from 4 to 8.29. Within a certain range, with an increase in the ethanol concentration (23%
26%), the anthocyanin partition coefficient increased, which is also seen in Fig. 6.7A. However, with a further increase in the ethanol concentration, a decline in the partition coefficient was observed (Fig. 6.7B). This may be because the anthocyanin content in the top phase was close to saturation, and the distribution in bottom phase was more than in the top phase, reducing the anthocyanin partition coefficient. With increases in pH (3
4) and ethanol concentration (23%
27%), the partition coefficient increased from 9 to 17. The impact of the ethanol concentration was similar to that seen in Fig. 6.7A. When the pH changed from 3 to 4, the partition coefficient increased from 9.52 to 17.46, which may have been because as the pH increased, anthocyanins gradually combined with H1, weakening the binding force of the top phase, which decreased the partition coefficient. However, an increase in pH can increase the electric potential (Rito-Palomares, 2004), and the electric potential is proportional to the partition coefficient, but the impact of this effect was not great. Our results were supported by the results of Wu et al. (2011) and Guo et al. (2012). The effects of pH and ammonium sulfate concentration on the anthocyanin partition coefficient are shown in Fig. 6.7D. With an increase in the ammonium sulfate concentration, the partition coefficient increased from 7 to 18, which are shown in Fig. 6.7B and C.

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2.1.5.2.3 Extraction Condition Optimization and Method Validation The optimum extraction conditions were determined by the canonical response surface analysis, which predicted that the conditions of a 47.34:1 liquid
solid ratio, 23.20% ethanol concentration, 21.56% ammonium sulfate concentration, and pH 3.33 would lead to the maximum anthocyanin yield, while the conditions of a 43.29:1 liquid
solid ratio, 27.00% ethanol concentration, 22.00% ammonium sulfate concentration, and pH 3.34 would result in the maximum anthocyanin partition coefficient. Considering the cost and process requirements comprehensively, we determined that a 45:1 liquid
solid ratio, 25% ethanol, 22% concentration of ammonium sulfate, and pH 3.3 were the best conditions for PSPAs. The verification experiment was performed using the selected optimal conditions. The experimental anthocyanin yield was 90.02 6 0.01%, and anthocyanin partition coefficient was 19.62 6 0.02, which was in agreement with the predicted values (90.12% and 19.73, respectively) using the equation. This indicated that the model was adequate for describing the extraction process.

2.1.6 Comparing ATPS to the Other Methods The extraction yield of ATPS was similar to those of traditional methods (Table 6.8), but the color value was significantly higher than in the other methods, the sugar content was significantly reduced by B30%
40%, and the level of protein was reduced B40%
70% compared with the other extraction methods. In conclusion, response surface methodology is a useful tool for determining the optimal extraction conditions of anthocyanins from purple sweet potato. ATPE was a mild method compared with the conventional solvent extraction. Meanwhile, this method can maintain the original composition and structure. These results demonstrated the successful extraction of anthocyanins with ATPE, providing potential benefits for the industrial extraction of anthocyanins from purple sweet potato. Table 6.8 Comparing ATPS to the Other Methods Method

Yield (%)

Color Value

Protein (w/w)

Sugar (w/w)

Citric acid

90.23

4.98

5.94

17.27

Formic acid

91.74

4.27

4.12

18.23

Acidified ethanol

90.78

5.42

4.88

15.93

ATPS

90.02

9.81

2.93

6.06

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2.2 PURIFICATION OF SWEET POTATO ANTHOCYANINS Macroporous resins are durable polar, nonpolar, or slightly hydrophilic polymers with high adsorption capacities for organic molecules (Fu et al., 2006). They can selectively adsorb the targeted constituents from aqueous and nonaqueous systems using electrostatic force, hydrogen bonding interactions, complexation, or size sieving (Gao et al., 2007). Therefore, macroporous resins have been widely used in the separation and purification of biologically active substances because of their physicochemical stability, high adsorption selectivity, and easy recycling (Wan et al., 2014). AB-8 macroporous resins are weak polar resins and have been widely used in the purification of plant polyphenols because of their appropriate surface area and nuclear pore size (Gao et al., 2013; Zhao et al., 2013). In this study, the adsorption properties of AB-8 macroporous resins for sweet potato anthocyanins was investigated.

2.2.1 The Adsorption and the Desorption Ratios of AB-8 Resin 2.2.1.1 Static Adsorption of Sweet Potato Anthocyanins on the AB-8 Resin Resin (2 g) was added to 200 mL sweet potato anthocyanins (absorbance 5 0.465). Sample were taken every 20 min and the absorbance of the anthocyanin solution was measured using the following formula: Adsorption rate 5

ðA0 2 Aend Þ ; A0

(6.6)

where A0 represents the absorbance at the beginning, and Aend represents the absorbance at the end. The static adsorption properties of sweet potato anthocyanins are shown in Fig. 6.8. The absorbance of sweet potato anthocyanin decreased as the time increased. When the time was 120 min, the absorbance was stable, and the final adsorption rate was 82%. 2.2.1.2 The Effects of pH on the Adsorption Rate of the AB-8 Resin The crude anthocyanin solutions (absorbance 5 0.465) were prepared, and each of them was adjusted to a different pH value (1.0, 1.5, 2.0, 2.5, and 3.0) using 1.0 mol/L HCl and 1.0 mol/L NaOH solutions. The adsorption rate was not significantly affected by pH values ranging from 1.0 to 3.0 (data not shown).

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0.8

Absorption

0.6 0.4 0.2 0

0

20

40

60 80 Time (min)

100

120

720

Figure 6.8 Static adsorption of sweet potato anthocyanins on the AB-8 resin.

0.5

Absorption

0.4 0.3 0.2 0.1 0

0

20

40 60 Ethanol (%)

80

100

Figure 6.9 The effects of ethanol on the desorption of the AB-8 resin.

2.2.1.3 The Effects of Ethanol on the Desorption of the AB-8 Resin Different ethanol concentrations (20%, 40%, 50%, 60%, 70%, 80%, and 90%) were used to study the desorption rate. The effects of ethanol on the desorption of the AB-8 resin are displayed in Fig. 6.9. The absorption value of desorption solution was the highest (Absorption 5 0.450) when 70% ethanol was used. If the ethanol concentration increased continuously, then the desorption decreased. Thus, 70% ethanol was chosen as the desorption solution.

2.2.2 Dynamic Adsorption and Desorption 2.2.2.1 The Outflow Curve of Anthocyanins and Adsorption Capacity of the Resin The conditions could be regarded as reaching absorption saturation when the absorbance values of the added and effluent solutions were similar. The adsorption capacity was determined when the adsorption reached saturation. Then, the column was washed using a 0.01%

Sweet Potato Anthocyanins

311

0.12 0.1 Absorption

0.08 0.06 0.04 0.02 0 0

1

2

3

4

5

6 7 8 Volume (l)

9

10 11 12 13 14

Absorption × dilution

Figure 6.10 The outflow curve of the anthocyanins.

80 70 60 50 40 30 20 10 0 0

100

200 Volume (ml)

300

400

500

Figure 6.11 Dynamic desorption curve of anthocyanins.

aqueous hydrochloric acid solution, and the resolved anthocyanins were washed with 70% aqueous ethanol. The desorption solution was collected, concentrated by rotary evaporation, lyophilized, and weighed to calculate the saturated absorption capacity: Absorption saturation 5

the weight of anthocyanins ðgÞ : the volume of resin ðmLÞ

(6.7)

The outflow curve of the anthocyanins is shown in Fig. 6.10. As the volume of the effluent solution increased, the absorption increased. In addition, the absorption was highest when the volume was 7 L, and absorption saturation was 0.06 g/mL of wet resin. 2.2.2.2 The Desorption Curve of Anthocyanins from the Resin After the resin reached the condition of saturated adsorption, the anthocyanins were resolved using 70% ethanol. The eluent flow rate was 1 mL/min. The dynamic desorption curve is shown in Fig. 6.11. In the desorption process, the absorption of the desorption solution was

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highest when the volume reached 70 mL, and the greatest amount of anthocyanins was desorbed when the volume reached 150 mL. The purified performance of AB-8 macroporous resin was examined by static adsorption, desorption and dynamic adsorption, and desorption tests. The AB-8 macroporous resin had a strong adsorption capacity for sweet potato anthocyanins, pH values in the range of 1.0
3.0 had no significant impact on the adsorption (P , 0.05), and the best desorption solution was 70% ethanol.

SECTION 3: THE STABILITIES OF ANTHOCYANINS FROM SWEET POTATO Sweet potato anthocyanins are easily oxidized and denatured; therefore, their biological activities easily decrease during processing. Thus, the stability of sweet potato anthocyanin is a key factor influencing its applications. The number of substituents, the binding site, the presence of organic acids, and the molecular weight all can affect the stability of an anthocyanin. High molecular weight anthocyanins acylated with organic acids showed greater stabilities. Acylated anthocyanins are the main components of PSPAs, which exhibited good thermal stability, and heat and light resistance during the production and storage process, so, they can be widely used as sources of food coloring agents. Moreover, their byproducts can be integrated in foods, chemicals, feed, and other raw materials for processing. This will improve the utilization value of purple sweet potato and has great potential for the development and utilization of sweet potato anthocyanins. This section examines the effects of different conditions on the stability of anthocyanins from purple sweet potato (Yan 176), and studies their degradation in different solvents (water, 10% ethanol, 20% ethanol, and 50% ethanol). The information provides a theoretical basis for the application of sweet potato anthocyanins in the food industry.

3.1 THE EFFECTS OF TEMPERATURE ON THE THERMAL STABILITIES OF ANTHOCYANINS Using 15 25
mL volumetric flasks, the samples were divided into five groups, and each group was run in triplicate. Then, 10 mL anthocyanin crude extract was added to each flask, and the pH was adjusted to 3 in a citric acid
disodium hydrogen phosphate buffer solution. Next,

Sweet Potato Anthocyanins

Preservation rate (%)

25°C

40°C

100°C

80°C

60°C

313

1.2 0.9 0.6 0.3 0

0

1

2

3

4

5

6

Time (h) Figure 6.12 The effects of temperature on the thermal stabilities of anthocyanins.

20 mL of each solution was placed in a 20 mL glass test tube and the tubes were incubated in 25, 40, 60, 80, and 100 C water baths. The absorbance values of the samples were measured every 1 h, and the preservation rate of the pigment was calculated as thermal stability using the following formula: Preservation rate 5

A ; A0

(6.8)

where A represents the absorbance after treatment, and A0 represents the absorbance before treatment. The results are shown in Fig. 6.12. Over time, the preservation rate was not significantly different when the temperature was 25 or 40 C. After heating for 1 h at 60 C, the preservation rate of anthocyanins decreased slightly, but with prolonged heating, the preservation rate did not change significantly. However, when the treatment temperature was raised to 80 or 100 C, the anthocyanin preservation rate decreased significantly. These results suggested that the anthocyanins were stable when the temperature was below 60 C. In addition, although the preservation rates of anthocyanins at 80 and 100 C were lower, they were still above 80% after heating 4 h, indicating that the PSPAs had good thermal stability. The anthocyanins in production and storage processes should not be exposed to a high temperature environment.

3.2 THE EFFECTS OF HIGH PRESSURE ON THE THERMAL STABILITY OF ANTHOCYANINS The anthocyanin crude extract was diluted 10-fold into 15 parts, and 10 mL of each was separately placed into pressurized plastic bags. Then, 100, 200, 300, 400, and 500 MPa were applied for 15 min at

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25°C

100°C 2h

1.5

Preservation rate (%)

1.2 0.9

0.6 0.3 0 0.1

100

200

300

400

500

Pressure (MPa) Figure 6.13 The effects of pressure on the thermal stability of anthocyanins.

Table 6.9 Effects of Different pH Levels on Color Changes During Heating Process Before treatment

pH

Color

pH

Color

2

Red

After treatment

2

Red

3

Red

3

Red

4

Purple
red

4

Slight purple
red

5

Purple
red

5

Slight purple
red

6

Slight purple

6

Light brown

7

Purple

7

brown

8

Blue
green

8

Light green

25 C after sealing. These were then sampled to determine the preservation rate. The remains were than heated at 100 C for 2 h, and the preservation rate was determined again (Fig. 6.13). The preservation rates of sweet potato anthocyanins are similar to those of the controls at different pressures (100
500 MPa). However, when the samples were heated for 2 h at 100 C, the preservation rate fell to B60%. These results indicated that the anthocyanins from sweet potato were pressure resistant. High pressure treatments are undoubtedly effective in producing sterilized anthocyanin pigments (Table 6.9).

3.3 THE EFFECTS OF DIFFERENT pH LEVELS ON THE THERMAL STABILITY OF ANTHOCYANINS Certain amounts of potato anthocyanin crude extract were placed in beakers, and the pH levels were adjusted to 2, 3, 4, 5, 6, 7, and 8, using

Sweet Potato Anthocyanins

Preservation rate(%)

100

100ć 1h

315

100ć 2h

80 60 40 20 0

0

2

4

pH

6

8

10

Figure 6.14 Effects of heating on the stability of sweet potato anthocyanins at different pH levels.

0.1 mol/L NaOH and 0.1 mol/L HCl. These samples were then transferred to a 50-mL volumetric flask with a corresponding pH buffer solution to a constant volume. Aliquots of 20 mL were placed into test tubes with stoppers, and the test tubes were placed at 100 and 25 C and heated for 1 and 2 h. The color change was recorded (Table 6.10) and the preservation rate calculated (Fig. 6.14). The color change associated with sweet potato anthocyanins at different pH values during heating 2 h at 100 C are shown in Table 6.10. Different pH values had different influences on the color change associated with the anthocyanins. At pH # 3, the color (red) did not change significantly before and after treatment, while the colors of the anthocyanin solutions at pH 4 and 5 were light red, becoming slightly purple after heating. At pH $ 6, the anthocyanin color changed, turning from light purple, purple, and blue
green to light brown, brown, and light green. These changes may be due to the anthocyanins being relatively stable under lower pH conditions (pH # 4). Thus, anthocyanins as the food colorant should always be stored at pH # 4 (Shi et al., 1992). The effects of different pH levels on anthocyanin stability during heating are shown in Fig. 6.14. With increasing pH values, the anthocyanin preservation rate decreased significantly. At pH # 4, the preservation rates were not significantly different after heating for 1 or 2 h While under alkaline conditions, longer heating times resulted in lower preservation rates. At pH # 6, the preservation rates of the anthocyanins decreased as the pH increased. When the pH was 6, the lowest preservation rates of 68% and 57% occurred after 1 h and 2 h, respectively, while at pH . 6 the preservation rate of anthocyanins increased as the pH increased. Anthocyanins had different structures at different

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pH levels. Under acidic conditions, anthocyanins showed a relatively stable red flavylium cation, which changed to colorless chalcone as the pH and temperature increased. At pH 6, alcoholic bases formed, an intermediate of paradox salt and yellow chalcone, which are unstable structures that can easily be destroyed.

3.4 EFFECTS OF DIFFERENT SOLVENT TREATMENTS ON THE THERMAL DEGRADATION OF SWEET POTATO ANTHOCYANINS 3.4.1 Thermal Degradation Experiment Samples (0.5 g) of sweet potato anthocyanins were weighed, dissolved using a pH 3 solution of deionized water, 10% ethanol, 20% ethanol, and 50% ethanol, then independently placed at 4, 25, 75, and 100 C for 30 min. Then, a certain amount of sample was cooled in ice water for the determination of degradation kinetics.

3.4.2 Thermal Degradation Model Many food components are affected by various factors during the storage process. The degradation kinetics of these components is always fitted by zero-order or first-order reaction kinetic models. Many references showed that the degradation kinetics of anthocyanins from black currant juice (Harbourne et al., 2008), purple corn (Yang et al., 2008a,b), raspberry (Verbeyst et al., 2011), purple potato (Nayak et al., 2011), and bilberry flower (Moldovan et al., 2012) fitted a first-order reaction kinetics model. This article assumed that sweet potato anthocyanin also followed first-order kinetics at different temperatures under storage solvent treatment. Thus: 2d ½C  5 k ½C m ; dt

(6.9)

where k represents the rate constant, m represents the reaction order, C represents the concentration of total anthocyanins, and t represents the reaction time.   ½Ct  Log 5 2k; (6.10) ½C0  where C0 represents the concentration of total anthocyanins before treatment, and Ct represents the concentration of total anthocyanins

Sweet Potato Anthocyanins

317

after treatment. The half-lives (t1/2) of the anthocyanins were calculated as follows: t1=2 5 2ln 0:5 3 k21 :

(6.11)

According to the activated complex theory for chemical reaction rates, for first-order reactions, the Arrhenius equation relates the reaction rate constants to the absolute temperature, as seen below:   2Ea k 5 A0 exp ; (6.12) RT where Ea represents the activation energy (kJ/mol), A0 represents a preexponential factor/frequency factor (1/s), T represents the absolute temperature (K), and R represents the gas constant (8.314 J/mol K). The reaction rate constant k and the activation energy, Ea were determined graphically from a plot of ln(C/C0) versus time and ln k versus 1/T, respectively. The thermal death time method (D
Z model) was used to estimate the decimal reduction time (D value), which is the heating time required to reduce the anthocyanin concentration by 90%, and the z value, which is the temperature change necessary to alter the thermal death time by one log cycle, with the following relationships: D 5 ln 

D Log Dref



10 k

(6.13)

ðT 2 Tref Þ ; Z

(6.14)

52

where Dref represents the D value at temperature Tref. The enthalpy of activation (ΔH) and entropy of activation (ΔS) were estimated using the Eyring
Polanyi model based on the transition state theory: k 5 kb 3

h 3 e2ΔH2TΔS=RT ; T

(6.15)

where T represents the absolute temperature (K), kb represents the Boltzmann constant (1.381 3 10223 J/K), h represents the Planck constant (6.626 3 10234 Js), and R represents the gas constant (8.314 J/mol K).

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3.4.3 Degradation Kinetic Parameters of Anthocyanins in Deionized Water The kinetic equation of log(Ct/C0) based on the anthocyanin changes over time in deionized water under different temperature conditions was used to construct Fig. 6.15. In deionized water, the logarithm of the concentration of anthocyanins was plotted against time, resulting in a straight line, which indicated pseudo first-order reaction kinetics for anthocyanin degradation. The fit coefficients of 75 and 100 C were higher than those of 4 and 25 C, suggesting that anthocyanin degradation under high temperature was better fitted to first-order reaction kinetics than degradation under low temperature. According to the kinetic model, we can determine a reaction rate constant (K), and thus can estimate half-life (t1/2), and D and Z values. According to Arrhenius, k 5 A0exp(2Ea/RT); therefore, we can calculate the activation energy (Ea) and can estimate the value of ΔH and ΔS using the Eyring
Polanyi model. The results are shown in Table 6.10.

0

Time (h) 6

3

9

12

0

log (c/c0)

–0.1

4°C

–0.2

25°C

–0.3

75°C 100°C

–0.4 –0.5 Figure 6.15 Relationship between the concentration of anthocyanins and time in deionized water.

Table 6.10 Degradation Kinetic Parameters of Anthocyanins in Deionized Water Temp

K (h21)

R2

t1/2 (h)

D (h)

Z ( C)

Ea (kJ/mol)



( C) 4

0.0009

0.9666

770

2558.43

25

0.003

0.9867

231

767.53

75

0.0316

0.9945

21.93

72.87

100

0.0442

0.9948

15.68

52.09

56.77

83.46

ΔH

ΔS

(kJ/mol/K)

(J/mol/K)

77.33

180.81

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319

The degradation rate constant k increased as the heating temperature increased, whereas the half-life sharply decreased as the heating temperature increased (Table 6.10). For example, the reaction rate constant at 75 C was 0.0316 h21, and the half-life was 21.93 h, whereas the degradation rate constant at 100 C increased to 0.0442 h21 and the half-life decreased to 15.68 h, indicating that the degradation of anthocyanins increased as the temperature increased and that the D value trend was similar to that of the half-life. Kirca et al. (2007) reported that the thermal stabilities of black carrot anthocyanins were stronger than those of sour cherries and red
oranges, possibly because black carrot anthocyanins presents many acylated anthocyanins. The half-life of black carrot anthocyanins at 70
90 C was 25.1
6.3 h at pH 4.0, and the stabilities of sweet potato anthocyanins were higher than black carrot anthocyanins, which could be attributed to the pH of the present study being 3.0. Sweet potato anthocyanins are more stable conducive anthocyanins, possibly because the acylated levels of sweet potato anthocyanins is higher than black carrot anthocyanins. The relationship of the degradation Ea and temperature was fitted to the Arrhenius equation (R2 5 0.9873), and the Ea is 83.46 kJ/mol. Reyes and Cisneros (2007) reported that the thermal degradation Ea values of purple potatoes, red potatoes, carrots, and purple grapes at pH 3 were 72.49, 66.7, 75.03, and 81.34 kJ/mol, respectively. In addition, the results for sweet potato anthocyanins were similar to those reported above. The reaction Ea of sweet potato anthocyanins was slightly higher than in the study by Reyes and Cisneros (2007), probably because the acylated level of sweet potato anthocyanins was higher than potato anthocyanins. Cisse et al. (2009) estimated ΔH and ΔS according to the Eyring
Polanyi model, and the ΔH and ΔS values of orange and blueberry roselle anthocyanins were 34.24
63.11 and 149
233 kJ/ mol/K, respectively. Thus, the degradation ΔH and ΔS of sweet potato anthocyanins in deionized water were 33.58 and 180.81 kJ/mol/K, respectively, which was within the scope of the previous report.

3.4.4 Degradation Kinetic Parameters of Anthocyanins in 10% Ethanol The thermal degradation of sweet potato anthocyanins in 10% ethanol solvent was similar to in deionized water, which were both fitted to first-order reaction kinetics, and the coefficients at 75 and 100 C were

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0

3

Time (h) 6

9

12

0 –0.1 log (c/c0)

4°C 25°C

–0.2

75°C

–0.3

100°C

–0.4 Figure 6.16 Relationship between the anthocyanin concentration in 10% ethanol and time.

Table 6.11 Degradation Kinetic Parameters of Anthocyanins in 10% Ethanol Temp ( C)

K (h21)

R2

t1/2 (h)

D (h)

4

0.001

0.9497

693

2302.59

25

0.0027

0.955

256.66

852.79

75

0.0244

0.9972

28.40

94.37

100

0.039

0.997

17.77

59.04

Z ( C)

60.34

Ea (kJ/mol)

78.53

ΔH

ΔS

(kJ/mol/K)

(J/mol/K)

72.41

188.37

higher than at 4 and 25 C (Fig. 6.16). The degradation kinetic parameters of anthocyanins in 10% ethanol are displayed in Table 6.11. The values of thermal degradation parameters in 10% ethanol were similar to those in deionized water. The reaction rate constants of these two stored solvents were not significantly different at 4 C. However, compared with deionized water, the reaction rate constant was decreased, and the half-life and D value were increased at 25, 75, and 100 C. This may be because the anthocyanins were easier dissolved in 10% ethanol than in water, indicating that the binding between ethanol and anthocyanins is strong. The relationship of anthocyanin thermal degradation Ea, and the temperature was fitted to the Arrhenius equation (R2 5 0.9937), the ΔH and ΔS values corresponded to the Eyring
Polanyi model (R2 5 0.9923), and the Ea, ΔH and ΔS values of anthocyanins in 10% ethanol were 78.53 kJ/mol, 72.41 kJ/mol/K, and 188.37 kJ/mol/K, respectively. Compared with the deionized water parameters, the Ea and ΔH were slight decreased, whereas the ΔS value was increased, indicating that the energy, which is necessary to convert nonactivated molecules into activated molecules, was reduced,

Sweet Potato Anthocyanins

321

and the degree of confusion was increased, possibly because the boiling point of ethanol is lower than that of water. The elevated temperatures increase the movements of molecules, leading to changes in the thermal degradation reaction constants.

3.4.5 Degradation Kinetic Parameters of Anthocyanins in 20% Ethanol The thermal degradation of sweet potato anthocyanins in 20% ethanol solvent was fitted to the first-order reaction kinetics, and the coefficients at 75 and 100 C were higher than at 4 and 25 C (Fig. 6.17). The degradation kinetic parameters of anthocyanins in 20% ethanol are displayed in Table 6.12. The regulation of thermal degradation in 20% ethanol storage solvent was similar to that in 10% ethanol and deionized water, and the reaction rate constants for the three storage solvents showed no significant differences at 4 and 25 C, but did at 75 and 100 C. The reaction rate constant in 20% ethanol was reduced, and the half-life and D value were increased compared with those in deionized water and 10% 0

3

Time (h) 6

9

12

0 4°C

log (c/c0)

–0.1

25°C

–0.2

75°C 100°C

–0.3 –0.4 Figure 6.17 Relationship between the anthocyanin concentration in 20% ethanol and time.

Table 6.12 Degradation Kinetic Parameters of Anthocyanins in 20% Ethanol Temp ( C)

K (h21)

R2

t1/2 (h)

D (h)

4

0.0008

0.9437

866.25

2878.23

25

0.0026

0.9644

266.54

885.61

75

0.0257

0.9973

26.97

89.59

100

0.029

0.9991

23.90

79.40

Z ( C)

61.56

Ea (kJ/mol)

78.09

ΔH

ΔS

(kJ/mol/K)

(J/mol/K)

71.97

189.94

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ethanol. The relationship of anthocyanin thermal degradation Ea and the temperature in 20% ethanol was fitted to the Arrhenius equation (R2 5 0.9749), and the ΔH and ΔS values corresponded to those of the Eyring
Polanyi model (R2 5 0.9698). The Ea, ΔH, and ΔS values of anthocyanins in 20% ethanol were 78.09 kJ/mol, 71.97 kJ/mol/K, and 189.94 J/mol/K, respectively.

3.4.6 Degradation Kinetic Parameters of Anthocyanins in 50% Ethanol The thermal degradation of sweet potato anthocyanins in 50% ethanol solvent was fitted to the first-order reaction kinetics, and the coefficients at 75 and 100 C were also higher than those at 4 and 25 C (Fig. 6.18). The degradation kinetic parameters of anthocyanins in 20% ethanol are displayed in Table 6.13. The regulation of thermal degradation in 50% ethanol storage solvent was similar to that in deionized water, 10% ethanol, and 20% ethanol, and the reaction rate constants of four storage solvents were not significantly different at 4 and 25 C. The reaction rate constant of

0

0

2

4

Time (h) 6

8

10

12

log˄c/c0˅

–0.1 4°C

–0.2

25°C 75°C

–0.3

100°C

–0.4 –0.5 Figure 6.18 Relationship between the anthocyanin concentrations in 50% ethanol and time.

Table 6.13 Degradation Kinetic Parameters of Anthocyanins in 50% Ethanol Temp ( C)

K (h21)

R2

t1/2 (h)

D (h)

4

0.009

0.9503

770

2558.43

25

0.0028

0.9774

247.5

822.35

75

0.0197

0.9872

35.18

116.88

100

0.0333

0.9941

20.81

69.15

Z ( C)

61.22

Ea (kJ/mol)

75.33

ΔH

ΔS

(kJ/mol/K)

(J/mol/K)

69.21

193.57

Sweet Potato Anthocyanins

323

50% ethanol was reduced at 75 C, while the reaction rate constant, half-life, and D value in 50% ethanol were decreased compared with those in 20% ethanol. The relationship of anthocyanin thermal degradation Ea and the temperature in 50% ethanol was fitted to the Arrhenius equation (R2 5 0.9958), and the ΔH and ΔS corresponded to those of the Eyring
Polanyi model (R2 5 0.9946). The Ea, ΔH and ΔS of anthocyanins in 20% ethanol were 75.33 kJ/mol, 69.21 kJ/mol/K, and 193.57 J/mol/K, respectively. The Ea and ΔH were reduced and ΔS was increased compared with in the other solvents, which may be because the association of anthocyanins is reduced in 50% ethanol. Tseng et al. (2006) found that the reaction Ea values of mallow pigment in 10%, 30%, and 50% ethanol were 24.45, 24.35, and 22.75 kcal/mol, respectively, and the reaction Ea decreased as the ethanol concentration increased, which was similar to our findings. In conclusion, sweet potato anthocyanins had good thermal stability below 60 C, at which the preservation rate was still 100%. Even after heating for 4 h at 100 C, the anthocyanin preservation rate was still greater than 80%. The anthocyanins of sweet potato had good pressure resistances, and the preservation rate remained unchanged after treating for 15 min at 500 MPa. However, the preservation rate fell to 61% after pressure treatment and then heating for 2 h. Anthocyanins from sweet potato were stable under strong acidic conditions (pH # 3), and color change was not obvious. However, anthocyanins showed unstable behaviors under alkaline conditions. The reaction rate constant increased and the half-life decreased as the temperature increased, and the thermal degradation laws were in line with first-order kinetics, whether the solvent was deionized water, or 10%, 20%, or 50% ethanol, and the fit coefficients at 75 and 100 C were larger than those at 4 and 25 C, indicating anthocyanin degradation under high temperature conditions was better fit to the first-order reaction kinetics than degradation under low temperature. In addition, the results of Arrhenius equation and the Eyring
Polanyi model showed that Ea and ΔH decreased, while ΔS increased, as the ethanol concentration increased. These results provided a theoretical basis for the application of anthocyanins in the food, pharmaceutical, and cosmetic industries.

