Natural polysaccharides in drug delivery and biomedical applications 9780128170564, 0128170565, 9780128170557

Table of contents :
Content: 1. Natural polysaccharides: sources and extraction methodologies 2. Pharmaceutical applications of natural polysaccharides 3. Sodium alginate in drug delivery and biomedical areas 4. Chitosan in drug delivery applications 5. Xanthan gum in drug delivery applications 6. Gellan gum in drug delivery applications 7. Guar gum in drug delivery applications 8. Locust bean gum in drug delivery application 9. Sterculia gum in drug delivery applications 10. Pectin in drug delivery applications 11. Cashew gum in drug delivery applications 12. Tamarind gum in drug delivery applications 13. Hyaluronic acid in drug delivery applications 14. Gum odina as pharmaceutical excipient 15. Alginate-chitosan combinations in controlled drug delivery 16. Synthesis of micro- and nanoparticles of alginate and chitosan for controlled release of drugs 17. Polysaccharides nanoparticles as oral drug delivery systems 18. Natural polysaccharide-based composites for drug delivery and biomedical applications 19. Natural polysaccharides for the delivery of anticancer therapeutics 20. Organic nanocomposites for the delivery of bioactive molecules 21. Natural polysaccharides for growth factors delivery 22. Marine polysaccharides for drug delivery in tissue engineering 23. Natural polysaccharides in tissue engineering applications 24. Natural polysaccharides in wound dressing applications 25. Polysaccharides from leafy vegetables: chemical, nutritional and medicinal properties 26. Electrospun natural polysaccharide for biomedical application

Citation preview

Natural Polysaccharides in Drug Delivery and Biomedical Applications Edited by Md Saquib Hasnain Amit Kumar Nayak

Academic Press is an imprint of Elsevier 125 London Wall, London EC2Y 5AS, United Kingdom 525 B Street, Suite 1650, San Diego, CA 92101, United States 50 Hampshire Street, 5th Floor, Cambridge, MA 02139, United States The Boulevard, Langford Lane, Kidlington, Oxford OX5 1GB, United Kingdom Copyright © 2019 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. Library of Congress Cataloging-in-Publication Data A catalog record for this book is available from the Library of Congress British Library Cataloguing-in-Publication Data A catalogue record for this book is available from the British Library ISBN: 978-0-12-817055-7 For information on all Academic Press publications visit our website at https://www.elsevier.com/books-and-journals

Publisher: Andre Gerhard Wolff Acquisition Editor: Erin Hill-Parks Editorial Project Manager: Sandra Harron Production Project Manager: Sreejith Viswanathan Cover Designer: Greg Harris Typeset by TNQ Technologies

List of contributors Yasir Faraz Abbasi AIMST University, Semeling, Kedah, Malaysia Syed Anees Ahmed Department of Pharmacology, Hygia Institute of Pharmaceutical Application and Research, Lucknow, India Tejraj M. Aminabhavi Department of Pharmaceutical Engineering and Polymer Science, SET’s College of Pharmacy, Dharwad, India Yubia De Anda-Flores Biopolymers-CTAOA, Research Center for Food and Development (CIAD, A.C.), Hermosillo, Sonora, Mexico Sukumaran Anil Department of Dentistry, Hamad Medical Corporation, Doha, Qatar Mohammad Tahir Ansari Faculty of Pharmacy and Health Sciences, Universiti Kuala Lumpur Royal College of Medicine, Perak, Ipoh, Malaysia Mona F. Arafa Department of Pharmaceutical Technology, College of Pharmacy, University of Tanta, Tanta, Egypt Esha Bala Centre for Rural Technology, Indian Institute of Technology, Guwahati, Assam, India Hriday Bera Faculty of Pharmacy, AIMST University, Semeling, Kedah, Malaysia Branko Bugarski Faculty of Technology and Metallurgy, University of Belgrade, Belgrade, Serbia Alma C. Campa-Mada Research Center for Food and Development, CIAD, A.C., Hermosillo, Sonora, Mexico Elizabeth Carvajal-Millan Research Center for Food and Development, CIAD, A.C., Hermosillo, Sonora, Mexico Pedro M. Castro CBQF e Centro de Biotecnologia e Quı´mica Fina e Laborato´rio Associado, Escola Superior de Biotecnologia, Universidade Cato´lica Portuguesa/Porto, Porto, Portugal; CESPU, Instituto de Investigacaeo e Formacaeo Avancada em Cieˆncias e Tecnologias da Saude, Gandra-PRD, Portugal Bankim Chandra Nandy Department of Pharmaceutics, School of Pharmacy, Techno India University, Kolkata, West Bengal, India Kiran Chaturvedi Department of Pharmaceutical Engineering and Polymer Science, SET’s College of Pharmacy, Dharwad, India Bor Shin Chee Athlone Institute of Technology, Materials Research Institute, Athlone, Co. Westmeath, Ireland

xvii

List of contributors Norma B. D’Accorso Universidad de Buenos Aires, Facultad de Ciencias Exactas y Naturales, Departamento de Quı´mica Orga´nica, Ciudad Universitaria, Ciudad Auto´noma de Buenos Aires, Argentina; Centro de Investigaciones en Hidratos de Carbono (CIHIDECAR)-CONICET-UBA, Ciudad Auto´noma de Buenos Aires, Argentina Arnab De Division of Microbiology & Biotechnology, Department of Pharmaceutical Technology, Jadavpur University, Kolkata, India Mabilly Cox Holanda de Barros Dias Athlone Institute of Technology, Biosciences Research Institute, Athlone, Co. Westmeath, Ireland Sanjay Dey Department of Pharmaceutics, School of Pharmacy, Techno India University, Kolkata, West Bengal, India Tanavi Dugge Shri Dharmasthala Manjunath Dental College, Dharwad, India Gamal M. El Maghraby Department of Pharmaceutical Technology, College of Pharmacy, University of Tanta, Tanta, Egypt Classius Ferreira da Silva Instituto de Cieˆncias Ambientais, Quı´micas e Farmaceˆuticas, Universidade Federal de Sa˜o Paulo, Diadema, Brazil Kuntal Ganguly Department of Pharmaceutical Engineering and Polymer Science, SET’s College of Pharmacy, Dharwad, India Gabriel H. Gomez-Rodriguez Research Center for Food and Development, CIAD, A.C., Hermosillo, Sonora, Mexico Neha Hans Department of Pharmacy, Birla Institute of Technology and Science, Pilani, Rajasthan, India Md Saquib Hasnain Department of Pharmacy, Shri Venkateshwara University, Gajraula, India Ankit Jain Institute of Pharmaceutical Research, GLA University, Mathura, Uttar Pradesh, India Sanjay K. Jain Pharmaceutics Research Projects Laboratory, Department of Pharmaceutical Sciences, Dr Hari Singh Gour Central University, Sagar, Madhya Pradesh, India Sougata Jana Department of Pharmaceutics, Gupta College of Technological Sciences, Asansol, West Bengal, India; Department of Health and Family Welfare, Directorate of Health Services, Kolkata, West Bengal, India Subrata Jana Department of Chemistry, Indira Gandhi National Tribal University, Amarkantak, Madhya Pradesh, India Sunil Kumar Dubey Department of Pharmacy, Birla Institute of Technology and Science, Pilani, Rajasthan, India Nurul Asmak Md Lazim Dept. of Bioprocess & Polymer Engineering, School of Chemical and Energy Engineering, Faculty of Engineering, Universiti Teknologi Malaysia, Johor Bahru, Johor, Malaysia Yiyang Liu Department of Pharmaceutics, School of Pharmacy, Fudan University & Key Laboratory of Smart Drug Delivery, Ministry of Education, Shanghai, China Jaime Lizardi-Mendoza Biopolymers-CTAOA, Research Center for Food and Development (CIAD, A.C.), Hermosillo, Sonora, Mexico Baboucarr Lowe School of Dentistry, The University of Queensland, Brisbane, QLD, Australia

xviii

List of contributors Ana Raquel Madureira CBQF e Centro de Biotecnologia e Quı´mica Fina e Laborato´rio Associado, Escola Superior de Biotecnologia, Universidade Cato´lica Portuguesa/Porto, Porto, Portugal Sabyasachi Maiti Department of Pharmacy, Indira Gandhi National Tribal University, Amarkantak, Madhya Pradesh, India Vero´nica E. Manzano Universidad de Buenos Aires, Facultad de Ciencias Exactas y Naturales, Departamento de Quı´mica Orga´nica, Ciudad Universitaria, Ciudad Auto´noma de Buenos Aires, Argentina; Centro de Investigaciones en Hidratos de Carbono (CIHIDECAR)-CONICET-UBA, Ciudad Auto´noma de Buenos Aires, Argentina Milan Milivojevic Faculty of Technology and Metallurgy, University of Belgrade, Belgrade, Serbia Ana Moira Mora´s Laboratory of Genetic Toxicology, Federal University of Health Sciences of Porto Alegre e UFCSPA, Porto Alegre, Rio Grande do Sul, Brazil Uttam A. More Department of Pharmaceutical Chemistry, Shree Dhanvantary Pharmacy College, Kim, Surat, India Dinara Jaqueline Moura Laboratory of Genetic Toxicology, Federal University of Health Sciences of Porto Alegre e UFCSPA, Porto Alegre, Rio Grande do Sul, Brazil S M Mozammil Hasnain Department of Mechanical Engineering, Birla Institute of Technology, Mesra, Ranchi, India Ida Idayu Muhamad Dept. of Bioprocess & Polymer Engineering, School of Chemical and Energy Engineering, Faculty of Engineering, Universiti Teknologi Malaysia, Johor Bahru, Johor, Malaysia; IJN-UTM Cardiovascular Engineering Centre, Universiti Teknologi Malaysia, Johor Bahru, Malaysia Balaram Naik Shri Dharmasthala Manjunath Dental College, Dharwad, India Sitansu Sekhar Nanda Department of Chemistry, Myongji University, Yongin, South Korea Jayanta Narayan De Medical College and Hospital, Kolkata, West Bengal, India Amit Kumar Nayak Department of Pharmaceutics, Seemanta Institute of Pharmaceutical Sciences, Mayurbhanj, Odisha, India Malleshappa N. Noolvi Department of Pharmaceutical Chemistry, Shree Dhanvantary Pharmacy College, Kim, Surat, India Michael Nugent Athlone Institute of Technology, Materials Research Institute, Athlone, Co. Westmeath, Ireland Marı´a Natalia Pacho Universidad de Buenos Aires, Facultad de Ciencias Exactas y Naturales, Departamento de Quı´mica Orga´nica, Ciudad Universitaria, Ciudad Auto´noma de Buenos Aires, Argentina; Centro de Investigaciones en Hidratos de Carbono (CIHIDECAR)-CONICET-UBA, Ciudad Auto´noma de Buenos Aires, Argentina Ivana Pajic-Lijakovic Faculty of Technology and Metallurgy, University of Belgrade, Belgrade, Serbia Pritish Kumar Panda Pharmaceutics Research Projects Laboratory, Department of Pharmaceutical Sciences, Dr Hari Singh Gour Central University, Sagar, Madhya Pradesh, India

xix

List of contributors Zhiqing Pang Department of Pharmaceutics, School of Pharmacy, Fudan University & Key Laboratory of Smart Drug Delivery, Ministry of Education, Shanghai, China Versha Parcha Dolphin (PG) Institute of Biomedical & Natural Sciences, Dehradun, Uttarakhand, India Sanjukta Patra Centre for Rural Technology, Indian Institute of Technology, Guwahati, Assam, India Cristiana Maria Pedroso Yoshida Instituto de Cieˆncias Ambientais, Quı´micas e Farmaceˆuticas, Universidade Federal de Sa˜o Paulo, Diadema, Brazil Manuela E. Pintado CBQF e Centro de Biotecnologia e Quı´mica Fina e Laborato´rio Associado, Escola Superior de Biotecnologia, Universidade Cato´lica Portuguesa/Porto, Porto, Portugal Sneha S. Rao Yenepoya Research Centre, Yenepoya (Deemed to be University), Mangalore, Karnataka, India M.E. Bhanoji Rao Calcutta Institute of Pharmaceutical Technology & Allied Health Sciences, Kolkata, West Bengal, India Agustin Rasco´n-Chu Research Center for Food and Development, CIAD, A.C., Hermosillo, Sonora, Mexico Kakarla Raghava Reddy School of Chemical and Biomolecular Engineering, The University of Sydney, Sydney, NSW, Australia P.D. Rekha Yenepoya Research Centre, Yenepoya (Deemed to be University), Mangalore, Karnataka, India Amalesh Samanta Division of Microbiology & Biotechnology, Department of Pharmaceutical Technology, Jadavpur University, Kolkata, India Farheen Sami Faculty of Pharmacy and Health Sciences, Universiti Kuala Lumpur Royal College of Medicine, Perak, Ipoh, Malaysia Shivani Saraf Pharmaceutics Research Projects Laboratory, Department of Pharmaceutical Sciences, Dr Hari Singh Gour Central University, Sagar, Madhya Pradesh, India Manoj Kumar Sarangi School of Pharmaceutical Sciences & Technology, Sardar Bhagwan Singh University, Dehradun, Uttarakhand, India Bruno Sarmento CESPU, Instituto de Investigacaeo e Formacaeo Avancada em Cieˆncias e Tecnologias da Saude, Gandra-PRD, Portugal; i3S - Instituto de Investigac¸a˜o e Inovac¸a˜o em Sau´de, Universidade do Porto, Porto, Portugal; INEB e Instituto de Engenharia Biome´dica, Universidade do Porto, Porto, Portugal Suguna Selvakumaran Department of Biotechnology, School of Science & Engineering, Manipal International University (MIU) No 1, Putra Nilai, Negeri Sembilan, Malaysia Kalyan Kumar Sen Department of Pharmaceutics, Gupta College of Technological Sciences, Asansol, West Bengal, India Patrı´cia Severino Universidade Tiradentes, Aracaju´, Brazil Zahra Shariatinia Department of Chemistry, Amirkabir University of Technology (Tehran Polytechnic), Tehran, Iran

xx

List of contributors Niharika Shiva Department of Pharmacy, Birla Institute of Technology and Science, Pilani, Rajasthan, India Siddhartha Singha Centre for Rural Technology, Indian Institute of Technology, Guwahati, Assam, India Gautam Singhvi Department of Pharmacy, Birla Institute of Technology and Science, Pilani, Rajasthan, India Eliana Maria Souto Faculty of Pharmacy, University of Coimbra, Coimbra, Portugal; Centre of Biological Engineering (CEB), University of Minho, Braga, Portugal Luiza Steffens Athlone Institute of Technology, Materials Research Institute, Athlone, Co. Westmeath, Ireland; Laboratory of Genetic Toxicology, Federal University of Health Sciences of Porto Alegre e UFCSPA, Porto Alegre, Rio Grande do Sul, Brazil Mohammad Tabish Department of Pharmacology, College of Medicine, Saqra University, Kingdom of Saudi Arabia Judith Tanori-Cordova Department of Polymers and Materials Research, University of Sonora, Hermosillo, Sonora, Mexico Ankita Tiwari Pharmaceutics Research Projects Laboratory, Department of Pharmaceutical Sciences, Dr Hari Singh Gour Central University, Sagar, Madhya Pradesh, India Jayachandran Venkatesan Yenepoya Research Centre, Yenepoya (Deemed to be University), Mangalore, Karnataka, India Amit Verma Pharmaceutics Research Projects Laboratory, Department of Pharmaceutical Sciences, Dr Hari Singh Gour Central University, Sagar, Madhya Pradesh, India Patrı´cia Hissae Yassue-Cordeiro Universidade Tecnolo´gica Federal do Parana´, Londrina, Brazil Dong Kee Yi Department of Chemistry, Myongji University, Yongin, South Korea Yuefei Zhu Department of Pharmaceutics, School of Pharmacy, Fudan University & Key Laboratory of Smart Drug Delivery, Ministry of Education, Shanghai, China

xxi

Preface Modern drifts toward the utilization of natural polymeric materials in numerous healthcare applications demand the replacement of the synthetic polymers with the natural ones because of their biocompatibility, biodegradability, economic extraction, and readily availability. The reality behind the rise in importance of these natural polymeric materials is that these sources are renewable if grown in a sustainable means and these can tender incessant supply of raw materials. Among these natural polymers, natural polysaccharides are considered as excellent biopolymers because these are nontoxic, stable, swellable, and biodegradable. Natural polysaccharides are extracted from various natural resources such as plants, animals, fungus, seaweeds, etc. Several research innovations have already been made on the applications of these natural polysaccharides in drug delivery and biomedical applications. In this context, thorough understanding of extraction, purification, characterization, and applications of natural polysaccharides and their derivatives for their utility in various drug delivery and different biomedical applications need to be thoroughly understood. The book “Natural Polysaccharides in Drug Delivery and Biomedical Applications” provides a fundamental overview of various natural polysaccharides, their sources, extraction methodologies, characterizations, and their applications in drug delivery and biomedical uses. It covers specific natural polysaccharides of various types and their effective applications in drug delivery and biomedical uses. This volume is a collection of 26 chapters by the academicians and researchers of pharmaceutical and biomedical fields across the world. A concise account on the contents of each chapter has been described to provide a glimpse of the book to the readers. Chapter 1 entitled “Natural Polysaccharides: Sources and Extraction Methodologies” describes various sources and extraction methodologies of natural polysaccharides. Chapter 2 entitled “Pharmaceutical Applications of Natural Polysaccharides” deals with properties of natural polysaccharides and their applications in cell encapsulation, drug/gene delivery, protein binding, wound healing, tissue engineering, bioimaging, contact lenses and implants, antibacterial textiles/papers, and antibacterial food additives/packaging materials. Chapter 3 entitled “Sodium alginate in drug delivery and biomedical areas” presents an account on different physicochemical properties, regulatory status, and various applications in drug delivery either alone or in combination with other polymers for drug delivery.