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SECTION 4: BIOLOGICAL ACTIVITY OF SWEET POTATO ANTHOCYANIN 4.1 THE EFFECTS OF SWEET POTATO ANTHOCYANIN ON ACUTE ALCOHOLIC LIVER DAMAGE AND DEALCOHOLIC EFFECTS Alcohol is a common unwanted liver substances, and long-term drinking will cause different degrees of damage to the digestive, circulatory, urinary, and blood systems. Alcoholic liver damage (ALD) caused by excessive alcohol consumption is a serious threat to human health. Alcoholic liver disease includes alcoholic hepatitis, alcohol hepatic fibrosis, alcoholic fatty liver, alcoholic liver cirrhosis (ALC), and slight ALD (Tuma et al., 2004). In the United States of America, ALC is one of the top seven higher mortality-rate diseases, accounting for more than 80% of liver cirrhosis (Galligan et al., 2012). In China, the morbidity of alcoholic liver disease is also increasing yearly, and has become the second largest cause of liver damage after the hepatitis virus (Zhuang et al., 2003). Alcohol consumption results in an imbalance in the body’s antioxidant system, which causes the formation of oxidative stress. Continuous oxidative stress will cause liver steatosis, resulting in hepatitis, hepatic fibrosis, liver cirrhosis, and liver cancer (Dey et al., 2006; Tulassay et al., 2008). In recent years, purple sweet potato has received attention because of its special color, high-nutritional value, and vital influences on human health. The anthocyanins in purple sweet potato are more stable than those in strawberry, perilla, and red cabbage (He et al., 2010). PSPAs can scavenge oxygen free radicals, reduce the lipid peroxide in blood and liver, and prevent liver damage induced by DMA, T-BHP, and APAP (Hwang et al., 2011a,b; Choi et al., 2009; Choi and Hwang, 2010). However, the pathological process of liver damage induced by chemicals is not the same as that induced by alcohol consumption, and thus could not simulate the process of ALD. In addition, there are still no reports on the preventive effect of PSPAs on acute and subacute ALD. Therefore, we carried out loss of righting reflex and recovery experiments on mice suffering alcohol-induced intoxication to investigate the potential dealcoholic effects of PSPAs.

4.1.1 Experimental Model for Investigating Acute ALD and Dealcoholic Effects The drunk prevention experiment. For the loss of righting reflex experiment on alcoholic intoxication, C57BL/6 mice were separated into

Sweet Potato Anthocyanins

325

four groups, control group (same volume of saline), low-dose group (50 mg PSPAs/kg body weight), middle-dose group (125 mg PSPAs/kg body weight), and high-dose group (375 mg PSPAs/kg body weight), with 10 animals in each group. All of the groups were fasted for 12 h, and then PSPAs in saline were administered intragastrically. After 30 min, the mice were treated with 50% ethanol solution (18 mL/kg body weight). The duration of the ethanol-induced loss of righting reflex was recorded. The judgment standards of loss of righting reflex were as follows: The ethanol treated mice were placed on their backs, and the righting reflex was judged as being lost if the above pose was maintained more than 30 s. For the righting reflex recovery experiment, the grouping method was same as that used in the loss of righting reflex experiment. All of the groups were fasted for 12 h and were then treated with 50% ethanol solution (18 mL/kg body weight). After 30 min, all of the groups were administered one dose of PSPAs in saline intragastrically. The duration of the recovery of the righting reflex was recorded. The drunk therapy experiment. The righting reflex for each control or treatment mouse was performed in triplicate. The duration of the ethanol-induced loss of righting reflex and the duration of the recovery of the righting reflex for each mouse were recorded as the means of three experiments. Acute ALD mice models. For the experiment on PSPA prevention of acute ALD, C57BL/6 mice were divided into five groups: negative control (without anthocyanins and ethanol treatments), acute ALD mice model group (with ethanol treatment only), low-dose group (50 mg PSPAs/kg body weight), middle-dose group (125 mg PSPAs/kg body weight), and high-dose group (375 mg PSPAs/kg body weight) of 12 animals. Equal volumes of saline were administered intragastrically for the negative control and the acute ALD mice model group, while PSPAs were administered intragastrically for low, middle, and high dose groups once daily for 30 consecutive days. The food and water intakes of mice were recorded once a day. Then, 4 h after the last administration, the negative control group was fed by gavage using an equal amount of purified water, while the other groups were treated with 50% ethanol solution (12 mL/kg body weight). All of the groups were fasted for 16 h, and then anesthesia was administered by injecting 60 mg/kg body weight of pentobarbital sodium into the abdominal cavities of the mice. Blood was sampled from abdominal aortas to

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Table 6.14 Effect of PSPAs on the Durations of Ethanol-Induced Loss and Recovery of the Righting Reflex (min) Treatment Group

Duration of Loss of Righting Reflex

Duration of Recovery of Righting Reflex

Control

5.97 6 2.83

445.55 6 24.87

Low-dose PSPAs

10.22 6 3.16a

423.51 6 15.12

Middle-dose PSPAs

11.77 6 2.10a

383.76 6 13.22a

High-dose PSPAs

13.69 6 2.75b

382.73 6 18.08a

Values are means 6 standard deviations of 10 determinations; compared with control. a P , 0.05, b P , 0.01.

determine the activities of serum AST, ALT, TG, TCH, LDL-C, and lactate dehydrogenase (LDH). Livers were weighed and cut into slices, some of which were kept in buffered formalin for histological observation, and 10% of liver tissue homogenates were obtained from the remaining liver sections and stored at 275 C.

4.1.2 Effect of Sweet Potato Anthocyanins on Alcoholic Intoxication The loss of righting reflex indicates that the mice are intoxicated, while the recovery of the righting reflex indicates that the mice are no longer impaired (Kimura et al., 2013). Table 6.14 shows the effects of the PSPAs on durations of ethanol-induced loss and recovery of the righting reflex. Compared with the control, the intake of PSPAs extended the duration of ethanol-induced loss of righting reflex significantly (P , 0.05), especially for the high-dose PSPAs group (P , 0.01). In addition, in the middle and high-dose PSPAs groups, the durations until recovery of the righting reflex were both shortened significantly (P , 0.05), indicating that PSPAs may have potential curative effects on alcoholic intoxication.

4.1.3 Effect of Sweet Potato Anthocyanins on Growth and the Liver Index The liver index can partly reflect the level of ALD (Xiang et al., 2011). Table 6.15 shows the effects of PSPAs on the original body weight, final body weight, liver weight, and liver index of mice in the acute ALD experiment. No significant differences were found for body weight among different treatment groups, and no significant differences were found for the final body weight among the different treatment groups. Compared with controls, acute alcohol intake resulted in

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Table 6.15 Effects of PSPAs on the Body Weight (g), Liver Weight (g), and Liver Index (%) of Acute ALD Mice Treatment Group

Weight

Liver Weight

Liver Index

Initial Weight

Final Weight

Control

19.15 6 0.32

22.41 6 0.91

1.08 6 0.10

4.81 6 0.22

Model

18.72 6 0.37

21.39 6 0.78

1.27 6 0.12b

5.92 6 0.17b

Low-dose PSPAs

19.00 6 0.56

22.11 6 1.12

1.23 6 0.17b

5.55 6 0.14b

Middle-dose PSPAs

18.70 6 0.43

22.35 6 1.04

1.23 6 0.08

b

5.50 6 0.19a,c

High-dose PSPAs

18.88 6 0.46

22.17 6 1.06

1.12 6 0.07

a

5.47 6 0.12a,c

Values are mean 6 standard deviation of 10 determinations; compared with the control, aP , 0.05, bP , 0.01; compared with the model, cP , 0.05.

a significant increase in the liver weight and liver index (P , 0.01), and the liver indices in low, middle, and high-dose groups were reduced by PSPA additions. Our results are in agreement with those reported by McKim et al. (2002), who found that antioxidants, such as cocoa flavonoids, could reduce acute ALD.

4.1.4 Effects of Sweet Potato Anthocyanins on Serum ALT, AST, and LDH Activities Change in serum biochemical indices are important symbols of liver damage. Serum transaminases, such as ALT and AST, are essential catalysts in human metabolic processes. An increase in ALT is a mark of liver cell membrane damage, and an increase in AST is a mark of liver cell mitochondrial damage. When damage, caused by inflammation and necrosis, occur in liver cells, ALT and AST are released into blood, causing increases in serum transaminases (Rajesh et al., 2004). The effects of PSPAs on the activity levels of serum ALT and AST in acute ALD mice are shown in Fig. 6.19. Compared with in the control, the activity levels of ALT and AST in the model group increased significantly (P , 0.01), indicating that acute alcoholism caused damage to the liver cell membrane and mitochondria. Compared with the model group, the activity level of ALT in low-dose, middle-dose, and high-dose groups of PSPAs decreased by 47.28%, 35.15%, and 45.04%, respectively, and the activity of AST decreased by 17.67%, 28.39%, and 30.21%, respectively, indicating that PSPAs had preventative effects on acute ALD. Our result was similar to the effect of genistein on ALD (Huang et al., 2013).

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Activities of ALT and AST (U/L)

180

**

Control Model Low dose group Middle dose group High dose group

150

## ## ##

120

90 ** 60 ## ## ## 30

0

ALT

AST

Figure 6.19 Effects of PSPAs on the activity levels of serum ALT and AST. Compared with control,  P , 0.05,  P , 0.01; Compared with the model, #P , 0.05, ##P , 0.01.

**

Activity of LDH (U/L)

500

**

*

400 300 200 100 0 Control

Model

Low dose Middle dose High dose group group group

Figure 6.20 Effects of PSPAs on the activity level of serum LDH. Compared with the control,  P , 0.05,  P , 0.01; Compared with the model, #P , 0.05, ##P , 0.01.

In addition, under normal circumstances, the LDH contained in living cells could not penetrate the cell membrane. When cells are affected by outside poisonous substances, cell membranes are damaged, and the permeability of cell membrane is changed. Thus, the intracellular LDH is released into the extracellular fluid, resulting in an increase in LDH activity (Mathew et al., 1985). The effects of PSPAs on the activity level of serum LDH in acute ALD mice is shown in Fig. 6.20. Compared with the control, the LDH level of the model group showed a significant increase (P , 0.01). The intake of

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PSPAs in advance could inhibit the increase in the LDH, especially in the high-dose PSPAs group. There was no significant difference between the LDH level of the high-dose PSPAs group and that of the control (P . 0.05).

4.1.5 Effects of Sweet Potato Anthocyanins on Serum TG, TCH, and LDL-C Activity Levels At the primary phase of alcoholic liver disease, large quantities of TG and TCH will accumulate, representing their roles as sensitive indicators of lipometabolism. LDL-C is endogenous TG, and the excessive intake of alcohol will cause an increase in the synthesis of LDL-C. The effects of PSPAs on the serum TCH, TG, and LDL-C levels in the acute ALD mice are shown in Figs. 6.21 and 6.22. Compared with the control, the levels of TCH and TG in the model group increased significantly, may be because the metabolism of alcohol will mostly result in the accumulation of TG and TCH. Compared with the model group, the levels of serum TCH and TG in low-dose, middle-dose, and high-dose anthocyanin groups decreased significantly (P , 0.05). Our results were in accordance with those reported by Zhang et al. (2011), who found that ostruthin could inhibit the increases in TCH, TG, and LDL-C induced by alcohol, and alleviate 3 Control

*

Model

Levels of TCH and TG (mmol/L)

2.5 *#

*# Low dose group

**# 2

Middle dose group *

1.5

High dose group #

#

#

1

0.5

0 TCH

TG

Figure 6.21 Effects of the PSPAs on the levels of serum TCH and TG. Compared with the control,  P , 0.05,  P , 0.01; Compared with the model, #P , 0.05, ##P , 0.01.

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0.3

Level of LDL-C(mmol/L)

0.25

**

**# *##

0.2

##

0.15 0.1 0.05 0

Control

Model

Low dose group

Middle dose group

High dose group

Figure 6.22 Effects of PSPAs on the serum LDL-C level. Compared with the control,  P , 0.05, Compared with the model, #P , 0.05, ##P , 0.01.



P , 0.01;

acute alcohol intoxication and hepatic lipid metabolism, possibly because anthocyanins and ostruthin are flavonoids, which possess strong antioxidant activities.

4.1.6 Effects of Sweet Potato Anthocyanins on the Hepatic MDA Content After the intake of a large quantity of alcohol in a brief period, it is metabolized in the liver, producing some aldehyde compounds that can combine with proteins, releasing a large amount of ROS, and increase the internal oxidative stress level and lipid peroxidation. MDA is the main product of lipid peroxidation induced by ROS, and the MDA content reflects the oxidative stress level of the liver (Wu et al., 2009). The effects of PSPAs on the hepatic MDA level of acute ALD mice are shown in Fig. 6.23. Compared with the control, MDA in the model group increased significantly (P , 0.05), indicating that alcohol intake increased the oxidative stress level in the liver. Compared with in the model group, MDA levels in the low-, middle- and high-dose PSPAs groups decreased significantly (P , 0.05). This may be because the PSPAs possess strong antioxidant activities. Our results were similar to those of Bharrhan et al. (2011), who found that catechin could decrease MDA in alcohol-induced damaged rat livers.

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Levlel of hepatic MDA(mg/g liver)

30 * 25

20

# ##

##

15

10

Control

Model

Low dose roup

Middle dose roup

High dose roup

Figure 6.23 Effects of PSPAs on the activity level of hepatic MDA. Compared with the control,  P , 0.05,  P , 0.01; Compared with the model, #P , 0.05, ##P , 0.01.

4.1.7 Effects of Sweet Potato Anthocyanins on Hepatic Superoxide Dismutase (SOD) and Glutathione S-transferases (GST) Activity Levels Recently, more attention is being paid to the effects of oxidative stress and lipid peroxidation on ALD. Alcohol can produce a large amount of radicals, such as O22, H2O2, and OH2, during its metabolism in the liver. When the radicals go cannot be fully eliminated by the human body, liver cells are damaged (Huang et al., 2009). GST and SOD are important internal antioxidases that can alleviate oxidative stress caused by alcohol. The SOD activity in the model group was lower than that in the control, indicating that the ability of mice to eliminate O22 was weakened by alcohol intake. Compared with the model, the SOD activity in the high-dose PSPAs group increased significantly, and was even higher than the normal level. Thus, the ability to eliminate O22 was also improved, and the damaging effects were alleviated (Fig. 6.24). The intake of alcohol increases the internal oxidation products, and GST can eliminate oxygen-free radicals effectively, prevent oxidative stress, and inhibit lipid peroxidation. Acetaldehyde is an alcohol metabolite that can combine with GST, resulting in changes in membrane permeability and fluidity, and thus, the structure and function of the cells being damaged. Moreover, when the structures and functions of cells are damaged, free radicals are not eliminated, resulting in aggravated liver damage. At present, the decrease in the GST content is a mechanism of ALD.

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Activity of hepatic SOD (U mg/g protein

2 **## 1.5

**##

* *#

1

0.5

0 Control

Model

Low dose group

Middle dose group

High dose group

Figure 6.24 Effects of PSPAs on the activity level of hepatic SOD. Compared with the control,  P , 0.01; Compared with the model, # P , 0.05, ## P , 0.01.



P , 0.05,

Compared with the control, the GST level in the model group decreased significantly, indicating that the liver cell membrane was damaged. The GST contents in the anthocyanins groups were significantly increased in a dose-dependent manner, indicating that PSPAs could alleviate alcohol-induced oxidative stress, thus alleviating alcohol-induced liver damage (Fig. 6.25). Our results are in agreement with those reported by Tang et al. (2012), who found that taurine could decrease alcohol-induced dyslipidemia and oxidative damage.

4.1.8 Effects of Sweet Potato Anthocyanins on the Hepatic ADH Activity Level ADH is a metalloenzyme, which exists in the cytoplasm and mitochondria of liver cells, that can transform ethanol into acetaldehyde, and then oxidize acetaldehyde into acetic acid. In mitochondria, some acetic acid can enter into the tricarboxylic acid cycle and be oxidized into CO2 and H2O, which are removed from the body (Oyama et al., 2005). The effects of PSPAs on the hepatic ADH level of the acute ALD mice is shown in Fig. 6.26. Compared with the control, the ADH level of the model group decreased significantly (P , 0.01), indicating that a large amount of alcohol could not be metabolized into

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200 **## 160

*#

* 120

Activity of hepatic GST

U mg/g protein

**##

80

40

0 Control

Model

Low dose group

Middle dose group

High dose group

Figure 6.25 Effects of PSPAs on the activity of hepatic GST. Compared with the control,  P , 0.05, Compared with the model, #P , 0.05, ##P , 0.01.



P , 0.01;

Activity of hepatic ADH (U mg/g protein)

50

40

30

**# **# **

20

**

10

0

Control

Model

Low dose group

Middle dose group

High dose group

Figure 6.26 Effects of PSPAs on the activity level of hepatic ADH. Compared with the control,  P , 0.05,  P , 0.01; Compared with the model, #P , 0.05, ##P , 0.01.

CO2 and H2O, causing hepatotoxicity. Compared with the model group, the ADH activity levels in the low-dose, middle-dose, and highdose PSPAs groups increased significantly. However, the ADH activity in the high-dose PSPAs group was still lower than that of the control.

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Table 6.16 Effects of PSPAs on the Histopathology of Acute ALD Mice Treatment Group

Parallel 1

Parallel 2

Parallel 3

Control

2

2

2

Model

Hepatocyte edema11 1

Hepatocyte edema 11

Hepatocyte edema 11 1

Low-dose PSPAs

Hepatocyte edema 11

Hepatocyte edema 1

Hepatocyte edema 11

Middle-dose PSPAs

Hepatocyte edema 1

2

2

High-dose PSPAs

2

2

2

“ 2 ” indicates no lesions; “ 1 ” minor lesions; “ 11 ” mild lesions; “ 11 1 ” moderate lesions; “ 11 1 1 ” severe lesions.

4.1.9 Effects of Sweet Potato Anthocyanins on the Histopathological Analysis The liver cells of C57BL/6J mice in the control group were of similar sizes, in a tight arrangement, with complete structures. They underwent uniform staining and showed no pathological liver changes. In the model group, the mice liver cells showed pathological changes, such as a loose arrangement and cellular edema. In the anthocyanin low-dose group, the pathological changes of the mice liver cells were similar to those in the model group, including a disordered arrangement of cells and cellular edema, but the degrees were reduced. In the anthocyanin middle-dose and high-dose groups, no pathological changes were observed, and the cellular morphology was close to that of the control group (Table 6.16). In conclusion, the study indicated that sweet potato anthocyanins had some effects on C57BL/6J mice as seen using acute ALD and the dealcoholic model. The results showed that sweet potato anthocyanins reduced the ALT, AST, TCH, TG, and LDL-C contents and liver lipid peroxidation, while increasing the activities of antioxidant enzymes in the liver. In short, sweet potato anthocyanins can reduce oxidative stress and the lipid peroxidation level, which may have potential inhibitory effects on acute alcoholic liver injury.

4.2 THE EFFECTS OF SWEET POTATO ANTHOCYANIN ON SUBACUTE ALCOHOLIC LIVER DAMAGE Normal cell metabolism generates ROS. As a secondary messenger, ROS regulates several different normal physiological functions in organism and participates in a variety of cellular redox-regulatory mechanisms to defend against oxidative stress and maintain redox

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balance. However, only at low or medium levels will the beneficial effects of ROS occur, and high levels of ROS and/or an inadequate antioxidant defense will cause oxidative stress, inducing damage to cellular structures, including phospholipid bilayers, proteins, and DNA (Valko et al., 2007). The cellular antioxidant system consists of antioxidases, such as SOD and GST, and some small molecular substances, such as glutathione, uric acid, and coenzymes. In addition, natural bioactive compounds, including polyphenols, carotenoids, and ascorbic acids in vegetables, fruits, tea, and other products of plant origin, also play important roles in the prevention of oxidation-related diseases (Cuevas-Rodríguez et al., 2010). Alcohol is a common unwanted substance in the liver, and longterm drinking will cause different degrees of damage to the digestive, circulatory, and urinary systems. After absorption, 90% of alcohol will be oxidized and decomposed in the liver. Alcohol metabolism leads to the generation of free radicals and the consumption of cellular antioxidants, increasing the occurrence of alcohol-related diseases (Nanji et al., 1995). Anthocyanins, as a category of natural polyphenols, are widespread throughout vegetables and fruits. Anthocyanins can scavenge oxygenfree radicals, reduce the lipid peroxide levels in blood and liver, and prevent liver damage induced by DMA, T-BHP, and APAP (Choi et al., 2009, 2010). Here, we studied the effects of sweet potato anthocyanins on serum ALT, AST, LDH, TG, TCH, LDL-C, liver MDA, GST, SOD, and ADH, as well as other indicators of liver pathology.

4.2.1 Subacute ALD Mice Models For the experiment on the preventative effects of PSPAs on subacute ALD, the subject grouping method was same as that used in the experiment on acute ALD. The same volumes of saline were administered intragastrically for the negative control and the acute ALD model group, while the appropriate level of PSPAs were administered intragastrically in low-dose, middle-dose, and high-dose groups once daily for 30 consecutive days. Then, 30% of ethanol solution was fed by gavage at 10 mL/kg body weight once a day to the model and anthocyanins groups for 14 days. More than 4 h occurred between the administration of saline or anthocyanins and the administration of

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ethanol. All groups were fasted for 4 h, and then the mice were anesthetized by injecting 60 mg/kg body weight of pentobarbital sodium into the abdominal cavity. Blood was sampled from the abdominal aorta to determine the activities of serum ALT, AST, LDH, TCH, TG, and LDL-C. Livers were weighed and cut into slices, some of which were kept in buffered formalin for histological observation. In addition, 10% of liver tissue homogenates were obtained from the remaining liver sections and stored at 275 C.

4.2.2 Effects of Sweet Potato Anthocyanins on Growth and Liver Index Table 6.17 shows the effects of PSPAs on original body weight, final body weight, liver weight, and the liver index of mice in the subacute ALD experiment. Compared with the control, the average body weight of mice in the model group decreased significantly (P , 0.05). This may be because alcohol metabolism changed the digestive functions of the stomach and intestine, resulting in changes in food consumption. The liver index can partly reflect the severity of ALD. Compared with the control, subacute alcohol intake resulted in significant increases in liver weights and the liver index (P , 0.01). Compared with the model, the liver weights in the middle-dose and high-dose PSPAs groups decreased significantly, and the liver indices of the middle- and highdose PSPAs groups decreased significantly (P , 0.01). In particular, no significant difference was found between the liver index of the highdose PSPAs group and that of the control.

Table 6.17 Effects of Anthocyanins on the Weight (g) and Liver Index (%) of Subacute ALD Mice Treatment Group

Weight

Liver Weight

Liver Index

Initial Weight

Final Weight

Control

19.05 6 0.32

22.31 6 0.91

1.08 6 0.10

4.81 6 0.22

Model

18.82 6 0.37

20.82 6 0.78a

1.16 6 0.09b

5.57 6 0.33b

Low-dose PSPAs

19.00 6 0.56

22.01 6 1.12

1.15 6 0.06b

5.23 6 0.12b

Middle-dose PSPAs

18.95 6 0.43

22.46 6 1.04

1.13 6 0.17

5.02 6 0.68c

High-dose PSPAs

18.88 6 0.46

22.43 6 1.06

1.11 6 0.14a,c

a,c

4.95 6 0.42c

Values are means 6 standard deviations of 10 determinations; compared with the control, P , 0.05, bP , 0.01; compared with the model, c P , 0.01. a

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4.2.3 Effects of Sweet Potato Anthocyanins on Serum ALT, AST, and LDH Activity Levels The changes in the serum biochemical indices are important indicators of liver damage. When damage, such as inflammation and necrosis, occur in liver cells, ALT and AST are released into the blood (Je et al., 2013). Compared with the control, the activity of AST in the model group increased, but not significantly, indicating that the intake of alcohol had caused slight damage to liver cell membranes and mitochondria. Compared with the model group, the activity levels of ALT and AST in low-dose, middle-dose, and high-dose groups of PSPAs decreased significantly, indicating that PSPAs had a preventive effect on subacute ALD (Fig. 6.27). The effects of PSPAs on activity of serum LDH in subacute ALD mice is shown in Fig. 6.28. Compared with the control, the LDH level of the model group increased significantly (P , 0.01), indicating that the intake of alcohol caused damage to the cell membrane. The intake of PSPAs in advance could inhibit the LDH activity, especially in middle-dose and high-dose groups, as there was no significant differences between their LDH activities and that of the control (P . 0.05). Our results were in agreement with those reported by Tang et al. (2012), who studied the effect of quercetin on ALD and found that quercetin could alleviate the release of liver-specific aminotransferases induced by ethanol. Anthocyanins and quercetin are both flavonoids,

Activity levels of ALT and AST U/L

180 150 120

Control Model Low-dose group Middle-dose group High-dose group

** ##

## ##

90 60

*

# #

#

30 0

ALT

AST

Figure 6.27 Effect of PSPAs on the activity levels of ALT and AST. Compared with the control,  P , 0.05,  P , 0.01; Compared with the model, #P , 0.05, ##P , 0.01.

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Activity of LDH U/L˅

500 ** **#

400

*##

##

Middle-dose group

High-dose group

300 200 100 0 Control

Model

Low-dose group

Figure 6.28 Effects of PSPAs on the activity level of LDH. Compared with the control,  P , 0.05, Compared with the model, #P , 0.05, ##P , 0.01.



P , 0.01;

which possess strong antioxidant activity, and one mechanism of alleviating ALD may be inhibiting the increase in oxidative stress.

4.2.4 Effects of Sweet Potato Anthocyanins on Serum TG, and LDL-C Levels and TCH Activity The long-term intake of alcohol increase the NADH/NAD 1 ratio, upregulate lipogenesis, and downregulate the transport capacity of fatty acids, causing hepatitis (Sozio et al., 2008). Therefore, the chronic intake of alcohol is often accompanied by a blood lipid increase, but antioxidants can reduce this trend. Taurine (Fang et al., 2011) and fermented barley (Puspo et al., 2010) significantly reduce the extent of the blood lipid increase induced by ALD because of their antioxidant activities. Our results were similar to these reported results. The subacute intake of alcohol increased the levels of serum TCH, TG (Fig. 6.29) and LDL-C (Fig. 6.30), while the addition of anthocyanins decreased their contents.

4.2.5 Effects of Sweet Potato Anthocyanins on the Hepatic MDA Activity Level Alcohol metabolism may lead to an increase in active oxygen radicals, these free radicals interact with the lipid membrane rapidly, causing a series of lipid peroxidation reactions (Rong et al., 2012). MDA is an important product of lipid peroxidation and can reflect the level of liver injury. The effects of PSPAs on the hepatic MDA level in subacute ALD mice are shown in Fig. 6.31. Compared with the control, the MDA level in the model group increased significantly. When pretreated with PSPAs, the lipid peroxidation in mice was inhibited

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3.5 Control

* *#

Levels of TCH and TG

mmol/L

3

*#

Model **#

Low-dose group

2.5

Middle-dose group High-dose group

2

* 1.5

# #

1

#

0.5 0 TCH

TG

Figure 6.29 Effects of PSPAs on the levels of TCH and TG. Compared with the control,  P , 0.05, Compared with the model, #P , 0.05, ##P , 0.01.



P , 0.01;

0.3 ** Level of serum LDL-C (mmol/L)

0.25 0.2 **#

0.15

*## ##

0.1 0.05 0

Control

Model

Low-dose group

Middle-dose group

Figure 6.30 Effects of PSPAs on the LDL-C level. Compared with control,  P , 0.05, with the model, #P , 0.05, ##P , 0.01.

High-dose group 

P , 0.01; Compared

significantly, and thus, MDA decreased significantly (P , 0.01). In particular, the MDA level in the high-dose PSPAs group was lower than that in the control, which could be attributed, at least in part, to the capacity to eliminate the active oxygen radicals of anthocyanins. Our results are in agreement with the findings of You et al. (2010) and López et al. (2011), who found that the antioxidants in taraxacum and amaranth had some preventive effects on ALD.

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Levels of hepatic MDA (mg/g liver)

30

24 * 18 #

##

##

12

6

0 Control

Model

Low-dose Middle-dose High-dose group group group

Figure 6.31 Effects of PSPAs on the MDA activity level. Compared with the control,  P , 0.05, Compared with the model, #P , 0.05, ##P , 0.01.



P , 0.01;

4.2.6 Effects of Sweet Potato Anthocyanins on Hepatic GST and SOD Activity Levels Recently, more attention has been paid to the effects of oxidative stress and lipid peroxidation. Alcohol can produce a large amount of radicals during its metabolism in the liver. When the radicals cannot be fully eliminated by the human body, liver cells are damaged (Albano, 2008). GST and SOD are important internal antioxidases that can alleviate the oxidative stress caused by alcohol. The intake of antioxidants, like vitamins E and C, in advance can alleviate alcohol-induced liver damage (McDonough et al., 2003). Compared with the control, the SOD and GST levels in the model group decreased. The intake of PSPAs in advance increased the SOD and GST activity levels significantly (P , 0.01) to normal levels. In addition, compared with the control, the addition of anthocyanins increased the SOD and GST levels, and the antioxidant activity in mice (Figs. 6.32, 6.33).

4.2.7 Effects of Sweet Potato Anthocyanins on the Hepatic ADH Activity Level ADH is a metalloenzyme, which exists in the cytoplasm and mitochondria of liver cells, that can transform ethanol into acetaldehyde, and then oxidize acetaldehyde into acetic acid. In mitochondria, some acetic acid can enter into the tricarboxylic acid cycle and be oxidized into CO2 and H2O, which are removed from the body (Eom et al., 2007).

Sweet Potato Anthocyanins

Hepatic GST level

U mg/g protein

250

341

**##

200

*

*#

**##

Model

Low-dose group

Middle-dose group

150 100 50 0 Control

High-dose group

Figure 6.32 Effects of PSPAs on the GST activity level. Compared with the control,  P , 0.05, Compared with the model, #P , 0.05, ##P , 0.01.

Activity of hepatic SOD (U mg/g protein)

3

**##



P , 0.01;



P , 0.01;

**##

2.4 *

1.8

*#

1.2

0.6

0 Control

Model

Low-dose group

Middle-dose group

High-dose group

Figure 6.33 Effects of PSPAs on the SOD activity level. Compared with the control,  P , 0.05, Compared with the model, #P , 0.05, ##P , 0.01.

Compared with the control, the ADH activity level in the model group decreased significantly (P , 0.01), indicating that large amount of ethanol could not be metabolized to CO2 and H2O, causing hepatotoxicity. Compared with the model group, the ADH activity level of the anthocyanin groups increased significantly (P , 0.05). However, the ADH activity level of the high-dose PSPAs group was still lower than that of the control (P , 0.01) (Fig. 6.34).

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Activity of ADH

U mg/g protein

50 40 30 20

**#

**#

Middle-dose group

High-dose group

**

** 10 0 Control

Model

Low-dose group

Figure 6.34 Effects of PSPAs on the ADH activity level. Compared with the control,  P , 0.05, Compared with the model, #P , 0.05, ##P , 0.01.



P , 0.01;

Table 6.18 Effects of PSPAs on the Histopathology of Subacute ALD Mice Treatment Group

Parallel 1

Parallel 2

Parallel 3

Control

2

2

2

Model

Hepatocyte edema 11 1

Hepatocyte edema 1111

Hepatocyte edema 11 1

Low-dose PSPAs

Hepatocyte edema 11

Hepatocyte edema 1

Hepatocyte edema 11

Middle-dose PSPAs

Hepatocyte edema 1

2

2

High-dose PSPAs

Hepatocyte edema 1

2

2

“ 2 ” indicates no lesions; “ 1 ” minor lesions; “ 11 ” mild lesions; “ 11 1 ” moderate lesions; “ 11 1 1 ” severe lesions.

4.2.8 Effects of Sweet Potato Anthocyanins on the Histopathological Analysis The effect of PSPAs on the histopathology of subacute ALD mice is shown in Table 6.18. Compared with the control, pathologic changes, such as liver cell swelling and fatty degeneration, occurred in mice livers of the model group, in which the mice were treated with alcohol continuously. After the intake of PSPAs, liver cell swelling was significantly alleviated, and the improving pathological effect was dosedependent. The liver tissue morphology after high-dose anthocyanin treatment was similar to that of normal tissue. In conclusion, the study indicated that sweet potato anthocyanins had some effects on C57BL/6J mice having subacute ALD. The results indicated that sweet potato anthocyanins could reduce

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oxidative stress and the lipid peroxidation level, which may have potential inhibitory effects on acute alcoholic liver injury by determining the content of enzymes (ALT, AST, and LDH), serum indices (TG, TCH, and LDL-C) and liver indices (MDA, ADH, SOD, and GST).

SECTION 5: APPLICATIONS OF SWEET POTATO ANTHOCYANINS Sweet potato anthocyanins are natural pigments characterized by their broad sources, multispecies, nontoxicity, and safety, which have antioxidative and hypotensive effects, as well as anticancer and other special physiological functions. Therefore, anthocyanins have important applications in the pharmaceutical, food, and cosmetics, industries.

5.1 PHARMACEUTICAL INDUSTRY Anthocyanins may function as antioxidants, improving blood circulation and reducing inflammation, as well as having anticancer and some other special pharmacological effects. Thus, the healthcare market is greatly interested in anthocyanins. Although pure anthocyanin is not sold on the market, many anthocyanin-related healthcare products exist in the form of capsules, tablets, and oral solutions. Clinically, anthocyanins have also been added to certain drugs to help patients. However, due to the high cost of drug development, anthocyanin application in medicine is restricted, but it has a bright future because of its strong physiological effects.

5.2 FOOD INDUSTRY There are two applications of anthocyanins in the food industry, one is to directly develop a health food, and the second is as a food additive. At present, people are paying increased attention to their health and a certain amount of anthocyanins taken daily could enhance the body’s resistance to damage and stress. Purple sweet potato is a highly nutritious food that could positively influence physical health. Anthocyanins can also be added to many foods and beverages, including purple sweet potato wine and milk, which are popular in the market place.

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Su et al. (2011) used purple sweet potato and fresh bovine milk as the raw materials to develop a fermented milk beverage with a low viscosity, relatively stable emulsion level, and pleasant taste. Li et al. (2011) studied the conditions to make purple sweet potato juice yogurt based on purple sweet potatoes and skim milk powder. Streptococcus thermophiles and Lactobacillus bulgaricus were mixed as fermentation agents in the yogurt. The amount of sugar added was 7%, the fermentation temperature was 41 C, and the fermentation time was 6 h. Under these conditions, the purple potato yogurt had a uniform color, good consistency, and delicate taste. Zheng et al. (2012) optimized the recipes of purple sweet potato-black sesame yogurt and purple sweet potato yogurt. The best formulae were as follows: (1) a ratio of black sesame and purple potato cubes of 1:4.56, inoculation amount of 4.12%, and a sucrose content of 7.75%, and (2) 3% purple sweet potato pulp, inoculation amount of 4%, and a sugar content of 7.5%. The amount of milk powder used was 13%. The products are characterized by slight purple colors with unique, moderately sweet and sour tastes, and are preservative-free, nutritious foods. The purple sweet potato can be used as a raw material for wine, and anthocyanins remain in the wine to form a beautiful purple color (very attractive product appearance) during the brewing process. The wine has a rice wine flavor and has a low-grade alcohol content, which is in line with the developing trend in alcohol production and consumption. Song (2009) invented a purple sweet potato wine, which has antiaging, anticancer, and antiatherosclerosis effects, and can improve immunity. Moreover, the production process was economic and highly efficient. A multifunctional healthy vinegar was developed by Shi et al. (2012), with the optimal conditions for alcohol fermentation being determined as an initial sugar content of 16%, with fermentation for 72 h at 30 C. The best conditions for acetic acid fermentation include a temperature of 32.01 C, an initial alcohol content of 7.25% vol., an inoculation amount of 9.79%, and an initial pH of 4.54. These conditions produced the greatest amount of acetic acid. Yang et al. (2012) investigated the fermentation conditions of purple potato wine using single factor experiments and response surface methodology. The gelatinized purple sweet potatoes were beaten into a slurry, liquefied for 2.5 h with 0.15% α-amylase at 65 C and pH 6.5, saccharified for 2.5 h with 0.15% glucoamylase at 60 C and pH 4.0, and fermented for 7 days with 0.0845% Angel alcohol active dry yeast at 20.72 C and pH

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3.355. These conditions produced a novel purple potato wine of B10 with a yield of 126.3%. Liu et al. (2012) developed a new composite beverage based on soybean and purple sweet potato using single factor and orthogonal experiments. The factor of the highest influence on the beverage’s sensory appeal was the ratio of purple sweet potato paste to soy milk, followed by the amount of sodium carboxymethyl cellulose, the amount of sugar, and the amount of citric acid. The best formula for this compound beverage included an 80/20 ratio of sweet potato pulp and milk, 7% sugar content, 0.15% citric acid%, and 0.2% sodium carboxymethyl cellulose. The drink had a natural color, pleasant flavor, and nutritionally functional ingredients, and has market potential. Sun et al. (2012) studied the color protection technology of potato anthocyanins in carbonated beverages. The best formulae were as follows: a 0.01/0.18 ratio of phosphoric acid to citric acid, 4 g sugar, 0.015 g each of vitamin C and sodium, and 0.015 g tannin. Using this process, when the product was stored at 20 C in the dark, the pigment retention rate was 98.2% after 25 days, the t0.9 (the time required to degrade 10%) was 144 days, and the half-life was 949 days. Purple sweet potato pigment in carbonated drinks is highly stable and, therefore, valuable. Sweet potato anthocyanins have significant free-radical scavenging, antioxidative, antimutational, and hypoglycemic effects, and can also protect the liver. If the anthocyanins were added into candy, staple foods, such as bread and pasta, pastries, such as purple sweet potato tarts and cakes, and jam and jelly, then they could provide better color, desirable taste, and functional ingredients. Yu et al. (2009) studied the use of purple sweet potato as the main raw materials in jelly. They obtained a sweet and sour tasting pale pink jelly, having no preservatives or synthetic colors, which is a healthy food. Although natural pigments are more safe and reliable than synthetic pigments, the former has some drawbacks, including susceptibility to outside temperature, pH, metal ions, light, and other factors. To maintain a stable color and good taste, it must be kept at pH 3
5. Anthocyanins have antibacterial effects, which can be used instead of synthetic preservatives, such as benzoic acid, as safe additives. The long-term intake of such synthetic preservatives is harmful to the body,

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therefore, using anthocyanins as a preservative is in line with the consumer’s food safety requirements. With the growing awareness of food safety, food coloring safety issues have received wide attention. Regulations have been introduced to limit the use of synthetic pigments in foods, such as in the United States where the kinds of synthetic pigments available were decreased from 24 to 7. China has decreased the available synthetic pigments, from the original 49 species to 17 species. The use of natural pigment substitutes for synthetic pigments is imperative. In recent years, the growth rate of the natural pigments in the international market was greater than 10%. Natural pigments on the current market are expensive, especially in Japan, the United States, Germany, and other developed countries. Therefore, an urgent need associated with pigment production is to find a high-quality, high-yield, and low-cost natural resource.

5.3 COSMETICS INDUSTRY With aesthetic awareness increasing, many people, especially women, are very focused on skin care. Human skin gradually loses its original elasticity and shine, and becomes wrinkled, with age. This is due to the body’s enzyme system being attacked by free radicals, which release collagenase and hard elastase. Moreover, under the catalysis of these two enzymes, the skin produces too much collagenase and hard elastase, leading to their crosslinking interaction and degradation, resulting in the reduction in skin elasticity. As the market related to human appearance increases, the demand for skin care products is also increasing, but the long-term use of synthetic cosmetics, especially some chemical substances, produces many unwanted side effects, such as skin irritations. Phthalates, which are contained in cosmetics and fragrances, are harmful to human body (Hauser and Calafat, 2005). Thus, the development of natural skin care products, characterized as safe, with an antiaging effect, has become an important research topic, and anthocyanins have these qualities, being classified as edible cosmetics and natural vitamins in Europe. They will have a vast market because of their applications in nontoxic makeup and their antioxidant capacity. Sweet potato anthocyanin, as a natural food coloring, is safe, nontoxic, odorless, colorful, and nutritious. Moreover, it had better light

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and heat resistance levels, as well as certain desirable pharmacological effects. High-yield production can be achieved at a low-cost. It is difficult to find a plant, as a new natural pigment resource, that can compete with the high anthocyanin-containing sweet potato.

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84. Verbeyst, L., Crombruggen, K.V., Plancken, I.V.D., et al., 2011. Anthocyanin degradation kinetics during thermal and high pressure treatments of raspberries. J. Food Eng. 105, 513
521. Wan, P., Sheng, Z., Han, Q., et al., 2014. Enrichment and purification of total flavonoids from Flos Populi extracts with macroporous resins and evaluation of antioxidant activities in vitro. J. Chromatogr. B 945, 68
74. Wang, G.L., Yue, J., Li, H.Y., et al., 2005. Extraction of anthocyanin from sweet potato by macroporous resin and its bacteriostatic mechanism. Sci. Agric. Sin. 38 (11), 2321
2326 (in Chinese). Wang, H., Dong, Y.S., Xiu, Z.L., 2008. Microwave-assisted aqueous two-phase extraction of piceid, resveratrol and emodin from Polygonum cuspidatum by ethanol/ammonium sulphate systems. Biotechnol. Lett. 30 (12), 2079
2084. Wu, D., Cederbaum, A.I., 2009. Oxidative stress and alcoholic liver disease. Semin Liver Dis. 29, 141
154. Wu, X.Y., Liang, L.H., Zou, Y., et al., 2011. Aqueous two-phase extraction, identification and antioxidant activity of anthocyanins from mulberry(Morus atropurpurea Roxb.). Food Chem. 129 (2), 443
453. Xiang, J.L., Zhu, W.X., Li, Z.X., et al., 2012. Effect of juice and fermented vinegar from Hovenia dulcis peduncles on chronically alcohol-induced liver damage in mice. Food Funct. 3, 628
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Yamakawa, O., Suda, I., Yoshimoto, M., 1998. Development and utilization of sweet potato cultivars with high anthocyanin content. J. Food Food Ingred. 178, 69
77. Yang, X.S., Yang, Z.M., Gao, F., 2006. Advances in anthocyanin pigments from purple sweet potato [J]. Chin. Agric. Sci. Bull. 26 (4), 94
97 (in Chinese). Yang, Y.L., Kan, J.Q., Shen, H.L., et al., 2012. Optimization of fermentation conditions for purple potato wine. Food Sci. 3 (33), 157
162 (in Chinese). Yang, Z.D., Fan, G.J., Gu, Z.X., et al., 2008a. Optimization extraction of anthocyanins from purple corn（Zea mays L.) cob using tristimulus colorimetry. Eur. Food Res. Technol. 227, 409
415. Yang, Z.D., Han, Y.B., Gu, Z.X., et al., 2008b. Thermal degradation kinetics of aqueous anthocyanins and visual color of purple corn (Zea mays L) cob. Innov. Food Sci. Emerg. Technol. 9, 341
347. Yao, Y.R., 2009. The purification, stability, and antioxidant of sweet potato. Hebei Agric. Univ. (in Chinese). Yin, Q.H., Liu, Y.Z., Xie, Y.Z., et al., 2002. Conditions of extracting anthocyanins from purple sweet potato. Jiangsu J. Agric. Sci. 18 (4), 236
240 (in Chinese). You, Y., Yoo, S., Yoon, H.G., et al., 2010. In vitro and in vivo hepatoprotective effects of the aqueous extract from Taraxacum officinale (dandelion) root against alcohol-induced oxidative stress. Food Chem. Toxicol. 48, 1632
1637. Yu, X., Zhao, F., Li, X., et al., 2009. Study on the development of purple sweet potato health jelly. Sci. Technol. Food Ind 30 (1), 246
251 (in Chinese). Zhai, Y.Q., Li, C.Q., Li, J.G., et al., 2012. Study the extraction process of purple sweet potato. J. Beijing Inst. Petrochem. Technol. 2 (20), 1
4 (in Chinese). Zhang, J.J., Xue, J., Wang, H.B., et al., 2011. Improves alcohol-induced fatty liver in mice by reduction of hepatic oxidative stress. Phytotherrapy Res. 25, 638
643. Zhang, Z.F., Fan, S.H., Zheng, Y.L., et al., 2009. Purple sweet potato color attenuates oxidative stress and inflammatory response induced by D-galactose in mouse liver. Food Chem. Toxicol. 47, 496
501. Zhang, Z.F., Lu, J., Zheng, Y.L., et al., 2010a. Purple sweet potato color protects mouse liver against D-galactose-induced apoptosis via inhibiting caspase-3 activation and enhancing PI3K/ Akt pathway. Food Chem. Toxicol. 48, 2500
2507. Zhang, Y., Shi, G.Q., Zhao, F.S., 2010b. Hydrolysis of casein catalyzed by papain in n-Propanol/ NaCl two-phase system. Enzyme Microbial. Technol. 46 (6), 438
443. Zhao, Z., Zhang, J., Chen, X., Liu, X., Li, J., Zhang, W., 2013. Separation of tungsten and molybdenum using macroporous resin: Equilibrium adsorption for single and binary systems. Hydrometallurgy 140, 120
127. Zheng, Q., Yuan, C.B., 2012. Optimization the recipe of solid purple sweet potato-black sesame yogurt. Food Ind. 9 (33), 295
297 (in Chinese). Zhuang, H., Zhang, J.H., 2003. Epidemiology of alcohol liver disease. Chin. J. Gastroenterol. 8 (5), 294
297.

FURTHER READING Andersen, O.M., Markham, K.R., 2006. Flavonoids: Chemistry, Biochemistry, and Applications. CRC Taylor and Francis, Boca Raton, USA. AOAC. Methods of analysis (15th ed.). Washington: Association 243 of Official Agriculture Chemistry, 1990.

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Cisse, M., Vaillant, F., Acosta, O., et al., 2009. Thermal degradation kinetics of anthocyanins from blood orange, blackberry, and roselle using the arrhenius, eyring, and ball models. J. Agric. Food Chem. 57, 6285
6291. Corrales, M., Garcia, A.F., Butz, P., et al., 2009. Extraction of anthocyanins from grape skins assisted by high hydrostatic pressure. J. Food Eng. 90 (4), 415
421. Galligan, J.J., Smathers, R.L., Shearn, C.T., et al., 2012. Oxidative stress and the ER stress response in a murine model for early-stage alcoholic liver disease. J. Toxicol. 8, 1
12. Gutiérrez, I.H., 2003. Influence of ethanol content on the extent of copigmentation in a cencibel young red wine. J. Agric. Food Chem. 51, 4079
4083. Hosseinian, F.S., Li, W., Beta, T., 2008. Measurement of anthocyanins and other phytochemicals in purple wheat. Food Chem. 109 (4), 916
924. Jing, Pu, Zhao, S.J., Xie, Z.H., et al., 2012. Anthocyanin and glucosinolate occurrences in the roots of Chinese red radish (Raphanus sativus L.), and their stability to heat and pH. Food Chem. 133 (4), 1569
1576. Kimura, Y., Sumiyoshi, M., Tamaki, T., 2012. Effects of the extracts and an active compound curcumenone isolated from Curcuma zedoaria rhizomes on alcohol-induced drunkenness in mice. Fitoterapia 8, 1
7. Kirca, A., Ozkan, M., Cemeroglu, B., 2007. Effects of temperature, solid content and pH value on the stability of black carrot anthocyanins. Food Chem. 101 (1), 212
218. Kobayashi, M., Oki, T., Masuda, M., et al., 2005. Hypotensive effect of anthocyanin-rich extract from purple-fleshed sweet potato cultivar Ayamurasaki in spontaneously hypertensive mice. Nippon Shokuhin Kagaku Kogaku Kaishi 52 (1), 41
44. Lee, J.M., Durst, R.W., Wrolstad, R.E., 2005. Determination of total monomeric anthocyanin pigment content of fruit juices, beverages, natural colorants, and wines by the pH differential method: Collaborative study. J. AOAC Int. 88 (5), 1269
1278. Liu, B., Yuan, L.P., 2012. Processing technology research of purple sweet potato-soy mixed beverage. J. Guangdong AIB Polytechnic Coll. 2 (28), 78
81 (in Chinese). Liu, C., Wang, Z., Li, X., et al., 2008. HPLC analysis and determination of anthocyanidin composents in different varieties of purple sweet potato. Chin. Food Nutr. 8, 19
21 (in Chinese). Liu, J., Ye, T.T., 2012. Study on the extraction technology of anthocyanins from purple sweet potato by cellulose-microwave assisted method. Guangzhou Chem. Ind. 40 (12), 60
62 (in Chinese). Lu, G.Q., Li, X.L., 2001. Stability of red pigments from purple sweet potato and other five red pigments. J. Zhejiang Univ. 27 (6), 635
638 (in Chinese). Lu, J., Wu, D.M., Zheng, Y.L., et al., 2012. Purple sweet potato color attenuates domoic acidinduced cognitive deficits by promoting estrogen receptor-α-mediated mitochondrial biogenesis signaling in mice. Free Rad. Biol. Med. 52, 646
659. Odake, K., Terahara, N., Toshio, H., et al., 1992. Chemical structures of two anthocyanins from purple sweet potato, Ipomoea batatas. Phytochemistry 31 (6), 2127
2130. Oki, T., Suda, I., Terahara, N., et al., 2006. Determination of acylated anthocyanin in human urine after ingesting a purple-fleshed sweet potato beverage with various contents of anthocyanin by LC-ESI-MS/MS. Biosci. Biotechnol. Biochem. 70 (10), 2540
2543. Paredes, L.O., Cervantes, C.M.L., Vigna, P.M., et al., 2010. Berries: improving human health and healthy aging, and promoting quality life: a review. Plant Foods Human Nutr. 65, 299
308. Pascual-Teresa, S.D., Sanchez-Ballesta, M.T., 2008. Anthocyanins: from plant to health. Phytochem. Rev. 7, 281
299.

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Patras, A., Brunton, N.P., O0 Donnell, C., et al., 2010. Effect of thermal processing on anthocyanin stability in foods, mechanisms and kinetics of degradation. Trends Food Sci. Technol. 21, 3
11. Pompeu, D.R., Silva, E.M., Rogez, H., 2009. Optimisation of the solvent extraction of phenolic antioxidants from fruits of Euterpe oleracea using Response Surface Methodology. Bioresour. Technol. 100 (23), 6076
6082. Prior, R.L., Wu, X., 2006. Anthocyanins: Structural characteristics that result in unique metabolic patterns and biological activities. Free Rad. Res. 40, 1014
1028. Reyes, L., Fernando, Z.L., 2007. Degradation kinetics and colour of anthocyanins in aqueous extracts of purple- and red-flesh potatoes (Solanum tuberosum L.) [J]. Food Chem. 100, 885
894. Sariburun, E., Sahin, S., Demir, C., et al., 2010. Phenolic content and antioxidant activity of raspberry and blackberry cultivars. J. Food Sci. 4 (75), C328
C335. Simeone, M., Alfani, A., Guido, S., 2004. Phase diagram, rheology and interracial tension of aqueous mixtures of Na-caseinate and Na-algihate. Food Hydrocoll. 18 (3), 463
470. Stephen, E.M., Akira, K., Erwin, G., et al., 2002. Cocoa extract protects against early alcoholinduced liver injury in the rat. Arch. Biochem. Biophys. 406, 40
46. Suda, I., Oki, T., Masuda, M., et al., 2003. Physiological functionality of purple-fleshed sweet potatoes containing anthocyanins and their utilization in foods. Jpn. Agric. Res. Q. 37 (3), 167
173. Verbeyst, L., Crombruggen, K.V., Plancken, I.V., et al., 2011. Anthocyanin degradation kinetics during thermal and high pressure treatments of raspberries. J. Food Eng. 105, 513
521. Viviana, R.L., Gabriela, S.R., María, S.G., et al., 2011. Antioxidant properties of Amaranthus hypochondriacus Seeds and their effect on the liver of alcohol-treated mice. Plant Foods Hum. Nutr. 66, 157
162. Yang, Z.D., Zhai, W.W., 2010. Optimization of microwave-assisted extraction of anthocyanins from purple corn (Zea mays L.) cob and identification with HPLC-MS. Innov. Food Sci. Emerg. Technol. 11 (3), 470
476. Zou, T.B., Wang, D.L., Guo, H.H., et al., 2012. Optimization of microwave-assisted extraction of anthocyanins from mulberry and identification of anthocyanins in extract using HPLC
ESI
MS [J]. J. Food Sci. 77 (1), C46
C50.

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CHAPTER

7

Chlorogenic Acids From Sweet Potato

SECTION 1: OVERVIEW OF CHLOROGENIC ACIDS FROM SWEET POTATO 1.1 Composition and Structure of Chlorogenic Acids 1.2 Biological Activities of Chlorogenic Acids 1.2.1 Antioxidant Activity 1.2.2 Antimicrobial Activity 1.2.3 Antimutation and Anticancer Activities 1.2.4 Hypoglycemic, Serum Lipid-Lowering, and Antihypertensive Activities 1.2.5 Protecting the Cardiovascular and Central Nervous Systems 1.2.6 Protection Against Diabetes 1.2.7 Other Biological Activities 1.3 Extraction, Separation, and Purification Methods of Chlorogenic Acids 1.3.1 Extraction Methods of Chlorogenic Acids 1.3.2 Chlorogenic Acid Separation and Purification Methods 1.4 Qualitative and Quantitative Analyses of Chlorogenic Acids 1.4.1 UV Spectrophotometry 1.4.2 Thin-Layer Chromatography 1.4.3 High-Performance Liquid Chromatography (HPLC) and its Related Techniques SECTION 2: TECHNOLOGY TO PREPARE CHLOROGENIC ACIDS FROM SWEET POTATOES 2.1 Pretreatment of Sweet Potato Leaves 2.2 Total Polyphenol Contents 2.3 Extraction of Chlorogenic Acids From Sweet Potato Leaves Sweet Potato Processing Technology. DOI: http://dx.doi.org/10.1016/B978-0-12-812871-8.00007-6 Copyright © 2017 China Science Publishing & Media Ltd. Published by Elsevier Inc. All rights reserved.