xxiii

Preface Chapter 4 entitled “Chitosan in Drug Delivery Applications” gives an account on the latest research on chitosan-based drug delivery systems along with its modifications and their future applications. Chapter 5 entitled “Xanthan Gum in Drug Delivery Applications” describes about the biochemistry, properties, and production of xanthan gum with its applications in pharmaceutical drug delivery, cosmetic, biomedical, and food industries. Chapter 6 entitled “Gellan gum in Drug Delivery Applications” presents a detailed account on sources, structure, and properties of gellan gum with emphasizing recent advances related to the development of different gellan gumebased drug delivery systems. Chapter 7 entitled “Guar Gum in Drug Delivery Applications” gives a detailed account on different types of guar gumebased drug delivery carriers with special focuses on different dosage forms such as tablets, beads, microspheres, nanoparticles, and hydrogels in single or in combination with other variety of biodegradable polymers, e.g., copolymers, grafts, blends, and interpenetrating polymer networks. Chapter 8 entitled “Locust Bean Gum in Drug Delivery Application” presents a detailed account on the processing of the locust bean gum, their properties, synergistic interaction, and its applications in pharmaceutical drug delivery. Chapter 9 entitled “Sterculia Gum in Drug Delivery Applications” gives a detailed account on the source, composition, properties, and applications of sterculia gum, with a comprehensive overview on the current development of different sterculia gum-based preparations in drug delivery. Chapter 10 entitled “Pectin in Drug Delivery Applications” deals with the text reports on prebiotic effect, hypoglycemic effect, hypocholesterolemic effect, the alleged effect on metastasis and apoptosis in cancer cells, and matrices for controlled and target-directed release of therapeutic compounds applications. Chapter 11 entitled “Cashew Gum in Drug Delivery Applications” presents a comprehensive and useful discussion about cashew gum in designing and development of biodegradable, biocompatible dosage forms. In addition, the chapter comprises a brief discussion about the source, isolation, chemical constituents, and properties of cashew gum. Chapter 12 entitled “Tamarind Gum in Drug Delivery Applications” includes a comprehensive and useful discussion about the applications of tamarind gum in the designing of various drug delivery systems. In addition, the article also briefly discusses about source, isolation, chemical composition, and properties of tamarind gum. Chapter 13 entitled “Hyaluronic Acid in Drug Delivery Applications” gives a detailed account on recent updates of hyaluronic acid in the formulation of various drug delivery systems. This chapter also presents an insight into the importance of hyaluronic acid uses in the development of targeted drug vectors.

xxiv

Preface Chapter 14 entitled “Gum Odina as Pharmaceutical Excipient” deals with a comprehensive and useful discussion of gum odina as pharmaceutical excipients. In addition, the article also briefly discusses about source and properties of gum odina. Chapter 15 entitled “Alginate-chitosan combinations in controlled drug delivery” gives a detailed account on the applications of alginateechitosan combination with emphasis on controlled drug delivery. Chapter 16 entitled “Synthesis of Micro- and Nanoparticles of Alginate and Chitosan for Controlled Release of Drugs” presents a detailed account on the combination of alginate and chitosan with emphasis on controlled drug delivery. Chapter 17 entitled “Polysaccharides Nanoparticles as Oral Drug Delivery Systems” gives a detailed account on basic and applied aspects of polysaccharides and their potential to develop nanoparticles to be employed as oral drug delivery systems. Chapter 18 entitled “Natural Polysaccharide-based Composites for Drug Delivery and Biomedical Applications” gives a detailed account on the development of polysaccharide-based composites with nanomaterials that include concise discussion on the types of polysaccharide, methods to prepare polysaccharide nanocomposites, and their applications for drug delivery and biomedicine. Chapter 19 entitled “Natural Polysaccharides for the Delivery of Anticancer Therapeutics” presents a useful discussion on the treatment of cancer by delivering the drugs utilizing polysaccharide-based drug delivery systems. Chapter 20 entitled “Organic nanocomposites for the Delivery of Bioactive Molecules” gives a detailed account on the major methods of composite nanoparticles production, major uses in key areas, potential toxicity, and future perspectives with special emphasis to a critical review of available information regarding their use in biomedical engineering. Chapter 21 entitled “Natural Polysaccharides for Growth Factors Delivery” describes about the usage of various natural polysaccharides for the delivery of different growth factors such as BMP-2, VEGF, PRP, FGF, etc. Chapter 22 entitled “Marine Polysaccharides for Drug Delivery in Tissue Engineering” deals with the marine polysaccharides obtained from the different sources for drug delivery in tissue engineering applications. Chapter 23 entitled “Natural Polysaccharides in Tissue Engineering Applications” deals with the applications of various natural polysaccharides in tissue engineering applications. Chapter 24 entitled “Natural Polysaccharides in Wound Dressings Applications” presents a useful discussion on natural polysaccharide-based dressings along with the leading mathematical models for the release of active compounds from flat systems.

xxv

Preface Chapter 25 entitled “Polysaccharides from Leafy Vegetables: Chemical, Nutritional and Medicinal Properties” gives a detailed account not only on the structural properties of the polysaccharide molecules but also on the broader features of their functionality. This understanding may instigate advanced research on the application of leaf polysaccharides and improved use of leafy vegetables as functional food. Chapter 26 entitled “Electrospun Natural Polysaccharide for Biomedical Application” presents a useful discussion on the electrospinning technique for fabricating nanofibers scaffolds for tissue regenerations, drug delivery systems, wound healing, and cancer therapy. We would like to thank all the contributing authors for their timely and valuable contributions in this book. We also thank the publisher, Elsevier’s Academic Press, Narmatha Mohan, and Sreejith Viswanathan for their invaluable help in organization of the editing process. We especially thank Erin HilleParks and Sandra Harron for their invaluable support throughout the completion of this book. We thankfully acknowledge the permissions to reproduce copyright materials from various sources. Finally, we would like to thank our family members, all respected teachers, friends, colleagues, and dear students for their continuous encouragements, inspirations, and moral supports during the preparation of the book. Together with our contributing authors and the publishers, we will be extremely pleased if our efforts fulfill the needs of academicians, researchers, students, polymer engineers, biomedical experts, pharmaceutical students, and formulators. Dr. Md Saquib Hasnain Dr. Amit Kumar Nayak

xxvi

CHAPTER 1

Natural polysaccharides: sources and extraction methodologies S M Mozammil Hasnain1, Md Saquib Hasnain2, Amit Kumar Nayak3 1

Department of Mechanical Engineering, Birla Institute of Technology, Mesra, Ranchi, India; Department of Pharmacy, Shri Venkateshwara University, Gajraula, India; 3Department of Pharmaceutics, Seemanta Institute of Pharmaceutical Sciences, Mayurbhanj, Odisha, India

2

Chapter Outline 1. Introduction 1 2. Sources of natural polysaccharides 2.1 2.2 2.3 2.4

2

Plant polysaccharides 2 Animal polysaccharides 3 Microbial polysaccharides 4 Algal polysaccharides 4

3. General extraction methodologies of natural polysaccharides 3.1 3.2 3.3 3.4

5

Hot water extraction method 6 Dilute alkali-water extraction method 6 Enzymolysis method 6 Other methods 7

4. Conclusion 7 References 8

1. Introduction In the past few decades, materials from the natural origin have gained wide acceptability in almost all fields of human lives more or less [1e8]. Currently, due to the exceptional biodegradability, sustainability, and higher abundances, as well as cost-effectiveness, natural materials are being widely used as the replacement for synthetic materials [9e13]. Natural polysaccharides, among a variety of natural origin materials, are the major industrial raw materials and have been the focus of thorough research due to above mentioned qualities [1,14e16]. Owing to a broad array of pharmacological activities like antitumor, immune-modulation, antioxidation, and antiinflammatory effects, natural polysaccharides extracted from various natural resources such as plants, animals, fungus, seaweeds, etc., are gaining increasing attention during the past few decades [17]. Natural Polysaccharides in Drug Delivery and Biomedical Applications. https://doi.org/10.1016/B978-0-12-817055-7.00001-7 Copyright © 2019 Elsevier Inc. All rights reserved.

1

2 Chapter 1 Natural polysaccharides comprise of numerous monosaccharide residues, which are interconnected with each other by means of the O e glycosidic linkages [17e19]. These polysaccharides yield simple sugar units like glucose, galactose, mannose, arabinose, xylose, uronic acids, etc., when hydrolyzed [1,20]. Natural polysaccharides, at the cellular level, are either present as the reserve materials in the cytoplasm (e.g., starch), or structural substances of the cell membranes or cell walls (e.g., cellulose) [9]. In general, extraction, purification, and uses of natural polysaccharides depend on their structural characteristics. The core structures of natural polysaccharides are extremely multifaceted, complex, and diverse. Nevertheless, the basic structures of the backbone chain of these natural polysaccharides are mannan, galactan, glucan, xylan, fructan, etc., or a combination of two or more monosaccharidic units (e.g., galactomannan, pectin) [1,17]. The structures of the polysaccharidic branched chains represent the great diversity of the biomacromolecular structural features. The natural polysaccharides possess different physicochemical characteristics as well as functional groups [14,21e24]. These also have some useful advantageous properties such as biocompatibility, biodegradability, nontoxicity, solubility in water, stability, higher degrees of swelling, capability by means of simple chemical modifications, etc [25e27]. These biomaterials also possess an extroverted array of chemical structures, which may be the potential sites for the chemical modifications/functionalizations [23,24,28e34]. The proneness of the natural polysaccharides to the microbial degradations and/or enzymatic actions can be employed as the useful advantages in some cases (e.g., colon targeted drug delivery systems) [26,29]. These polysaccharides are capable of forming a variety of three-dimensional (3D) interconnected structures of molecular networking, well-known as “gels.” The gel strength of natural polysaccharides depends on their molecular structures, polysaccharide concentration in gel compositions, ionic strength, pH, temperature, etc. [16,22,35]. The current chapter describes various sources and extraction methodologies of natural polysaccharides.

2. Sources of natural polysaccharides Depending on the origins (sources), these natural polysaccharides are classified as plant polysaccharides, animal polysaccharides, algal polysaccharides, and microbial polysaccharides [36].

2.1 Plant polysaccharides At present, polysaccharides extracted from plant sources are expansively employed in a number of industrial purposes because of their biodegradability, sustainability, lower processing charge, and higher abundances in nature [37e40]. Various plant polysaccharides

Natural polysaccharides: sources and extraction methodologies 3 Table 1.1: Some important plant polysaccharides and their sources. Sources Plant polysaccharides Gum Arabica Guar gum Sterculia (karaya) gum Locust bean gum Tamarind gum Konjac glucomannan Okra gum Gum kondagogu Cashew tree gum Dillenia fruit gum Moringa gum Abelmoschus gum Albizia gum Terminalia gum Gum odina Gum cordia Ispaghula mucilage Fenugreek seed mucilage Aloe mucilage Mimosa pudica seed mucilage Spinacia oleracea L. leaves mucilage Basella alba L. leaves mucilage Linseed polysaccharide Potato starch Jackfruit seed starch

Botanical names

Family

References

Acacia Arabica Cyamompsis tetraganolobus Sterculia urens Ceratonia siliqua Tamarindus indica Amorphophallus konjac Hibiscus esculentus Cochlospermum gossypium Anacardium occidentale Dillenia indica L. Moringa oleifera Abelmoschus esculentus Albizia procera Terminalia randii Lannea woodier Cordia oblique Plantago ovate Trigonella foenum-graecum L. Aloe barbadensis Mimosa pudica Spinacia oleracea L.

Leguminosae Leguminosae Sterculiaceae Fabaceae Leguminosae Araceae Malvaceae Colchospermaceae Anacardiaceae Dilleniaceae Moringaceae Malvaceae Leguminosae Combretaceae Anacardiaceae Boraginaceae Plantaginaceae Fabaceae Liliaceae Mimosaceae Amaranthaceae

[48,49] [50] [22,51e53] [54] [41,55e61] [62] [63e65] [28] [66e70] [71] [72] [73] [74] [75] [76] [77] [27,78,79] [80,81] [82] [83] [46]

Basella alba L. Linum usitatissimum L., Solanum tuberosum L. Artocarpus heterophyllus

Basellaceae Linaceae Solanaceae Moraceae

[47] [84,85] [86e88] [89e92]

comprise structurally different sets of polysaccharidic macromolecular biomaterials with a broader range of physicochemical characteristics [41e45]. Presently, the search for new sources of plant polysaccharides has been advanced continuously in terms of their variety and properties [46,47]. Some important plant polysaccharides and their sources are mentioned in Table 1.1.

2.2 Animal polysaccharides The polysaccharides of animal origin play an imperative role as their tissue composition demonstrates significant biomedical effect [93]. The extracellular matrix in the animal tissues consisting of an interconnecting lattice of heteropolysaccharides and fibrous proteins are packed with a jellylike substance that supports the cell adhesion as well as cell growth [93,94]. This also facilitates porous pathways for the diffusion of oxygen and nutrients to the individual cells [94]. For example, various heteropolysaccharides are well-known as the glycosaminoglycans, belonging to a group of linear polymers

4 Chapter 1 Table 1.2: Some important animal polysaccharides and their sources. Animal polysaccharides Chitin and chitosan

Chondroitin sulfate Hyaluronic acid

Heparin and heparan sulfate Dermatan sulfate Keratin sulfate

Sources Chitin and chitosan (deacetylated derivative of chitin) extracted from the arthropods; generally exists as powders, granules, or sheets. Chondroitin sulfate (a mucopolysaccharide) occurs in ligaments, cartilage, tendons, etc. Hyaluronic acid (a nonsulfated polysaccharide) is the component of extracellular matrix. In the human body, hyaluronic acid is found in most of the connective tissues. Heparin is biosynthesized and is stored in the mast cells. Heparan sulfate (a proteoglycan) usually occurs on the cellular surface and in the extracellular matrix. Dermatan sulfate (a glycosaminoglycan) occurs mostly in skin, blood vessels, heart valves, and tendons. Keratan sulfate (a sulfated glycosaminoglycan) is occurs especially in the cartilage, cornea, and bone.

References [95,96]

[97,98] [99]

[100]

[101] [102]

comprised of repeating units of disaccharide [93,94]. Different glycosaminoglycans comprise hyaluronic acid, chondroitin sulfate, heparin and heparan sulfate, keratin sulfate, dermatan sulfate, etc. [93,95]. In addition to the glycosaminoglycans, chitin and chitosan (i.e., deacetylated derivative of chitin) also belong to the animal polysaccharides, which are widely exploited in different biomedical uses including drug delivery, tissue engineering, wound healing, etc. [93,96]. Some important animal polysaccharides and their sources are listed in Table 1.2.

2.3 Microbial polysaccharides A huge quantity of polysaccharides is produced by the microorganisms in presence of additional resource of carbon [103]. Some of the polysaccharides work as storage substances and are excreted by the cells. These polysaccharides obtained from the microbes may be neutral (e.g., dextran) or acidic (e.g., gellan gum) in character [104]. These microbial polysaccharides are recently explored as well as exploited for the diverse biomedical applications. Microbial sources, in comparison with plant sources and algae sources, at the commercial scale, are favored for the economical and sustainable production of polysaccharides as the microbial sources facilitate prompt and high yields under fully controlled environment of fermentation [104]. Some important microbial polysaccharides and their sources are listed in Table 1.3.

2.4 Algal polysaccharides Marine algae are very rich in polysaccharides, for instance, glycosaminoglycans (such as agar-agar, alginates, etc.) [115,116]. A large amount of attention has already been directed

Natural polysaccharides: sources and extraction methodologies 5 Table 1.3: Some important microbial polysaccharides and their sources. Microbial polysaccharides Dextrans Xanthan gum Gellan gum Pullulan Curdlan Scleroglucan Emulsan Bacterial alginate

Sources Produced by Leuconostoc mesenteroides, Streptococcus mutans, Acetobacter sp. Produced by the bacterium Xanthomonas campestris Produced by the bacterium Pseudomonas elodea Produced by Aureobasidium pullulans Produced by nonpathogenic bacteria Agrobacterium biovar. Obtained by means of fermentation of Sclerotium rolfsii Produced by Acinetobacter calcoaceticus Produced by the bacteria Pseudomonas aeruginosa and Azotobacter vinelandii

References [105,106] [107,108] [109] [110] [111] [112] [113] [40,114]

Table 1.4: Some important algal polysaccharides and their sources. Algal polysaccharides Agar-agar Alginate

Carrageenan Fucoidan

Sources Agar-agar is extracted from Gelidium amansii, Gelidium cartilagineum, Gelidium conferoides, Gelidium pristoides, etc. From various species of kelp, alginates are extracted. Various brown marine algae such as Laminaria hyperborea, Ascophyllum nodosum, Macrocystis pyrifera, etc., are identified as important primary sources of commercially available alginates. The most common sources are: Eucheuma cottonii and E.spinosum. Produced from Laminaria saccharina, Fucus evanescens, Fucus serratus, Fucus distichus, Fucus spiralis, Fucus vesiculosus, Ascophyllum nodosum, and Cladosiphon okamuranus.

References [118] [119e123]

[124,125] [126]

to the extraction and characterization of marine algal polysaccharides on account of their various health advantages [115]. For this reason, these natural polysaccharides (algal polysaccharides) have already received a rising interest for the use in various biomedical applications [115,116]. These algal polysaccharides are often intimately associated with various pharmacological activities like anticoagulant, antioxidant, antitumor, and immunomodulatory activities [116,117]. Some important algal polysaccharides and their sources are listed in Table 1.4.

3. General extraction methodologies of natural polysaccharides The extraction techniques for these natural origin polysaccharides are somewhat different with respect to small molecules [127]. Moreover, different extraction techniques are employed for diverse kinds of polysaccharides as the different polysaccharides comprise of different characteristics. There are several techniques for the extraction of polysaccharides [127]. The principle behind the selection of extraction method of polysaccharide is to uphold the intrinsic properties of these natural origin polysaccharides without any changes

6 Chapter 1 throughout the progression of extraction. However, in animals/plants, some polysaccharides are present outside of the cell wall (called extracellular polysaccharides or exopolysaccharides), whereas the majority of the polysaccharides are present in the cell wall (called intracellular polysaccharides). Hence, for the extraction of these polysaccharides primary step is to crush the animal/plant materials to release intracellular polysaccharides. In addition to the customary crushing techniques, nowadays, ultrasonic gas flow crushing technology is employed to break the animal/plant cell wall or fungal spores significantly augmenting the extraction efficacy of polysaccharide. By the mechanical crushing, the elimination of lipid has to be done as the animal/plant cell walls are mainly enclosed by lipids. The universal technique employed for removal of lipid is the use of ethanol reflux for a period of 6e8 h in the Soxhlet extractor [128]. Following the elimination of lipids, the raw materials are subjected to the extraction of polysaccharides. There are several extraction methodologies of natural polysaccharides available and among these, four methods are often utilized.

3.1 Hot water extraction method This method is extensively used for the extraction of polysaccharides nowadays and the principle involved behind this methodology is that the majority of the polysaccharides has greater solubility in hot water and are stable enough. Hence, by employing this technique of extraction, polysaccharides can get the minimum destruction. The customary practice for the extraction is to extract in hot water for a period of 2e6 h [129]. If extract is too viscous, then the residue can be separated by means of centrifugation.