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2.4 Purification of Chlorogenic Acids From Sweet Potato Leaves using AB-8 Macroreticular Adsorbent Resin 2.4.1 Activation Pretreatment of AB-8 Macroreticular Adsorbent Resin 2.4.2 Static Adsorption and Desorption test of AB-8 Macroreticular Adsorbent Resin 2.4.3 Dynamic Adsorption and Desorption of AB-8 resin 2.5 Qualitative and Quantitative Analyses of Chlorogenic Acids From Sweet Potato Leaves by HPLC SECTION 3: BIOLOGICAL ACTIVITIES OF CHLOROGENIC ACIDS FROM SWEET POTATOES 3.1 In vitro Antioxidant Activity of Chlorogenic Acids From Sweet Potatoes 3.1.1 2,2-Diphenyl-1-Picrylhydrazyl (DPPH) Radical-Scavenging Activity of Chlorogenic Acids From Sweet Potatoes 3.1.2 Hydroxyl Radical-Scavenging Activity of Chlorogenic Acids From Sweet Potatoes 3.1.3 Superoxide (O22•)-Scavenging Activities of Chlorogenic Acids From Sweet Potatoes 3.1.4 Peroxy Radical-Scavenging Activities of Chlorogenic Acids From Sweet Potatoes 3.1.5 Ferric-Reducing Antioxidant Power of Chlorogenic Acids From Sweet Potatoes 3.1.6 Lipid Peroxidation-Inhibiting Activities of Chlorogenic Acids From Sweet Potatoes 3.2 Antimicrobial Activities of Chlorogenic Acids From Sweet Potatoes 3.3 Aldose Reductase Inhibitory Activities of Chlorogenic Acids From Sweet Potatoes 3.4 Anticancer Activities of Chlorogenic Acids From Sweet Potatoes 3.5 Other Biological Activities of Chlorogenic Acids From Sweet Potatoes SECTION 4: THE STABILITY OF CHLOROGENIC ACIDS FROM SWEET POTATOES 4.1 Effect of pH on Chlorogenic Acids From Sweet Potato Leaves

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4.2 Effects of Heat Treatments on Chlorogenic Acids From Sweet Potato Leaves 4.3 Effects of Light on Chlorogenic Acids From Sweet Potato Leaves SECTION 5: THE APPLICATIONS OF CHLOROGENIC ACIDS FROM SWEET POTATOES 5.1 Food Industry 5.2 Medicine and Health-Protection Industry 5.3 Daily Chemical Industry References

Abstract This chapter gives a relatively comprehensive introduction about chlorogenic acids from sweet potato. It starts by presenting a wellrounded overview on chlorogenic acids. It then presents the preparing technology of chlorogenic acids from sweet potato, including pretreatment, extraction, purification, and qualitative and quantitative analyses. Next, the biological activities of chlorogenic acids from sweet potato are introduced and followed by the effects of pH, heat, and light treatments on the total polyphenol content and antioxidant activities of chlorogenic acids from sweet potato. By the end of this chapter, the applications of chlorogenic acids from sweet potato in different products are introduced.

SECTION 1: OVERVIEW OF CHLOROGENIC ACIDS FROM SWEET POTATO In developing countries, desertification has contributed to a reduction in cultivable land and, therefore, to an increase in food shortage. Crops that are resistant to different environmental, soil, and temperature conditions are required. Sweet potato (Ipomoea batatas L.) is a highly resistant crop that originated in Central America. In Japan, where sweet potato is considered to be a hardy plant, both roots and leaves are consumed (Ishida et al., 2000). However, in China, sweet potato leaves are only used in livestock feed. Furthermore, studies focusing on the bioactive components of sweet potato leaves are scarce.

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Sweet potato leaves can be harvested several times during the year, and their yields are much greater than those of green leafy vegetables (An et al., 2003). Furthermore, compared with green leafy vegetables, sweet potato leaves are more tolerant of diseases, pests, and high moisture conditions. Sweet potato leaves constitute an alternative source of green leafy vegetables during their off-season and could potentially alleviate food shortages caused by natural disasters, such as tsunamis, floods, and typhoons (Taira et al., 2013). Sweet potato leaves contain a large amount of phenolic compounds, of which more than 70% are chlorogenic acids and their derivatives, and 10% to 20% are flavonoid compounds. Phenolic compounds, which are a phenylpropanoid class of secondary metabolites generated during aerobic respiration in plants through the shikimic acid pathway, widely exist in higher dicotyledonous plants and ferns. Chlorogenic acids possess many human health-related activities, such as radicalscavenging, antibacterial and antiinflammatory, tumor-inhibiting, liver-protecting and cholagogic, and blood-activating and antihypertensive, and have become a focus of natural products-related research.

1.1 COMPOSITION AND STRUCTURE OF CHLOROGENIC ACIDS Chlorogenic acids belong to the ester compound family and are formed by the condensation of quinic acid and trans-cinnamic acids, which include coffee acid, p-cumaric acid, and ferulic acid (Marques et al., 2009; Michael et al., 2006). Because of the different positions, varieties, and quantities of the condensation between quinic acid and transcinnamic acids, there are a variety of chlorogenic acids (molecular structures are shown in Fig. 7.1 and Table 7.1). 3-O-caffeoylquinic acid is the most common of the chlorogenic acids, with the molecular formula C16H18O9, and a molar mass of 354.30 g/mol. In addition, the esterified chlorogenic acids (esterification reactions occur at R1 of the molecular structure of quinic acid in Fig. 7.1) are also chlorogenic acids, such as methyl chlorogenate and chlorogenic ethyl ester.

1.2 BIOLOGICAL ACTIVITIES OF CHLOROGENIC ACIDS Chlorogenic acids, which are the main functional components of many Chinese herbal medicines, possess strong biological activities and are important indicators of quality control in many medicines.

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Figure 7.1 Composition and molecular structure of chlorogenic acids.

Table 7.1 Molecular Structure and Plant Sources of Chlorogenic Acids Name

R1

R2

R3

Plant Sources

3-O-caffeoylquinic acid

C

H

H

Existing widely

4-O-caffeoylquinic acid

H

C

H

Eucommia ulmoides

5-O-caffeoylquinic acid

H

H

C

Sweet potato leaves, coffee

3-O-coumaroyl guinic acid

p-Co

H

H

Hemerocallis fulva

4-O-coumaroyl guinic acid

H

p-Co

H

Coffee, Hemerocallis fulva

5-O-coumaroyl guinic acid

H

H

p-Co

Hemerocallis fulva

3-O-feruloylquinic acid

F

H

H

Eucommia ulmoides, honeysuckle

4-O-feruloylquinic acid

H

F

H

Eucommia ulmoides, coffee

5-O-feruloylquinic acid

H

H

F

Tea, coffee

4,5-O-caffeoylquinic acid

H

C

C

Eucommia ulmoides, tea

3,4-O-caffeoylquinic acid

C

C

H

Honeysuckle, sweet potato

3,5-O-caffeoylquinic acid

C

H

C

Honeysuckle, Eucommia ulmoides

3,4-O-coumaroyl guinic acid

p-Co

p-Co

H

Coffee

3,5-O-coumaroyl guinic acid

p-Co

H

p-Co

Coffee

4,5-O-coumaroyl guinic acid

H

p-Co

p-Co

Coffee

3-O-feruloyl,4-O-caffeoylquinic acid

F

C

H

Sweet potato leaves, coffee

3-O-caffeoyl,4-O-feruloylquinic acid

C

F

H

Sweet potato leaves, coffee

3-O- feruloyl,5-O-caffeoylquinic acid

F

H

C

Sweet potato leaves

3-O- caffeoyl,5-O-feruloylquinic acid

C

H

F

Sweet potato leaves

3-O- feruloyl,5-O-caffeoylquinic acid

H

F

C

Sweet potato leaves

3-O- caffeoyl,5-O-feruloylquinic acid

H

C

F

Sweet potato leaves

3,4,5-O-caffeoylquinic acid

C

C

C

Sweet potato leaves, coffee

Note: R1, R2, and R3 represent the three different sites in the molecular structure of quinic acid in Fig. 7.1, respectively. H represents hydrogen. C, F, and Co represent caffeic acid, ferulic acid, and cumaric acid, respectively, in Fig. 7.1.

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1.2.1 Antioxidant Activity The phenolic hydroxyl of chlorogenic acids easily reacts with free radicals, making the free radicals lose activity. Thus, chlorogenic acids have remarkable antioxidant activity levels. Chlorogenic acids in plants have high antioxidant activities, such as reducing power, metalchelating activity, and lipid peroxidation-inhibiting activity (Liao et al., 2011). Antioxidant activity is an important activity of chlorogenic acids and is the basis of antitumor, antiaging, and other physiological functions. Li et al. (2012) reported that chlorogenic acids possess high antioxidant activity levels and could protect bone marrow stem cells from oxidative stress injuries induced by reactive oxygen species. By promoting Akt phosphorylation, chlorogenic acids increase the expression of the FOXO gene family and the antiapoptotic protein Bcl-2, which inhibits the damage caused by reactive oxygen species to cells. Sato et al. (2011) found that because of the high antioxidant activity, chlorogenic acids showed significant protective effects on cells having ischemic-reperfusion injuries. Although there are many studies on the antioxidant activities of chlorogenic acids at home and abroad, these studies are cursory. Additionally, there are few studies on the differences in antioxidant activities between different chlorogenic acids, thus failing to provide sufficient theoretical support for the application of antioxidant activities.

1.2.2 Antimicrobial Activity At present, domestic and foreign research indicates that chlorogenic acids have significant inhibitory effects on common pathogenic bacteria. Continuing to improve the antibacterial research using chlorogenic acids and elucidating the inhibitory mechanism will promote their application as additives during food processing.

1.2.3 Antimutation and Anticancer Activities Chlorogenic acids can generate anticancer effects through several mechanisms, including adjusting the cell cycle, inducing apoptosis, and inhibiting cell growth. Research shows that the molecular mechanism responsible for chlorogenic acids adjusting cell survival and apoptosis involved the following: (1) Reducing the intensity of signal factors in the signal transduction pathway of cell growth and reproduction, thereby reducing the viability of cancer cells; (2) Improving the activity of quinone oxidoreductase-glutathione transferase and NAD(P)H, thereby opposing the induction of oxidants having carcinogenic effects

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on cell carcinogenesis; (3) Reducing the activity of oncogenic kinases like MAPK, thereby preventing the cancerization of cells; and (4) Stimulating the expression of cancer suppressor genes, like Nrf2, thereby inhibiting the growth of cancer cells (Granado-Serrano et al., 2007; Feng et al., 2005). Jiang et al. (2000) found that in an alkaline environment chlorogenic acids were prooxidants, which could induce tumor cells that generated large DNA fragments and led to nuclear agglutination, thereby inhibiting of tumor cell growth. Aryl hydrocarbon hydroxylase is an important metabolic enzyme involved with polynuclear aromatic hydrocarbons, and chlorogenic acids can enhance the activity of aryl hydrocarbon hydroxylase, thereby enhancing the antimutagenic effects of tissue cells against polynuclear aromatic hydrocarbons (Matsunaga et al., 2002). Thus, chlorogenic acids have the potential to be widely used in the research and development of antitumor drugs.

1.2.4 Hypoglycemic, Serum Lipid-Lowering, and Antihypertensive Activities Cho et al. (2010) added chlorogenic acids (0.02%) to the diets of obese mice and found that the triglyceride and cholesterol levels in the blood, livers, and hearts of mice were significantly decreased, whereas the levels of adiponectin and leptin in the blood were increased. Thus, chlorogenic acids could significantly inhibit the activity of fatty acid synthase, 3-hydroxy-3-methyl pentyl acyl coenzyme A, and cholesterol acyltransferase, and increase the activity of fatty acid β-oxidase. In addition, compared with caffeic acid, chlorogenic acids are more effective in regulating lipid metabolism. Ong et al. (2013) found that chlorogenic acids could adjust the glucose tolerance level and insulin sensitivity in mouse liver cells, thereby improving gluconeogenesis and the lipid metabolism of liver. In addition, the long-term feeding of a certain amount of chlorogenic acids to mice inhibited the activity of glucose 6-phosphatase and then weakened the pathological changes in liver cells and blood lipid levels, and its mechanism was related to the activation of 50 AMP-activated protein kinase. Andersen et al. (2006) confirmed that chlorogenic acids had hypoglycemic effects in animals and that there was no significant difference in the hypoglycemic effects of chlorogenic acids and glibenclamide (the main efficacy component of hypoglycemic drugs) after 3 h. The mechanism might be related to inhibiting the absorption of glucose-6-phosphate and glucose. The

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hypoglycemic and serum lipid-lowering activities of chlorogenic acids have been confirmed many times, but research on their mechanisms is limited and fails to provide a reliable theoretical basis for the application of chlorogenic acids in hypoglycemic, serum lipid-lowering, and antihypertensive activities.

1.2.5 Protecting the Cardiovascular and Central Nervous Systems The antioxidant activities of chlorogenic acids can increase the potassium ion concentration in blood and decrease the levels of triacylglycerol and cholesterolin in the blood, preventing cardiovascular disease (Frank et al., 2003). Chlopcikova et al. (2004) found that chlorogenic acids could protect myocardial cells from the oxidative damage induced by doxorubicin and significantly inhibit the lipid peroxidation of myocardial cells and mitochondrial membranes induced by doxorubicin, with median inhibitory concentrations (IC50) of 8.04 6 0.74 and 6.87 6 0.52 μmol/L, respectively. Chlorogenic acids can reduce cerebral infarctions, and blood
brain barrier and brain edema by inhibiting lipid peroxidation and the activity of matrix metalloproteinases, thereby significantly inhibiting cerebral lesions (Lee et al., 2012). Kim et al. (2012) found that the hydrogen peroxide (H2O2)-induced apoptosis of neurons was strongly inhibited by treatments with chlorogenic acids and that the activities of intracellular antioxidant enzymes were increased. Thus, chlorogenic acids can protect vascular endothelial cells by scavenging free radicals and by antilipid peroxidation, thus playing roles in the prevention and treatment of atherosclerosis, thrombosis, and hypertension.

1.2.6 Protection Against Diabetes Chlorogenic acids, as antioxidants, can inhibit the activity of glucose 6-phosphatase in the liver and can regulate glucose’s passive absorption driving force, a Na1 electrochemical gradient, thus generating antagonistic effects on glucose transport, decreasing the rate of the intestinal absorption of glucose, and preventing diabetes (Riksen et al., 2009). Kurata et al. (2011) found that chlorogenic acids from sweet potato leaves had inhibitory effects on aldose reductase and that 3,4,5-O-caffeoylquinic acid (3,4,5-O-CQA) had the highest inhibitory effect. Aldose reductase inhibitors play important roles in diabetic complications, such as cataracts, neuropathy, retinopathy, and

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nephropathy, indicating that 3,4,5-O-CQA had a potential medicinal value for treating diabetic complications.

1.2.7 Other Biological Activities Wang et al. (2009a,b) found that chlorogenic acids could significantly inhibit the activity of the hepatitis B virus. Chlorogenic acids have antiinflammatory effects that inhibit the activity of hyaluronidase. The antiinflammatory and antiinfection activities are stronger than aspirin. Chlorogenic acids also effectively inhibit H2O2-induced red blood cell hemolysis (Wang et al., 2008a,b). Nilufer et al. (2013) found that chlorogenic acids had radiation-protective effects, and the comet assay showed that chlorogenic acids could significantly reduce X-ray-induced DNA damage in human peripheral lymphocytes, with the damage reduction of 5.99%
53.57%. At present, domestic and foreign scholars study the biological activities of chlorogenic acids, mainly those involving antioxidants, free radical scavenging, antiinflammation, tumor inhibition, and hyperglycemic, antihypertensive, cardiovascular protection effects. The researchers tried to explain the mechanisms of a variety of biological activities involving chlorogenic acids, but some of the mechanisms are not yet conclusive, and thus, need to be further studied.

1.3 EXTRACTION, SEPARATION, AND PURIFICATION METHODS OF CHLOROGENIC ACIDS 1.3.1 Extraction Methods of Chlorogenic Acids There are many methods for extracting chlorogenic acids, including water extraction, organic solvent extraction, enzymatic hydrolysis, supercritical fluid extraction, microwave-assisted extraction, and ultrasonic-assisted method. The organic solvent extraction method is a classical method, and ultrasonic- and microwave-assisted organic solvent extraction methods are widely used at present because of their short extraction times and simple operations. The principles, application examples, and advantages and disadvantages of each method are shown in Table 7.2.

1.3.2 Chlorogenic Acid Separation and Purification Methods There are many chlorogenic acid separation and purification methods, such as metal salt precipitation, recrystallization, β-cyclodextrin

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Table 7.2 Extraction Methods of Chlorogenic Acids Methods

Principles

Advantages and Disadvantages

Water extraction

Chlorogenic acids are soluble in water, thus hot water (B50 C) can be used as the extracting agent

Advantages: simple process and low cost

Chlorogenic acids are soluble in ethanol, methanol, acetone, and other organic solvents, and can be extracted in certain concentrations of organic solvents. Ethanol is a widelychosen organic solvent because it is cheap and environmentally nontoxic

Advantages: lower energy consumption, simple processes, and high-product yields

Pretreatments of raw materials with cellulase and pectinase can break the plant cell wall and macromolecules, making the chlorogenic acids in cells more soluble

Advantages: high product yield and high purity

Supercritical fluid extraction

When the extraction agent (usually CO2) is above the critical temperature and pressure, the solubility is enhanced and components with different polarities, boiling points and molecular weights can be successively extracted by the extraction agent

Advantages: high purity

High hydrostatic pressure extraction

At room temperature with 100 to 1000 MPa hydrostatic pressure in the extraction system, the extraction solvent quickly penetrates to the interior of the cell, promoted by the pressure, and the pressure can also cause cell deformations and ruptures, resulting in intracellular chlorogenic acids dissolving

Advantages: short extraction time

Microwaveassisted extraction

During microwaving the inner tissue of the plant is rapidly heated and broken, so that the chlorogenic acids in the tissue cells are fully expelled

Advantages: simple, fast and efficient process, conserves energy and is environmentally conscious

The mechanical effect of ultrasound is used to break the cell, and the chlorogenic acids can penetrate into the solution from the plant cell

Advantages: short extraction time, high yield, simple process, large handling capacity, and easily realized industrial production

Organic solvent extraction

Enzymatic hydrolysis

Ultrasonicassisted method

Disadvantages: long production cycle and large amount of waste water

Disadvantages: long extraction time, large solvent usage amount, and difficult to recycle

Disadvantages: high cost, effect of the reaction system on enzyme activities and poor reproducibility

Disadvantages: high technology and equipment costs, large operational energy consumption and low yields

Disadvantages: expensive equipment, and inability to realize industrial production

Disadvantages: expensive equipment

Disadvantages: poor equipment stability

inclusion, organic solvent, membrane separation, resin, and chromatography, and some methods cannot be widely used because of cumbersome processes, complex operations, poor separations, and purification effects. Seven commonly used methods in the field of chlorogenic acid separation and purification are introduced in this section, and the principles, application examples, and advantages and disadvantages of each method are shown in Table 7.3.

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Table 7.3 Chlorogenic Acid Separation and Purification Methods Methods

Principles

Advantages and Disadvantages

Organic solvent

The different solubilities of chlorogenic acids in organic solvents are used, and one or more compounds can be enriched and purified by multiple extractions. This method is usually used with other separation methods to realize full separation

Advantages: low cost, simple process

The permselectivity of the membrane was used to separate small molecular weight chlorogenic acids from proteins and carbohydrates, thereby realizing separation

Advantages: low-energy consumption, no pollution, and high yield

Chlorogenic acids are phenolic acid type compounds. By adjusting the pH, chlorogenic acids can exist as anions or ionic forms, which can be adsorbed and separated by strong basic anion exchange or strong acid cation resins (fiber)

Advantages: great adsorption capacity and simple process

The millipore gel sieves substances with different molecular weights, resulting in different diffusion rates, which can separate and purify the substances by molecular weight. Dextran gel chromatography is often used to separate and purify chlorogenic acids

Advantages: simple equipment, easy operation, and high purity

High speed countercurrent chromatography

This is liquid
liquid chromatography. The separation of substances is based on the different distribution coefficients in the solvent system

Advantages: high purity

Macroreticular resin absorption

Macroreticular resin does not contain ion exchange groups but has polymeric adsorbents with macroporous structures, which can separate and purify chlorogenic acids by adsorption and screening

Advantages: simple process, high production, convenient for resin regeneration, and fit for industrial production

Based on the different distribution coefficients of solute molecules in fixed and mobile phases, HPLC realizes the separation at the equilibrium point after multiple distributions. HPLC is divided into analysis and preparation types. Analytical HPLC is mainly used for identification, detection and smallscale preparations, and preparation HPLC is mainly used for the separation and preparation of materials

Advantages: high separation effect, high purity, and high degree of automation

Membrane separation

Ion exchange separation

Gel chromatography

High performance liquid chromatography

Disadvantages: poor solvent selectivity, and low purification efficiency

Disadvantages: long production cycle, easily affected by filtration conditions, and easily polluted

Disadvantages: long production cycle, hard to clean and regenerate of resin, and not fit for large-scale production

Disadvantages: high production cost, low yield, and high investment for industrial production

Disadvantages: high purity requirements for substances, difficult to determine the separation method, and high cost

Disadvantages: complicated resin types

Disadvantages: expensive instruments, low production, and high technical requirements

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1.4 QUALITATIVE AND QUANTITATIVE ANALYSES OF CHLOROGENIC ACIDS The compositions of chlorogenic acids are relatively complex, and there is no characteristic group that is obviously different from other phenolic acids. Therefore, there are no simple and easy methods for the qualitative and quantitative analyses. At present the commonly used qualitative and quantitative chlorogenic acid analyses methods include ultraviolet (UV) spectrophotometry, thin-layer chromatography and high-performance liquid chromatography.

1.4.1 UV Spectrophotometry Chlorogenic acids have UV absorption peaks in a certain wavelength range; thus, UV spectrophotometry can be used for quantitative analyses. Chlorogenic acids have maximum absorption peaks in the wavelength range of 300 to 400 nm, and in a certain concentration range (10 to 80 μg/mL), there is a good linear relationship between chlorogenic acid concentration and absorbance. Although the UV spectrophotometric method is simple and fast, the results are often high, and it is mainly used for initial quantifications because the measured object usually contains impurities, such as caffeic acid, and these impurities have the same absorption at the same wavelength.

1.4.2 Thin-Layer Chromatography Thin-layer chromatography is used to separate, identify, and quantify mixed samples by coating a holder on the support plate during the stationary phase and a suitable solvent acts during the mobile phase. At present, in the qualitative and quantitative analyses of chlorogenic acids, thin-layer chromatography is mainly used for preliminary qualitative analyses. Although thin-layer chromatography is simple, the separation difficulties and the vulnerability to interference by impurities result in it being not often used in the qualitative and quantitative analyses of chlorogenic acids.

1.4.3 High-Performance Liquid Chromatography (HPLC) and its Related Techniques High-performance liquid chromatography is the most common analytical method reported in the literature. It has the characteristics of obvious separation effects, high sensitivity and good reproducibility.

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Jung et al. (2011) identified a variety of chlorogenic acids from sweet potato leaf polyphenols using an HPLC
UV
Vis detection system, with acetonitrile and 0.5% of formic acid aqueous solution as the mobile phase for the gradient elution and a column temperature of 30 C. The chlorogenic acid qualitative and quantitative analyses obtained by HPLC and its related techniques are accurate and reliable, reproducible, quick, and have a high degree of automation. Thus, at present it is widely used in the qualitative and quantitative analyses of chlorogenic acids. At home and abroad, chlorogenic acid research has become more and more mature, and with more in-depth research, its applications will be more extensive; however, the following aspects can be improved: 1. Developing new raw material resources for chlorogenic acid extractions. Some studies have found that chlorogenic acids can be extracted from agricultural by-products, food processing waste residues and waste liquids. The development of new resources for chlorogenic acid extractions, which may aid in the comprehensive utilization of waste materials, has great environmental significance; 2. Studying the separation and purification of chlorogenic acids and their structural
functional relationships. Chlorogenic acids are composed of a variety of components and, at present, the separation and purification processes of each component are still not efficient enough. The activity differences between different components needs further study. In addition, the mechanisms behind the activity levels of chlorogenic acids, especially in vivo activity levels, is not clear, resulting in a lack of reliable theoretical support for their use. 3. Expanding the application range and increasing the application depth. Chlorogenic acids have many biological activities. At present, the uses for the chlorogenic acid activities in food, medicine, and chemical industries are focused on aspects of free-radical scavenging, and antibacterial and antiinflammatory properties, as well as on the development and application of tumor inhibition-, cardiovascular protection-, blood circulation activation-, blood pressure depression-, and diabetes prevention-related activities, which need to be further expanded. Thus, much work on chlorogenic acids is still necessary.

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SECTION 2: TECHNOLOGY TO PREPARE CHLOROGENIC ACIDS FROM SWEET POTATOES The preparation of chlorogenic acids from sweet potato leaves includes two steps: Extraction and purification. The methods to extract chlorogenic acids from sweet potato leaves include water extraction, organicsolvent extraction, the enzyme-hydrolysis method, the microwave-assisted method, and the ultrasonic-assisted method, of which the ultrasonicassisted ethanol solvent extraction is widely used because it is a simple and highly efficient process (Li et al., 2007). Compared with the relatively mature extraction methods, there are limited studies on the purification of chlorogenic acids from sweet potato leaves. At present, the commonly used purification methods for plant chlorogenic acids include the metal-salt precipitation method, recrystallization method, β-cyclodextrin-inclusion method, organicsolvent method, membrane-separation method, resin method, and chromatography method; however, some methods cannot be widely used because of cumbersome processes, complex operations, or poor separations and purification effects. Because of its simple process, low cost, and efficient separation and purification results, macroporous adsorption resin is the most extensively applied. Macroporous adsorption resin is a polymer adsorbent that does not have an ion exchange group, but has a macroporous structure, and it can separate and purify chlorogenic acids through adsorption and screening. Because of its fast adsorption, high desorption rate, large adsorption capacity, high elution rate, and simple resin regeneration, macroporous adsorption resin is widely used in the purification of chlorogenic acids from plants, such as eucommia bark, and honeysuckle. In this section, sweet potato leaves were used as raw materials, chlorogenic acids were extracted from sweet potato leaves by ultrasonicassisted ethanol solvent extraction, and the AB-8 macroporous adsorption resin was used as adsorbent. The purification process of chlorogenic acids from sweet potato leaves was studied. Because there is no accurate and efficient method for the quantitative analysis of chlorogenic acids, which are the main components of sweet potato leaves, the total phenolic content (TPC) was taken as the index to determine the optimal process parameters. The methods for the preparation of chlorogenic acids from sweet potato leaves are introduced as follows.

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2.1 PRETREATMENT OF SWEET POTATO LEAVES Two sweet potato leaf cultivars, “Yuzi No.7” and “Ximeng No.1,” were collected in August 2012 from the Sweet Potato Research Institute in Xuzhou, Jiangsu Province, China. The 10
15 cm long leaf parts at the tips were selected. The fresh sweet potato leaves were washed, drained, freeze-dried, and ground to a powder with a grinder. After the samples were sifted through a 40 mesh sieve, the leaf powder was packed into aluminum foil bags and stored at 4 C for further use.

2.2 TOTAL POLYPHENOL CONTENTS The TPC was measured using the Folin
Ciocalteu method. Briefly, a 0.5 mL sample solution was mixed with 1.0 mL of Folin
Ciocalteu reagent (103dilution) and allowed to react at 30 C for 30 min. Then 2.0 mL of saturated Na2CO3 (10% w/v) was added and kept at 30 C for 30 min. The absorbance was measured at 736 nm using a UV-3010 spectrophotometer (Hitachi, Ltd., Tokyo, Japan). A calibration curve for the chlorogenic acid standards (at concentrations of 0.02, 0.04, 0.06, 0.08, and 0.10 mg/mL) was prepared. The linear regression equation was y 5 8.7671x 1 0.0068 and R2 5 0.9994. The TPC was expressed as mg chlorogenic acid equivalent per mL of sample solution (mg CAE/mL).

2.3 EXTRACTION OF CHLOROGENIC ACIDS FROM SWEET POTATO LEAVES The sweet potato leaf powder (10 g) was extracted with 70% (v/v) ethanol solution (200 mL) for 30 min at 50 C using ultrasonic (59 kHz) assistance. After the solution was centrifuged at 8711 3 g for 10 min, the residue was reextracted twice with 70% ethanol as described above. The supernatants were combined and concentrated in a rotary evaporator to obtain the crude chlorogenic acid extraction.

2.4 PURIFICATION OF CHLOROGENIC ACIDS FROM SWEET POTATO LEAVES USING AB-8 MACRORETICULAR ADSORBENT RESIN 2.4.1 Activation Pretreatment of AB-8 Macroreticular Adsorbent Resin The resins were soaked four times with (w/v) 95% (v/v) ethanol for 24 h, washed thoroughly with distilled water until the water clarified

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Table 7.4 The Physical Parameters of AB-8 Macroreticular Adsorbent Resin Type of

Specific Surface

Moisture

Average Pore

Particle

Wet Vacuum

Resin

Area (m2/g)

Content (%)

Size (A)

Size (mm)

Density (g/mL)

AB-8

480
520

60
70

130
140

$95

1.05
1.09

Polarity

Weak polar

and were then soaked four times with (w/v) 2 mol/L HCl and 2 mol/L NaOH solutions successively for 4 h for each solution. They were thoroughly washed in distilled water until the washing fluid became neutral. The resins were filtered to remove the water before use. The physical parameters of AB-8 macroreticular adsorbent resin are shown in Table 7.4.

2.4.2 Static Adsorption and Desorption Test of AB-8 Macroreticular Adsorbent Resin 2.4.2.1 Determination of Static Adsorption and Desorption Performance Exactly 2 g of the pretreated AB-8 macroporous resin was put into a 250 mL triangular flask and 50 mL of crude chlorogenic acid solution (TPC, 1.0 mg CAE/mL) was added. The flask was shaken on an immersion oscillator (130 r/min) at 25 C for 24 h to reach the adsorption equilibrium. The TPC of the solution after adsorption was determined using the Folin
Ciocalteu method. The adsorption capacities were calculated using formula (7.1). After the adsorption equilibrium was reached, the resins were washed twice using 50 mL distilled water and then desorbed by 50 mL 70% (v/v) ethanol solution in a flask, which was shaken on an immersion oscillator (130 r/min) at 25 C for 24 h. The TPC of the desorption solution was then determined. The desorption ratio was calculated using formula (7.2): Qt 5

V0 ðC0
Ct Þ ; M

(7.1)

where Qt is the adsorption capacity of the resin at time t (mg/g); C0 and Ct are the TPCs of the sample solution at the beginning and at time t, respectively (mg CAE/mL); V0 is the initial volume of the solution added to the flask (mL), and M is the weight of the resin (g). D5

100Cd Vd ½V0 ðC0
Ce Þ

(7.2)

where D is the desorption ratio (%); Cd is the TPC of the desorption solution (mg CAE/mL); Vd is the desorption solution volume (mL); Ce is the

Chlorogenic Acids From Sweet Potato

Adsorption curve

30.0

373

100.0

Desorption Curve 80.0

20.0 60.0 15.0 40.0

D (%)

Qt (mg CAE/g)

25.0

10.0 20.0

5.0 0.0

0.0 0

5

10

15 Time (h)

20

25

Figure 7.2 The static adsorption and desorption curve of AB-8 resin.

equilibrium TPC in the sample solution, and C0 and V0 are as defined in formula (7.1). The adsorption and desorption kinetics of AB-8 macroporous resins on chlorogenic acids from sweet potato leaves are shown in Fig. 7.2. The chlorogenic acids from sweet potato leaves were adsorbed rapidly by AB-8 macroporous resins. Within 1 h, the adsorption level had increased sharply. After 3 h, the adsorption level did not show any further significant changes, which suggested that the adsorption equilibrium occurred at 3 h when the adsorption amount was 20.06 mg CAE/g. As shown by the desorption curve, the chlorogenic acids absorbed into the resin were desorbed effectively by 70% (v/v) ethanol solution and the desorption ratio increased rapidly during the initial stage (within 1 h). After 2 h, the desorption ratio did not change significantly and equilibrium occurred when the desorption ratio was 82.51%. 2.4.2.2 Effect of Sample pH on the Adsorption Capacity of AB-8 Resin The crude chlorogenic acid solutions (TPC, 2.0 mg CAE/mL) were prepared, and each of them was adjusted to a different pH value (2.0, 3.0, 5.0, 7.0, and 8.0) using 1.0 mol/L HCl and 1.0 mol/L NaOH solutions. Then, 50 mL of each solution was adsorbed by 2 g resin as described above, and the TPC of the solution was determined when adsorption had finished. The adsorption amounts were calculated to investigate the relationship between adsorption capacity and sample pH value.

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30

Qe (mg CAE/g)

25 20

15 10 5

0

0

2

4

6

8

10

pH

Figure 7.3 Effect of sample pH on the adsorption capacity of AB-8 resin.

The effect of the pH value on the adsorption of AB-8 resin is shown in Fig. 7.3. The adsorption capacity of AB-8 resin decreased as the pH value rose, especially when the pH value was higher than 6.0. The phenolic molecules are slightly polar and acidic, which makes them sensitive to the pH value. In low pH value solutions, the chlorogenic acids existed as molecules that were easily identified and adsorbed by the resin. In contrast, in relatively high pH solutions, chlorogenic acids may exist as ions due to the ionization reaction, which are more difficult to adsorb. As the adsorption capacities did not change significantly at pH 2.0 and 3.0, pH 3.0 was chosen as the optimum pH value of the sample solution. 2.4.2.3 Effect of Initial Sample TPC on the Adsorption Capacity of AB-8 Resin The crude chlorogenic acid solutions were diluted by distilled water to different TPCs (0.2, 1.0, 1.5, 2.0, and 2.5 mg CAE/mL). About 50 mL of each solution was adsorbed by 2 g of resin and the TPC of the solution was determined when adsorption had finished. The adsorption amounts at different sample concentrations were calculated to detect the effects of sample concentration on the adsorption capacity of the AB-8 resin. The effect of the initial TPC on static adsorption is shown in Fig. 7.4. The adsorption capacity rose as the original TPC increased if the original TPC was less than 2.0 mg CAE/mL. The highest adsorption capacity was observed when the original TPC was 2.0 mg CAE/mL. However, the adsorption capacity decreased if the original TPC was

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375

30

Qe (mg CAE/g)

25 20 15 10 5 0 0.0

0.5

1.0 1.5 2.0 C0 (mg CAE/mL)

2.5

3.0

Figure 7.4 Effect of sample concentration on the adsorption capacity of AB-8 resin.

higher than 2.0 mg CAE/mL. When the sample concentration was low, the adsorption capacity increased as the TPC rose because the number of active sites related to the chlorogenic acids increased. However, with further increases in the TPC, more impurities were adsorbed on the AB8 resin, resulting in competition for active sites between the chlorogenic acids and the impurities, which led to a slight drop in the adsorption capacity. 2.4.2.4 Effect of Ethanol Concentration on the Desorption Ratio of AB-8 Resin After reaching equilibration, the resins were washed twice with 50 mL distilled water to remove impurities and each of the 2 g resin samples were desorbed as described in 2.4.1 using 50-mL ethanol solutions at different concentrations [30%, 50%, 70%, 90%, and 100% (v/v)]. The desorption ratios were calculated to investigate the relationship between desorption and the ethanol concentration. As shown in Fig. 7.5, the desorption ratio increased as the ethanol concentration rose from 30% (v/v) to 70% (v/v), and the highest desorption ratio of 90.9% was observed when the ethanol concentration was 70% (v/v). The desorption ratio did not significantly change when the ethanol concentration continued to increase. Chlorogenic acids do not dissolve in low concentration ethanol solutions, whereas some impurities are desorbed in high ethanol concentrations. Therefore, the 70% (v/v) ethanol solution was selected as the optimum ethanol concentration.

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100

D (%)

90 80 70 60 50 20

40 60 80 Ethanol concentration (%)

100

Figure 7.5 Effect of ethanol concentration on the desorption ratio of AB-8 resin.

2.4.3 Dynamic Adsorption and Desorption of AB-8 resin 2.4.3.1 Effect of Flow and Elution Rates on the Adsorption Capacity and the Desorption Ratio of AB-8 Resin Dynamic adsorption and desorption experiments were carried out in a glass column (diameter of 1 cm, length of 10 cm) wet-packed with pretreated AB-8 resin. The bed volume (BV) of the resin was 10 mL (equal to 5 g resin). Then 50-mL sample solutions (TPC, 2.0 mg CAE/ mL, pH 3.0) were allowed to flow through the glass column at different flow rates (0.5, 1.0, 1.5, 2.0, 2.5, and 3.0 BV/h). The TPCs of the effluent solution were determined, and the resin adsorption amounts were calculated to investigate the relationship between adsorption capacities and flow rate. After the adsorption equilibrium had been reached, the column was first washed with 100 mL distilled water at a flow rate of 1.0 BV/h and then eluted using a 50 mL 70% (v/v) ethanol solution at different flow rates (0.5, 1.0, 1.5, 2.0, 2.5, and 3.0 BV/h). After desorption, the TPC of the effluent solution was determined, and the desorption ratio of the resin was calculated so that the effects of elution rate on the desorption ratio of the resin could be determined. Fig. 7.6 shows that the adsorption capacity was maintained at a high level when the flow rate was lower than 1.0 BV/h, but decreased when the rate was higher than 1.0 BV/h. This might suggest that some of the chlorogenic acids leaked out without being adsorbed by the resin due to the high flow rate. Therefore, 1.0 BV/h was selected as the optimal flow rate because the adsorption capacity did not show any significant differences at rates of 0.5 and 1.0 BV/h, and the production

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30

377

100 Feeding

Eluting

80

20 60 15

D (%)

Q e (mg CAE/g)

25

40 10 20

5 0

0

1

2 Solution rate (BV/h)

3

4

0

Figure 7.6 Adsorption and desorption ratios of AB-8 resin at different flow rates.

efficiency was too low when the flow rate was lower than 1.0 BV/h. A similar regulation was observed between the elution rate and the desorption ratio. The desorption ratio was higher than 90% when the elution rate was less than 1.5 BV/h. However, the desorption ratio decreased when the elution rate was higher than 1.5 BV/h, which indicated that chlorogenic acids could be desorbed more completely at low elution rates. Chlorogenic acids could be dissolved and eluted more thoroughly at low elution rates if ethanol is entering the resin micropores. The desorption ratios at elution rates of 0.5, 1.0, and 1.5 BV/h were not significantly different; therefore, 1.0 BV/h was selected as the optimal elution rate. 2.4.3.2 Dynamic Adsorption and Desorption Properties Under Optimum Conditions Exactly 100 mL of the sample solution (TPC, 2.0 mg CAE/mL, pH 3.0) was allowed to flow through the glass column at a flow rate 1.0 BV/h. The BV was 10 mL. Every 5 mL of eluted solution was collected, and their TPCs were determined. The resin was defined as being at adsorption equilibrium when the TPC of the eluted solution was a tenth of the initial sample solution and the total volume of the eluted solution was at the dynamic handling capacity of the resin. After the adsorption equilibrium had been determined, the column was first washed with 100 mL distilled water and then eluted by 70% (v/v) ethanol solution at a flow rate of 1.0 BV/h. Every 5 mL of eluted solution was collected and their TPCs were determined. The resin was defined as reaching desorption equilibrium when the TPC of the eluted solution was at its lowest level and did not substantially change.

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TPC after adsorption (mg CAE/mL)

0.25

8.0

Dynamic adsorption Dynamic desorption

0.20 6.0 0.15 4.0 0.10 2.0 0.05

0.00

0.0

1.0

2.0 3.0 4.0 Solution volume (BV)

5.0

6.0

TPC after desorption (mg CAE/mL)

378

0.0

Figure 7.7 Dynamic adsorption and desorption properties at the optimal parameters.

A sample solution (TPC, 2.0 mg CAE/mL, pH 3.0) was injected into the AB-8 resin column, with a BV of 10 mL, at a rate of 1.0 BV/h. The dynamic adsorption curve is shown in Fig. 7.7. The TPC of the effluent solution was less than 0.15 mg CAE/mL when the injection volume was less than 3 BV, which indicated that the chlorogenic acids leakage stayed at a low level. When the injection volume was 5 BV, the TPC of the effluent solution was 0.2 mg CAE/mL, which was a tenth of the initial concentration. This indicated that the dynamic adsorption equilibrium had been reached. The 5 BV sample solution of chlorogenic acids from sweet potato leaves could be dynamically adsorbed by AB-8 resin, and the adsorption capacity reached 26.8 mg CAE/g. The saturated resin was eluted by 70% (v/v) ethanol solution at 1.0 BV/h. Fig. 7.7 shows that the elution peak did not possess a tail. The desorbed polyphenols were mainly concentrated in the 0
2.0 BV effluent solutions, which suggested that most of the chlorogenic acids were desorbed by low-concentration ethanol solutions. The desorption equilibrium occurred when the resin was eluted by a 3-BV ethanol solution with a desorption ratio of 90.9%.

2.5 QUALITATIVE AND QUANTITATIVE ANALYSES OF CHLOROGENIC ACIDS FROM SWEET POTATO LEAVES BY HPLC “Yuzi No.7” and “Ximeng No.1” crude chlorogenic acids solutions were dynamically adsorbed and desorbed by an AB-8 macroporous resin column. The eluted solution was collected and concentrated in a

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379

rotary evaporator at 45 C to remove the ethanol and then freeze-dried. From 100 g of “Yuzi No.7” and “Ximeng No.1” sweet potato leaf powders, 5.56 6 0.47 and 6.03 6 0.61 g of purified products were independently obtained, respectively, producing 87.33 6 1.53% and 82.67 6 4.51% of purified chlorogenic acid, respectively. The qualitative and quantitative analyses of the purified chlorogenic acids from sweet potato leaves were carried out using an Agilent1200 series HPLC, which consisted of a Model G1322A degasser, a Model G1311A quat pump, a Model G1329A auto injector, a Model G1316 column oven and a Model C1315D diode array detector. The separation was completed using a ZORBAX Eclips Plus C18 (4.6 3 150 mm, 5 μm). The mobile phase consisted of ultrapure water containing 0.5% (v/v) phosphoric acid (A) and acetonitrile (B). The elution was performed with the following linear gradient: 0
15 min: 20%
65% B; 15
15.1 min: 65%
80% B; and 15.1
20 min: 80% B. The flow rate was 1 mL/min, the injection volume was 20 μL, the column oven temperature was 40  C and the constituents were detected at 326 nm. The standards were accurately weighed, dissolved in methanol to prepare a stock solution (1 mg/mL) and stored in a refrigerator until needed. Each of the standard stock solutions was diluted to 50 μg/mL with 80% (v/v) methanol. Mixed standard solutions (0.5, 1.0, 5.0, 10.0, and 50.0 μg/mL) were also prepared so that a corresponding standard curve could be created. Each peak in the mixed standard chromatogram was identified by comparing the retention time with that of the single standard chromatogram. All of the solutions were filtered through a 0.45 μm membrane and analyzed as described above. The purified products of the chlorogenic acids from sweet potato leaves from the two cultivars (“Yuzi No.7” and “Ximeng No.1”) were accurately weighed and then dissolved in 80% (v/v) methanol to prepare sample solutions (200 μg/mL), which were then analyzed as described above. The purified chlorogenic acids identified by HPLC are shown in Fig. 7.8 and Table 7.5. Chlorogenic acids from sweet potato leaves consisted mainly of seven caffeoylquinic acids and a small amount of caffeic acid. The contents of three di-caffeoylquinic acids were relatively higher in “Yuzi No.7” than in “Ximeng No.1.” The 3,5-O-CQA

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50

5 Yuzi No.7

40

mUA

30 20

4

6

10 1 23

7

0 0

2

4

6

8 Time (min)

10

12

14

16

30 Ximeng No.1

5 20 mUA

3

4

10 1

6

2

7 0 0

2

4

6

8 Time (min)

10

12

14

16

Figure 7.8 The HPLC chromatographs of chlorogenic acids from sweet potato leaves.

Table 7.5 The Constituents of Chlorogenic Acids Purified From Two Sweet Potato Leaf Cultivars (%) Cultivars

5-O-CQA

3-O-CQA

4-O-CQA

4,5-O-CQA

3,5-O-CQA

3,4-O-CQA

3,4,5-OCQA

Yuzi No. 7

2.46 6 0.03

0.87 6 0.00

1.02 6 0.01

20.96 6 0.27

31.39 6 0.26

13.04 6 0.11

2.64 6 0.03

Ximeng No. 1

5.77 6 0.25

1.73 6 0.02

5.69 6 0.01

21.29 6 0.04

19.45 6 0.06

4.72 6 0.00

2.98 6 0.01

Note: The peaks 1
7 in the HPLC chromatography in Fig. 7.8 represent 5-O-CQA, 3-O-CQA, 4-O-CQA, 4,5-O-CQA, 3,5-O-CQA, 3,4-O-CQA, and 3,4,5-O-CQA, respectively. CQA, caffeoylquinic acid.

[31.39% 6 0.26% dry weight (DW)] content was highest. The content of 3,4,5-O-CQA was 2.64% 6 0.03% DW in “Yuzi No.7,” which was lower than in the di-caffeoylquinic acids, but higher than 3-, 4-, and 5-O-CQA, whereas the 3-O-CQA content (0.87% 6 0.00% DW) was the lowest among the caffeoylquinic acids. The 4,5-O-CQA content

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(21.29% 6 0.04% DW) was highest in “Ximeng No.1,” followed by 3,5-O-CQA (19.45% 6 0.06% DW). The 5-O-CQA (5.77% 6 0.25% DW) and 4-O-CQA (5.69% 6 0.01% DW) contents in “Ximeng No.1” were higher than in “Yuzi No.7.” The total content of these eight phenolic constituents in “Yuzi No.7” was 72.74% 6 0.99%, which was lower than the TPC (87.33% 6 1.53%) determined by the Folin
Ciocalteu method. This also occurred with “Ximeng No.1.” It is possible that the detection range of the HPLC was different from that of the Folin
Ciocalteu method. Specifically, some unknown phenolic constituents, indicated by some small absorption peaks in the HPLC chromatograms, were not identified by the HPLC method but were detected by the Folin
Ciocalteu method. In conclusion, AB-8 macroreticular adsorbent resin can adsorb chlorogenic acids from sweet potato leaves rapidly and has good adsorption and desorption properties for chlorogenic acids from sweet potato leaves. The optimum purification parameters are as follows: the TPC of the solution is 2.08 mg CAE/mL, pH value is 3.0, the ethanol desorption solution concentration is 70% (v/v), and the sample injection and elution flow rates are 1 mL/min. The 5-BV sample solution can be dynamically adsorbed by an AB-8 resin column, and the resin column at adsorption equilibrium can be desorbed thoroughly by a 2-BV ethanol solution. More than 5 g of purified polyphenols were obtained from 100 g of sweet potato leaf powder under optimal conditions. The purified polyphenols mainly consisted of caffeoylquinic acids, including three types of di-caffeoylquinic acid, and possessed strong O22 radical scavenging activities and oxygen radical absorbance capacities. The process of purifying chlorogenic acids from sweet potato leaves by AB-8 macroporous resin has the characteristics of a large adsorption capacity, rapid adsorption rate, easy desorption, simple purification process, high continuous degree, and high product purity; therefore, it has the potential to be used in large-scale production.

SECTION 3: BIOLOGICAL ACTIVITIES OF CHLOROGENIC ACIDS FROM SWEET POTATOES Chlorogenic acids possess biological activities, such as those required for free radical-scavenging, antiinflammation, tumor-inhibition, liver-protection, blood-activation, and antihypertensivity, as well as

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in antibacteriocides and cholagogues. The reputed roles of sweet potato leaves, as indicated by their nicknames, such as anticancer vegetables, space food, and longevity greens, are directly related to their abundant chlorogenic acids contents (Islam et al., 2003; Ishida et al., 2000). As mentioned earlier, sweet potato, especially leaves of sweet potato, is rich in chlorogenic acids, of which the three disubstituted caffeoylquinic acids, 3,5-, 4,5-, and 3,4-O-CQA, show the highest content levels (Islam et al., 2002; Wang et al., 2008a,b). Some studies have shown that the free radical-scavenging activities of the chlorogenic acids are positively correlated with the number of caffeoyl groups in the molecules (Iwai et al., 2004). Therefore, the component composition gives the chlorogenic acids in sweet potatoes higher antioxidant, antimicrobial, aldose reductase inhibitory, and anticancer activities.

3.1 IN VITRO ANTIOXIDANT ACTIVITY OF CHLOROGENIC ACIDS FROM SWEET POTATOES 3.1.1 2,2-Diphenyl-1-Picrylhydrazyl (DPPH) Radical-Scavenging Activity of Chlorogenic Acids From Sweet Potatoes DPPH is a relatively stable organic nitrogen-free radical in organic solvents, which has deep purple colors and a strong absorption at 517 nm. When a free radical-scavenging agent is present, the DPPH single electron is paired, presenting a light yellow color, and the absorbance at the maximum wavelength decreases (He et al., 2012). The DPPH radical-scavenging activity of chlorogenic acids from two cultivars, “Yuzi No. 7” and “Ximeng No. 1,” were determined and compared with that of vitamin C. Fig. 7.9 showed that, when the concentration was greater than 15 μg/mL, the DPPH radicalscavenging activity of chlorogenic acids from “Yuzi No. 7” and “Ximeng No. 1” increased with an increasing concentration, and the dose
effect relationship was obvious. At the sample concentration of 15 μg/mL, the DPPH radical-scavenging ratio reached 80%. At the sample concentration of 20 μg/mL, the DPPH radical scavenging ratios of chlorogenic acids from “Yuzi No. 7” and “Ximeng No. 1” reached their highest values, 85.5% and 88.4%, respectively.

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Figure 7.9 DPPH radical-scavenging activity of chlorogenic acids from sweet potato leaves. Note: labeled with a different letter in the same concentration are significantly different (P , .05).

a2b

Values

3.1.2 Hydroxyl Radical-Scavenging Activity of Chlorogenic Acids From Sweet Potatoes Among the currently known reactive oxide species, hydroxyl radicals are free radicals possessing the strongest toxicity and the greatest ability to harm the living body. They can interact with a variety of moleculars in vivo through electron transfer, addition, and dehydrogenation, causing oxidative damage to substances such as sugars, amino acids, proteins, nucleic acids, and lipids (Halliwell et al., 1999). The Fe31/ascorbic acid/EDTA/H2O2 system was used to generate hydroxyl radicals. The hydroxyl radical-scavenging activities of chlorogenic acids in “Yuzi No. 7” and “Ximeng No. 1” were determined and compared with that of 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (Trolox). Fig. 7.10 shows that the activities the samples showed significant dose
effect relationships, and at a concentration of 0.50 mg/ mL, the hydroxyl radical-scavenging activities of “Yuzi No. 7” and “Ximeng No. 1 reached their highest values (80.29% and 76.29%, respectively). At a low concentration (0.01 mg/mL), the hydroxyl radicalscavenging activity of chlorogenic acids from sweet potato leaves was higher than that of Trolox. At concentrations of 0.05 and 0.20 mg/mL, the hydroxyl radical-scavenging activities of chlorogenic acids from sweet potato leaves were equal to that of Trolox.

3.1.3 Superoxide (O22•)-Scavenging Activities of Chlorogenic Acids From Sweet Potatoes

O22• is the free radical in the body having the longest lifetime, and it usually acts as an initiator of the free radical chain reaction, which can

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Figure 7.10 Hydroxyl radical-scavenging activities of chlorogenic acids from sweet potatoes. Note: labeled with a different letter in the same concentration are significantly different (P , 0.05).

Figure 7.11 O22-•-scavenging activities of chlorogenic acids from sweet potato leaves. Note: with a different letter in the same concentration are significantly different (P , .05).

a2b

a2b

Values

Values labeled

produce other free radicals through a series of reactions, causing further harm to the body. Therefore, the O22•-scavenging activity is usually considered an important index for measuring the antioxidant activities of samples (Gryglewski et al., 1986). In our lab, a photochemiluminescence assay was used to determine the O22•-scavenging activities of chlorogenic acids from sweet potato leaves (“Yuzi No. 7” and “Ximeng No. 1”), and the results are shown in Fig. 7.11. Chlorogenic acids from sweet potato leaves showed strong concentration-dependent scavenging activities against O22•. When the sample concentration was 20 μg/mL, the O22•-scavenging activity of

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“Yuzi No.7” was 62.61 μg ACE/mL, which was 3.1 times that of ascorbic acid. At the same concentration, the activity of “Ximeng No.1” was 47.06 μg ACE/mL, which was 2.4 times that of ascorbic acid. The above results indicated that chlorogenic acids possessed high O22•-scavenging activities.

3.1.4 Peroxy Radical-Scavenging Activities of Chlorogenic Acids From Sweet Potatoes Lipid peroxidation is a major cause of many pathological phenomena. Peroxy radicals are the intermediate products of lipid peroxidation, which can attack lipids, causing the lipid peroxidation chain reaction. The oxygen radical absorbance capacity (ORAC) method was used to determine the peroxy radical-scavenging activity of chlorogenic acids from sweet potato leaves, and the results are shown in Fig. 7.12. The ORAC of chlorogenic acids from sweet potato leaves increased as the sample concentrations rose. When the concentration of chlorogenic acids from sweet potato leaves was 20 μg/mL, the activity of “Yuzi No.7” was 55.78 μg Trolox equivalent (TE)/mL, which was significantly higher than the other samples, and was 2.8 times that of Trolox. The activity of “Ximeng No.1” was 43.72 μg TE/mL, which was 2.2 times that of Trolox.

Figure 7.12 Oxygen radical absorbance capacity of chlorogenic acids from sweet potato leaves. Note: labeled with a different letter in the same concentration are significantly different (P , 0.05).

a2b

Values

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3.1.5 Ferric-Reducing Antioxidant Power of Chlorogenic Acids From Sweet Potatoes Ferric-reducing antioxidant power is used to evaluate the total antioxidant capacity of samples, which is based on the colorimetric analysis method of redox reactions. In solutions having low pH values, Fe312,4,6-tri(2-pyridyl)-s-triazine complexes can be reduced to ferrous forms by the reducing substances in the sample. Fe21-2,4,6-tri(2-pyridyl)-s-triazine appears blue and has a maximum absorption at 593 nm (Benzie et al., 1996). In our lab, the ferric-reducing antioxidant powers of chlorogenic acids from sweet potato leaves were determined, and the results are shown in Fig. 7.13. The ferric-reducing antioxidant powers of chlorogenic acids from sweet potato leaves increased in a dose-dependent manner. At the sample concentration of 100 μg/mL, the ferric-reducing antioxidant power of the chlorogenic acids from “Yuzi No. 7” and “Ximeng No. 1” were greatest, at 92.07 and 82.22 μg TE/mL, respectively. The reduction activity of Fe31 is mainly determined by the hydrogen-donating abilities of the samples; the stronger the hydrogen donating ability, the stronger the Fe31-reduction activity (Benzie et al., 1996). Chlorogenic acids from sweet potato leaves possess good hydrogen-donating abilities, and when the concentration was greater than 50 μg/mL, the hydrogen-donating ability was equivalent to that of Trolox.

Figure 7.13 Ferric-reducing antioxidant powers of chlorogenic acids from sweet potato leaves. Note: labeled with a different letter in the same concentration are significantly different (P , .05).

a2b

Values

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387

3.1.6 Lipid Peroxidation-Inhibiting Activities of Chlorogenic Acids From Sweet Potatoes Lipid oxidation has an important influence on food and human health. Lipid oxidation is an important chemical factor causing a decline in food quality, especially foods rich in unsaturated fatty acids, which is very easily oxidated into lipid peroxides. Then, corruption occurs until toxicity emerges after decomposition and polymerization reactions. In living bodies, the occurrence of cancer and the aging of cells are also related to the oxidation of lipids in vivo. Metal ions are important factors that induce lipid oxidation. Therefore, the metal ion-chelating activity is an important index for determining the lipid peroxidationinhibiting activity of a sample (Huang et al., 1996). Huang et al. (2010) studied the inhibitory effects of the aqueous extracts of sweet potato leaves on the lipid-oxidation system induced by the Fe21/H2O2 system. The ferrous ion-chelating activity of the aqueous extract was also determined. At a concentration of 0.1
0.4 mg/mL, the inhibitory effect of the extract on the lipid peroxidation of the system occurred in a dose-dependent manner, with the inhibition rate ranging from 42.22% to 83.36%, and the chelating rate ranging from 4.66% to 14.52%, indicating that the aqueous extracts of sweet potato leaves possess good lipid peroxidationinhibiting activities. After the qualitative analysis of the aqueous extract by HPLC, the main components were found to be chlorogenic acids. Chlorogenic acids from sweet potato leaves had strong antioxidant activities in vitro, which were closely related to the compositions and the molecular structures of the chlorogenic acids. The main chlorogenic acids in sweet potato leaves may be three disubstituted caffeoylquinic acids: 3,5-, 4,5, and 3,4-O-CQA (Islam et al., 2002; Wang et al., 2008a,b). The radical-scavenging activities of chlorogenic acids were positively related to the numbers of caffeoyl groups in the molecules, and the radical-scavenging activities of disubstituted CQAs was 2.0 times that of monosubstituted CQAs and 1.0 to 1.8 times that of ascorbic acid (Iwai et al., 2004). The antioxidant activities of chlorogenic acids were not only related to the numbers of phenolic hydroxyl groups, but also to the electron-donating capabilities of the molecules (Rice-Evans et al., 1995). The phenolic hydroxyl

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groups of chlorogenic acids are good hydrogen donors, and the caffeoyl group possesses a significant electron-donating capability (Medina et al., 2007; Rice-Evans et al., 1996). Chlorogenic acids from sweet potato leaves, especially the disubstituted CQAs have two caffeoyl groups, which is why chlorogenic acids from sweet potato leaves show greater antioxidant activities and possess a developmental value as a natural antioxidant.