3.2 Dilute alkali-water extraction method A number of the acidic or higher molecular weight polysaccharides are not soluble in hot water, whereas solubility of these is greater in the solution of dilute alkali as compared to that in hot water. Consequently, for the extraction of these polysaccharides, 5%e15% (w/w) solution of NaCO3 or NaOH is often used instead of hot water. The temperature for the extraction must be kept lower than 10
C when dilute alkali solution is used to extract as the polysaccharides are vulnerable to degradation. Generally, the extraction of polysaccharides is firstly done in hot water and after that dilute alkali solution is employed for the extraction of residual polysaccharides [127]. Thus, the majority of the polysaccharides from animals/plants are extracted by this technique.

3.3 Enzymolysis method In this method of polysaccharide extraction, the raw plant materials are crushed and then crushed materials are suspended in water. As per the optimum condition of reaction for

Natural polysaccharides: sources and extraction methodologies 7 Table 1.5: Benefits and drawbacks of extraction methods of polysaccharides. Extraction methods of polysaccharides

Benefits

Hot water extraction method

Most widely employed; easy to carry out

Dilute alkali-water extraction method Enzymolysis method

Some acidic polysaccharides can be extracted Mild reaction conditions

Other methods (DMSO; 2-methoxyethanol-LiCl; acidic aqueous solution, etc.)

Seldom employed

Drawbacks Time-consuming In hot water some polysaccharides are not dissolved easily Extraction temperature should be kept below 10
C Seldom employed alone; generally combined with other extraction methods Expensive and low yield

composite enzymes to be employed, first of all optimum temperature and pH are required. After that, the required quantity of composite enzymes is supplemented in the suspension and left to react for a certain time and then, the residue is filtered and the filtrate solution is the polysaccharide extract. This procedure has been used in a number of polysaccharide-based healthcare products [128]. The extensively used method of polysaccharide extraction is the combination of hot water extraction and enzymolysis techniques, that is, firstly, the use of hot water and then enzymolysis to extract polysaccharides [127]. In this combination method, the extracted amounts of polysaccharides are augmented.

3.4 Other methods For the extraction of polysaccharides, the above mentioned methods are employed. There are few other methods apart from these three which are not frequently utilized due to high expense and minimum yield, for example, dimethyl sulfoxide (DMSO) or some organic solvents of alkali metal salts such as 2-methoxyethanol-LiCl or acidic aqueous solution are often utilized to extract out the polysaccharides, etc. [127]. The benefits and drawbacks of all the above mentioned polysaccharide extraction methods are listed in Table 1.5.

4. Conclusion The sources and extraction of polysaccharides are very important issues in polymer science. Natural polysaccharides extracted from various natural resources such as plants, animals, fungus, seaweeds, etc., have gained increasing attention for their various biomedical uses including drug delivery, tissue engineering, wound healing, etc. It is quite

8 Chapter 1 difficult to get the fractions of homogeneous active polysaccharides. This is one of the most important factors to impede the research on polysaccharide developments. The literature studies have enumerated various extraction methodologies to extract different natural polysaccharides, which are somewhat different with respect to small molecules. There are a number of available methods and approaches for extraction and purification of natural polysaccharides. The researchers must cautiously select proper methods for the extraction of natural polysaccharides based on specific properties/characteristics of the polysaccharide to be researched. The present chapter will be helpful to the polysaccharide researchers to provide information about various sources and extraction methodologies of natural polysaccharides.

References [1] Yu Y, Shena M, Song Q, Xie J. Biological activities and pharmaceutical applications of polysaccharide from natural resources: a review. Carbohydr Polym 2018;183:91e101. [2] Sinha Mahapatra SS, Mohanta S, Nayak AK. Preliminary investigation on angiogenic potential of Ziziphus oenoplia M. Root ethanolic extract by chorioallantoic membrane model. Sci Asia 2011;37:72e4. [3] Hasnain MS, Nayak AK, Singh R, Ahmad F. Emerging trends of natural-based polymeric systems for drug delivery in tissue engineering applications. Sci. J. UBU. 2010;1(2):1e13. [4] Jena BK, Ratha B, Kar S, Mohanta S, Nayak AK. Antibacterial activity of the ethanol extract of Ziziphus xylopyrus willd. (Rhamnaceae). Int J Pharm Rev Res 2012;1:46e50. [5] Jena BK, Ratha B, Kar S, Mohanta S, Tripathy M, Nayak AK. Wound healing potential of Ziziphus xylopyrus willd. (Rhamnaceae) stem bark ethanol extract using in vitro and in vivo model. J Drug Deliv Ther 2012;2:41e6. [6] Hati M, Jena BK, Kar S, Nayak AK. Evaluation of anti-inflammatory and anti-pyretic activity of Carissa carandas L. Leaf extract in rats. Int J Pharm Chem Biol Sci 2014;1(1):18e25. [7] Hasnain MS, Ahmad S, Chaudhary N, Hoda MN, Nayak AK. Biodegradable polymer matrix nanocomposites for bone tissue engineering. In: Asiri I,AM, Mohammad A, editors. Applications of nanocomposite materials in orthopedics, a volume in woodhead publishing series in biomaterials. Elsevier Inc.; 2019. p. 1e37. [8] Saini P, Sharma N. Natural polymers used in fast disintegrating tablets: a review. Int J Drug Dev Res 2012;4:18e27. [9] Malafaya PB, Silva GA, Reis RL. Natural-origin polymers as carriers and scaffolds for biomolecules and cell delivery in tissue engineering applications. Adv Drug Deliv Rev 2007;59:207e33. [10] Nayak AK, Pal D, Pradhan J, Hasnain MS. Fenugreek seed mucilage-alginate mucoadhesive beads of metformin HCl: design, optimization and evaluation. Int J Biol Macromol 2013;54:144e54. [11] Nayak AK, Pal D. Natural starches-blended ionotropically-gelled micrparticles/beads for sustained drug release. In: Thakur VK, Thakur MK, Kessler MR, editors. Handbook of composites from renewable materials, volume 8, nanocomposites: advanced applications. USA: WILEY-Scrivener; 2017. p. 527e60. [12] Pal D, Nayak AK, Saha S. Interpenetrating polymer network hydrogels of chitosan: applications in controlling drug release. In: Mondal IH, editor. Cellulose-based superabsorbent hydrogels, polymers and polymeric composites: a reference series. Cham: Springer; 2018. p. 1e41. [13] Nayak AK, Pal D, Santra K. Screening of polysaccharides from tamarind, fenugreek and jackfruit seeds as pharmaceutical excipients. Int J Biol Macromol 2015;79:756e60. [14] Nayak AK, Pal D. Functionalization of tamarind gum for drug delivery. In: Thakur VK, Thakur MK, editors. Functional biopolymers. Switzerland: Springer International Publishing; 2018. p. 35e56.

Natural polysaccharides: sources and extraction methodologies 9 [15] Hasnain MS, Nayak AK. Chitosan as responsive polymer for drug delivery applications. In: Makhlouf ASH, Abu-Thabit NY, editors. Stimuli responsive polymeric nanocarriers for drug delivery applications. Types and triggers, vol. 1. Woodhead Publishing Series in Biomaterials, Elsevier Ltd.; 2018. p. 581e605. [16] Pal D, Nayak AK. Interpenetrating polymer networks (IPNs): natural polymeric blends for drug delivery. In: Mishra M, editor. Encyclopedia of biomedical polymers and polymeric biomaterials, vol. VI. New York, NY 10017, U.S.A.: Taylor & Francis Group; 2015. p. 4120e30. [17] Xie JH, Jin ML, Morris GA, Zha XQ, Chen HQ, Yi Y. Advances on bioactive polysaccharides from medicinal plants. Crit Rev Food Sci Nutr 2016;56:S60e84. [18] Nayak AK, Pal D. Plant-derived polymers: ionically gelled sustained drug release systems. In: Mishra M, editor. Encyclopedia of biomedical polymers and polymeric biomaterials, vol. VIII. New York, NY 10017, U.S.A.: Taylor & Francis Group; 2016. p. 6002e17. [19] Nayak AK, Pal D, Das S. Calcium pectinate-fenugreek seed mucilage mucoadhesive beads for controlled delivery of metformin HCl. Carbohydr Polym 2013;96:349e57. [20] Prajapati VD, Jani GK, Moradiya NG, Randeria NP. Pharmaceutical applications of various natural gums, mucilages and their modified forms. Carbohydr Polym 2013;92:1685e99. [21] Nayak AK, Hasnain MS, Pal D. Gelled microparticles/beads of sterculia gum and tamarind gum for sustained drug release. In: Thakur VK, Thakur MK, editors. Handbook of springer on polymeric gel. Switzerland: Springer International Publishing; 2018. p. 361e414. [22] Nayak AK, Pal D. Sterculia gum-based hydrogels for drug delivery applications. In: Kalia S, editor. Polymeric hydrogels as smart biomaterials, springer series on polymer and composite materials. Switzerland: Springer International Publishing; 2016. p. 105e51. [23] Nayak AK, Pal D. Chitosan-based interpenetrating polymeric network systems for sustained drug release. In: Tiwari A, Patra HK, Choi J-W, editors. Advanced theranostics materials. USA: WILEY-Scrivener; 2015. p. 183e208. [24] Jana S, Saha A, Nayak AK, Sen KK, Basu SK. Aceclofenac-loaded chitosan-tamarind seed polysaccharide interpenetrating polymeric network microparticles. Colloids Surfaces B Biointerfaces 2013;105:303e9. [25] Pal D, Nayak AK. Alginates, blends and microspheres: controlled drug delivery. In: Mishra M, editor. Encyclopedia of biomedical polymers and polymeric biomaterials, vol. I. New York, NY 10017, U.S.A.: Taylor & Francis Group; 2015. p. 89e98. [26] Nayak AK. Tamarind seed polysaccharide-based multiple-unit systems for sustained drug release. In: Kalia S, Averous L, editors. Biodegradable and bio-based polymers: environmental and biomedical applications. USA: WILEY-Scrivener; 2016. p. 471e94. [27] Guru PR, Bera H, Das M, Hasnain MS, Nayak AK. Aceclofenac-loaded Plantago ovata F. Husk mucilage-Znþ2-pectinate controlled-release matrices. Starch e Sta¨rke. 2018;70:1700136. [28] Malik S, Kumar A, Ahuja M. Synthesis of gum kondagogu- g-poly (N-vinyl-2-pyrrolidone) and its evaluation as a mucoadhesive polymer. Int J Biol Macromol 2012;51(5):756e62. [29] Nayak AK, Bera H, Hasnain MS, Pal D. Synthesis and characterization of graft-copolymers of plant polysaccharideses. In: Thakur VK, editor. Biopolymer grafting, synthesis and properties. Elsevier Inc.; 2018. p. 1e62. [30] Bera H, Boddupalli S, Nayak AK. Mucoadhesive-floating zinc-pectinate-sterculia gum interpenetrating polymer network beads encapsulating ziprasidone HCl. Carbohydr Polym 2015;131:108e18. [31] Jana S, Manna S, Nayak AK, Sen KK, Basu SK. Carbopol gel containing chitosan-egg albumin nanoparticles for transdermal aceclofenac delivery. Colloids Surfaces B Biointerfaces 2014;114:36e44. [32] Malakar J, Nayak AK, Das A. Modified starch (cationized)-alginate beads containing aceclofenac: formulation optimization using central composite design. Starch e Sta¨rke. 2013;65:603e12. [33] Jana S, Das A, Nayak AK, Sen KK, Basu SK. Aceclofenac-loaded unsaturated esterified alginate/gellan gum microspheres: in vitro and in vivo assessment. Int J Biol Macromol 2013;57:129e37.

10 Chapter 1 [34] Jana S, Maji N, Nayak AK, Sen KK, Basu SK. Development of chitosan-based nanoparticles through inter-polymeric complexation for oral drug delivery. Carbohydr Polym 2013;98:870e6. [35] Nayak AK, Ara TJ, Hasnain MS, Hoda N. Okra gum-alginate composites for controlled releasing drug delivery. In: Asiri I,AM, Mohammad A, editors. Applications of nanocomposite materials in drug delivery, a volume in woodhead publishing series in biomaterials. Elsevier Inc.; 2018. p. 761e85. [36] Nayak AK, Pal D. Natural polysaccharides for drug delivery in tissue engineering. Everyman’s Sci 2012;46:347e52. [37] Nayak AK, Hasnain MS. Plant polysaccharides in drug delivery applications. In: Nayak AK, Hasnain MS, editors. Plant polysaccharides-based multiple-unit systems for oral drug delivery. Singapore: Springer; p. 19e23. [38] Nayak AK, Hasnain MS. Some other plant polysaccharide based multiple units for oral drug delivery. In: Nayak AK, Hasnain MS, editors. Plant polysaccharides-based multiple-unit systems for oral drug delivery. Singapore: Springer; p. 123e128. [39] Beneke CE, Viljoen AM, Hamman JH. Polymeric plant-derived excipients in drug delivery. Molecules 2009;14:2602e20. [40] Pal D, Nayak AK. Plant polysaccharides-blended ionotropically-gelled alginate multiple-unit systems for sustained drug release. In: Thakur VK, Thakur MK, Kessler MR, editors. Handbook of composites from renewable materials, volume 6, polymeric composites. USA: WILEY-Scrivener; 2017. p. 399e400. [41] Nayak AK, Pal D. Tamarind seed polysaccharide: an emerging excipient for pharmaceutical use. Indian. J. Pharm. Edu. 2017;51:S136e46. [42] Nayak AK, Pal D. Trigonella foenum-graecum L. Seed mucilage-gellan mucoadhesive beads for controlled release of metformin HCl. Carbohydr Polym 2014;107:31e40. [43] Nayak AK, Pal D, Santra K. Development of pectinate-ispagula mucilage mucoadhesive beads of metformin HCl by central composite design. Int J Biol Macromol 2014;66:203e21. [44] Nayak AK, Pal D, Santra K. Tamarind seed polysaccharide-gellan mucoadhesive beads for controlled release of metformin HCl. Carbohydr Polym 2014;103:154e63. [45] Nayak AK, Pal D, Santra K. Development of calcium pectinate-tamarind seed polysaccharide mucoadhesive beads containing metformin HCl. Carbohydr Polym 2014;101:220e30. [46] Nayak AK, Pal D, Pany DR, Mohanty B. Evaluation of Spinacia oleracea L. Leaves mucilage as innovative suspending agent. J Adv Pharm Technol Res 2010;1(3):338e41. [47] Pal D, Nayak AK, Kalia S. Studies on Basella alba L. Leaves mucilage: evaluation of suspending properties. Int. J. Drug Discov. and Technol. 2010;1(1):15e20. [48] Nayak AK, Das B, Maji R. Calcium alginate/gum Arabic beads containing glibenclamide: development and in vitro characterization. Int J Biol Macromol 2012;51:1070e8. [49] Nayak AK, Hasnain MS. Gum Arabic based multiple units for oral drug delivery. In: Nayak AK, Hasnain MS, editors. Plant polysaccharides-based multiple-unit systems for oral drug delivery. Singapore: Springer, p. 25e30. [50] Krishnaiah YSR, Rama Rao T, Ushasree M, Satyanarayana S. A study on the in vitro evaluation of guar gum as a carrier for oral controlled drug delivery. Saudi Pharmaceut J 2001;9(2):91e8. [51] Nayak AK, Hasnain MS. Sterculia gum based multiple units for oral drug delivery. In: Nayak AK, Hasnain MS, editors. Plant polysaccharides-based multiple-unit systems for oral drug delivery. Singapore: Springer, p. 67e82. [52] Bera H, Kandukuri SG, Nayak AK, Boddupalli S. Alginate-sterculia gum gel-coated oil-entrapped alginate beads for gastroretentive risperidone delivery. Carbohydr Polym 2015;120:74e84. [53] Guru PR, Nayak AK, Sahu RK. Oil-entrapped sterculia gum-alginate buoyant systems of aceclofenac: development and in vitro evaluation. Colloids Surfaces B Biointerfaces 2013;104:268e75. [54] Nayak AK, Hasnain MS. Locust bean gum based multiple units for oral drug delivery. In: Nayak AK, Hasnain MS, editors. Plant polysaccharides-based multiple-unit systems for oral drug delivery. Singapore: Springer, p. 61e66.

Natural polysaccharides: sources and extraction methodologies 11 [55] Nayak AK, Hasnain MS. Tamarind polysaccharide based multiple units for oral drug delivery. In: Nayak AK, Hasnain MS, editors. Plant polysaccharides-based multiple-unit systems for oral drug delivery. Singapore: Springer, p. 31e59. [56] Nayak AK, Pal D, Santra K. Swelling and drug release behavior of metformin HCl-loaded tamarind seed polysaccharide-alginate beads. Int J Biol Macromol 2016;82:1023e7. [57] Bera H, Boddupalli S, Nandikonda S, Kumar S, Nayak AK. Alginate gel-coated oil-entrapped alginateetamarind gumemagnesium stearate buoyant beads of risperidone. Int J Biol Macromol 2015;78:102e11. [58] Nayak AK, Pal D, Malakar J. Development, optimization and evaluation of emulsion-gelled floating beads using natural polysaccharide-blend for controlled drug release. Polym Eng Sci 2013;53:338e50. [59] Nayak AK, Pal D. Ionotropically-gelled mucoadhesive beads for oral metformin HCl delivery: formulation, optimization and antidiabetic evaluation. J Sci Ind Res 2013;72:15e22. [60] Pal D, Nayak AK. Novel tamarind seed polysaccharide-alginate mucoadhesive microspheres for oral gliclazide delivery. Drug Deliv 2012;19:123e31. [61] Nayak AK, Pal D. Development of pH-sensitive tamarind seed polysaccharide-alginate composite beads for controlled diclofenac sodium delivery using response surface methodology. Int J Biol Macromol 2011;49:784e93. [62] Fan J, Wang K, Liu M, He Z. In vitro evaluations of konjac glucomannan and xanthan gum as the sustained release material of matrix tablets. Carbohydr Polym 2008;73(2):241e7. [63] Sinha P, Ubaidulla U, Hasnain MS, Nayak AK, Rama B. Alginate-okra gum blend beads of diclofenac sodium from aqueous template using ZnSO4 as a cross-linker. Int J Biol Macromol 2015;79:555e63. [64] Sinha P, Ubaidulla U, Nayak AK. Okra (Hibiscus esculentus) gum-alginate blend mucoadhesive beads for controlled glibenclamide release. Int J Biol Macromol 2015;72:1069e75. [65] Nayak AK, Hasnain MS. Okra gum based multiple units for oral drug delivery. In: Nayak AK, Hasnain MS, editors. Plant Polysaccharides-Based Multiple-Unit Systems for Oral Drug Delivery. Singapore: Springer, p. 83e92. [66] Hasnain MS, Rishishwar P, Ali S. Use of cashew bark exudate gum in the preparation of 4% lidocaine HCl topical gels. Int J Pharm Pharm Sci 2017;9:146e50. [67] Hasnain MS, Rishishwar P, Ali S. Floating-bioadhesive matrix tablets of hydralazine HCl made of cashew gum and HPMC K4M. Int J Pharm Pharm Sci 2017;9:124e9. [68] Hasnain MS, Rishishwar P, Rishishwar S, Ali S, Nayak AK. Extraction and characterization of cashew tree (Anacardium occidentale) gum; use in aceclofenac dental pastes. Int J Biol Macromol 2018;116:1074e81. [69] Das B, Dutta S, Nayak AK, Nanda U. Zinc alginate-carboxymethyl cashew gum microbeads for prolonged drug release: development and optimization. Int J Biol Macromol 2014;70:505e15. [70] Das B, Nayak AK, Nanda U. Topical gels of lidocaine HCl using cashew gum and carbopol 940: preparation and in vitro skin permeation. Int J Biol Macromol 2013;62:514e7. [71] Ketousetuo K, Bandyopadhyay AK. Development of oxytocin nasal gel using natural mucoadhesive agent obtained from the fruits of Dellinia indica. L. Sci. Asia. 2007;33:57e60. [72] Panda DS, Choudhury NSK, Yedukondalu M, Si S, Gupta R. Evaluation of gum of Moringa oleifera as a binder and release retandant in tablet formulation. Indian J Pharm Sci 2008;70(5):614e8. [73] Ofoefule SI, Chukwu A. Application of Abelmoschus esculantus gum as a mini-matrix for furosemide and diclofenac sodium tablets. Indian J Pharm Sci 2001;63(6):532e5. [74] Pachuau L, Mazumdar B. Albizia procera gum as an excipient for oral controlled release matrix tablets. Carbohydr Polym 2012;90(1):284e95. [75] Bamiro OA, Odeku OA, Sinha VR, Kumar R. Terminalia gum as a directly compressable excipient for controlled drug delivery. AAPS PharmSciTech 2012;13(1):16e24. [76] Jena AK, Nayak AK, De A, Mitra D, Samanta A. Development of lamivudine containing multiple emulsions stabilized by gum odina. Future J Pharmaceut Sci 2018;4:71e9.