3.2 ANTIMICROBIAL ACTIVITIES OF CHLOROGENIC ACIDS FROM SWEET POTATOES Chlorogenic acid has a strong antibacterial action, which can be used as a food preservative, and, when combined with nitrites in food, can reduce the final nitrite content in food. Wang et al. (2010) studied the antibacterial activities of chlorogenic acids from sweet potato leaves, and the minimum inhibitory concentrations of chlorogenic acids from sweet potato leaves towards five common pathogens were determined. Table 7.6 shows that the chlorogenic acids from sweet potato leaves have inhibitory effects on Lactobacillus plantarum, Staphylococcus, Hansenula polymorpha, Escherichia coli, and Staphylococcus aureus, and the minimum inhibitory concentrations were 1000, 1000, 500, 250, and 200 μg/mL, respectively. Chlorogenic acids from sweet potato leaves showed stronger inhibitory effects on E. coli and S. aureus.

Table 7.6 The Experimental Results of the Minimum Inhibitory Concentration of Chlorogenic Acids From Sweet Potato Leaves The Selected Strains

The Concentration of Chlorogenic Acids From Sweet Potato Leaves (μg/mL) 0

50

125

200

250

500

1000

Lactobacillus plantarum

1

1

1

1

1

1

2

Staphylococcus

1

1

1

1

1

1

2

Hansenula polymorpha

1

1

1

1

1

2

2

Escherichia coli

1

1

1

1

2

2

2

Staphylococcus aureus

1

1

1

2

2

2

2

Note: “ 1 ” indicates bacterial growth, while “ 2 ” indicates no bacterial growth.

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3.3 ALDOSE REDUCTASE INHIBITORY ACTIVITIES OF CHLOROGENIC ACIDS FROM SWEET POTATOES Aldose reductase catalyzes the conversion from glucose to sorbitol in mammals, which is the main cause of diabetic complications, such as cataracts and neurological diseases (Nishimura et al., 1991). Aldose reductase inhibitors can effectively inhibit the abnormal increase in the sorbitol content of the organs of diabetic patients, which can be used as active ingredients in the prevention and treatment of diabetes complications. Aldose reductase inhibitors of plant origin consist of flavonoids, phenolic acids and their derivatives, terpenes, and alkaloids, which may have been recognized with the role of inhibition of aldose reductase (Asano et al., 2002). Kurata et al. (2011) isolated a plurality of chlorogenic acid components from sweet potato leaves, and the aldose reductase inhibitory activity of each component was determined and compared with the common aldose reductase inhibitors quercetin and sulfuretin. The results are shown in Table 7.7. Among the chlorogenic acid components, 3,4,5-O-CQA showed the greatest inhibitory effect on aldose reductase, with a IC50 value of 0.25 6 0.011 μmol/L, which was significantly greater than those of quercetin (IC50 5 4.13 6 0.66 μmol/L) and sulfuretin (IC50 5 1.08 6 0.13 μmol/L). The IC50 values of the disubstituted CQAs 3,4-CQA and 3,5-O-CQA were 0.81 6 0.10 and 0.86 6 0.03 μmol/L, respectively, which were significantly greater than that of sulfuretin. The IC50 value of the inhibitory effect of 4,5-O-CQA on aldose reductase was 1.15 6 0.06 μmol/L, which was similar to that of sulfuretin. The IC50 value of the inhibitory effect of 5-O-CQA on aldose reductase was 4.55 6 0.11 μmol/L, which was similar to that of Table 7.7 The Aldose Reductase Inhibitory Activities of Chlorogenic Acids From Sweet Potato Leaves (Kurata et al., 2011) Components

IC50 (μmol/L)

5-O-caffeoylquinic acid

4.55 6 0.113

3,4-O-caffeoylquinic acid

0.81 6 0.096

3,5-O-caffeoylquinic acid

0.86 6 0.033

4,5-O-caffeoylquinic acid

1.15 6 0.063

3,4,5-O-caffeoylquinic acid

0.25 6 0.011

Quercetin

4.13 6 0.657

Sulfuretin

1.08 6 0.127

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quercetin. Thus, the aldose reductase inhibitory activities of chlorogenic acids from sweet potato leaves were related to the number of coffee anhydrides in the molecules; the greater the number of coffee anhydrides, the stronger the aldose reductase inhibitory activity.

3.4 ANTICANCER ACTIVITIES OF CHLOROGENIC ACIDS FROM SWEET POTATOES The anticancer activities of chlorogenic acids, which have significant inhibitory effects on colorectal, liver and laryngeal cancers, have been widely confirmed and are considered to be effective chemical protective agents against cancer. Chlorogenic acids can inhibit variation in carcinogens, such as aflatoxin B and benzopyrene, by inhibiting kinases, and they also can achieve anticancer effects by reducing the utilization rate of carcinogens and their transportation to the liver (SangUn et al., 2000). The single- and double-substituted CQAs in chlorogenic acids can inhibit the secretion of histamine, thus inhibiting the activity of HIV synthase, which then inhibits cancer cell reproduction (Mahmood et al., 1993). 3,4,5-O-CQA and 4,5-O-CQA can inhibit the proliferation of liver cancer cells. In addition, 3,4,5-O-CQA can significantly and selectively inhibit HIV activity (Matsui et al., 2004; Maruta et al., 1995). Kurata et al. (2007) reported that chlorogenic acids from sweet potato leaves had inhibitory activities on the proliferation of cancer cells, including those of human gastric cancer, colon cancer and promyelocytic leukemia. The inhibitory effects of three double-substituted CQAs isolated from sweet potato leaves on the proliferation of cancer cells was dose dependent. The sensitivities of cancer cells to different chlorogenic acid components were different. 3,4,5-O-CQA showed significant inhibitory effects on the proliferation of three types of cancer cells. Additionally, the mechanism of the 3,4,5-O-CQA inhibitory effect on the proliferation of cancer cells was related to promoting cell apoptosis, indicating that 3,4,5-O-CQA from sweet potato leaves had the potential to prevent cancer. Huang et al. (2004) reported that the water extracted from sweet potato leaves (mainly chlorogenic acids) could effectively inhibit the proliferation of human lymphoma NB4 cell, with an IC50 value of 449.6 6 27.73 μg/mL. Chlorogenic acids from sweet potatoes can effectively inhibit the proliferation of cancer cells, which is associated with the high content of double- and triplesubstituted CQAs.

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3.5 OTHER BIOLOGICAL ACTIVITIES OF CHLOROGENIC ACIDS FROM SWEET POTATOES Chlorogenic acids from sweet potatoes can increase the activities of superoxide dismutase in the sera and skin tissue homogenates of mice. Superoxide dismutase is a bioactive substance that exists in animal sera and is the most direct antioxidase in blood. It is considered to be the most valuable material in free radical-scavenging, antitumor and antiaging processes, immunity enhancing processes, and inflammation inhibition (McCord et al., 1969). Thus, chlorogenic acids from sweet potatoes have a variety of potential physiological activities. Sasaki et al. (2013) studied the effects of chlorogenic acids from purple sweet potatoes on the spatial learning and memory of rapidly aging mice. The expression levels of antioxidant-activated proteins, energy metabolism proteins and neural plasticity-related proteins in mice brains were significantly higher than those in the control group. Thus, chlorogenic acids from purple sweet potatoes have neuroprotective activities in mice brains, and can improve the spatial learning ability and memory of rapidly aging mice. In conclusion, chlorogenic acids from sweet potatoes are important in many biological processes, having in vitro antioxidant, antibacterial, antiinflammatory, and anticancer activities, as well as roles in preventing and treating cardiovascular disease, improving superoxide dismutase activity, and promoting the expression of useful proteins in the body. The above biological activities indicate that the chlorogenic acids from sweet potatoes have great development and application values. However, at present, there is not enough research into the biological activities of chlorogenic acids from sweet potatoes, and the mechanisms of various biological activities are not well enough understood. Thus, an adequate theoretical basis for the development and application of sweet potato chlorogenic acids does not exist. In addition, there are far more biological activities of chlorogenic acids than those presented here; therefore, the biological activities of chlorogenic acids from sweet potatoes still need to be explored.

SECTION 4: THE STABILITY OF CHLOROGENIC ACIDS FROM SWEET POTATOES Chlorogenic acids from sweet potatoes are generally sensitive to adverse environmental conditions, including unfavorable temperatures,

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light conditions, and pH levels, and they are, thus, susceptible to degradative reactions during product processing and storage. In Section 2, we introduced that the polyphenols from sweet potato leaves are mainly composed of chlorogenic acids. Therefore, in this section, the effects of pH, heat, and light treatments on the TPC and antioxidant activities of chlorogenic acids from sweet potato leaves are introduced to determine the stability of chlorogenic acids from sweet potato leaves, and provide a theoretical basis for the application of chlorogenic acids from sweet potato leaves in food, medicine, and other fields.

4.1 EFFECT OF pH ON CHLOROGENIC ACIDS FROM SWEET POTATO LEAVES Sweet potato leaf polyphenols from “Jishu No. 04150” and “Shangshu No. 19” purified by AB-8 macroreticular resin according to the method described in Section 2 were used as test samples. The TPC of sweet potato leaf polyphenols from “Jishu No. 04150” and “Shangshu No. 19” were 84.63% 6 3.07% and 83.15% 6 2.22%, respectively. Determining the effects of different pH solvent systems on the stability of chlorogenic acids from sweet potato leaves was carried out using the method reported by Majo et al. (2011). Briefly, phosphate buffered solutions of different pH values (3.0, 5.0, 7.0, and 8.0) were prepared with disodium hydrogen phosphate and citric acid. Chlorogenic acids from sweet potato leaves were dissolved in different pH buffer solutions, resulting in 1.0 mg/mL sample solutions. Then, the TPCs and antioxidant activities of sample solutions were determined using the Folin
Ciocalteu and ORAC methods, respectively. The effects of pH values on the TPCs and antioxidant activities of chlorogenic acids from sweet potato leaves is shown in Fig. 7.14. For “Jishu No. 04150,” there was no significant difference among the TPCs in pH 3, 5, and 7 solvent systems, but the TPC of the pH 8-sample solution was significantly lower than those of the other samples. For “Shangshu No. 19,” none of the TPCs of the samples in different pH-solvent systems showed any significant difference. For “Jishu No. 04150,” the antioxidant activity in the pH 7-solvent system was 2.71 mg TE/mL, which was significantly greater than those of the other solvent systems, followed by that of the pH 5-solvent system (2.13 mg TE/mL). The chlorogenic acids in the pH 8-solvent system

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Figure 7.14 Effects of different pH values on the TPCs and antioxidant activities of chlorogenic acids from sweet potato leaves. Values were means 6 SD of three determinations. Data on the same broken lines that were not significantly different are represented by the same letter (P , .05).

(1.65 mg TE/mL) showed the lowest antioxidant activity level. For “Shangshu No. 19,” the chlorogenic acids in the pH 7-solvent system also showed the greatest antioxidant activity (1.89 mg TE/mL), and there were no significant differences among the values of the pH 3-, 5-, and 7-solvent systems. Chlorogenic acids in the pH 8-solvent solution showed the lowest antioxidant activity (1.43 mg TE/mL). Thus, the TPCs and antioxidant activities of chlorogenic acids from sweet potato leaves were greater in neutral- and weak acid-solvent systems, and the optimum pH range was 5.0
7.0.

4.2 EFFECTS OF HEAT TREATMENTS ON CHLOROGENIC ACIDS FROM SWEET POTATO LEAVES The effects of heat treatments on the stability of chlorogenic acids from sweet potato leaves were determined following the method reported by Lee et al. (2006). Briefly, 1.0 mg/mL sample solutions were prepared by dissolving chlorogenic acids from “Jishu No. 04150” and “Shangshu No. 19” in distilled water. Then, the sample solutions were incubated in 50, 65, 80, and 100 C water baths. The TPCs and antioxidant activities were determined after 0, 10, 30, 60, and 90 min, and the retention rates of TPCs and antioxidant activities were calculated according to the following formulas: R1 ð%Þ 5

C1 3 100%; C0

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where R1 is the retention rate of TPC, C0 is the TPC before heat treatment (mg CAE/mL), and C1 is the TPC after heat treatment (mg CAE/mL). R2 ð%Þ 5

A1 3 100%; A0

where R2 is the retention rate of antioxidant activity, A0 is the antioxidant activity before heat treatment (mg TE/mL), and A1 is the antioxidant activity after heat treatment (mg TE/mL). The effects of heat treatment on the TPCs and antioxidant activities of chlorogenic acids from sweet potato leaves are shown in Fig. 7.15. For “Jishu No. 04150,” the retention rates of TPCs from samples treated at 50, 65, 80, and 100 C for 90 min were all greater than 91%, which indicated that heat treatments had little effects on the TPCs of the samples. The antioxidant activities of samples treated at 50 and 60 C did not show significant differences, and after heat treatments of 90 min, the retention rates of the antioxidant activities were 81.33% and 94.55%, respectively. During the heat treatment processes at 80 and 100 C, the retention rates of antioxidant activities in the samples decreased significantly, and after 90 min, the retention rates of the antioxidant activities were 62.14% and 61.86%, respectively. For the same treatment time, a 100 C-heat treatment showed the greatest effect on the antioxidant activities of the samples, and the retention rate of the antioxidant activity was significantly lower than those of other treatment temperatures, indicating that heat treatments at lower temperatures had little effect on the antioxidant activities of chlorogenic acids from sweet potato leaves and that heat treatments at high temperatures could induce a sharp decrease in antioxidant activity. For “Shangshu No. 19,” the retention rates of the TPCs of the samples at all of the heat-treatment temperatures were higher than 91%. The heat treatment at 65 C showed little effect on the antioxidant activities of the samples, and after treatment for 90 min, the retention rate of the antioxidant activity was 100.15%. During the heattreatment process at 80 and 100 C, the retention rates of the antioxidant activities of the samples decreased significantly, and after 90 min, the retention rates of the antioxidant activities were 70.65% and 65.25%, respectively. For the same treatment time, 80- and 100 C-heat treatments showed greater effects on antioxidant activities, and after

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Figure 7.15 Effects of different heat treatments on the retention rates (%) of the total polyphenol contents and antioxidant activities of chlorogenic acids from “Jishu No. 04150” at 50 C (A), 65 C (B), 80 C (C), and 100 C (D), and from “Shangshu No. 19” at 50 C (E), 65 C (F), 80 C (G), and 100 C (H). Values were means 6 SD of three determinations. Data on the same broken lines that were not significantly different are represented by the same letter (P , .05).

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30 min, the retention rates of the antioxidant activities were significantly lower than those of other treatment temperatures. Thus, heat treatments at lower temperatures had limited effects on the TPCs and antioxidant activities of chlorogenic acids from sweet potato leaves, and heat treatments at high temperatures had greater effects on the antioxidant activities.

4.3 EFFECTS OF LIGHT ON CHLOROGENIC ACIDS FROM SWEET POTATO LEAVES The effects of light on the stability of chlorogenic acids from sweet potato leaves were determined according to the method reported by Wang et al. (2009a,b). Briefly, 1.0 mg/mL sample solutions were prepared by dissolving chlorogenic acids from “Jishu No. 04150” and “Shangshu No. 19” in distilled water. A set of sample solutions was maintained in a location receiving direct sunlight from 10:00 am to 3:00 pm, and another set of sample solutions was covered with tin foil and kept in the same place. The TPCs and antioxidant activities of the sample solutions were determined every hour, and the retention rates of the TPCs and antioxidant activities were calculated according to the following formulas: R1 ð%Þ 5

C1 3 100%; C0

where R1 is the retention rate of the TPC, C0 is the TPC before light treatment (mg CAE/mL), and C1 is the TPC after light treatment (mg CAE/mL). R2 ð%Þ 5

A1 3 100%; A0

where R2 is the retention rate of antioxidant activity, A0 is the antioxidant activity before light treatment (mg TE/mL), and A1 is the antioxidant activity after light treatment (mg TE/mL). The effects of light on the TPCs and antioxidant activities of chlorogenic acids from sweet potato leaves are shown in Fig. 7.16. The TPCs of chlorogenic acids from “Jishu No. 04150” and “Shangshu No. 19” did not show significant changes under light treatments within 5 h, and there were no significant differences between the light treatment and dark groups within the same treatment time. After 5 h, the retention

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Figure 7.16 Effects of different light treatments on the retention rates (%) of total polyphenol contents of sweet potato leaf polyphenols from “Jishu No. 04150” (A) and “Shangshu No. 19” (B), and the effects of different light treatments on the retention rates (%) of antioxidant activities of sweet potato leaf polyphenols from “Jishu No. 04150” (C) and “Shangshu No.19” (D). Values were means 6 SD of three determinations. Data on the same broken lines that were not significantly different are represented by same letter (P , .05).

rates of the TPCs of “Jishu No. 04150” and “Shangshu No. 19” with the light treatment were 97.55% and 94.87%, respectively. For “Jishu No. 04150,” the antioxidant activity did not change significantly under light treatment within 5 h, and there was no significant difference between the two treatment groups. After 5 h, the retention rate of the antioxidant activity with the light treatment was 92.56%. For Shangshu No. 19, although the antioxidant activity showed a significant decrease during the first 2 h under the light treatment, there were no significant changes after 2 h, and the light-treatment group was not significantly different from the dark group. After 5 h, the retention rate of the antioxidant activity was 91.43%. Thus, within the testing time, light had a limited effect on the TPCs and antioxidant activities of chlorogenic acids from sweet potato leaves. Chlorogenic acids are esterifications of caffeic acid and quinic acid that can be hydrolyzed into caffeic acid and quinic acid under both acidic and alkaline conditions. Caffeic acid is a small phenolic acid, thus its hydrolyzation does not induce significant changes in the TPCs of sample solutions. However, there is a definite positive correlation

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between the antioxidant activity and the number of caffeoyl groups in the molecular structures of the chlorogenic acids (Iwai et al., 2004). The hydrolyzation of chlorogenic acids under alkaline and strong acidic conditions induces a decrease in the number of caffeoyl groups, and further decreases the antioxidant activities of the molecules. In the present study, we corroborated these results, finding that chlorogenic acids from sweet potato leaves in a pH 8-solvent system showed lower antioxidant activity levels, while those in neutral and weak acid systems showed higher antioxidant activity levels. Some studies reported that the orthophthalic carboxylic structures of chlorogenic acids were very easily resolved under high temperature heat treatments, which further decreased the antioxidant activities. In the present study, the antioxidant activities of tested samples after a 100 C-heat treatment for 90 min decreased significantly, suggesting that during the process of purifying chlorogenic acids from sweet potato leaves, high temperatures and long heat-treatment periods should be avoided. In conclusion, chlorogenic acids from sweet potato leaves possessed high antioxidant activities and processing stability, giving them the potential to be a new type of natural antioxidant.

SECTION 5: THE APPLICATIONS OF CHLOROGENIC ACIDS FROM SWEET POTATOES 5.1 FOOD INDUSTRY Chlorogenic acids are a new type of high-efficiency phenolic antioxidant that can fully or partially replace the commonly used synthetic antioxidants in some foods. Chlorogenic acids can greatly improve the stability of fruit juices, such as strawberry juice. Adding a small amount of chlorogenic acids can increase the oxidation resistance of lard and prolong the storage time. Chlorogenic acids can efficiently maintain the sensory and nutritional qualities of grapes during the storage period. Thus, chlorogenic acids can be used in the field of food processing and storage as antioxidants and preservatives.

5.2 MEDICINE AND HEALTH-PROTECTION INDUSTRY The “Drug standards” issued by the Ministry of Health of the People’s Republic of China includes more than 170 kinds of patented Chinese

Chlorogenic Acids From Sweet Potato

399

medicines, having actions related to clearing heat and removing toxins, and antibacterial and antiinflammatory properties, all of which contain chlorogenic acids. At present, chlorogenic acids are important indices of quality control in the medical production of Yinhuang and Shuanghuanglian preparations. At present, the antibacterial and antiinflammatory properties of chlorogenic acids have been well researched and developed in the pharmaceutical industry, while biological activities, such as tumor inhibition, lowering blood pressure and blood glucose lowering, and cardiovascular protection, have not been thoroughly developed or applied in the pharmaceutical industry. Therefore, there are still broad application prospects for chlorogenic acids in the development of medicines and health products with high performance levels and high added values.

5.3 DAILY CHEMICAL INDUSTRY Chlorogenic acids can promote the synthesis and decomposition of collagen in human skin, bones, and muscles, which can promote metabolism and prevent aging. Japan has made use of the antioxidant properties of chlorogenic acids to develop antiaging skin care products. Chlorogenic acids in the UV region have strong absorption abilities, which can reduce UV damage to human skin. In addition, chlorogenic acids can inhibit the activities of tyrosinase and catalase, and scavenge reactive oxygen species in skin cells. These properties can be applied to beauty products. Cosmetics containing chlorogenic acids can effectively dissipate blood stasis, and remove toxins from the body by promoting blood circulation, decreasing acne and spots on human skin. Additionally, chlorogenic acids can activate enzymes in cells, maintain the physiological activity of epidermal cells, and promote cell metabolism, resulting in skin lightening. At present, research on the applications of chlorogenic acids in the daily chemical industry in China is relatively limited, and the gap with the developed countries is relatively large. China has a large population, and the demand for daily chemical products is great. Thus there is a large market demand for the development of daily chemical products containing chlorogenic acids. Research on chlorogenic acids has grown at home and abroad, and with its increase, applications are becoming more extensive. However,

400

Sweet Potato Processing Technology

there is still room for improvement in some areas, such as: (1) Developing new raw materials for chlorogenic acids: chlorogenic acids could be extracted from agricultural by-products and food processing waste water and residues. This comprehensive utilization of waste materials is of great environmental significance; (2) In-depth studies on the separation, purification, and the structure
activity relationships of chlorogenic acids: chlorogenic acids are composed of a variety of components. At present, the separation and purification processes of individual components are limited, and the differences in activity levels among different components needs further study. Additionally, the mechanisms responsible for chlorogenic acid activities, especially the in vivo mechanisms are still not clear, and, as a result, there is no reliable theoretical support for the utilization of chlorogenic acid activities; and (3) Expanding the application range, and improving the application depth: Chlorogenic acids have a variety of biological activities. At present, the applications of their activities in food, medicine, and daily chemical fields are mainly focused on free radical-scavenging, antisepsis, and antiinflammation. The development and application of activities, such as anticancer, cardiovascular protection, and diabetes prevention, needs to be expanded. Thus, there is still a lot of work to be done in the study of chlorogenic acids.

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956. Riksen, N.P., Rongen, G.A., Smit, P., 2009. Acute and long-term cardiovascular effects of coffee: implications for coronary heart disease. Pharmacol. Therap. 121 (2), 185
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Sasaki, K., Han, J., Shimozono, H., et al., 2013. Caffeoylquinic acid-rich purple sweet potato extract, with or without anthocyanin, imparts neuroprotection and contribute to the improvement of spatial learning and memory of SAMP8 mouse. J. Agric. Food Chem. 61 (21), 5037
5045. Sato, Y., Itagaki, S., Kurokawa, T., et al., 2011. In vitro and in vivo antioxidant properties of chlorogenic acid and caffeic acid. Int. J. Pharm. 403 (1), 136
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684. Wang, Z., Michael, N.C., 2008b. Profiling the chlorogenic acids of sweet potato (Ipomoea batatas) from China. Food Chem. 106 (1), 147
152.

FURTHER READING Ardestani, A., Yazdanparast, R., 2007. Antioxidant and free radical scavenging potential of Achilleasantolina extracts. Food Chem. 104 (1), 21
29. Galvan d’Alessandro, L., Kriaa, K., Nikov, I., et al., 2012. Ultrasound assisted extraction of polyphenols from black chokeberry. Sep. Purif. Technol. 93, 42
47. Guifeng, W., Liping, S., Yudan, R., et al., 2009. Anti-hepatitis B virus activity of chlorogenic acid, quinic acid and caffeic acid in vivo and in vitro. Antiviral Res. 83 (2), 186
190. Maqsood, S., Benjakul, S., 2010. Comparative studies of four different phenolic compounds on in vitro antioxidative activity and the preventive effect on lipid oxidation of fish oil emulsion and fish mince. Food Chem. 119 (1), 123
132. Prior, R.L., Hoang, H., Gu, L., et al., 2003. Assays for hydrophilic and lipophilic antioxidant capacity (oxygen radical absorbance capacity (ORACFL)) of plasma and other biological and food samples. J. Agric. Food Chem. 51 (11), 3273
3279. Rumbaoa, R.G.O., Cornago, D.F., Geronimo, I.M., 2009. Phenolic content and antioxidant capacity of Philippine sweet potato (Ipomoea batatas L) varieties. Food Chem. 113 (4), 1133
1138. Upadhyay, R., Ramalakshmi, K., Jagan Mohan Rao, L., 2012. Microwave-assisted extraction of chlorogenic acids from green coffee beans. Food Chem. 130 (1), 184
188. Zhang, B., Yang, R., Zhao, Y., et al., 2008. Separation of chlorogenic acid from honeysuckle crude extracts by macroporous resins. J. Chromatogr. B 867 (2), 253
258. ´ ´ Zielinski, H., Zielinska, D., Kostyra, H., 2012. Antioxidant capacity of a new crispy type food products determined by updated analytical strategies. Food Chem. 130 (4), 1098
1104.

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APPENDIX

1

Production Line of Sweet Potato Protein

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APPENDIX

2

Production Line of Sweet Potato Dietary Fiber

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APPENDIX

3

Production Line of Sweet Potato Pectin

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INDEX Note: Page numbers followed by “f ” and “t” refer to figures and tables, respectively.

A AB-8 macroreticular adsorbent resin, 370, 381 purification of chlorogenic acid, 371
378 AB-8 resin macroporous resins, 309 macroreticular resin, 392 pH effects on adsorption rate, 309 static adsorption of sweet potato anthocyanins, 309 Abrasive peeling, 268 Absorption of cholesterol, 160 of Na1, K1, Ca21, Fe31, and Pb21, 160
161 saturation, 310
311 ACE. See Angiotensin converting enzyme (ACE) Acetaldehyde, 331 Acetic acid ester starch, 21
22 Acetone, 126
127 Acetyl coenzyme A, 130 Acid extraction, 139, 192 Acid precipitation methods, 55 Acidified ethanol, 305
306 Acidity regulator, 63
64 Active peptides, raw material of, 114 Acute ALD, sweet potato anthocyanin on, 324
334 on alcoholic intoxication, 326 experimental model for, 324
326 on growth and liver index, 326
327 on hepatic ADH activity level, 332
333 on hepatic MDA content, 330 on hepatic SOD and GST, 331
332 on serum ALT, AST, and LDH activities, 327
329 on serum TG, TCH, and LDL-C activity levels, 329
330 3-O-(6-O-(E)-caffeyl-2-O-(6-O-Acyl-β-Dglucopyranosyl)-β-D-glucopyranoside)5-O-β-D-gluco-pyranosides, 290
291 Acylated anthocyanins, 312 Additives on quality of sweet potato starch noodles and vermicelli, 20
21

ADH, 332
333, 340 sweet potato anthocyanins effects on hepatic ADH activity level, 332
333, 340
341 Adiponectin, 144
145, 363
364 concentration of plasma, 144
146 plasma, 154
155 Adipose cell metabolic disorders, 140 Adsorption equilibrium, 372
373 ratios of AB-8 resin, 309 effects of pH on adsorption rate of AB-8 resin static adsorption of sweet potato anthocyanins on AB-8 resin, 309 Adsorptive bubble separation. See Foam separation method Aggregation, 61 Agro-products, 266 AI. See Atherosclerosis index (AI) Air-flow rate, 68
69 Akt phosphorylation, 362 Alanine aminotransferase (ALT), 295 sweet potato anthocyanins effect on, 327
329, 328f, 337
338, 337f ALC. See Alcoholic liver cirrhosis (ALC) Alcohol, 324, 335. See also Dealcoholic effects, sweet potato anthocyanin on alcohol-soluble protein, 58 alcoholic hepatitis, 324 alcoholic intoxication, sweet potato anthocyanins, 326 hepatic fibrosis, 324 Alcoholic liver cirrhosis (ALC), 324 Alcoholic liver damage (ALD), 324. See also Acute ALD, sweet potato anthocyanin on Aldose reductase inhibitors, 364
365 activities of chlorogenic acids, 389t, 390 Alkali extraction method, 139 Alkaline environment, 363 peeling, 268 α-amylase, 127
128 α-glucosidase inhibitor, 293
294

412

Index

α-L-Araf, 189f [(1,2)-α-L-rhap-(1,4)-α-D-GalpA], 188
189 ALT. See Alanine aminotransferase (ALT) Alum, 19
21 American Cereal Chemists Association, 124 Amino acid, 276
277 sequences of sporamin, 54t Amino-ethyl phenol (APAP), 295 Ammonium bicarbonate, 273 Ammonium sulfate, 61
62, 307 ethanol
ammonium sulfate, 288
289 treatment, 60 50 AMP-activated protein kinase, 363
364 Amylase, 126
127 Amylopectin, 4
6 structure of, 5
6, 5f sweet potato, 11 Amylose, 4
6, 12
13 content, 7
8 structure of, 4
5 Analysis of variance (ANOVA), 276
277, 302
303 of anthocyanin partition coefficient test, 303t yield test, 302t PSPAs for extraction yield of, 300
302 for partition coefficient of, 302
303 of quadratic model, 300 Angiotensin converting enzyme (ACE), 294 Animal experimental study on losing weight and reducing lipid levels, 91
92 of obesity prevention, 91 Animal sources, 125 ANOVA. See Analysis of variance (ANOVA) Anthocyanin(s), 335 applications of sweet potato anthocyanins, 343
347 biological activity of sweet potato anthocyanin, 324
343 copigmentation effect, 284
285 degradation, 283, 284f interactions, 285f structure, 282
283, 282f, 282t sweet potato, 285
297 bioactivity of, 292
297 effects of pH levels on thermal stability of anthocyanins, 314
316, 315f extraction method of purple sweet potato, 286
290 high pressure effects on thermal stability of anthocyanins, 313
314, 314f

preparation, 297
308 purification methods for, 290, 291t stabilities of anthocyanins from, 312
323 structures, 290
292, 292f temperature effects on thermal stabilities of anthocyanins, 312
313, 313f thermal degradation, 316
323 Antiatherosclerosis, 294 Antibacterial, 296, 388. See also Chlorogenic acids from sweet potato Anticancer activities of chlorogenic acids, 390 inhibitory effects of sweet potato protein, 86
88 on cancer cell proliferation, 86 on in vitro migration and invasion of cancer cells, 87 on tumor metastasis, 87
88 Antihyperglycemia, 293
294 Antihypertensive activities, 363
364 Antiinflammatory effects, 365 Antimicrobial activity of chlorogenic acids, 362, 388, 388t Antimutagenic effects, 363 Antimutation and anticancer activities, 362
363 Antioxidant(s), 80, 292
293 activity of chlorogenic acids, 362, 364, 382
388 DPPH radical-scavenging activity, 382, 383f ferric-reducing antioxidant power, 386, 386f hydroxyl radical-scavenging activity, 383, 384f lipid peroxidation-inhibiting activities of, 387
388 peroxy radical-scavenging activities, 385, 385f superoxide-scavenging activities, 383
385, 384f activity of sweet potato protein, 80
83, 83f DPPH radical scavenging activity, 82, 83f hydroxyl radical scavenging activity, 81
82 linoleic acid peroxidation-inhibiting effect, 82
83 superoxide anion radical scavenging activity, 80
81 Antioxidases, 80 AOAC. See Association of Official Analytical Chemists (AOAC)

Index

APAP. See Amino-ethyl phenol (APAP) Apoptosis, 362
363 Aqueous two-phase extraction (ATPE), 288
289, 297 Aqueous two-phase system (ATPS), 288, 308, 308t Aryl hydrocarbon hydroxylase, 363 Aspartate aminotransferase (AST), 295 sweet potato anthocyanins, 327
329, 328f, 337
338, 337f Association of Official Analytical Chemists (AOAC), 124 AOAC-SPDF, 136
137, 137t AST. See Aspartate aminotransferase (AST) Atherosclerosis index (AI), 294 ATPE. See Aqueous two-phase extraction (ATPE) ATPS. See Aqueous two-phase system (ATPS)

B Baking biscuits with sweet potato granules, 274 sweet potato granules in bread, 271 Bcap-37 breast cancer cells, 208, 226 effects of pectin concentrations on, 212f effects of thermal-modified sweet potato pectin on, 216
217, 216f effects of unmodified pectin on adhesive capabilities of, 215f inhibitory effects of thermal-modified pectin on, 221f inhibitory effects of unmodified pectin on, 220f in McCoy’s 5 A culture medium, 209 effect of pH-modified pectin on, 217
218, 217f in PMA-treated group, 216
217 sweet potato pectin effects on cancer cell proliferation, 211
213 Bed volume (BV), 376
377, 381 Bend-to-break rate, 275
276 β-1,4-glucosidic bands, 124
125 β-D-Galp, 189f β-D-GalpA, 189f β-mercaptoethanol, 58, 79 β-sheet structures, 53
54 β-turn structures, 53
54 Beverages sweet potato dietary fiber in, 174 sweet potato pectin, 256
257

413

Bifidobacterium, 128
129 Biological activities of sweet potato anthocyanins, 296
297 on acute ALD and dealcoholic effects, 324
334 antiatherosclerosis, 294 antibacterial, 296 antihyperglycemia and hypertension, 293
294 antioxidant, 292
293 enhanced memory, 295
296 liver protection, 294
295 on subacute ALD, 334
343 of sweet potato pectin, 208
226 cell cultures, 209 effects on cancer cell metastasis, 214
226 effects on cancer cell migration, 218
222 effects on cancer cell proliferation, 210
214 effects on cancer cell survival rates, 209
210, 209t effects on uPA, 223
226 pH-modified pectin, preparation of, 208 thermal-modified pectin, preparation of, 209 of sweet potato protein, 55
56, 80 Biological medicine, 115 Biological method fermentation method, 138 mixed enzymes method, 138
139 single enzyme method, 135
137 Biscuits, sweet potato granules effects in, 272
275 arrangement and packing, 274 baking, 274 biscuit production, 275 cooling, 274 dough concoction, 274 mixing, 274 procedure of making biscuits, 272
274 recommended ratio of granules, 273
274 rolling, 274 supplemental ingredient treatment, 273 Black spot pathogen, 267 Blood glucose and blood lipid concentrations, 142
143, 142t, 152
153 regulation, 193
194 Blood index blood glucose and blood lipid concentrations, 142
143, 152
153 concentration of plasma adiponectin and GLP-1, 144
146, 145f

414

Index

Blood index (Continued) plasma adiponectin and GLP-1 concentrations, 154
155, 155f plasma inflammatory factor concentrations, 146
147, 146t, 156, 156t plasma leptin and insulin concentrations, 143
144, 144f, 153
154, 154f Blood lipid concentrations, 142
143, 142t, 152
153 levels regulation, 193
194 Bowman-Birk soybean trypsin inhibitors, 85
86 Bread, 255 bread production, 272 sensory qualities, 170
171, 173t phenotypic evaluation of bread, 171t quality evaluation of bread, 172t sweet potato dietary fiber in, 170
174 physicochemical properties of bread, 171, 173t sensory qualities of, 170
171 texture of bread, 171
174, 173t sweet potato granules in, 270
272 activation of dry yeast, 271 baking, 271 cooling and packing, 272 flour mixing, 271 formation, 271 optimization of recipe for, 272 procedure of making, 270
272 proofing, 271 static fermentation, 271 Breakdown viscosity of starch, 13 Browan-Birk-type inhibitors, 83
84 BV. See Bed volume (BV)

C C chain, 5
6 Ca21 effect on viscosity of pectin solution, 228
230 concentrations effects on particle sizes of pectin emulsions, 242
243 on texture parameters of pectin gel, 238
239 absorption of, 160
161 Caffeic acid, 361f, 397
398 3-O-Caffeoylquinic acid, 360 3,4,5-O-Caffeoylquinic acid (3,4,5-O-CQA), 364
365, 389
390 Caffeoylquinic acids, 379
381, 387
388 6-O-Caffeoylsophorose (CS), 293
294 Calcium concentration, 268
269

Cancer, protecting intestines from, 160 Cancer cell, 208 adhesion, 214
216 pH-modified sweet potato pectin effects on, 215
216 thermal-modified sweet potato pectin effects on, 216
218 unmodified sweet potato pectin effect on, 215
216 apoptosis, 195
196 migration, sweet potato pectin effects on, 218
222 pH-modified sweet potato pectin effects on, 221
222 thermal-modified sweet potato pectin effects on, 219
221 unmodified sweet potato pectin effect on, 219 sweet potato pectin effects, 211
213 Bcap-37 breast cancer cell proliferation on cancer cell proliferation, 210
214 effects on uPA, 223
226 HT-29 colon cancer cells, 210
211 SMMC-7721 liver cancer cell proliferation, 213
214 on survival rates, 209
210, 209t Cancer metastasis inhibiting effects on, 195 sweet potato pectin effects on, 214
226 cancer cell adhesion, 214
216 Candied fruit, 255 Carbazole sulfuric acid, 202
203 Carbinol pseudobase, 283, 283f Carbohydrate, 125 metabolism, 129
131 Cardiovascular system, protection, 364 CCFS. See Commercial sweet potato starch made by centrifugation (CCFS) Cecum, organic acid content in, 150, 150t, 159
160, 159t Cellular antioxidant system, 334
335 Cellulose, 124
125 Central nervous system, protection, 364 Centrifugation, 18, 135
136 Centrifugation separation (CFS) method, 14 quality comparison of SLPS and, 36
41 color of sweet potato starches and noodles, 37, 37t cooking quality, 39
40, 39t microstructure, 40
41 retrogradation of starch and noodles, 37
39 textural properties, 39t, 40, 40t

Index

structure and physicochemical properties, 24
36 physicochemical characteristics analysis, 31
36 proximate composition, 24
25 structural analysis, 26
31 Fourier transform infrared spectrometry, 28
29 particle size distributions, 26 pasting properties, 33
34 starch color effects, 31 swelling power and solubility, 32
33 thermal properties, 30
31 XRD, 29
30 CFS method. See Centrifugation separation (CFS) method Chain length, 5
6 Chemical reagent in combination with enzymatic hydrolysis method, 126
127 in combination with enzymatic method, 139
140 Chemical separation method, 126, 139
140 acid and alkali extraction, 139 chemical reagent in combination with enzymatic method, 139
140 water extraction, 139 Chlorogenic acids from sweet potato, 359
369 applications daily chemical industry, 399
400 food industry, 398 medicine and health-protection industry, 398
399 biological activities, 360
365, 381
391 aldose reductase inhibitory activities, 389t, 390 anticancer activities, 390 antimicrobial activity, 362, 388, 388t antimutation and anticancer activities, 362
363 antioxidant activity, 362, 382
388 hypoglycemic, serum lipid-lowering, and antihypertensive activities, 363
364 protecting cardiovascular and central nervous systems, 364 protection against diabetes, 364
365 composition and structure, 360, 361f, 361t extraction methods, 365, 366t qualitative and quantitative analyses, 368
369 HPLC, 368
369 thin-layer chromatography, 368 UV spectrophotometry, 368

415

separation and purification methods, 365
367, 367t stability, 391
398 effects of heat treatments, 393
396, 395f effects of light, 396
398, 397f effect of pH, 392
393, 393f technology to preparation, 370
381 extraction, 371 pretreatment of sweet potato leaves, 371 purification, 371
378 qualitative and quantitative analyses by HPLC, 378
381, 380f, 380t TPC, 371 Cholesterol, 127
128, 363
364 absorption, 160 Cholesterolin, 364 Chymus, 161 Citric acid, 63, 192 CL 9
11 glucose residues/chain. See Shortbranched chain (CL 9
11 glucose residues/chain) CL 12
14 glucose residues/chain. See Shortbranched chains (CL 12
14 glucose residues/chain) CoA carboxylase, 130 Coarse separation method, 126 Cold extraction methods, 286 Cold paste viscosity (CPV), 34 Cold paste viscosity to HPV (CPV/HPV), 34 Color, 31, 31t Color of sweet potato starches and noodles, 37, 37t Colorimetric method, 189
190 Commercial sweet potato starch made by centrifugation (CCFS), 24
25, 25t, 26t, 31
32 particle size distributions, 26 pasting properties, 33
34 polarizing microscopy, 27
28 starch color effects, 31 sweet potato starch production, 26
27, 27f swelling power and solubility, 32
33 thermal properties, 30
31 XRD, 29
30 Commercial sweet potato starch made by sour liquid processing (CSLPS), 24
25, 25t, 26t particle size distributions, 26 pasting properties, 33
34 polarizing microscopy, 27
28 starch color effects, 31 sweet potato starch production, 26
27, 27f

416

Index

Commercial sweet potato starch made by sour liquid processing (CSLPS) (Continued) swelling power and solubility, 32
33 thermal properties, 30
31 XRD, 29
30 Concentration, foam separation method, 66 Condiments, sweet potato dietary fiber in, 175 Constipation, 161 Cooking quality, 39
40, 39t Cooling biscuits with sweet potato granules, 274 sweet potato granules in bread, 272 temperature, 77 Copigmentation effect, 284
285 Cosmetics, 399 industry, 346
347 Coumaric acid, 361f CPV. See Cold paste viscosity (CPV) 3,4,5-O-CQA. See 3,4,5-O-Caffeoylquinic acid (3,4,5-O-CQA) 4,5-O-CQA, 379
381 5-O-CQA, 379
381, 389
390 C-reactive protein (CRP), 146
147 Crops, 359 CRP. See C-reactive protein (CRP) Crude chlorogenic acid solutions, 373
374 Crystal region, 6
7 CS. See 6-O-Caffeoylsophorose (CS) CSLPS. See Commercial sweet potato starch made by sour liquid processing (CSLPS) Cultivars, sweet potato, 266

D Daily chemical industry, 399
400 DEAE-52 ion-exchange chromatography, 79 Dealcoholic effects, sweet potato anthocyanin on, 324
334 on alcoholic intoxication, 326 experimental model for, 324
326 on growth and liver index, 326
327, 327t on hepatic ADH activity level, 332
333 on hepatic MDA content, 330 on hepatic SOD and GST, 331
332 on histopathological analysis, 334, 334t on serum ALT, AST, and LDH activities, 327
329 on serum TG, TCH, and LDL-C activity levels, 329
330 Degree of esterification, 190 Degree of number-average DP (DPn), 4
5 Degree of polymerization (DP), 4
5 Degree of weight-average DP (DPw), 4
5

Deionized water, degradation kinetic parameters of anthocyanins in, 318
319, 318t Denaturation, 75 time, 76
77 Desertification, 359 Design Expert software, 206 Desorbed polyphenols, 378 Desorption AB-8 resin effects of ethanol on desorption of, 310 effect of ethanol concentration on, 375, 376f curve of anthocyanins from resin, 311
312 performance, 372
373 ratio, 373 Di-caffeoylquinic acid, 379
381 Diabetes. See also Insulin aldose reductase, 389 protection against, 364
365 Dialysis, 192
193 Dietary fiber, 124, 270. See also Sweet potato dietary fiber classification, 125 composition, 124
125 extraction methods, 125
127 chemical reagent in combination with enzymatic hydrolysis method, 126
127 chemical separation method, 126 coarse separation method, 126 enzymatic hydrolysis method, 126 physicochemical and functional properties of dietary fiber, 127
128 in preventing obesity, 128
131 Differential scanning calorimetry (DSC), 9
10, 10t analysis of sweet potato starches, 30t Dimethyl nitrosamine (DMA), 295 2,2-Diphenyl-1-picrylhydrazyl (DPPH), 292
293, 382 radical-scavenging activity, 382 Disubstituted caffeoylquinic acids, 382 DMA. See Dimethyl nitrosamine (DMA) Dough concoction, 274 DP. See Degree of polymerization (DP) DPn. See Degree of number-average DP (DPn) DPPH. See 2,2-Diphenyl-1-picrylhydrazyl (DPPH) DPPH radical scavenging activity, 82 DPw. See Degree of weight-average DP (DPw) “Drug standards”, 398
399 Dry stool. See Constipation

Index

Dry weight (DW), 379
381 Dry yeast activation, 271 Drying, 132, 134
136 pectin, 193 DSC. See Differential scanning calorimetry (DSC) DW. See Dry weight (DW) Dynamic adsorption and desorption of AB-8 resin effect of flow and elution rates on, 376
377, 377f properties under optimum conditions, 377
378, 378f anthocyanins desorption curve from resin, 311
312 outflow curve and adsorption capacity of resin, 310
311, 311f D
Z model. See Thermal death time method (D
Z model)

E EAI. See Emulsifying activity index (EAI) Edible protein powder, 112
113 Electrophoresis, 58
59 Electrostatic interaction, 160
161 Elution rate, AB-8 resin, 376
377, 379 Emulsifier, 113
114 Emulsifying activity index (EAI), 95
96 Emulsifying properties of pectin, 199
200 of sweet potato pectin, 240
255 Ca21 concentrations effects on, 242
243 effects on adsorption of pectin, 243
245 emulsifying activity, 245
246 emulsion micro imaging, 251
255 emulsion stability, 249
251 emulsion viscosity, 247
248 NaCl concentrations effects on, 242
243 oil-phase volume fraction effects on, 241
242 pectin concentration effects on, 241
242 pH value concentrations effects on, 242
243 of sweet potato protein, 95
105 effects of pH on, 98
101 high hydrostatic pressure treatment, 103
105 NaCl on, 101
103 oil volume fraction on, 95
98 protein concentration, 95
98 Emulsion stability index (ESI), 96 Enhanced memory, 295
296 Enrichment ratio, 64
65

417

Enzymatic hydrolysis method, 126 chemical reagent in combination with, 126
127 sieve combination operation steps, 134
135 results, 135, 135t technological processes, 134 Enzymatic method, chemical reagent in combination with, 139
140 Escherichia coli, 84 ESI. See Emulsion stability index (ESI) Esterification(s), 397
398 degree of, 190 Esterified starch, sweet potato, 21
22, 42
43 Ethanol, 192
193, 307 concentration on desorption ratio of AB-8 resin, 375, 376f degradation kinetic parameters of anthocyanins in 10% ethanol, 319
321, 320t of anthocyanins in 20% ethanol, 321
322, 321t of anthocyanins in 50% ethanol, 322
323, 322t ethanol
ammonium sulfate, 288
289 solution, 373 Experimental animal study. See Animal experimental study External factors, pectin, 199 Extraction of ATPE, 298 experimental design for optimizing parameters, 299
308, 299t analysis of response surface, 303
308, 304f statistical analysis and model fitting, 300
303 experimental design for selection of parameters, 298
299 method of purple sweet potato, 286
290 ATPE, 288
289 chemical analysis of powders, 297
298 microwave extraction, 287 solvent extraction, 286
287 supercritical fluid extraction, 287 ultrasonic extraction, 288 of pectin, 191
193 drying pectin, 193 pectin hydrolysis and extraction, 192 pectin purification method, 192
193 pretreatment of raw materials, 191

418

Index

Extraction (Continued) of sweet potato anthocyanins, 297
308 comparing ATPS to other methods, 308, 308t experimental design for optimizing extraction parameters, 299
308, 299t experimental design for selection of extraction parameters, 298
299 extraction of ATPE, 298 Plackett
Burman experiment, 299, 299t sweet potato dietary fiber, 131
140 biological method, 135
139 chemical separation method, 139
140 sieve combined enzymatic hydrolysis method, 134
135 sieve method, 132
134 from sweet potato leaves, 371 from sweet potato pectin, 201
202 of sweet potato protein, 60
61, 63 solvent effects on extraction of, 58
59 technologies, 55 Eyring
Polanyi model, 320
321

F Fatty acid synthase, 130, 363
364 Fe31 absorption of, 160
161 Fe31/ascorbic acid/EDTA/H2O2 system, 383 Feces, accelerating output speed of, 161 Fermentation method, 138 microbial fermentation, 138 static fermentation, 271 Ferric-reducing antioxidant power of chlorogenic acids, 386 Ferrous ion-chelating activity, 387 Ferulic acid, 361f FFA. See Free fatty acid (FFA) Flavylium cation, 283, 283f Flour mixing, 271 Foam separation method, 64. See also Isoelectric point precipitation method; Thermal denaturation method; Ultrafiltration method factors influencing air-flow rate, 68
69 concentration, 66 foaming time, 68 inclined angle of foam separation tower, 69
70 influent volume, 67
68 pH value, 67

foaming process, 65f principle of, 64
65 process of sweet potato protein, 65
66 Foam stability of sweet potato protein, 110 Foaming sweet potato protein, 110 time, 68 Folin
Ciocalteu method, 372
373, 379
381 Food applications of sweet potato starch in, 42 food-processing technology, 57 industry, 343
346, 398 processing, 268 science, 284
285 Food and Drug Administration, 282 Fourier transform infrared spectrometry, 28
29 FOXO gene, 362 Free fatty acid (FFA), 129 Freeze
drying procedure, 269 Freeze
thaw stability, 35
36 Freshly peeled sweet potatoes, 63 Frozen food, 256 Fructo-oligosaccharide, 128
129 Functional properties of dietary fiber, 127
128 of pectin physicochemical functions, 198
200 physiological functions, 193
198 of sweet potato dietary fiber absorption of cholesterol, 160 absorption of Na1, K1, Ca21, Fe31, and Pb21, 160
161 accelerating output speed of feces, 161 protecting intestines from cancer, 160 of sweet potato protein, 92

G 4-GalA(1,2)-Rhap backbone, 189f Galactooligosaccharide, 128
129 D-Galactose, 295
296 Galacturonic acid(s), 187
190, 199 content determination in pectin solution, 202
203 factors affecting content of sweet potato pectin, 203
206 extraction time effects on, 205 pH value effects on, 204, 204f, 205f solid/liquid ratio effects on, 205
206 temperature effects on, 203
204, 203f Gas
liquid chromatography, 189
190 Gel properties of pectin, 198
199

Index

Gel texture analysis of sweet potato pectin Ca21 concentrations effects on, 238
239 pectin concentration effect on, 236
237 pH value effects on, 239
240 sucrose concentration effect on, 237 Gelatinization, 134 enthalpy, 11 temperature, 9
10 Gelation properties of sweet potato pectin, 231
240 Ca21 concentrations effects on, 234 gel’s linear viscoelastic region determination, 231
232 pectin concentration effects on, 232
233 pH value effects on, 235
236 sucrose concentration effects on, 233
234 Gelling properties of sweet potato protein, 105
106 Glibenclamide, 363
364 Glucagon-like peptide 1 (GLP-1), 130
131, 144
146, 145f, 154
155, 155f Glucokinase, 131 Gluconeogenesis, 363
364 Glucose tolerance level, 363
364 test, 141
142, 142t, 151, 152t Glucose-6-phosphatase, 131, 363
365 Glutathione S-transferases (GST), 331
332 Glycoprotein extraction technologies, 55 staining, 58 Gout, 296 Grease, 273
274 Grinding, 132, 134
136 Growth, sweet potato anthocyanins, 326
327, 336 GST. See Glutathione S-transferases (GST) Guinea pigs, 193
194

H HCl. See Hydrochloric acid (HCl) HDL. See High density lipoprotein (HDL) HDL cholesterol (HDL-C), 151 Health foods, sweet potato dietary fiber in, 175 Heat treatment on chlorogenic acids, 393
396 on soluble sweet potato protein, 107
109 Heating temperature, 76 Heavy metals, 194
195 Hemicellulose, 124
125 Hepatic ADH activity level, sweet potato anthocyanins, 332
333, 333f, 340
341 Hepatic GST activity levels, 340

419

Hepatic MDA activity level, sweet potato anthocyanins, 330, 338
339, 340f Hepatic SOD, sweet potato anthocyanins, 331
332, 332f, 333f HGA. See Homogalacturonan (HGA) High density lipoprotein (HDL), 143 High hydrostatic pressure treatment, 103
105, 105t, 109, 109t High-performance ion-exchange liquid chromatography, 189
190 High-performance liquid chromatography (HPLC), 368
369 qualitative and quantitative analyses, 378
381, 380f High-performance molecular exclusion chromatography, 191 Histopathological analysis, sweet potato anthocyanins, 334, 334t, 342
343, 342t HMG CoA. See 3-Hydroxy-3-methyl-glutaryl coenzyme A (HMG CoA) Homogalacturonan (HGA), 188 Hot extraction methods, 286 Hot PV (HPV), 34 Hot PV to PV (HPV/PV), 34 HPLC. See High-performance liquid chromatography (HPLC) HPLC
UV
Vis detection system, 368
369 HPV. See Hot PV (HPV) HT-29 human colon cancer cells, 209 inhibitory effects of unmodified pectin on, 219f sweet potato pectin effects on cancer cell proliferation, 210
211 Human intestinal tract, 124 Human metabolic processes, 292 Humectant, 114 Hydrochloric acid (HCl), 63 Hydrogen peroxide (H2O2), 364 Hydrolysis solvents, 189
190 Hydrolyzation of chlorogenic acids, 397
398 6-Hydroxy-2,5,7,8-tetramethylchroman-2carboxylic acid, 383 3-Hydroxy-3-methyl pentyl acyl coenzyme A, 363
364 3-Hydroxy-3-methyl-glutaryl coenzyme A (HMG CoA), 130 Hydroxyl radical-scavenging activity, 383 Hypertension, 293
294 Hypoglycemic effects, 363
364

I I. batatas cv Yamagawamurasaki. See Purple sweet potato IDF. See Insoluble dietary fiber (IDF)

420

Index

IL-6. See Interleukin-6 (IL-6) In vitro migration and invasion of cancer cells, 87 Inclined angle of foam separation tower, 69
70 Inflammatory cytokine tumor necrosis factor-α (TNF-α), 129 Influent protein concentration, 73 Influent volume, 67
68 Insoluble dietary fiber (IDF), 125 Insulin, 363
364. See also Diabetes concentrations, 143
144, 144f, 153
154, 154f insulin-containing molecules, 130
131 Interface-adsorbed protein, 97
98 Interleukin-6 (IL-6), 129, 146
147 Intermolecular copigmentation, 284
285 Intestine intestinal microbes, 128
129 SCFAs in, 129 Inulin, 128
131 Ion-exchange chromatography method, 79 resin, 192
193 Ionization reaction, 374 Ions, 61
62 Ipomeamarone, 267 Ipomoea batatas.. See Sweet potato (Ipomoea batatas) Ipomoein, 52
53 Isoelectric point precipitation method, 62. See also Foam separation method; Thermal denaturation method; Ultrafiltration method extraction of sweet potato protein by, 63 factors influencing, 63
64 acidity regulator, 63
64 metal ions, 64 proteins, 64 principle of, 62 Isoelectric precipitation method. See Isoelectric point precipitation method