12 Chapter 1 [77] Ahuja M, Yadav M, Kumar S. Application of response surface methodology to formulation of ionotropically gelled gum cordial/gellan beads. Carbohydr Polym 2010;80(1):161e7. [78] Nayak AK, Pal D, Santra K, Plantago ovata F. Mucilage-alginate mucoadhesive beads for controlled release of glibenclamide: development, optimization, and in vitro-in vivo evaluation. J Pharm 2013. Article ID 151035. [79] Nayak AK, Hasnain MS, Beg S, Alam MI. Mucoadhesive beads of gliclazide: design, development and evaluation. Sci Asia 2010;36(4):319e25. [80] Nayak AK, Hasnain MS. Fenugreek seed mucilage based multiple units for oral drug delivery. In: Nayak AK, Hasnain MS, editors. Plant polysaccharides-based multiple-unit systems for oral drug delivery. Singapore: Springer; p. 93e112. [81] Nayak AK, Pal D, Pradhan J, Ghorai T. The potential of Trigonella foenum-graecum L. Seed mucilage as suspending agent. Ind J Pharm Educ Res 2012;46:312e7. [82] Jani GK, Shah DP, Jani VC. Evaluating mucilage from Aloe barbadensis millar as a pharmaceutical excipient for sustained release matrix tablets. Pharmaceut Technol 2007;31(11):90e8. [83] Singh K, Kumar A, Langyan N, Ahuja M. Evaluation of Mimosa pudica seed mucilage as sustainedrelease excipient. AAPS PharmSciTech 2009;10(4):1121e7. [84] Nayak AK, Hasnain MS. Linseed polysaccharide based multiple units for oral drug delivery. In: Nayak AK, Hasnain MS, editors. Plant polysaccharides-based multiple-unit systems for oral drug delivery. Singapore: Springer; p. 117e121. [85] Hasnain MS, Rishishwar P, Rishishwar S, Ali S, Nayak AK. Isolation and characterization of Linum usitatisimum polysaccharide to prepare mucoadhesive beads of diclofenac sodium. Int J Biol Macromol 2018;116:162e72. [86] Nayak AK, Hasnain MS. Potato starch based multiple units for oral drug delivery. In: Nayak AK, Hasnain MS, editors. Plant polysaccharides-based multiple-unit systems for oral drug delivery. Singapore: Springer; p. 113e116. [87] Malakar J, Dutta P, Purokayastha SD, Dey S, Nayak AK. Floating capsules containing alginate-based beads of salbutamol sulfate: in vitro-in vivo evaluations. Int J Biol Macromol 2014;64:181e9. [88] Malakar J, Nayak AK, Jana P, Pal D. Potato starch-blended alginate beads for prolonged release of tolbutamide: development by statistical optimization and in vitro characterization. Asian J Pharm 2013;7:43e51. [89] Nayak AK, Pal D, Santra K. Artocarpus heterophyllus L. Seed starch-blended gellan gum mucoadhesive beads of metformin HCl. Int J Biol Macromol 2014;65:329e39. [90] Nayak AK, Pal D. Blends of jackfruit seed starch-pectin in the development of mucoadhesive beads containing metformin HCl. Int J Biol Macromol 2013;62:137e45. [91] Nayak AK, Pal D. Formulation optimization of jackfruit seed starch-alginate mucoadhesive beads of metformin HCl. Int J Biol Macromol 2013;59:264e72. [92] Nayak AK, Pal D, Hasnain MS. Development and optimization of jackfruit seed starch-alginate beads containing pioglitazone. Curr Drug Deliv 2013;10:608e19. [93] Li Q, Niu Y, Xing P, Wang C. Bioactive polysaccharides from natural resources including Chinese medicinal herbs on tissue repair. Chin Med 2018;13:7. [94] De Angelis PL. Glycosaminoglycan polysaccharide biosynthesis and production: today and tomorrow. Appl Microbiol Biotechnol 2012;94(2):295e305. [95] Ray S, Sinha P, Laha B, Maiti S, Bhattacharyya UK, Nayak AK. Polysorbate 80 coated crosslinked chitosan nanoparticles of ropinirole hydrochloride for brain targeting. J Drug Deliv Sci Technol 2018;48:21e9. [96] Verma A, Dubey J, Verma N, Nayak AK. Chitosan-hydroxypropyl methylcellulose matrices as carriers for hydrodynamically balanced capsules of moxifloxacin HCl. Curr Drug Deliv 2017;14:83e90. [97] Juqun X, Ling Z, Hua D. Drug-loaded chondroitin sulfate-based nanogels: preparation and characterization. Colloids Surfaces B Biointerfaces 2012;100:107e15.

Natural polysaccharides: sources and extraction methodologies 13 [98] Mohtashamiana S, Boddohia S, Hosseinkhani S. Preparation and optimization of self-assembled chondroitin sulfate-nisin nanogel based on quality by design concept. Int J Biol Macromol 2018;107B:2730e9. [99] Papakonstantinou E, Roth M, Karakiulakis G. Hyaluronic acid: a key molecule in skin aging. Derm Endocrinol 2012;4(3):253e8. [100] Casu B, Naggi A, Torri G. Re-visiting the structure of heparin. Carbohydr Res 2015;403:60e8. [101] Trowbridge JM, Gallo RL. Dermatan sulfate: new functions from an old glycosaminoglycan. Glycobiology 2002;12(9):117R. 25R. [102] Funderburgh JL. Keratan sulfate: structure, biosynthesis, and function. Glycobiology 2000;10(10):951e8. [103] Fiume MM, Heldreth B, Bergfeld WF, Belsito DV, Hill RA, Klaassen CD, Liebler DC, Marks Jr JG, Shank RC, Slaga TJ, Snyder PW, Andersen FA. Safety assessment of microbial polysaccharide gums as used in cosmetics. Int J Toxicol 2016;35(1):5Se49S. [104] Bhatia S. Microbial polysaccharides as advance nanomaterials. In: Bhatia S, editors. Systems for drug delivery, safety, animal, and microbial polysaccharides. Switzerland: Springer International Publishing; p. 29e54. [105] Ochrimenko S, Vollrath A, Tauhardt L. Dextran-graft-linear poly(ethylene imine)s for gene delivery: importance of the linking strategy. Carbohydr Polym 2014;113(9):597e606. [106] Anirudhan TS, Binusreejayan. Dextran based nanosized for the controlled and targeted delivery of curcumin to liver cancer cells. Int J Biol Macromol 2016;88:222e35. [107] Shiledar RR, Tagalpallewar AA, Kokare CR. Formulation and in vitro evaluation of xanthan gum-based bilayered mucoadhesive buccal patches of zolmitriptan. Carbohydr Polym 2014;101:1234e42. [108] Shalviri A, Liu Q, Abdekhodaie MJ, Wu XY. Novel modified starch-xanthan gum hydrogels for controlled drug delivery: synthesis and characterization. Carbohydr Polym 2010;79:898e907. [109] Nayak AK, Pal D, Santra K. Ispaghula mucilage-gellan mucoadhesive beads of metformin HCl: development by response surface methodology. Carbohydr Polym 2014;107:41e50. [110] Guo H, Liu Y, Wang Y, Wu J, Yang X, Li R, Wang Y, Zhang N. pH-sensitive pullulan-based nanoparticle carrier for adriamycin to overcome drug-resistance of cancer cells. Carbohydr Polym 2014;111:908e17. [111] Han J, Cai J, Borjihan W, Ganbold T, Rana TM, Baigude H. Preparation of novel curdlan nanoparticles for intracellular siRNA delivery. Carbohydr Polym 2015;117:324e30. [112] Coviello T, Grassi M, Rambone G, Alhaique F. A crosslinked system from scleroglucan derivate: preparation and characterization. Biomaterials 2001;22:1899. 909. [113] Rubinovitz C, Gutnick DL, Rosenberg E. Emulsan production by Acinetobacter calcoaceticus in the presence of chloramphenicol. J Bacteriol 1982;152(1):126e32. [114] Hasnain MS, Nayak AK. Alginate-inorganic composite particles as sustained drug delivery matrices. In: Asiri I,AM, Mohammad A, editors. Applications of nanocomposite materials in drug delivery, a volume in woodhead publishing series in biomaterials. Elsevier Inc.; 2018. p. 39e74. [115] Xu SY, Huang X, Cheong KL. Recent advances in marine algae polysaccharides: isolation, structure, and activities. Mar Drugs 2017;15(12):388. [116] de Jesus Raposo MF, de Morais AM, de Morais RMSC. Marine polysaccharides from algae with potential biomedical applications. Mar Drugs 2015;13:2967e3028. ¨ zogul F, O ¨ zogul Y, Regenstein JM. Marine bioactive compounds and their health benefits: a [117] Hamed I, O review. Compr Rev Food Sci Food Saf 2015;14:446e65. [118] Armise´n R, Agar GF. Handbook of hydrocolloids. 2nd ed. Woodhead publishing; 2009. p. 82e107. [119] Nayak AK, Beg S, Hasnain MS, Malakar J, Pal D. Soluble starch-blended Ca2þ-Zn2þ-alginate composites-based microparticles of aceclofenac: formulation development and in vitro characterization. Fut. J. Pharma. Sci. 2018;4:63e70. [120] Hasnain MS, Nayak AK, Singh M, Tabish M, Ansari MT, Ara TJ. Alginate-based bipolymericnanobioceramic composite matrices for sustained drug release. Int J Biol Macromol 2016;83:71e7.

14 Chapter 1 [121] Jana S, Gangopadhaya A, Bhowmik BB, Nayak AK, Mukhrjee A. Pharmacokinetic evaluation of testosterone-loaded nanocapsules in rats. Int J Biol Macromol 2015;72:28e30. [122] Malakar J, Das K, Nayak AK. In situ cross-linked matrix tablets for sustained salbutamol sulfate release e formulation development by statistical optimization. Polym Med 2014;44:221e30. [123] Malakar J, Nayak AK, Pal D. Development of cloxacillin loaded multiple-unit alginate-based floating system by emulsionegelation method. Int J Biol Macromol 2012;50(1):138e47. [124] Hezaveh H, Muhamad II . The effect of nanoparticles on gastrointestinal release from modified k-carrageenan nanocomposite hydrogels. Carbohydr Polym 2012;89(1):138e45. [125] Li L, Ni R, Shao Y, Mao S. Carrageenan and its applications in drug delivery. Carbohydr Polym 2014;103:1e11. [126] Isnansetyo A, Laili Lutfia FN, Nursid M, Trijoko SRA. Cytotoxicity of fucoidan from three tropical brown algae against breast and colon cancer cell lines. Phcog J 2017;9(1):14e20. [127] Shi L. Bioactivities, isolation and purification methods of polysaccharides from natural products: a review. Int J Biol Macromol 2016;92:37e48. [128] Chaplin MF, Kennedy JF. Carbohydrate analysis a practical approach. 2nd ed. Oxford: Oxford University Press; 1994. [129] Bao XF, Duan JY, Fang XY, Fang JN. Chemical modifications of the (1/3)-a-D-glucan from spores of Ganoderma lucidum and investigation of their physicochemical properties and immunological activity. Carbohydr Res 2001;336:127e40.

CHAPTER 2

Pharmaceutical applications of natural polysaccharides Zahra Shariatinia Department of Chemistry, Amirkabir University of Technology (Tehran Polytechnic), Tehran, Iran

Chapter Outline List 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12.

of abbreviations 15 Introduction 16 Application of natural polysaccharides Application of natural polysaccharides Application of natural polysaccharides Application of natural polysaccharides Application of natural polysaccharides Application of natural polysaccharides Application of natural polysaccharides Application of natural polysaccharides Application of natural polysaccharides Application of natural polysaccharides Application of natural polysaccharides additives/packaging materials 48 13. Conclusion 50 Acknowledgment 50 References 50

in in in in in in in in in in in

cell encapsulation 19 pharmaceutics/drug delivery 29 gene delivery 33 protein binding 36 wound healing 38 tissue engineering 41 bioimaging 42 preparation of contact lenses 45 preparation of implants 46 preparation of antibacterial textiles/papers preparation of antimicrobial food

List of abbreviations AgNPs ALG-PAAm AO/PI BADSCs BSA CMCel CMT CMV

Silver nanoparticles Alginate-polyacrylamide Acridine orange/propidium iodide Brown adipose-derived stem cells Bovine serum albumin carboxymethyl cellulose Carboxymethyl tamarind Cytomegalovirus

Natural Polysaccharides in Drug Delivery and Biomedical Applications. https://doi.org/10.1016/B978-0-12-817055-7.00002-9 Copyright © 2019 Elsevier Inc. All rights reserved.

15

47

16 Chapter 2 CO CS CS CS-NS Cur CVB3 DOX E. coli GAGs GEM GK GT HA HASCs HIV HMSN HSV IOLs MNPs MPECHs MR n-HAp/CS-TSP NS NT2 PGA PVP QDs QPG S. aureus SCL SiRNA SO SPI TCL TML

Clove oil Chitosan St-AgNPs: Chitosan:starch-silver nanoparticles Chitosaneneem seed Curcumin Coxsackievirus B3 Doxorubicin hydrochloride Escherichia coli Glycosaminoglycans Gemcitabine hydrochloride Gum karaya Gum tragacanth Hyaluronic acid Human adiposeederived stem cells Human immunodeficiency virus Hollow mesoporous silica nanoparticles Herpes simplex virus Intraocular lens Magnetic nanoparticles Magnetic responsive polyelectrolyte complex hydrogels Mauran Nano-hydroxyapatite/chitosan-tamarind seed polysaccharide Nanostarch NTera2 Propylene glycol alginate Poly(vinyl pyrrolidone) Quantum dots Quaternized analogs of pectic galactan Staphylococcus aureus Soft contact lens Small interference RNA Sandalwood oil Soybean protein isolate Therapeutic contact lens Timolol maleate

1. Introduction Natural polymers are more preferred compared to the synthetic ones because they are easily accessible, prone to chemical modification, renewable, economic, nontoxic, stable, hydrophilic, biocompatible, and biodegradable compared to expensive synthetic polymers that have shown environmental and toxicity problems along with long-time synthetic methods [1e4]. Polysaccharides are the most plentiful natural biopolymers which are of growing attraction as effective materials in various biomedical fields, and this can be due to their inherent exceptional valuable features [5e9]. Polysaccharides possess several functional groups and display variable physicochemical properties and essential biological activities that make them suitable materials in numerous pharmaceutical areas such as drug

Pharmaceutical applications of natural polysaccharides 17 delivery and tissue engineering [10e13]. Thus, the number of natural carbohydrates used to deliver desired materials for particular pharmaceutical applications is increasing [14e16]. Encapsulation of living cells into polymeric materials is usually done with the aim of protecting the cells against destruction by the immune system. This method was first introduced in 1933 when Bisceglie et al. investigated the encapsulation influence on the survival of tumor cells in the abdominal cavities of pigs and found that long cell survival was attained through enveloping cells by immunoprotective membranes [17]. Hence, Bisceglie used amnion tissue membrane; however, he did not distinguish the ability of such method to treat diseases. In 1950, Algire and coworkers presented the idea of the “diffusion chamber” in order to graft therapeutic cells [18]. Also, they emphasized on the significance of using biocompatible polymers having sustained and favored properties which are a requirement for therapeutic purposes [18]. Since then, numerous people employed the encapsulation route to treat diverse kinds of diseases [19]. Numerous diseases can be overcome using this technology including anemia [20], hemophilia B [21], dwarfism [22], pituitary disorders [23], liver [24] and kidney failure [25], diabetes mellitus [26], and central nervous system insufficiency [27]. Today, encapsulation in polymers is one of the most frequently utilized cell immobilization technologies because of its simple and mild preparation procedures [28]. As an example, alginate is widely used in cell encapsulation [29]. Polysaccharides are also attractive candidates for gene delivery. Moreover, the modification of nonionic and hydrophilic polysaccharides improves the half-life of polyplexes in blood circulation through avoiding their unwanted interactions with serum proteins thus preventing their clearance with the reticuloendothelial cells [30,31]. Recently, the interactions that occur between biodegradable natural polysaccharides and therapeutic proteins have broadly been examined [32e36]. Such interactions are very important to assess the macroscopic characteristics of processed foods like texture, flow, and stability [37]. Polysaccharides can be processed as ingredients of functional foods and serve as edible films [38], interfacial stabilizers [39,40], microcapsules [41,42], electrostatic gels [43,44], and alternatives for meat or fat [45]. Furthermore, the interactions that occur between polysaccharides and proteins are essential to determine the physical and structural properties of formulated foods [46,47]. It is known that limited drugs are accessible in modern medicine to stimulate wound healing process. Besides, despite the considerable progress in this area, highly safe and effective wound healing treatments are not available. Consequently, advanced investigations have been done to find innovative natural medicines with potential activities to hasten and increase wound healing effect during the healing progression [48]. Currently, polysaccharide biopolymers have been found as promising agents in different forms [49].

18 Chapter 2 For instance, it was exhibited that plant polysaccharides accelerated healing, modulated the inflammatory stage [50e52], and stimulated proliferation of dermal keratinocytes and fibroblasts [53]. Natural biomaterials are commonly exploited as scaffolds in tissue engineering applications in various forms such as bioceramics, hydrogels, and composites (nanocomposites/ biocomposites) [54e56]. The increased attention to the natural polymers is mostly related to the growing concerns and awareness to the environmental problems of plastics subsequent to their being discarded in the environment post usage. Indeed, natural and biobased polymers can decrease plastic waste production and CO2 emission [57]. Moreover, natural biodegradable polymers such as proteins and polysaccharides are significantly similar to the extracellular matrix, chemically adaptable, biodegradable, biocompatible, and reveal required biological potential [58,59] for making biobased polymers. After the pioneer application of chitosan (CS) and its derivatives in the aqueous colloidal synthesis of CdS quantum dots (QDs) [60], QDs capped with carbohydrate polymers were increasingly used in chemical analysis and biomedical applications [61e65]. For instance, carbon quantum dots possessing tunable and very strong fluorescence properties were employed in optronics, sensors, and biomedicines [66]. The systemic administration of medicines in ocular diseases is usually unsuccessful, primarily due to the bloodeocular barrier. The ocular barriers are blooderetinal and bloodeaqueous barriers that mainly prevent drug absorption from the blood [67,68]. Recently, drug-laden contact lenses are widely used to treat ocular illnesses. Contact lenses are curved and thin plastic lenses that are worn on the cornea for protection of the eye and/or to correct vision [69]. Currently, contact lenses can serve as drug carriers to control anterior segment diseases. Contact lenses are able to enhance the drugs’ residence time period on the eye to >30 min, and this is longer compared to 2 min when eye drops are used, confirming they can enhance bioavailability of drug on the cornea [70]. Additionally, the exposure of drug in the systemic circulation along with its side effects will be decreased. The drug-laden contact lenses could be utilized for longer times by patients to decrease required administration rate. Titanium and its alloys are commonly applied as implants in the dental and orthodontic areas because they have shown exceptional mechanical properties, osteoconductivity, and biocompatibility [71,72]. Nevertheless, infection of implants is a serious clinical problem which has led to implant failure, long hospitalization, and also death [73,74]. The bioinert characteristic is the inherent shortcoming of titanium-based implants [75]. As a result, scientists tried to modify the titanium surface by coatings and/or nanostructures in order to increase osteogenesis inducing capability and inhibit bacterial growth [76e78]. For example, coating the titanium surface by antibacterial materials such as chitosan is of great interest [79].

Pharmaceutical applications of natural polysaccharides 19 Production of antibacterial textiles is an imperative part in the textile manufacturing. Although several chemicals and approaches are existing to produce antibacterial textiles, some of them are toxic to humans [80,81]. Thus, in order to overcome this drawback, natural polymers as ecofriendly and biocompatible materials are employed in fabrication of antibacterial textiles. It is well known that papers are used for diverse applications including in paper indicators; sensors; filters; printing/writing, packaging, and household products [82e84]. Paper has unique features such as low weight, low cost, low environmental impact, and suitable mechanical properties [85]. However, paper has poor barrier properties along with poor grease/oil resistance [86]. They are susceptible to react with fat molecules because they have lipophilic properties that lead to damaging the printed papers. Several polymers like chitosan were examined to alter the surface of papers with the purpose of enhancing their characteristics [87,88]. Because application of synthetic polymers results in environmental problems and their recycling is hard, ecofriendly preparation methods are broadly explored to achieve antimicrobial papers [89,90]. The frequent usage of plastic packaging materials in the food industry has brought about pollution problems. Accordingly, application of biodegradable packaging biopolymers has been proposed in order to decrease the environmental contamination [91]. An antimicrobial packaging can interact with products or headspace in order to decrease, delay, and/or inhibit the development of microorganisms which can exist on food surfaces [92]. This strategy can tackle the contamination of food, reduce danger of pathogen growing, and extend the food shelf life. Also, the increasing demand for using natural preservatives and additive-free food motivate the introduction of natural antimicrobial agents to the packaging materials that can gradually migrate to the food medium and eliminate the requirement for consuming extra amounts of preservatives that are directly introduced into the food products [93,94]. In this chapter, the most recent research results on pharmaceutical applications of natural polysaccharides in miscellaneous biomedical areas including cell encapsulation, drug/gene delivery, wound healing, protein binding, tissue engineering, bioimaging, preparation of contact lenses and implants, antibacterial textiles/papers, and antibacterial food additives/ packaging materials (Scheme 2.1) will be presented.

2. Application of natural polysaccharides in cell encapsulation Cell-based therapy includes implantation or delivery of living cells or their sustained growth in patients to treat certain diseases [95]. On contrary to small drug molecules and biologicals like engineered antibodies and proteins that are the principal therapeutic materials for various diseases, the cell-based therapy transports complex living compounds

20 Chapter 2

Cell encapsulation ation

Drug delivery

Gene delivery

Wound healing g

P Protein binding

Bioimaging

Natural Polysaccharides Tissue engineering ing Implants

Antibacterial textiles/papers p p

Contact lens Food packaging

Food additives F

Scheme 2.1 The different pharmaceutical applications of natural polysaccharides.

to modulate their function, sensing, and responding to the environment. One of the foremost advantages of employing cell encapsulation is its potential to use allogenic cells for transplantation as it can protect encapsulated cells from being destroyed by the immune system [96]. Also, it can decrease usage of antiimmunoresponse therapeutics and subsequent influences of traditional transplants through avoiding the body immunoresponse [97]. Cell encapsulation leads to the sustained drug release from encapsulated cells in comparison to the common drug administration that caused underdosing or overdosing effects [96]. Besides, cell encapsulation is able to resolve the challenge of drug localization in a place because encapsulated cells are transplanted to desired site(s) with superior efficacy and lesser whole dosage than traditional approaches [98]. Some therapeutic applications for encapsulated cells are treatments of diabetes, degenerative bone/joint disease, hepatic and central nervous system diseases. It is known that brown adipose derived stem cells (BADSCs) are favorite stem cells for the treatment of myocardial infarction because they can effectively and spontaneously differentiate to cardiomyocytes [99]. In this context, a neotype three-dimensional cell expansion method was developed for BADSCs. For this purpose, “clickable” zwitterionic starch hydrogels were prepared from starch derived from methacrylate modified sulfobetaine using dithiol-functionalized poly(ethylene glycol) cross-linker through the “thiol-ene” Michael addition reaction. Furthermore, CGRGDS peptide was immobilized onto the hydrogels by a comparable “clickable” method. The Young’s moduli were

Pharmaceutical applications of natural polysaccharides 21 dependent on the concentration of precursor solutions and changed from 22.28 to 74.81 kPa. Outstanding antifouling property was observed upon the incorporation of zwitterionic fragments. The BADSCs were evenly encapsulated into the hydrogels and they were cultured for 10 days. High cell proliferation was seen and its extent was increased by decreasing the mechanical strength and addition of the CGRGDS. Interestingly, the cell “stemness” was well preserved during the encapsulation/culture and the released cells from the hydrogels excellently kept the capability of efficient spontaneous cardiomyogenic differentiation. Thus, it was found that zwitterionic starch hydrogel was suitable for growth and “stemness” preservation of BADSCs. The cellular viability and morphology within the 10-day culture were followed through acridine orange-propidium iodide (AO-PI) staining. Also, Fig. 2.1 reveals that cells preserve spherical shape in this period, as they are surrounded by hydrogel and cannot be stretched by growing over the TCP’s surface. Nevertheless, size of cells noticeably increases with time illustrating the cells are not restrained in hydrogel and they remold their neighboring environment by growing. Moreover, dramatic continuous cell proliferations happened so that little dead cells are observed in all hydrogels. Thus, hydrogels are very biocompatible and advantageous for the growth of cells. Fig. 2.2 exhibits that the BADSCs constantly express GATA-4 during the entire culture time that is increased in all hydrogels which confirms the cells expanded in the hydrogels outstandingly conserve the cardiac-differentiate potentiality. Moreover, introducing CGRGDS leads to GATA-4 expression by slightly more amount of cells when compared with that without CGRGDS. This designates that the CGRGDS causes cellular function preservation for adherent stem cells [99]. The cell encapsulation in alginate was carried out to study the differentiation process of embryonic cancer stem NTera2 (NT2) cells to dopamine generating cells [100]. Encapsulation of cells into polymer beads led to their isolation by immune system and made them suitable for transplantation and achieving an auspicious tool to deliver bioactive agents to the brain. The alginate polysaccharide was employed in this process because it is one of the most commonly used materials, which is well adopted by several tissues such as the brain. Also, two diverse initial cell concentrations (0.5
106 and 1.0
106/mL) were examined to recognize which one would more homogeneously distribute in the alginate beads and reveal superior cell viability at various culture stages. Two diverse CaCl2 concentrations of the gelling bath (1.0 and 0.1 M) were used to acquire beads with altered permeability including LP (low permeability) and HP (high permeability) beads for CaCl2 concentrations of 1.0 and 0.1 M, respectively. Then, the encapsulated cells were preserved inside a humid incubator with 95% humidity and 5% CO2 for 30 days and the cell solution was renewed every 2e3 days. Pure alginate beads were obtained as reference by the same method. The designations of the various samples are given in Table 2.1.

22 Chapter 2 Figure 2.1 AO/PI staining fluorescence images of BADSCs cultured in different S/P hydrogels with/without CGRGDS and ST/P-7.5 hydrogel for 1, 4, 7, and 10 days [99].

Pharmaceutical applications of natural polysaccharides 23

Figure 2.2 GATA-4 immunofluorescence staining of BADSCs cultured in different S/P hydrogels with/without CGRGDS and ST/P-7.5 hydrogel for 1, 4, 7, and 10 days [99].

24 Chapter 2 Table 2.1: Designation of the systems based on NT2 cells encapsulated within alginate beads in different conditions [100]. Sample 0.5NT2-HP 0.5NT2-LP 1.0NT2-HP 1.0NT2-LP

[NT2 cell] (106/mL) 0.5 0.5 1.0 1.0

[CaCl2] gelling bath (M) 0.1 1.0 0.1 1.0

It was found that higher number of cells stimulated clusters formation and caused better interactions of encapsulated cells and subsequently promoted mitotic effect. The live/dead cells distribution within the alginate polymeric beads was confirmed by the fluorescein diacetate/propidium iodide staining using fluorescence microscopy imaging which proved the existence of living neuronal positive cells. The activities of the encapsulated NT2 cells were established through dopamine formation. Hence, the NT2-alginate system was a valuable active platform that could produce/release dopamine and could be used for the treatment of Parkinson disease. One day after culture, all cells were evenly well encapsulated into the alginate beads (Fig. 2.3A). Greater mitotic effect was observed 8 days after culture (Fig. 2.3B) in capsules prepared using the maximum cell concentration (1.0
106/mL). Large amount of cells supported cluster creation and caused more efficient interactions among encapsulated cells and promoted mitotic effect. Cells in the LP capsules showed incomplete localized growth possibly because of uneven distribution of cells when the encapsulation is initiated. The fluorescence images of 1.0NT2-HP subsequent to diverse culture durations (7, 9, 11, 16, and 21 days, Fig. 2.4) reveal that the encapsulated cells are increased due to their strong mitotic activities. Nevertheless, a progressive rise in the mortality of cells was seen (nuclei were stained in red color) mainly apparent for the separated cells that could not be organized as clusters. In all cases, cell viability was preeminent relative to the moderate or little cell death [100]. The ever-growing clinical usage of cell-based therapy leads to development of systems in order to store and distribute cell therapy materials that are suitable for the clinical applications [101]. Recently, encapsulation to alginate was performed to increase the maintenance of human adiposeederived stem cells (hASCs) in the course of hypothermic storing and to find a large-scale production process. A dropwise technique was adopted to create scalable alginate beads using the calcium cross-linker which produced 3500 jelly beads in 1 minute. Also, the influence of alginate amount was assessed on viscosity properties of nonjelly sodium alginates, mechanical characteristics, and structures of Ca cross-linked alginate beads using diverse alginate and calcium quantities. The mechanical strength mainly depended on alginate quantity and 1.2% alginate cross-linked using 100 mM CaCl2 revealed a tensile stress of 35 kPa. Using the optimum parameters for

Pharmaceutical applications of natural polysaccharides 25

Figure 2.3 Optical microscopy images of encapsulated cells for different samples: (A) 0.5NT2-HP, 1.0NT2-HP, and 1.0NT2-LP, after 1 day of culture (bar ¼ 500 mm (left) and 200 mm (right)); (B) 0.5NT2-HP, 1.0NT2-HP, and 1.0NT2-LP, after 8 days of culture (bar ¼ 500 mm (left), 200 mm (middle), and 100 mm (right)) [100].

26 Chapter 2

Figure 2.4 Fluorescence microscopy images of cells encapsulated within 1.0NT2-HP beads, at different days of culture (bar ¼ 200 mm). Living cells are stained green with fluorescein diacetate, whereas dead cells are stained red using propidium iodide (for interpretation of the references to color in this figure legend, the reader is referred to the web version of this article) [100].

Pharmaceutical applications of natural polysaccharides 27 the beads, the human stem cells were immobilized and the encapsulated hASCs did not exhibit defeat in cell viability, and they also showed a homogeneous distribution after large-scale manufacture. After storing, the released cells were attached and revealed a normal shape once they were returned into the culture medium. Consequently, a scalable process was developed for encapsulation/storage of stem cells that was appropriate for the cell treatment supply chain. To explore the spatial cells distribution after encapsulation and their viability, some imaging techniques were carried out 48 h following the bead preparation. Beads displayed a spherical morphology along with a small tail (Fig. 2.5A) and such appearance was

Figure 2.5 The spatial distribution of cells through alginate beads. About 48 h after encapsulation, hASCs in beads were examined by phase microscopy (A), infrared imaging (B), and confocal microscopy (CeE). (CeE) hASCs were stained with calcein-AM (live indicator; green) and ethidium homodimer-1 (dead indicator: red) and maximal projection (C), Orthogonal projection (D) and 3D projection (E) images were captured. Arrowheads indicate the position of dead cells (for interpretation of the references to color in this figure legend, the reader is referred to the web version of this article) [101].

28 Chapter 2 preserved subsequent to the creation of most beads (Fig. 2.5B). Also, uniform cell distribution was confirmed through infrared images of encapsulated cells which evidently represented bead shapes with a greater intensity at overlap points (see Fig. 2.5B). Confocal images of live-dead stained encapsulated cells indicated great viabilities at earlier times so that cells exhibited appropriate distributions from projected Z-stacks (Fig. 2.5C) but little dead cells were observed (specified with arrow-heads). It was obvious in orthogonal XZ and YZ planes (Fig. 2.5D) with all dead and live cells dispersed in bead depth (the dead cells are shown by arrow-heads). Such cellular dispersion was revealed by volume analysis (see Fig. 2.5E). The uniform dispersal of deceased cells without viability loss approves that cell death was not stimulated by possible external stresses which cells may experience in the course of processing. The hASCs binding was examined using normal tissue culture for 24 h after hypothermic storing for 1 or 7 days. The hASCs cultured for 24 h one day after encapsulation revealed a regular spindle-shaped fibroblast resembling morphology which demonstrated suitable attachment capability (see Fig. 2.6). There were not any noticeable differences between nonencapsulated (control) and encapsulated hASCs stored in identical conditions. However, subsequent to 7-days storage, the alginate encapsulation preserved attachment (Fig. 2.6) but encapsulated cells demonstrated a practical attachment although with

Figure 2.6 Morphology and attachment of encapsulated ASCs following storage. hASCs were stored for either 1 or 7 days before plating at 5000 cells/cm2and returning to normal tissue culture conditions. Scale bars ¼ 200 mm [101].

Pharmaceutical applications of natural polysaccharides 29 slightly lower ability compared to 1 day; the control samples revealed significantly decreased attached cells along with more rounded dead cells [101].