K

K1, absorption of, 160
161 Kunitz-I type trypsin inhibitors, 85
86 Kunitz-type inhibitors, 83
84

L Laboratory sweet potato starch made by centrifugation (LCFS), 24
25, 25t, 26t Fourier transform infrared spectrometry, 28
29 particle size distributions, 26

pasting properties, 33
34 polarizing microscopy, 27
28 starch color effects, 31 sweet potato starch production, 26
27, 27f swelling power and solubility, 32
33 thermal properties, 30
31 Laboratory sweet potato starch made by sour liquid processing (LSLPS), 24
25, 25t, 26t Fourier transform infrared spectrometry, 28
29 particle size distributions, 26 pasting properties, 33
34 polarizing microscopy, 27
28 starch color effects, 31 sweet potato starch production, 26
27, 27f swelling power and solubility, 32
33 thermal properties, 30
31 XRD, 29
30 Lactate dehydrogenase (LDH), 325
326 sweet potato anthocyanins, 327
329, 328f, 337
338, 338f Lactobacillus, 128
129 L. bulgaricus, 344 Lactococcus lactis, 14
17 LCFS. See Laboratory sweet potato starch made by centrifugation (LCFS) LDH. See Lactate dehydrogenase (LDH) LDL. See Low-density lipoprotein (LDL) LDL-C. See Low-density lipoprotein cholesterol (LDL-C) Light on chlorogenic acids, 396
398 Lignin, 124
125 1,4-Linked α-D-galacturonic acid, 189 Linoleic acid peroxidation-inhibiting effect, 82
83 Lipid, 12
13. See also Obesity content, 8 peroxidation, 364, 384
385 activities of chlorogenic acids, 387
388 regulation, 194 Lipid-lowering, 363
364 Liquid flow rate, 72 Liver cells, 363
364 Liver indices concentrations of TG and TC in liver, 147
148, 147f pathological analysis of liver, 148
149, 149f pathological analysis of liver, 157
159, 158f sweet potato anthocyanins, 326
327, 327t, 336 TG and TC concentrations of liver, 156
157, 157f

Index

Liver protection, 294
295 Longkou vermicelli production, 14
15 Low-density lipoprotein (LDL), 294 Low-density lipoprotein cholesterol (LDL-C), 294 sweet potato anthocyanins, 329
330, 330f, 338, 339f LSLPS. See Laboratory sweet potato starch made by sour liquid processing (LSLPS)

M Macroporous adsorption resin, 370 Macroporous resins, 309 Malondialdehyde (MDA), 293, 330, 331f Mature extraction methods, 370 Meat products, sweet potato dietary fiber in, 174 Medicine and health-protection industry, 398
399 Membrane cleaning, 74 Metabolic enzyme, 363 Metal ion-chelating activity, 387 Metal ions, 64, 192
193 Mice models, subacute ALD, 335
336 Microbial fermentation, 138 Microporous starch, sweet potato, 22
23 Microstructure, 40
41 Microwave extraction, 287 Mitochondrial membranes, 364 Mixed enzymes method, 138
139 Mixing, biscuits with sweet potato granules, 274 Moisture Content, 9 Monosaccharide composition analysis, 189
190 Myocardial cells, 364

N

Na1 absorption, 160
161 electrochemical gradient, 364
365 NaCl concentrations effects on particle sizes of pectin emulsions, 242
243 on D4,3 of sweet potato protein emulsion, 101
102 effect on viscosity of pectin solution, 228
230 on gel structure of sweet potato protein, 106f on solubility of sweet potato protein, 93
94

421

sweet potato protein on emulsifying activity of, 102 on stability of, 103 Natural food-coloring pigments, 282 Natural trypsin inhibitor, 56 Neutral sugars, 189
190, 199 Nitric acids, 192 Nitrogen, 23
24 Noodles, sweet potato granule applications in, 275
276 manufacturing technologies, 275 noodle production, 276 procedure for making noodles, 275
276 mixing with water and flour and static aging, 276 proper amount of sweet potato granules for noodle production, 276 rolling, 276 suitable sweet potato granules, 275
276 Noodles and vermicelli. See Sweet potato starch noodles and vermicelli Nuclear magnetic resonance spectroscopy, 190 Nutrients, 193

O Obesity. See also Lipid dietary fiber in preventing altering metabolism, 129
131 intestinal microbes, 128
129 physical effects, 128 regulating SCFAs in intestine, 129 prevention, 88
92 experimental animal study, 91 sweet potato protein on 3T3-L1 preadipocytes, 88
91 effect of sweet potato dietary fiber on prevention, 140
150 blood index determination, 142
147 on body weights of rats, 140
141, 141t glucose tolerance test, 141
142, 142t liver indices, 147
149 organic acid content in cecum, 150, 150t effect of sweet potato dietary fiber on treating blood indices, 152
156 on body weights of rats, 151 glucose tolerance test, 151, 152t liver indices, 156
159 organic acid content in cecum, 159
160, 159t

422

Index

O
H stretching vibration, 28
29 Oil volume fraction effect on sweet potato protein concentration and composition, 97
98 on emulsifying activity and stability, 95
96 on size of emulsion particles, 95 Oil-holding capacity, 127, 161 of dietary fibers from varieties of sweet potato, 169
170, 169f of sweet potato protein, 110
111 temperature effects of sweet potato dietary fiber, 164
165, 165f Okra pectin RG-I region, 196 Oligosaccharides, 124, 128
129 Operating pressure, 72
73 Operational cycle time, 73 ORAC method. See Oxygen radical absorbance capacity (ORAC) method Organic acid content in cecum, 150, 150t, 159
160, 159t Organic solvent extraction method, 365 Orthogonal test, 276
277 Orthophthalic carboxylic structures, 397
398 Outflow curve of anthocyanins and adsorption capacity of resin, 310
311, 311f Oxygen radical absorbance capacity (ORAC) method, 385

P Packing, sweet potato granules in bread, 272 Pb21, absorption of, 160
161 PDGF. See Platelet-derived growth factor (PDGF) PDGF receptor-β (PDGFR-β), 294
295 Peak viscosity (PV), 33
34 Pectin, 124
125, 186
200 distribution, 186
188 cell wall structure, 187f polysaccharide domains, 188f extraction, 191
193 drying pectin, 193 pectin hydrolysis and extraction, 192 pectin purification method, 192
193 pretreatment of raw materials, 191 extraction methods, 192 optimization, 206
208 functional properties, 193
200 hydrolysis, 192 internal factors, 199 purification method, 192
193 research methods determining degree of esterification, 190

monosaccharide composition analysis, 189
190 relative molecular weight determination, 191 structure, 188
189 HGA, 188 RG-I, 188
189, 189f RG-II, 189 PEG. See Polyethylene glycol (PEG) Peroxy radical-scavenging activities, 385 pH, 307 on chlorogenic acids, 392
393 effects on sweet potato protein, 56
57 on adsorption rate of AB-8 resin, 309 on emulsifying activity of, 99 on particle size of emulsion particles, 98 on stability of emulsion, 99
101 levels effects on thermal stability of anthocyanins, 314
316, 315f on soluble sweet potato protein, 107
109 value, 227
228 concentrations effects on particle sizes of pectin emulsions, 242
243 effect on viscosity of pectin solution, 227
228 effects on texture parameters of pectin gel, 239
240 foam separation method, 67 on solubility of sweet potato protein, 93
94 salting-out method, 62 thermal denaturation method, 78 pH-modified pectin pH-modified pectin and anticancer activities, 196
197 preparation, 208 pH-modified sweet potato pectin effects on cancer cell adhesion, 215
216 on cancer cell migration, 221
222 on uPA, 225
226 Pharmaceutical industry, 343 Phenolic compounds, 360 Phenolic hydroxyl groups, 362, 387
388 Phenolic molecules, 374 Phenylpropanoid class, 360 Phosphoric acids, 192 Phosphorus content, 9 Photochemiluminescence assay, 384
385 Phthalates, 346 Physicochemical characteristics analysis color, 31, 31t freeze
thaw stability, 35
36

Index

paste transparency, 31
32 pasting properties, 33
34 swelling power and solubility, 32
33, 32f of sweet potato pectin, 227
255 gel texture analysis, 236
240 gelation properties, 231
240 microstructure, 240 viscosity analysis, 227
231 Physicochemical composition of sweet potato dietary fiber, 132
133, 133t Physicochemical functions of pectin emulsifying properties of pectin, 199
200 gel properties of pectin, 198
199 viscosity-associated properties of pectin, 198 Physicochemical properties of bread, 171 of dietary fiber, 127
128 of sweet potato dietary fiber, 161
170 of dietary fiber from varieties of sweet potato, 168 dietary fibers from sweet potato varieties, 167
168, 168f oil-holding capacities of dietary fibers, 169
170 temperature effects on oil-holding capacity, 164
165, 165f viscosity, 166
167, 166f, 167f water-holding capacity, 161
162, 162f, 163f waters-welling capacity of, 162
164, 164f, 165f of sweet potato protein, 56
57 of sweet potato starch, 24 comparison of quality of SLPS and CFS, 36
41 structure and physicochemical properties of SLPS and CFS, 24
36 Physiological functions of pectin, 193
198 effects on heavy metals, 194
195 effects on proliferation and apoptosis of cancer cells, 195
196 inhibiting effects on cancer metastasis, 195 lipid peroxidation regulation, 194 pH-modified pectin and anticancer activities, 196
197 regulation of blood glucose and blood lipid levels, 193
194 Physiological properties of sweet potato dietary fiber, 140
161 functional properties, 160
161 obesity effect on preventing, 140
150 effect on treating, 151
160

423

Plackett
Burman experiment, 299, 299t Plant cell wall, 124 Plant chlorogenic acids, 370 Plant sources, 125 Plasma adiponectin, 154
155, 155f concentration, 144
146, 145f Plasma inflammatory factor concentrations, 146
147, 146t, 156, 156t Plasma leptin, 143
144, 144f, 153
154, 154f Platelet-derived growth factor (PDGF), 294
295 Polarizing microscopy, 27
28, 28f Polyethylene glycol (PEG), 288 Polynuclear aromatic hydrocarbons, 363 Polyphenols, 381 Polysaccharide, 124
125 Potassium ion concentration, 364 Pressure extraction technology, 289 Pretreatment of feed liquid, 74 of sweet potato leaves, 371 Production line of sweet potato dietary fiber, 407 of sweet potato pectin, 409 of sweet potato protein, 405 Production technology of sweet potato pectin, 201
208 extraction process of pectin from sweet potato pectin, 201
202 factors affecting yield and galacturonic acid content, 203
206 galacturonic acid content determination, 202
203 of sweet potato starch, 16
18 noodle and vermicelli production process without alum, 19
21 noodles and vermicelli, 19 starch and series products, 16 Proofing, sweet potato granules in bread, 271 Propionic acid, 129 Protease, 139
140 Protein concentration, 61 sweet potato protein concentration and composition, 97
98 on emulsifying activity and stability, 95
96 on size of emulsion particles, 95 Protein-to-solvent ratio, 63 Proteins, 62, 64, 113
114 molecules, 62 protein kinase C, 193
194 salting out, 61
62 in sweet potato, 58

424

Index

PSPAs. See Purple sweet potato anthocyanins (PSPAs) Pulsed electric-assisted extraction, 289
290 Pumpkin-sweet potato starch noodles, 20 Purification, 371
378 of chlorogenic acid using AB-8 macroreticular adsorbent resin activation pretreatment, 371
372 dynamic adsorption and desorption of AB-8 resin, 376
378 static adsorption and desorption test, 372
375 method of chlorogenic acid, 365
367, 367t, 369 method of sweet potato protein, 78
79 ion-exchange chromatography method, 79 Sephadex G-75 gel chromatography method, 79 of sweet potato anthocyanins, 290, 291t, 309
312 adsorption and desorption ratios of AB-8 resin, 309
310 dynamic adsorption and desorption, 310
312 Purple sweet potato, 286, 290
291, 343 extraction method, 286
290 ATPE, 288
289 microwave extraction, 287 solvent extraction, 286
287 supercritical fluid extraction, 287 ultrasonic extraction, 288 powder chemical analysis, 297
298 Purple sweet potato anthocyanins (PSPAs), 293
294, 325
326, 326t, 342f analysis of variance for extraction yield, 300
302, 302t ANOVA for partition coefficient, 302
303, 303t PV. See Peak viscosity (PV) Pyruvate carboxykinase, 131

Q Quinic acid, 360, 361f, 397
398 Quinoidal base, 283, 283f Quinone oxidoreductase-glutathione transferase, 362
363

R Radical-scavenging activity of chlorogenic acids DPPH, 382, 383f hydroxyl, 383, 384f peroxy, 385, 385f

Rapid Visco Unit (RVU), 13 Rapidly digestible starch, 15
16 Raw materials decay, 266 Raw sweet potatoes, 266 Reactive oxygen species (ROS), 293 Reagent methods, 55 Resistance starch granules RS2, 15
16 Response surface analysis, 303
308, 304f extraction condition optimization and method validation, 308 interaction between variables on anthocyanin partition coefficient, 306
307 interaction between variables on Y1, 304
306 Response surface methodology, 308 Retrogradation, 35
36 rate, 12
13 of starch and noodles, 37
39 viscosity, 13
14 Retrograded starch RS3, 15
16 Rhamnogalacturonan (RG), 124
125 RG-I, 188
189, 189f RG-II, 188
189 Rhizomatous crop, 267 Rolling, biscuits with sweet potato granules, 274 ROS. See Reactive oxygen species (ROS) RVU. See Rapid Visco Unit (RVU)

S Salt saturation, 61 species on solubility of sweet potato protein, 94 type, 61
62 Salting-out method, 60 factors influencing protein salting out, 61
62 principle of, 60 sweet potato protein extraction technology, 60
61 Sanitary Standards of Using Food Additives, 19 Scanning electron microscopy, 26
27, 27f SCFAs. See Short-chain fatty acids (SCFAs) Schizophyllum commune, 138 SDF. See Soluble dietary fiber (SDF) SDS
polyacrylamide gel electrophoresis (SDS
PAGE), 53 Sephadex G-75 gel chromatography method, 79 glucan, 79

Index

Shear rate effect on viscosity of pectin solution, 230
231 Short-branched chain (CL 9
11 glucose residues/chain), 12
13 Short-branched chains (CL 12
14 glucose residues/chain), 12
13 Short-chain fatty acids (SCFAs), 129 in intestine, 129 Sieve method, 132
134 component analysis of sweet potato dietary fiber, 133
134, 133t operation steps, 132 physicochemical composition of, 132
133, 133t sieve combined enzymatic hydrolysis method, 134
135 technological processes, 132 Single enzyme method, 135
137 operation steps, 136 results, 136
137 technological process, 136 Slowly digestible starch, 15
16 SLPS. See Sweet potato starches isolated during sour liquid processing (SLPS) Slurry mixing, 132, 134, 136 SM-SPDF. See Sweet potato dietary fiber using sieve method (SM-SPDF) SMMC-7721 liver cancer cells, 209 sweet potato pectin effects on cancer cell proliferation, 213
214 SOD. See Superoxide dismutase (SOD) Sodium bicarbonate, 273 Solid-to-solvent ratio, 63 Solubility of sweet potato protein, 92
94 pH and NaCl on, 93
94 pH value on, 93 salt species on, 94 of sweet potato starch, 11
12 Soluble dietary fiber (SDF), 125 Soluble sweet potato protein, 107
109 Solvent extraction, 286
287, 290 on extraction of sweet potato protein on extraction yield, 58
59, 59t protein in sweet potato, 58 purity, 58
59, 59t Sorbitol, 389 Sour liquid, 16
17 Sporamin, 52
53, 89 amino acid sequences, 54t three-dimensional structure model, 54f

425

Spray drying method, 193, 269 Stability of chlorogenic acids, 391
398 effects of heat treatments, 393
396, 395f effects of light, 396
398, 397f effect of pH, 392
393, 393f ratio of starches, 34 Staple foods, sweet potato dietary fiber in, 174
175 Starch noodles and vermicelli. See Sweet potato starch noodles and vermicelli Starch-based super absorbent resin, sweet potato, 23
24 applications, 43 Static adsorption, 372
375 and desorption test of AB-8 macroreticular adsorbent resin effect of ethanol concentration, 375, 376f effect of initial sample TPC, 374
375, 375f performance, 372
373, 373f effect of sample pH, 373
374, 374f of sweet potato anthocyanins on AB-8 resin, 309 Static fermentation, 271 Statistical analysis and model fitting, 300
303 analysis of variance for extraction yield of PSPAs, 300
302, 302t ANOVA for partition coefficient of PSPAs, 302
303, 303t Steam peeling, 268 Streptococcus thermophiles, 344 Structural analysis Fourier transform infrared spectrometry, 28
29 particle size distribution, 26 polarizing microscopy, 27
28, 28f scanning electron microscopy, 26
27, 27f thermal properties, 30
31 XRD, 29
30, 29f Subacute ALD, sweet potato anthocyanin effect on, 334
343 on growth and liver index, 336 on hepatic ADH activity level, 340
341 on hepatic GST and SOD activity levels, 340 on hepatic MDA activity level, 338
339 on histopathological analysis, 342
343, 342t mice models, 335
336 on serum ALT, AST, and LDH activity levels, 337
338 serum TG, and LDL-C levels and TCH activity, 338

426

Index

Sucrose concentration effect on texture parameters of pectin gel, 237 effect on viscosity of pectin solution, 228
230 Sulfuric acids, 192 Super absorbent resin, sweet potato starch-based, 23
24 applications, 43 Supercritical fluid extraction, 287 Superoxide (O22•)-scavenging activities of chlorogenic acids, 383
385 Superoxide anion radical scavenging activity, 80
81 Superoxide dismutase (SOD), 293, 331, 391 activity levels, 340, 341f Supplemental ingredient treatment, 273
274 “SuShu8” starch, 14
15 SW480 cells, 86, 87t Sweet potato (Ipomoea batatas), 3, 285
286 ketoalcohol, 267 pulp, 186 residue, 131 Sweet potato acetylated starch, 22 applications, 43 Sweet potato anthocyanin applications, 343
347 cosmetics industry, 346
347 food industry, 343
346 pharmaceutical industry, 343 bioactivity, 292
297 on acute ALD and dealcoholic effects, 324
334 on subacute ALD, 334
343 preparation extraction, 297
308 purification, 309
312 status, 285
297 extraction method of purple sweet potato, 286
290 purification methods, 290 structures, 290
292 Sweet potato dietary fiber applications, 170
175 in beverages, 174 in bread, 170
174 in condiments, 175 in health foods, 175 in meat products, 174 in staple foods, 174
175 extraction technology, 131
140 physicochemical properties of, 161
170 physiological properties of, 140
161 production line, 407

Sweet potato dietary fiber using sieve method (SM-SPDF), 132 Sweet potato granules, 264
265 addition of emulsifier, 269 applications, 270 in biscuits, 272
275 in bread, 270
272 in noodles, 275
276 in thick slurries, 276
277 calcium soaking, 268
269 cleaning, 267 color preservation, 268 drying, 269 manufacturing technologies, 265 mashing, 269 peeling, 267
268 selection of materials, 267 selection of special cultivars, 266 slicing, 268 steaming, 269 storage of raw sweet potatoes, 266 Sweet potato leaves, 359
360 chlorogenic acids, 364
365, 370, 390 antibacterial activities, 388 extraction, 371 heat treatments effects, 393
396 inhibitory effects, 388 light effects, 396
398 O22•-scavenging activities, 384f pH effect, 392
393 purification using AB-8 macroreticular adsorbent resin, 371
378 qualitative and quantitative analyses by HPLC, 371
378, 380f pretreatment, 371 as raw materials, 370 Sweet potato pectin applications, 257 beverages, 256
257 bread, 255 candied fruit, 255 frozen food, 256 yogurt products, 256 biological activities, 208
226 emulsifying properties, 240
255 optimization of pectin extraction, 206
208, 207t physicochemical characteristics, 227
255 production line, 409 production technology, 201
208 Sweet potato protein, 52 applications, 111
112 biological medicine, 115

Index

edible protein powder, 112
113 emulsifier, 113
114 humectant, 114 raw material of active peptides, 114 biological activity, 55
56, 80 anticancer activity, 86
88 antioxidant activity, 80
83 obesity prevention and weight loss, 88
92 trypsin inhibitory activity, 83
86 functional assay results, 113t functional properties, 92 emulsifying properties, 95
105 gelling properties, 105
106 SDS
PAGE pattern, 99f solubility of sweet potato protein, 92
94 physicochemical properties, 56
57 production line, 405 production technologies, 57 effects of solvent on extraction of, 58
59 foam separation method, 64
70 isoelectric point precipitation method, 62
64 salting-out method, 60
62 thermal denaturation method, 75
78 ultrafiltration method, 70
74 purification method, 78
79 research status on technologies, 55 SDS
PAGE, 59f sources and structures, 52
54 structural properties, 107
109 foaming properties and foam stability, 110 heat treatment, 107
109 high hydrostatic pressure treatments, 109, 109t oil-holding capacities, 110
111 pH, 107
109 water-holding capacities, 110
111 Sweet potato resistant starch, 15
16 applications sweet potato acetylated starch applications, 43 sweet potato esterified starch applications, 42
43 sweet potato starch-based superabsorbent resin applications, 43 production process sweet potato acetylated starch, 22 sweet potato esterified starch, 21
22 sweet potato microporous starch, 22
23 sweet potato starch-based super absorbent resin, 23
24

427

Sweet potato starch, 3
4, 14 applications, 41
42 in food, 42 resistant starch, 42
43 characteristics gelatinization enthalpy, 11 gelatinization temperature, 9
10 retrogradation rate, 12
13 solubility, 11
12 swelling power, 11 viscosity, 13
14 chemical composition, 8t, 25t amylose content, 7
8 lipid content, 8 moisture content, 9 phosphorus content, 9 mineral compositions, 26t paste parameters, 10t physicochemical properties, 24 production technology, 16 structure and morphology, 4
7 amylose and amylopectin, 4
6 morphology and size of sweet potato starch, 7 X-ray diffraction type, 6
7, 6f sweet potato resistant starch, 15
16 sweet potato resistant starch production process, 21
24 Sweet potato starch noodles and vermicelli additives effects on quality, 20
21 in China, 14 introduction to, 14 production process, 19
20 without alum, 19
21 of pure sweet potato starch noodles and vermicelli, 20 research on, 14
15 starch color effects, 31 Sweet potato starches isolated during sour liquid processing (SLPS), 14 amylose content, 24
25 comparison of quality, 36
41 color of sweet potato starches and noodles, 37, 37t cooking quality, 39
40, 39t microstructure, 40
41 retrogradation of starch and noodles, 37
39 textural properties, 39t, 40, 40t Fourier transform infrared spectrometry, 28
29 particle size distributions, 26 pasting properties, 33
34

428

Index

Sweet potato starches isolated during sour liquid processing (SLPS) (Continued) physicochemical characteristics analysis, 31
36 proximate composition, 24
25 starch color effects, 31 structural analysis, 26
31 structure and physicochemical properties, 24
36 swelling power and solubility, 32
33 thermal properties, 30
31 XRD, 29
30 Swelling power and solubility of SLPS and CFS, 32
33, 32f of starch, 11

T t-butyl hydroperoxide (T-BHP), 295 3T3-L1 preadipocytes, proliferation and differentiation of, 88
91 Tartaric acids, 192 TC. See Total cholesterol (TC) TCH. See Total cholesterol potato anthocyanin (TCH) TDF. See Total dietary fiber (TDF) TE. See Trolox equivalent (TE) Temperature effect on protein salting out, 62 effect on trypsin inhibitory activity of sweet potato, 85 effect on ultrafiltration, 73 effect on viscosity of pectin solution, 230
231 heating, 76 Textural properties, 39t, 40, 40t TGs. See Triglycerides (TGs) Thermal death time method (D
Z model), 317 Thermal degradation in 10% ethanol, 319
321, 320t in 20% ethanol, 321
322, 321t in 50% ethanol, 322
323, 322t in deionized water, 318
319, 318t experiment, 316 kinetic parameters of anthocyanins model, 316
317 Thermal denaturation method, 75. See also Foam separation method; Isoelectric point precipitation method; Ultrafiltration method influencing factors cooling temperature, 77 denaturation time, 76
77

heating temperature, 76 pH value, 78 mechanism, 75 production process of sweet potato protein, 75
76, 76f Thermal stabilities of anthocyanins high pressure effects, 313
314, 314f pH level effects, 314
316, 315f temperature effects, 312
313, 313f Thermal-modified pectin preparation, 209 Thermal-modified sweet potato pectin effects on cancer cell adhesion, 216
218 migration, 219
221 on uPA, 224
225 Thick slurries, sweet potato granules in, 276
277 Thin-layer chromatography, 368 Thinly rolled noodles, 276 TNF-α. See Inflammatory cytokine tumor necrosis factor-α (TNF-α) Total cholesterol (TC), 143 concentrations of liver, 156
157, 157f in liver, 147
148, 147f Total cholesterol potato anthocyanin (TCH), 294 sweet potato anthocyanins effect, 329
330, 329f, 338, 339f Total dietary fiber (TDF), 132
133 Total phenolic content (TPC), 370
371 of desorption solution, 372
373 of effluent solution, 376 heat treatment effects on stability of chlorogenic acids, 393
394 initial sample TPC effect on adsorption capacity of AB-8 resin, 374
375 light effects on stability of chlorogenic acids, 396
397 pH value effects, 392
393 of sweet potato leaf polyphenols, 392 Traditional potato cleaning method, 267 Trans-cinnamic acids, 360 Transduction pathway, 362
363 Triacylglycerol, 364 Triglycerides (TGs), 128, 294, 363
364 concentrations of liver, 156
157, 157f in liver, 147
148, 147f sweet potato anthocyanins effects, 329
330, 329f, 338 Trolox, 383 Trolox equivalent (TE), 385 Trypsin inhibitors, 86

Index

Trypsin inhibitory activity of sweet potato protein, 83
86 determination, 84
85 factors influencing effects of growth conditions on, 85 protein content to, 85 structural changes on, 85
86 temperature effect on, 85 Tumor metastasis, 87
88 TαA-SPDF, 136
137, 137t

U Ultrafiltration method, 55, 57, 70, 71f. See also Foam separation method; Isoelectric point precipitation method; Thermal denaturation method influencing factors, 72
74 influent protein concentration, 73 liquid flow rate, 72 membrane cleaning, 74 operating pressure, 72
73 operational cycle time, 73 pore size of ultrafiltration membranes, 74 pretreatment of feed liquid, 74 temperature, 73 principle of, 70
71 of sweet potato protein, 71
72 Ultrasonic extraction, 288 Ultraviolet (UV) spectrophotometry, 368 Unmodified sweet potato pectin effect on cancer cell adhesion, 215
216 on cancer cell migration, 219 on uPA content of cancer cells, 224 Urokinase-type plasminogen activator (uPA), 223
226 pH-modified sweet potato pectin effects on, 225
226 sweet potato pectin effects in cancer cells, 223
226 thermal-modified sweet potato pectin effects on, 224
225 UV spectrophotometry. See Ultraviolet (UV) spectrophotometry

429

V Vacuum concentration method, 192
193 Vascular endothelial cells, 364 Viscosity, 13
14, 127, 161 analysis of sweet potato pectin, 227
231 effects of pectin concentration and pH value, 227
228 effects of sucrose, NaCl, and Ca21 concentrations, 228
230 effects of temperature and shear rate, 230
231 of sweet potato dietary fiber, 166
167, 166f, 167f viscosity-associated properties of pectin, 198

W Washing, 132, 134, 136 Water extraction method, 139 Water-holding capacity, 127, 161 dietary fibers from different sweet potato varieties, 167
168, 168f of sweet potato dietary fiber, 161
162, 162f, 163f of sweet potato protein, 110
111 Water-swelling capacity, 127
128, 161 of dietary fiber from varieties of sweet potato, 168 of sweet potato dietary fiber, 162
164, 164f, 165f Weight loss, 88
92 animal experimental study on, 91
92 sweet potato protein effect on 3T3-L1 preadipocytes, 88
91 White sugar, 271, 273
274

X X-ray diffraction (XRD), 6
7, 6f, 29
30, 29f X-ray-induced DNA damage, 365

Y Yogurt products, 256