3. Application of natural polysaccharides in pharmaceutics/drug delivery Natural polysaccharides (NP) are broadly used in drug delivery applications. Mauran (MR) is a polysaccharide macromolecule that is extracted from Halomonas maura halophilic bacterium. The antioxidant properties of MR-chitosan (CS) nanoparticles were examined to determine their usefulness for biomedical applications, and it was found that they could effectively defend against oxidative stress [102]. The ability of extremophilic mauran-based nanoparticles was investigated for in vitro and ex vivo scavenging of reactive oxygen species. The 5-fluorouracil incorporated MR-CS nanoparticles were assessed for cancer inhibition and their healing efficacy was compared using glioma and breast adenocarcinoma cells confirming they could specifically affect the cancer cells. The fluorescent labeled nanoparticles were utilized to evaluate the cell internalization by the nanocarriers by means of flow cytometry and confocal microscopic imaging indicating effective cellular uptake and absorption by such nanoparticles which were suitable for bioimaging as well as recognizing the cell binding by such nanoparticles. Moreover, the internalized fluorescent nanocarriers revealed a minor effect on the normal cells’ integrity. The cellular recognition and absorption of the MR-CS NPs were evaluated by fluorescent images of FITC-tagged L929 cells treated by MR-CS NPs. The FITC tagged CS was employed to create MR-CS NPs through polyelectrolyte complexation. The dye tagged NPs were introduced into the cells and their images were obtained by means of confocal microscope using a 488 nm laser. The fluorescent images of MR-CS-FITC NPs absorbed by the L929, GI-1, and MCF7 cells are indicated in Fig. 2.7A, E, and I show bright field imaginings and Fig. 2.7B, F, and J exhibit the DAPI stained nuclei. Fig. 2.7C, G, and K reveal green fluorescing MR-CS NPs and Fig. 2.7D, H, and L depict merged images of L929, GI-1, and MCF7 cells, respectively. The absorption tests in 24 h proved that MR-CS NPs had desirable cell acceptance without oxidative destruction and cell membrane interruption by the MR existence in L929 cells. Consequently, fluorescent images of normal cells supported the results achieved by antioxidant tests confirming MR was a useful biomaterial to promote the antioxidant mechanism up to the 500 g/mL concentration. Likewise, confocal microscopy images of cancer cells supported the flow cytometry data. Fig. 2.7 approved that the number of fluorescent NPs that was absorbed via GI-1 and MCF7 cells were rather lower compared to that of L929 cells after incubation for 24 h. Comparable results were observed for the SR-MR-CS NPs. Fig. 2.8 shows fluorescent images for the SR-MR/CS NPs absorbed by the L929, GI-1, and MCF7 cells. Fig. 2.8A, E, and I reveal bright field images, Fig. 2.8B, F, and J demonstrate the DAPI stained nuclei. Fig. 2.8C, G, and K illustrate the green fluorescing MR-CS NPs

30 Chapter 2

Figure 2.7 FITC tagged MR/CS nanoparticle absorption studies using confocal microscopy; (A), (E), and (I) bright field images of L929, MCF7, & G1 cells; (B), (F), and (J) DAPI stained nucleus; (C), (G), and (K) FITC tagged MR/CS nanoparticles accumulated in cells; (D), (H), and (L) merged images [102].

and Fig. 2.8D, H, and L depict the merged images of L929, GI-1, and MCF7 cells, respectively. The confocal and flow cytometry tests proved that the MR-CS was an effective carrier for drug molecules to release them at the target sites. The biocompatibility and antioxidant characteristics of such particles enable their applications as drug carriers for cancer therapy, and MR-CS NPs could be utilized as an effective targeting material [102]. The gum karaya (GK) was employed to synthesize gold nanoparticles (GNP) which were applied to deliver anticancer drug [103]. The GK nanoparticles displayed extraordinary biocompatibility throughout cell survival test using both normal Chinese hamster ovary

Pharmaceutical applications of natural polysaccharides 31

Figure 2.8 Sypro-ruby tagged MR/CS nanoparticle absorption studies using confocal microscopy; (A), (E), and (I), bright field images of L929, MCF7, & G1 cells; (B), (F), and (J), DAPI stained nucleus; (C), (G), and (K), Sypro-ruby tagged MR/CS nanoparticles accumulated in cells; (D), (H), and (L) merged images [102].

cells plus A549 human nonsmall cell lung cancer cells as well as in the course of hemolytic toxicity assessments. The anticancer drug gemcitabine hydrochloride (GEM) was incorporated into the nanoparticles indicating a drug loading efficiency of 19.2%. The GEM loaded nanoparticles presented superior inhibition against cancer cells growth in clonogenic and antiproliferation tests relative to the native GEM. This result was associated to greater generation of reactive oxygen species using the GEM containing nanoparticles in A549 cells compared to the native GEM confirming the GK had a noteworthy capacity to synthesize biocompatible gold nanoparticles, and it could be applied as an anticancer drug delivery vehicle. The influence of formulations on the A549

32 Chapter 2

Figure 2.9 Effect on the morphology of A549 human lung cancer cells incubated with gemcitabine hydrochloride (GEM) and GEM loaded gum karaya stabilized gold nanoparticles (GEM-GNP). Untreated cells are shown as control [103].

Figure 2.10 Clonogenic assay: GEM loaded gum karaya stabilized gold nanoparticles (GEM-GNP) showed greater inhibition of colony formation activity of A549 human lung cancer cells as compared to native GEM and untreated cells (control) [103].

cells morphologies are displayed in Fig. 2.9. Furthermore, colony formation inhibition test was accomplished to confirm the extended antiproliferation influence of GEM-GNP. Less colonies were detected for the cells treated by GEM-GNP sample compared to control and GEM-treated cells (Fig. 2.10). In another study, Cassia obtusifolia seed mucilage was extracted and used in drug delivery [104]. The seed mucilage was assessed for the occurrence of polysaccharide and a mucilage-based biodegradable film was achieved using seeds of C. obtusifolia. The in vitro oral acute toxicity assay and degradation in simulated body fluids revealed a great LD50 > 2 g/(kg of body weight) which established the benign nature of this excipient. The diclofenac incorporated film illustrated a continuous drug release that was attributed to the swelling of film and dug diffusion. Thus, it was validated that the C. obtusifolia seed mucilage could be used as a film-forming excipient showing improved features for drug delivery purposes. Phytochemical tests for the identification of seed mucilage proved the occurrence of carbohydrate and mucilage (Table 2.2). Nonetheless, the tests did not show existence of starches, alkaloids, glycosides, tannins, and steroids. The carbohydrate

Pharmaceutical applications of natural polysaccharides 33 Table 2.2: Phytochemical identification tests of isolated mucilage from Cassia obtusifolia seed [104]. Identification tests Test for carbohydrates Test for starches Test for proteins and amino acids Test for mucilage Test for glycosides Test for alkaloids Test for steroids and sterols Test for tannins

Name of tests

Observation

Molisch test Iodine test Ninhydrin test

þ  

Ruthenium red test Legal, Keller-Killiani, Borntrager tests Mayer, Dragendroff tests Libermann-Burchard test Ferric chloride, lead acetate tests

þ    

þ, Present; , Absent.

Table 2.3: Physical properties of Cassia obtusifolia seed mucilageebased film [104]. Physical property

Results

Film thickness (mm) Weight measurement (mg) Moisture content (%) pH Folding endurance Mechanical strength (kg/mm2) Mucoadhesive force (dynes/cm2)

0.14  0.01 0.92  0.02 0.92  0.02 6.96  0.05 >500 0.2 2400.60  89.45

chemical analysis exhibited that polysaccharides were present in the sample. Table 2.3 represents the physical properties of C. obtusifolia seed mucilage film.

4. Application of natural polysaccharides in gene delivery Genes are inherited units and their defects or malfunctioning results in several conditions and diseases including cystic fibrosis and cancer. Most of the traditional treatments involve drug-based methods focusing on symptoms/signs instead of the fundamental root reason of the disease which is the faulty genes. Therefore, in order to effectively treat genetic disorders, defective genes must be corrected or augmented at molecular level through the gene therapy approach [105,106]. Although clinical trials were started in early 1990s, this method is yet at the infancy step as there is trouble in development of a perfect vector to protect/deliver nucleic acids to the desired target sites without any negative influence [107e109]. Sugar-containing cationic polymers show high potential to deliver genes such as therapeutic RNA interference (RNAi).

34 Chapter 2 Table 2.4: Particle size and zeta potential of siRNA-loaded 6AC-100 nanoparticlesa [110]. Weight ratio (siRNA/6AC-100) 1:10 1:5

Charge ratio (¡/þ) 1:14 1:7

Particle size (nm) 92.9  3.3 182.7  5.1

Polydispersity index ([(m2)/G2]) 0.267 0.099

Zeta potential (mV) 21.7  0.1 17.5  1.1

a Particle preparation conditions: 6AC-100 concentration: 1.0 mg/mL (PBS pH7.4), siRNA concentration: for weight ratio 1:10, 0.1 mg/mL (7.0 mM) and for weight ratio 1:5,0.2 mg/mL (14 mM), T: 25  2 C. Data shown are the mean  standard deviation (n ¼ 3).

The RNAi can downregulate the gene expression as posttranscriptionally and this is an important therapeutic process which can highly decrease the amount of disease causing proteins which are not treated using common small drug molecules. Nonetheless, clinical usage of small interference RNA (siRNA) needs designing effective siRNA sequences and developing benign and proficient delivery formulations. In order to achieve a siRNA delivery system which is also biocompatible, natural polysaccharide curdlan was modified chemically by a regioselective method so that amine groups were introduced on the glucose groups [110]. The resultant 6-amino-curdlan was watersoluble that produced NPs after its complexation with siRNAs. The zeta potentials and particle sizes of siRNA-loaded 6AC-100 nanoparticles are provided in Table 2.4. The curdlan-based NPs powerfully delivered siRNAs to mouse primary cells and human cancer cells and decreased 70%e90% of the target mRNA amount. Furthermore, 6-amino-curdlan nanoparticle could deliver the siRNA targeting eGFP to the mouse embryonic stem cell that soundly expressed eGFP and substantially reduced the GFP protein concentration. Hence, the curdlan-based nanoparticle was proposed as an auspicious vehicle to deliver short RNAs in order to diminish endogenous mRNAs. It was investigated to find if the 6AC polymer could deliver siRNA to cells using the curdlan-based nanoparticles. For this purpose, A549 cells were treated with Cy3-labeled RNA (Cy3-siRNA) that was complexed to 6AC-100 NPs; then the siRNA cell internalization was tracked using fluorescence microscope. For most cells treated by 6AC-100 NPs, after 4 h transfection, strong fluorescence signals were seen signifying 6AC100 could effectively deliver siRNA into cytoplasm (see Fig. 2.11). In order to evaluate the transfection efficacy of 6AC-100 on stem cells, a complex of siRNA and 6AC-100 was used to target eGFP into mouse embryonic stem cells (mES cells) which continuously expressed eGFP. Fig. 2.12 exhibits that the GFP signal was greatly decreased 24 h after transfection using 25 nM siRNA complexed with 6AC-100 confirming 6AC-100 powerfully delivered siRNA into stem cells and reduced gene expression at the protein level [110]. Glycosaminoglycans (GAGs) are known as naturally occurring polymers that are generally employed in gene delivery to enhance stability and reduce toxicity and nonspecific interactions, thus improving transfection efficacy. The sorbitan esterebased lipid NPs functionalized by hyaluronic acid and GAGs chondroitin sulfate were used as gene

Pharmaceutical applications of natural polysaccharides 35

Figure 2.11 Internalization of siRNA in cancer cells by 6AC-100. A549 cells were transfected with Cy3-labeled siRNA complexed with 6AC-100 nanoparticles. Localization of dye-labeled siRNA after 4 h was monitored by fluorescence microscopy [110].

Figure 2.12 In vitro silencing of endogenous mRNA by 6AC-100 nanoparticles in mouse embryonic stem cells. Fluorescence microscope image of mES cells stably expressing eGFP treated with nontargeting siRNA complexed with 6AC-100 nanoparticles, or siRNA targeting GFP complexed with 6AC-100 nanoparticles. mRNA levels are expressed as percent of control [110].

delivery vehicles [111]. Such nanoplatforms incorporated with plasmid DNA were evaluated for the physicochemical properties and stability, protection capacity, and proficient transfection of cells using the improved green fluorescence of plasmid protein in vitro along with the in vivo and in vitro biocompatibility. It was established that compounds with extraordinary biological significance and targeting capacity (like hyaluronic acid and chondroitin sulfate) could fruitfully be introduced in the sorbitan esterebased nanoparticles to stabilize both nanosystems and protect the plasmid DNA. It was found that adding the hyaluronic acid and chondroitin sulfate caused long-standing stability of the nanoplatforms in both lyophilized and liquid states confirming they could be used in industry. Such functionalized nanosystems could transfect A549 cells without

36 Chapter 2 affecting the cells’ viability and revealed innocent safety profiles in vivo approving the ability of these nanoparticles as gene delivery platforms.

5. Application of natural polysaccharides in protein binding Proteins are natural polyelectrolytes having exceptional functional characteristics like the capacity to produce emulsions and gels. Thus, they can be used in combination with natural polysaccharides for biomedical applications like the encapsulation of bioactive materials [112]. Also, it is well known that functionality and stability of protein can be enhanced under unfavorable conditions through complex creation between protein and polysaccharide. The nature of interactions among polysaccharides and proteins can be electrostatic and/or covalent [113]. In a recent work, polysaccharide-protein-surfactant complexes were achieved through coprecipitation method using propylene glycol alginate (PGA), zein, as well as lecithin or rhamnolipid. Such ternary complexes were used as delivery platforms to increase the curcumin (Cur) bioavailability and stability [114]. The curcumin-containing zein-PGA, zein, zein-PGA-lecithin, and zein-PGA-rhamnolipid complexes were denoted as Z-P-Cur, Z-Cur, Z-P-L-Cur, and Z-P-R-Cur, respectively. Encapsulation efficiency of the curcumin was increased due to the existence of surfactants and polysaccharides in the complexes compared to the pure zein NPs as Z-P-Cur (67%), Z-Cur (21%), Z-P-R-Cur (92%), as well as Z-P-L-Cur (94%). Incorporating the surfactants to the complexes considerably enhanced the bioaccessibility and photostability of curcumin. Hence, the developed ternary complexes were promising means for encapsulation, protection, and delivery of hydrophobic nutraceuticals for usage in pharmaceuticals, supplements, and foods. The carrageenan and xanthan gum, respectively, indicated high and medium negatively charged polysaccharides. Diverse proportions of these biopolymers were heated along with soybean protein isolate (SPI) [115]. After mixing by simulated stomach juice, both carrageenan-SPI and xanthan-SPI immediately underwent self-assembled gelation using the biopolymer ratios greater than 0.01. At upper biopolymer ratios, a stronger gel was formed. Also, highly negatively charged carrageenan produced a stronger gel compared to that combined with xanthan gum. It was found that the SPI digestibility was postponed after mixing with the polysaccharides, and it is improved by increasing the biopolymer ratio. Fig. 2.13 exhibits the SEM images of xanthan-SPI (A and B) as well as carrageenanSPI (C and D) (polysaccharides/SPI ratio ¼ 0.1) which immediately were mixed with SGF (A and C) after 1 h digestion (B and D). Polysaccharides having higher negative charge could more strongly delay the SPI digestion. Besides, the microstructures of both the carrageenan-SPI and xanthan-SPI gels were observed by scanning electron microscopy before and after simulated stomach digestion which confirmed the gels delayed the SPI digestion [115].

Pharmaceutical applications of natural polysaccharides 37

Figure 2.13 SEM images of xanthan-SPI (A and B) and carrageenan-SPI (C and D) (polysaccharides: SPI ratio of 0.1) immediately mixed with SGF (A and C) and after 1 h digestion (B and D) [115].

In another research, CS and bovine serum albumin (BSA) were employed to fabricate BSA-CS nanogels through a green facile self-assembly procedure [116]. The nanogels were utilized to encapsulate doxorubicin hydrochloride (DOX) with 46.3% entrapment efficiency in order to understand both less cytotoxicity and gradual release of DOX. Also, the pure DOX and DOX-containing BSA-CS (DOX-BSA-CS) revealed IC50 amounts equal to 0.05 and 0.22 mg/mL, respectively, against growth of SGC7901 cells. The DOX cytotoxicity significantly declined in 24 h after its encapsulation into the nanogels demonstrating the loaded drug was slowly released during 24 h indicating BSA-CS was an appropriate sustained release platform. The cell uptake tests pointed out that the DOXBSA-CS was more rapidly diffused to the cancer cells compared to the pure drug. Also, it was evidenced that the DOX-BSA-CS induced apoptosis of gastric cancer cells 7901 was superior to the plain drug confirming it was favorable for the gastric cancer treatment.

38 Chapter 2

6. Application of natural polysaccharides in wound healing Cutaneous wound healing is one of the most important processes in numerous pathologies, which is required in postsurgery scars and burns. It is well recognized that the wound healing occurs as a dynamic interactive phenomenon that can substitute missing and devitalized tissue layers and cells through interacting processes classified as hemostasis, inflammation, proliferation, and matrix remodeling [117]. It is essential that an injured/ wound site immediately be covered with a dressing that is able to prevent microbial invasion, preserve wet medium for efficient skin regeneration, let gaseous passage, and adsorb exudates [118]. Nevertheless, commercially existing wound dressings only meet some of these standards. Consequently, the development of novel nanodressings using nanocomposites is of growing importance [119]. Recently, polymeric nanocomposites have been developed as wound dressing materials. Among such materials, polysaccharide nanocomposites are proficient candidates because of their exceptional tissue mimicking and biocompatibility characteristics [120]. They can mimic full-thickness skin wounds because these kinds of dressing have morphologies reminiscent of the native skin and appropriate features for a good wound healing process [120]. It is known that starch is a plentiful, relatively low-cost, and ecological material which can be prepared as nanoparticles and fillers to fabricate bionanocomposite wound dressing materials. In a recent effort, symmetric and asymmetric porous films were fabricated using chitosan (CS) and poly(vinyl pyrrolidone) (PVP) along with nanostarch (NS) as filler through salt leaching process for wound dressing usage [121]. Symmetric CS-PVPnanostarch (CSPNS) films containing 3 and 1%wt of nanostarch were achieved without their coating by stearic acid but the CS-PVP-nanostarch-stearic acid (CSPNSeS) asymmetric film was obtained through coating with stearic acid. It was found that the stearic acid covered surface had microporous, hydrophobic, bacterial antiadhesion characteristics whereas the hydrophilic stearic acid uncoated surface displayed higher bactericidal and noncytotoxic properties with a very porous structure. All of the asymmetric and symmetric films demonstrated nearly identical barrier; mechanical, hemolytic, and swelling features indicating the stearic acid did not influence the hemolytic and physical properties while the nanostarch amount significantly affected these features. The CSPNS1%-S film showed outstanding Staphylococcus aureus antiadhesion capacity. Moreover, excision wound healing in vivo verified that the CSPNS1%-S film illustrated improved healing process in addition to enhanced collagen development and reepithelialization. In vivo wound healing experiment was accomplished using adult albino rats; images indicating wound healing are displayed in Fig. 2.14. In the day wound was created, wound areas were identical in all groups. Granulation tissues were detected on day 7 in wounds covered by CPNS1%-S so that the healed rates were 32% and 20% for the CP-S dressing

Pharmaceutical applications of natural polysaccharides 39

Figure 2.14 In vivo wound healing analysis of control, CP-S, and CPNS1%-S dressing material [121].

and control, respectively. Wound areas were considerably decreased on day 14 in group dressed with CPNS1%-S and wound bleeding was stopped so that healing rates were improved to 81%, 75%, and 60% for the CPNS1%-S, CP-S, and control dressing, respectively. All wounds were entirely cured but the healing rate was very greater for the CPNS1%-S compared to those of the CP-S dressing and control. It is noteworthy that high rate of wound closure using CPNS1%-S was attributable to synergistic influence of CS, PVP, stearic acid, and nanostarch due to their mechanical, barrier, antibacterial, anticell adhesion, cytotoxicity, and hemolytic properties. The healing efficacy of the wound dressing was studied by histological analysis. Fig. 2.15 illustrates the histological analysis results for the CPNS1%-S dressing, CP-S, and control. Inflammatory cells seen on day 7 were greater in CP-S and control but fresh blood vessels were observed and inflammatory cells were highly reduced for the CPNS1%-S dressed sample. Collagen fiber was created on day 14 in CPNS1%-S that supported generation of additional granulation tissues. Hair follicle cells were seen on day 21 using the CPNS1%-S

40 Chapter 2

Figure 2.15 Histological Images of control, CP-S, and CPNS1%-S dressing material [121].

and CP-S materials. Such findings confirmed the powerful influence of CPNS1%-S as an asymmetric wound dressing to healing wounds [121]. In another study, the antioxidant, hemolytic activity and in vivo wound healing effect of FWEP polysaccharide which was extracted from fenugreek (Trigonella foenum-graecum) seeds were assessed [122]. The in vivo and in vitro antioxidant activities were estimated by different tests, and it was revealed that the FWEP had strong antioxidant potency whereas hemolytic activity was not witnessed to bovine erythrocytes. Also, the FWEP hydrogel was applied on a wound position in a rat model which substantially improved wound healing, as well as expedited wound closure, 14 days after the wound generation. The histological analysis validated formation of entirely reepithelialized wound and whole epidermal renewal. Overall, it was established that FWEP exhibited high wound healing capacity that was probably related to its antioxidant activity. Several dextran-based bionanocomposite films incorporated with sandalwood oil (SO) and clove oil (CO) were prepared which could prevent infection as a result of their intrinsic antibacterial potency and modify the wound healing process to accelerate scarfree healing [123]. The dextran-nanosoy-glycerol-chitosan (DNG-CS) nanocomposites were fabricated followed by adding SO and CO to acquire herbal DNG-CS-CO and DNG-CS-SO nanodressing materials which demonstrated >98% bactericidal effects against both Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) microorganisms only

Pharmaceutical applications of natural polysaccharides 41 using very low loading amounts of 10% and 5% for SO and CO, respectively. Such encapsulation approach led to controlled EO diffusion during 72 h measured for drug effectiveness by means of bacterial reduction counting and serial disk diffusion assays. The bacterial adherence test established the proficiency of these dressings to stop microbial invasion. The in vivo wound healing test by means of DNG-CS-CO dressings on male Swiss albino mice (BALB/c strain) revealed full healing in 14 days along with extraordinary efficiency in scar inhibition. Also, the histological analysis proved that CO and SO treatments brought about ordered collagen deposition together with fibroblast migration.

7. Application of natural polysaccharides in tissue engineering The structural and biological functions of polysaccharides have made them appropriate compounds to be exploited in tissue engineering. Such biomaterials exhibit appropriate biochemical and mechanical properties for tissue engineering applications [124]. The polysaccharides reveal numerous features as eligible biomaterials for tissue engineering including biodegradability, biocompatibility, and the cell delivery capacity [125]. An ideal biomaterial not only has a suitable chemical structure but also it shows favorable macroscopic structural properties, that is, the biomaterial scaffold has a porous structure to allow mass transportation (diffusion as well as permeability) [126]. Further, the biomaterial should reproduce the mechanical, elastic, and organizational characteristics of native tissues, and this is especially imperative for vital and very specific tissues like the cardiac tissues [127,128]. The best biomaterial should have a structure to stimulate cell attachment/growth once enabling its organization and perhaps differentiation to a very well-ordered biomimetic structure [129]. Also, it should be prone to resist high and permanent mechanical stresses. Another foremost role is related to the integration with host tissue and final substitution by the extracellular matrix of the host [130]. The biomaterials should have biological activities to accelerate the tissue repair. For instance, cell recruitment, angiogenesis, and cardiomyocyte protection properties are beneficial for the treatment of heart diseases [131]. Also, the tissue engineering products must be effective and economical considering their functionality and production simplicity [132]. Polysaccharides are auspicious materials meeting most of these criteria that can be used as eligible biomaterials for tissue engineering. Nanocomposites of nanohydroxyapatite, chitosan, and tamarind seed polysaccharide (n-HAp/CS-TSP) were fabricated in weight ratios equal to 70:10:10, 70:15:15, and 70:20:20, respectively [133]. The n-HAp/CS-TSP (70:10:20) exhibited the most rough and porous surface, improved thermal stability in addition to maximum compressive modulus, and strength. Also, the n-HAp/CS-TSP (70:10:20) showed greater swelling, satisfactory degradation, as well as enhanced biomineralization within simulated body fluid in

42 Chapter 2 comparison to the nanocomposites n-HAp/CS and n-HAp/CS-TSP (70:15:15 and 70:20:10). The n-HAp/CS-TSP (70:15:15) indicated greater nontoxic activity to MG-63 cells plus superior hemocompatibility. Thus, the n-HAp/CS-TSP nanocomposites were considered as more suitable biomaterials for bone tissue engineering relative to the n-HAp/ CS nanobiocomposite. Recently, a series of hydrogels were synthesized through incorporating the synthetic hydroxyethyl methacryate monomer along with a semisynthetic polymer backbone (carboxymethyl tamarind, CMT) in diverse molar ratios [134]. Such materials were denoted as CMT:HEMA hydrogels. The biocompatibility tests using NIH-3T3, HaCaT, in addition to mouse dermal fibroblast cells established that these materials are biocompatible and they were not cytotoxic. The mitochondrial functionality assays proved that they were safe and noncytotoxic. The hemolytic assessment using red blood cells plus acute skin irritation experiment on SD rats verified that they were appropriate for the ex vivo applications. Finally, it was suggested that these hydrogels were favorable for in vivo commercial and clinical applications to treat skin disorders. Skin edema and erythema were not detected during the experiment (Fig. 2.16A). Microscopic assessment of various skin sections which were stained using eosin and hematoxylin did not display morphological variations in dermal or epidermal skin layers in tested animals relative to the control (Fig. 2.16B). Weight gain and feed consumption were similar in both control and test groups. Based on gross, systemic notes and skin microscopic analysis, it was found that the CMT:HEMA was well accepted by all animals upon topical skin usage [134]. Xanthan gum and CS were blended using Fe3O4 magnetic nanoparticles (MNPs) to create magnetic responsive polyelectrolyte complex hydrogels (MPECHs) through in situ ionic complexation by means of D-(þ)-glucuronic acid d-lactone to acidifying the medium [135]. It was shown that adding Fe3O4 MNPs to the PECH highly enhanced mechanical characteristics as well as storage modulus. The in vitro cell culture using NIH3T3 fibroblast cells on MPECHs displayed increased growth and adhesion of cells under an external magnetic field compared to the native PECH confirming the MPECH can be utilized as a magnetic stimulated material in tissue regeneration.

8. Application of natural polysaccharides in bioimaging Nowadays, nanocomposites of natural polysaccharides and quantum dots (QDs) are widely used in bioimaging applications [136]. Recently, sodium carboxymethyl cellulose (CMCel) was used as a biocompatible and multifunctional polysaccharide to synthesize fluorescent ZnCdS alloyed quantum dot nanostructures at room temperature by an aqueous green method [137]. The core-shell nanoconjugates were comprised of the ZnCdS semiconductor

Pharmaceutical applications of natural polysaccharides 43

Figure 2.16 Ex-vivo corrosion/irritation test for CMT:HEMA hydrogel. (A) Pictographic representation of the area of the dorsal surface of SD rats where the control and hydrogel solution was topically applied. Images were taken at 24, 48, and 72 h after the application of hydrogel. (B) Histopathology of skin is represented through H & E staining of skin tissue of three independent animals after 15 days of application of hydrogel and control (top panel). Images were acquired by 40X objective. Lower panel represents enlarged view of the epidermis of the same tissue sections [134].

QD core along with the CMCel shell. Besides, CMCel functional groups controlled the diameters of colloidal wateresoluble nanocrystals and their hydrodynamic sizes. Also, the nanoconjugates were cytocompatible and luminescent which were used in bioimaging of human osteosarcoma cancer cells, approving that such polysaccharide-based fluorescent conjugates were auspicious nanoformulations for diagnosis and bioimaging of cancer cells. Quaternized analogs of pectic galactan (QPG) were prepared through the reaction of 3chloro-2-hydroxypropyl trimethyl ammonium chloride and pectic galactan in aqueous sodium hydroxide environment [138]. The QPG had electrostatic interactions with plasmid DNA in aqueous medium and formed complexes with spherical reduced shape showing nanometer sizes ranging from of 60e160 nm. These complexes were fluorescently labeled by 5-DTAF and added into the C6 rat glioma cells. It was indicated that they could eternalize the cells and reach close to the nucleus in 24 h justifying this modified natural polysaccharide could also be used as a biodegradable and biocompatible gene delivery and cell specific carrier.

44 Chapter 2

Figure 2.17 Representative confocal image of C6 cellular path of QPG/pGFP complexes 24 h after exposure to the complexes. Bars represent 10 mm [138].

QPG-pGFP labeled complexes were administered to C6 rat glioma cells. Fig. 2.17 indicates the confocal image. Cells were observed after 24 h contact with the complexes. The labeled QPG (QPG-5-DTAF) was used to obtain images of the complexes (green). The fluorescent labeled membrane was yellow but the nucleus had a blue color. Thus, after 24 h, the complexes were entered inside the cells so that they were adjacent to the nucleus [138]. An enzyme and redox dual-stimuli responsive carrier called HMSN-SS-CDPEI@HA was achieved using carbon dots coated hollow mesoporous silica nanoparticles (HMSN) to be used in targeted drug delivery and cell bioimaging [139]. The positively charged CDPEI NPs were obtained using polyethylenimine (PEI) and grafted onto the HMSN pore openings via disulfide bonds which were employed as gatekeepers in order to entrap drugs inside the hollow cavities. Natural polysaccharide hyaluronic acid (HA) was also grafted onto the HMSN surfaces in order to examine targeted drug delivery, enhanced stability, as well as controlled drug release. Doxorubicin was used as an anticancer drug in the experiments because of its extensive clinical applications. The in vitro drug release test established that doxorubicin-containing HMSN-SS-CDPEI@HA had enzyme and redox dual-responsive drug release characteristics. Besides, the HMSN-SS-CDPEI@HA illustrated exceptional biocompatibility and fluorescent properties. The confocal laser scanning microscopy and flow cytometry established that the HMSN-SS-CDPEI@HA had a greater cell uptake through the CD44-receptor mediated endocytosis using CD44-receptor

Pharmaceutical applications of natural polysaccharides 45 overexpressed A549 cells compared to the NIH-3T3 (receptor-negative) cells which led to superior cytotoxicity effect against A549 cells relative to the NIH 3T3 cells. Hence, the HMSN-SS-CDPEI@HA was prepared to be a dual-stimuli responsive, real-time imaging, and targeted drug delivery platform which could be a promising system for the cancer treatment.

9. Application of natural polysaccharides in preparation of contact lenses Therapeutic ophthalmic lenses with prolonged drug release have been developed with the aim of circumventing tedious and fruitless eye drop administration. Hence, coating the contact lenses with natural polymers containing desired drugs is usually carried out [140]. Recently, the layer-by-layer deposition was applied using alginate and CS natural polymers in order to regulate releasing diverse ophthalmic drugs using three kinds of lenses including one silicone-based hydrogel used as a soft contact lens (SCL) with the drug releasing ability plus two commercially existing lenses called Definitive 50 for SCLs and CI26Y for intraocular lens (IOLs) [141]. The optimized coating was composed of a (alginateeCaCl2)/(CS þ glyoxal) double layer on top and an alginate-CaCl2 final layer to stop the CS degradation with tear fluid proteins. It was found that this coating had outstanding properties in controlling the release of diclofenac antiinflammatory drug once maintaining or increasing the physical features of the lenses. It was found that the coating could control the diclofenac release from IOL and SCL lenses at least for 1 week. Also, it was very hydrophilic (water contact angle z 0) as well as biocompatible; thus, it could avoid using additional surface treatments to improve the user relief. A therapeutic contact lens (TCL) with extended wear was prepared to sustainably deliver timolol maleate (TML) through molecular imprinting method [142]. The TCL contained a TML imprinted in a copolymer made up of carboxymethyl chitosan-g-hydroxy ethyl methacrylate-g-polyacrylamide (CmCS-g-HEMA-g-pAAm) introduced into polyHEMA framework. The TML was reloaded into the lens with an exceptional reloading capacity (6.53 mg of TML/TCL). Furthermore, the drug release accomplished in lacrimal fluid well obeyed the Higuchi model, which revealed the diffusion mechanism happened without polymer degradation. The TML drug release kinetics showed a continuous release which was suitable to achieve TML therapeutic index; besides, it could provide an appropriate medicine for glaucoma. The hydrogel contact lenses have attracted great interest as carriers in oculopathy treatment but traditional hydrogels do not demonstrate outstanding drug loading and controlled release effects which is due to lack of extra interactions of simple hydrophilic polymeric chains with drug molecules. To overcome these problems, some functional hydrogels were synthesized for the delivery of ophthalmic drug in oculopathy treatment [143]. Thus, mono-GMA-b-CD monomer and MA-b-CD cross-linker were added into the

46 Chapter 2 hydrogel via copolymerization reaction. The equilibrium swelling ratios and contact angles of hydrogels were changed with variations in ratios of mono-GMA-b-CD and MA-b-CD. The hydrogels revealed similar water loss, suitable transparency, and elastomer rheological characteristics. The functional hydrogels containing b-CD presented improved protein resistance as well as noticeably higher drug encapsulation compared to traditional hydrogels. Also, the drug release from the hydrogels was changed depending on the MA-b-CD and mono-GMA-b-CD ratios. The in vivo tests proved that hydrogel contact lenses had higher efficiency in decreasing intraocular tension compared with commercial eye drops confirming they are favorable for application in oculopathy therapy.

10. Application of natural polysaccharides in preparation of implants Polysaccharide hydrogels form three-dimensional matrixes of hydrophilic polymeric chains which are able to preserve great amount of water within the macromolecular structure; thus, they are interesting materials for preparation of implants [144,145]. Also, they have suitable features like adjustable mechanical and antibacterial properties, enhanced fluid film lubrication, in addition to friction decrease which lead to their application as injectable materials to fill out defects with any shapes [146]. So far, metal covered implants are widely employed against dental pathogens that cause biofilm creation as well as dental implants failure. Such nanoparticles can be applied together with natural polysaccharide biomacromolecules to improve the potential of these biologically active compounds. Alginate-polyacrylamide (ALG-PAAm) hydrogels were fabricated as orthopedic prosthesis materials and indentation experiments were performed to evaluate their mechanical features [147]. Also, their tribological responses were assessed by means of reciprocating sliding movement alongside alumina ceramic ball. The hydrogels were prepared by means of two diverse amounts of cross-linker in order to investigate the cross-linker influence on their wear resistances. Different loads and sliding speeds were used in absence and presence of bovine serum as a lubricant to mimic human gait/running cycle. The mass loss of each dried sample was measured using thermogravimetric analysis before and after every experiment and the wear volume was examined by profilometry. Increasing the cross-linker amount improved the elastic modulus up to 21% and the hardness to 32%. The mass loss was enhanced using a greater loading irrespective of cross-linker ratio. Nevertheless, using a higher sliding speed, a smaller amount of material was removed as a result of wear. Upon lubrication using the utmost load, the lowest average friction coefficient was attained for hydrogels containing 0.06% cross-linking agent, which is favorable in comparison to that measured for the articular cartilage. The observation of worn surfaces by means of electron microscopy indicated that adhesion was the principal wear mechanism. Also, hydrogels having superior cross-linking densities exhibited greater tribological performance and stiffness under lubrication.

Pharmaceutical applications of natural polysaccharides 47 In another work, silver (Ag) conjugated CS nanoparticles were prepared to be used as an effective coating compound for the titanium dental implants [148]. The bioactive CS was prepared using Aspergillus flavus Af09 and then conjugated to Ag NPs. The Ag-CS nanoparticles exhibited satisfactory growth inhibition capacity against the two main dental pathogens, namely, Streptococcus mutans and Porphyromonas gingivalis. The Ag-CS NPs inhibited the bacterial adhesion and decreased biofilm creation. Moreover, the nanoparticles inhibited the quorum sensing generation in both microorganisms. The nanoparticles were biocompatible, as they did not display any cell cytotoxicity approving that they could be used as suitable coatings for the titanium dental implants that can result in corrosion resistant dental implants with greater passivating properties. Some CS-based hydrogel implants were fabricated and employed for regeneration of peripheral nervous tissue by electrodeposition technique by means of a solution containing CS and an organic acid [149]. To enhance mechanical strengths of implants, hydroxyapatite was introduced into the solution which was also utilized as a source for calcium ions. Effects of additive and polymer concentrations were evaluated on chemical, mechanical, and biological features of the implants. It was found that the physicochemical characteristics of the resulting structures were extremely related to the original composition of solution. The in vitro proinflammatory as well as cytotoxic assessments exhibited biocompatibility of developed implants which proposed that the animal tests could be carried out.

11. Application of natural polysaccharides in preparation of antibacterial textiles/papers Production of antibacterial textiles is significant in the textile industry. Although a lot of chemicals and methods exist to manufacture antibacterial textiles, all of them are not benign to humans [150e152]. In order to overcome these shortcomings, biocompatible and ecofriendly materials should be developed and used for the fabrication of antibacterial textiles. The natural biopolymeric polysaccharides which are commonly utilized in biomedical application are suitable candidates for the preparation of antibacterial textiles/papers [81]. An ecological green synthesis of chitosan/neem seed (CS/NS) composite was carried out using aqueous neem seed extract through coprecipitation technique [153]. Cotton fabrics were modified with glutaraldehyde and citric acid as cross-linkers and then the composite was coated on the cotton fabric through chemical linkages formed between cellulose and composite. The bactericidal potency of the CS/NS composite covered cotton fabric was assessed in absence and presence of cross-linkers using both Gram-negative and Gram-positive bacteria by means of agar well diffusion technique. It was validated that the CS/NS composite coated on the cotton fabric showed a higher bactericidal effect

48 Chapter 2 compared to the cotton fabric lacking cross-linkers. Accordingly, the CS-neem seed composite could be utilized to achieve medicinal textiles. A facile and economical method was developed to achieve antibacterial cotton fabric using biodegradable gum tragacanth (GT) and silver nanoparticles (AgNPs) [154]. Diverse GT concentrations (2, 4, and 6 g/L) and an Ag amount of 5% relative to weight of dry GT were employed to examine their influences on the physical, mechanical, biological properties and antibacterial efficacy (against E. coli and S. aureus) of cotton fabric. Adding low amount of Ag NPs in the composite enhanced the antibacterial effect of fabric relative to fabric only treated with GT. Additionally, the cotton treated with 4% GT-Ag showed appropriate stiffness and tensile strength in comparison to the fabric treated with 6% GT-Ag composite. It was found that the GT as well as GT-Ag treated fabrics were biocompatible to fibroblast cells. In another effort, papers coated with chitosan:starch-silver nanoparticles (CS:St-AgNPs) were prepared to be applied in antimicrobial packages [155]. The St-AgNPs had a spherical morphology and a mean diameter of 7 nm. CS was added to the St-AgNPs mixture in diverse ratios (9:1, 8:2, 7:3, and 5:5) and the effect of various CS:St-AgNPs ratios were assessed on the papers characteristics including oil and water resistance, mechanical properties, and antimicrobial capacities. The characteristics of the papers coated by nanoparticles were highly related to the CS:St-AgNPs ratio. The CS:St-AgNPs coated paper fabricated using the 9:1 ratio exhibited exceptional mechanical characteristics and suitable resistance to oil and water. The CS:St-AgNPs coated papers presented significant improvement in oil and water resistance, mechanical strength, antifungal and bactericidal efficacy confirming they were potential candidates for preparation of functional antimicrobial textiles/papers.

12. Application of natural polysaccharides in preparation of antimicrobial food additives/packaging materials Food safety is a major challenge; especially, it is vital due to the growing consumer demand for fast foods. Consequently, significance of food storage as well as preservation is enhanced. In fact, there is an increasing attention for natural and higher quality foods prepared without using chemical preservers. Application of natural antimicrobial agents to produce safe and healthy foods is a favorable outlook [156]. Recently, corn starch was employed as a polymeric material to develop antimicrobial packaging materials using pediocin or nisin as food preservatives [157]. Halloysite clay was utilized as nanofiller to reinforce the films and bacteriocins were adsorbed on nanoclay before its addition to the film forming solutions. The films were active against

Pharmaceutical applications of natural polysaccharides 49 Clostridium perfringens and Listeria monocytogenes and halloysite preserved antimicrobial efficacy compared to the films lacking nanofiller. It was found that the adsorption method was promising to retain a suitable crystallinity and uniform shape for the films. For the nanocomposite containing nisin, adsorption enhanced the water barrier property. The mechanical resistances of films loaded by the halloysite were increased. Also, the elongation at break was considerably improved for films containing pediocin or nisin in addition to those incorporated with nisin and halloysite. Thus, these antimicrobial nanocomposite films were suitable food packaging materials. Several antimicrobial films were achieved using calcium alginate and lysozyme [158]. In order to optimize the antimicrobial activity, ultrasonic irradiation was used to enhance the immobilization efficacy. Diverse ultrasonic duration and power were applied. It was found that sonication speeded up the lysozyme immobilization rate and increased the lysozyme amount immobilized on the supports. The catalytic performance of the microbicidal film was assessed by the turbidimetric test and showed the greatest value using 147.8 W. Also, the antimicrobial effect was enhanced by sonication and determined through the inhibition zone method. To discover the mechanism of ultrasonic influence on lysozyme immobilization, the changes in lysozyme structures were studied before and after the ultrasonic irradiation. The fluorescence and circular dichroism spectra demonstrated that sonication influenced both the secondary and tertiary structures of lysozyme. Also, structural variations in the enzyme improved the enzymatic activity. Sonication impacts on the films’ microstructures were followed by the scanning electron microscopy and it was observed that the film surface had several cracks after sonication. Nisin is a famous bacteriocin which is approved to be used as a food additive in food preservation. Some nisin-incorporated pectin-inulin particles were achieved in order to avoid interaction of this bacteriocin with food components [159]. To prepare particles, pectins with diverse esterification degrees were utilized. Combining pectin and inulin improved the effectiveness of nisin addition compared to nisin-pectin samples. For all pectins examined using nisin amounts of 0.1e1.0 mg/mL in pH values of 4.0e7.0, the loading efficiency was 100%. The inulin and pectin combination for particle preparation slightly enhanced the microbicidal potency of nisin compared to nisin-pectin particles. Also, the antimicrobial effects of nisin-containing pectin-inulin particles depended on the pectin esterification degree. All particles having low pectin esterification degree or no esterified pectic acid revealed greater activity than the particles with high pectin esterification degree. High nisin-loading and comparable antimicrobial potency of inulinpectin particles compared to those of the nisin-pectin particles proved that combining inulin with pectin was very effective to produce antimicrobial particles for application in food industry.

50 Chapter 2

13. Conclusion In this work, the pharmaceutical applications of natural polysaccharides were reviewed in numerous biomedical fields such as cell encapsulation, drug/gene delivery, protein binding, wound healing, tissue engineering, bioimaging, preparation of contact lenses and implants, antibacterial textiles/papers, and antibacterial food additives/packaging materials. In case of cell encapsulation, alginate was used to encapsulate the human adiposeederived stem cells in order to increase their maintenance during hypothermic storage and produce them in large scale. The gold nanoparticles were synthesized using gum karaya and applied to deliver the gemcitabine hydrochloride anticancer drug. For gene delivery application, 6-amino-curdlan was used to achieve nanoparticles through its complexation with siRNAs which delivered siRNAs to mouse primary cells and human cancer cells and decreased 70%e90% of the target mRNA level. As an example of protein binding, the polysaccharide-protein-surfactant complexes were obtained using propylene glycol alginate, zein along with lecithin or rhamnolipid and applied as curcumin delivery platforms. Also, the FWEP polysaccharide isolated from fenugreek (Trigonella foenumgraecum) seeds revealed antioxidant, hemolytic, and in vivo wound healing features. The blends of xanthan gum and CS using Fe3O4magnetic nanoparticles yielded magnetic responsive polyelectrolyte complex hydrogels which were appropriate materials for tissue engineering. The nanocomposites fabricated using natural polysaccharides and quantum dots were applied in bioimaging applications. Moreover, natural polysaccharides containing various drugs were employed to coat contact lenses in order to release the desired drug in a controlled rate. As implant materials, the alginate/polyacrylamide hybrid hydrogels were fabricated as orthopedic prosthesis compounds. Also, the natural polysaccharides were utilized to prepare antibacterial textiles and papers. The corn starch was applied to obtain antimicrobial packaging materials using nisin or pediocin for food preservation. Finally, it can be stated that natural polysaccharides are very valuable materials that have found significant pharmaceutical applications.

Acknowledgment The authors appreciatively acknowledge all financial supports of this research by the Research Office of Amirkabir University of Technology (Tehran Polytechnic).

References [1] Seidi F, Jenjob R, Phakkeeree T, Crespy D. Saccharides, oligosaccharides, and polysaccharides nanoparticles for biomedical applications. J Control Release 2018;284:188e212. [2] Shariatinia Z, Zahraee Z. Controlled release of metformin from chitosanebased nanocomposite films containing mesoporous MCM-41 nanoparticles as novel drug delivery systems. J Colloid Interface Sci 2017;501:60e76.

Pharmaceutical applications of natural polysaccharides 51 [3] Kohsari I, Shariatinia Z, Pourmortazavi SM. Antibacterial electrospun chitosan-polyethylene oxide nanocomposite mats containing ZIF-8 nanoparticles. Int J Biol Macromol 2016;91:778e88. [4] Simo´a G, Ferna´ndez-Ferna´ndeza E, Vila-Crespob J, Ruipe´rezb V, Rodrı´guez-Nogales JM. Research progress in coating techniques of alginate gel polymer forcell encapsulation. Carbohydr Polym 2017;170:1e14. [5] Kohsari I, Shariatinia Z, Pourmortazavi SM. Antibacterial electrospun chitosanepolyethylene oxidenanocomposite mats containing bioactive silver nanoparticles. Carbohydr Polym 2016;140:287e98. [6] Fazli Y, Shariatinia Z, Kohsari I, Azadmehr A, Pourmortazavi SM. A novel chitosan-polyethylene oxide nanofibrous mat designed for controlled co-release of hydrocortisone and imipenem/cilastatin drugs. Int J Pharm 2016;513:636e47. [7] Germershaus O, Lu¨hmann T, Rybak J-C, Ritzer J, Meinel L. Application of natural and semi-synthetic polymers for the delivery of sensitive drugs. Int Mater Rev 2015;60:101e30. [8] Shariatinia Z, Nikfar Z. Synthesis and antibacterial activities of novel nanocomposite films of chitosan/ phosphoramide/Fe3O4 NPs. Int J Biol Macromol 2013;60:226e34. [9] Shariatinia Z, Nikfar Z, Gholivand K, Abolghasemi Tarei S. Antibacterial activities of novel nanocomposite biofilms of chitosan/phosphoramide/Ag NPs. Polym Compos 2015;36:454e66. [10] Ngwuluka NC, Ochekpe NA, Aruoma OI. Naturapolyceutics: the science of utilizing natural polymers for drug delivery. Polymers 2014;6:1312e32. [11] Cordeiro AS, Jose´ Alonso M, de la Fuente M. Nanoengineering of vaccines using natural polysaccharides. Biotechnol Adv 2015;33:1279e93. [12] Shariatinia Z, Mazloom Jalali A. Chitosan-based hydrogels: preparation, properties and applications. Int J Biol Macromol 2018;115:194e220. [13] Shariatinia Z, Fazli M. Mechanical properties and antibacterial activities of novel nanobiocomposite films of chitosan and starch. Food Hydrocolloids 2015;46:112e24. [14] Fazli Y, Shariatinia Z. Controlled release of cefazolin sodium antibiotic drug from electrospun chitosan-polyethylene oxide nanofibrous mats. Mater Sci Eng C 2017;71:641e52. [15] Shelke NB, James R, Laurencin CT, Kumbar SG. Polysaccharide biomaterials for drug delivery and regenerative engineering. Polym Adv Technol 2014;25:448e60. [16] Ali A, Ahmed S. A review on chitosan and its nanocomposites in drug delivery. Int J Biol Macromol 2018;109:273e86. [17] Zamboni F, Collins MN. Cell based therapeutics in type 1 diabetes mellitus. Int J Pharm 2017;521:346e56. [18] Algire GH, Weaver JM, Prehn RT. Growth of cells in vivo in diffusion chambers. I. Survival of homografts in mice. J Natl Cancer Inst 1954;15:493e507. [19] Rokstad AMA, Lacı´k I, de Vos P, Strand BL. Advances in biocompatibility and physico-chemical characterization of microspheres for cell encapsulation. Adv Drug Deliv Rev 2014;67e68:111e30. [20] Koo J, Chang TSM. Secretion of erythropoietin from microencapsulated rat kidney cells. Int J Artif Organs 1993;16:557e60. [21] Liu HW, Ofosu FA, Chang PL. Expression of human factor IX by microencapsulated recombinant fibroblasts. Hum Gene Ther 1993;4:291e301. [22] Chang PL, Shen N, Westcott AJ. Delivery of recombinant gene products with microencapsulated cells in vivo. Hum Gene Ther 1993;4:433e40. [23] Colton CK. Implantable biohybrid artificial organs. Cell Transplant 1995;4:415e36. [24] Uludag H, Sefton MV. Microencapsulated human hepatoma (HepG2) cells: in vitro growth and protein release. J Biomed Mater Res 1993;27:1213e24. [25] Cieslinski DA, David Humes H. Tissue engineering of a bioartificial kidney. Biotechnol Bioeng 1994;43:678e81. [26] Lim F, Sun AM. Microencapsulated islets as bioartificial endocrine pancreas. Science 1980;210:908e10. [27] Aebischer P, Goddard M, Signore AP, Timpson RL. Functional recovery in hemiparkinsonian primates transplanted with polymer-encapsulated PC12 cells. Exp Neurol 1994;126:151e8.

52 Chapter 2 [28] Dulieu C, Poncelet D, Neufeld RJ. Encapsulation and immobilization techniques. In: Ku¨htreiber WM, Lanza RP, Chick WL, editors. Cell encapsulation technology and therapeutics. Bosto: Birkha¨user; 1999. p. 3e17. [29] De Vos P, Lazarjani HA, Poncelet D, Faas MM. Polymers in cell encapsulation from an enveloped cell perspective. Adv Drug Deliv Rev 2014;67:15e34. [30] Doh K-O, Yeo Y. Application of polysaccharides for surface modification of nanomedicines. Ther Deliv 2012;3:1447e56. [31] Passirani C, Barratt G, Devissaguet JP, Labarre D. Interactions of nanoparticles bearing heparin or dextran covalently bound to poly(methylmethacrylate) with the complement system. Life Sci 1998;62:775e85. [32] Shariatinia Z. Pharmaceutical applications of chitosan. Adv Colloid Interface Sci 2019;263:131e94. [33] Costa JR, Silva NC, Sarmento B, Pintado M. Delivery systems for antimicrobial peptides and proteins: towards optimization of bioavailability and targeting. Curr Pharmaceut Biotechnol 2017;18:108e20. [34] Gaber M, Medhat W, Hany M, Saher N, Fang J-Y, Elzoghby A. Protein lipid nanohybrids as emerging platforms for drug and gene delivery: challenges and outcomes. J Control Release 2017;254:75e91. [35] Shariatinia Z. Carboxymethyl chitosan: properties and biomedical applications. Int J Biol Macromol 2018;120:1406e19. [36] Song M, Li L, Zhang Y, Chen K, Wang H, Gong R. Carboxymethyl-b-cyclodextrin grafted chitosan nanoparticles as oral delivery carrier of protein drugs. React Funct Polym 2017;117:10e5. [37] Tolstoguzov V. Some thermodynamic considerations in food formulation. Food Hydrocolloids 2003;17:1e23. [38] Ferreira CO, Nunes CA, Delgadillo I, Lopes-Da-Silva JA. Characterization of chitosan-whey protein films at acid pH. Food Res Int 2009;42:807e13. [39] Dickinson E. Interfacial structure and stability of food emulsions as affected by protein-polysaccharide interactions. Soft Matter 2008;4:932e42. [40] Gu YS, Decker EA, Mcclements DJ. Application of multi-component biopolymer layers to improve the freeze-thaw stability of oil-in-water emulsions: b-Lactoglobulin-i-carrageenan-gelatin. J Food Eng 2007;80:1246e54. [41] Nori MP, Favaro-Trindade CS, Alencar SMD, Thomazini M, de Camargo Balieiro JC, Contreras Castillo CJ. Microencapsulation of propolis extract by complex coacervation. LWT - Food Sci Technol 2011;44:429e35. [42] Xiao JX, Huang GQ, Wang SQ, Sun YT. Microencapsulation of capsanthin by soybean protein isolatechitosan coacervation and microcapsule stability evaluation. J Appl Polym Sci 2014;131:257e65. [43] Laneuville SI, Turgeon SL, Sanchez C, Paquin P. Gelation of native b-lactoglobulin induced by electrostatic attractive interaction with xanthan gum. Langmuir 2006;22:7351. [44] Le XT, Turgeon SL. Rheological and structural study of electrostatic crosslinked xanthan gum hydrogels induced by b-lactoglobulin. Soft Matter 2013;9:3063e73. [45] Chen WS, Soucie WG. Edible fibrous serum milk protein/xanthan gum complexes. 1985. United States Patent, 4559233. [46] Schmitt C, Sanchez C, Desobrybanon S, Hardy J. Structure and technofunctional properties of proteinpolysaccharide complexes: a review. Crit Rev Food Sci Nutr 1998;38:689e753. [47] Strauss G, Gibson SM. Plant phenolics as cross-linkers of gelatin gels and gelatin-based coacervates for use as food ingredients. Food Hydrocolloids 2004;18:81e9. [48] Abdel-Azim NS, Shams KA, Shahat AA, El Missiry MM, Ismail SI, Hammouda FM. Egyptian herbal drug industry: challenges and future prospects. Res J Med Plant 2011;5:136e44. [49] Kulkarni AD, Joshi AA, Patil CL, Amale PD, Patel HM, Surana SJ, Belgamwar VS, Chaudhari KS, Pardeshi CV. Xyloglucan: a functional biomacromolecule for drug delivery applications. Int J Biol Macromol 2017;104:799e812. [50] Pereira LDP, Mota MRL, Brizeno LAC, Nogueira FC, Ferreira EGM, Pereira MG, Assreuy AMS. Modulator effect of a polysaccharide-rich extract from Caesalpinia ferrea stem barks in rat cutaneous wound healing: role of TNF-a IL-1b, NO, TGF-b. J Ethnopharmacol 2016;187:213e23.

Pharmaceutical applications of natural polysaccharides 53 [51] Trombetta D, Puglia C, Perri D, Licata A, Pergolizzi S, Lauriano ER. Effect of polysaccharides from Opuntia ficus-indica (L.) cladodes on the healing of dermal wounds in the rat. Phytomedicine 2006;13:352e8. [52] Bae JS, Jang KH, Park SC, Jin HK. Promotion of dermal wound healing by polysaccharides isolated from Phellinus gilvus in rats. J Vet Med Sci 2005;67:111e4. [53] Tabandeh MR, Oryon A, Mohammadalipour A. Polysaccharides of Aloe vera induce MMP-3 and TIMP-2 gene expression during the skin wound repair of rat. Int J Biol Macromol 2014;65:424e30. [54] Kumar A, Madhusudana Rao K, Han SS. Application of xanthan gum as polysaccharide in tissue engineering: a review. Carbohydr Polym 2018;180:128e44. [55] Ulery BD, Nair LS, Laurencin CT. Biomedical applications of biodegradable polymers. J Polym Sci B Polym Phys 2011;49:832e64. [56] Zohuri GH, Sandaroos R, Ahmadjo S, Damavandi S, Rabiee A, Shamekhi MA. Tissue engineering: biomaterial application, Encycl. Biomed. Polym. Polym. Biomater. CRC Press; 2015. p. 7901e32. [57] Salerno A, Pascual CD. Bio-based polymers, supercritical fluids and tissue engineering. Process Biochem 2015;50:826e38. [58] Silva AK, Juenet M, Meddahi-Pelle´ A, Letourneur D. Polysaccharide-based strategies for heart tissue engineering. Carbohydr Polym 2015;116:267e77. [59] Pina S, Oliveira JM, Reis RL. Natural-based nanocomposites for bone tissue engineering and regenerative medicine: a review. Adv Mater 2015;27:1143e69. [60] Mansur HS, Mansur AAP, Curti E, de Almeida MV. Bioconjugation of quantum-dots with chitosan and N,N,N-trimethyl chitosan. Carbohydr Polym 2012;90:189e96. [61] Luna-Martı´nez JF, Herna´ndez-Uresti DB, Reyes-Melo ME, Guerrero-Salazar CA, Gonza´lezGonza´lez VA, Sepu´lveda-Guzma´n S. Synthesis and optical characterization of ZnSesodium carboxymethyl cellulose nanocomposite films. Carbohydr Polym 2011;84:566e70. [62] Mansur AAP, Ramanery FP, Oliveira LC, Mansur HS. Carboxymethylchitosan functionalization of Bi2S3 quantum dots: towards eco-friendly fluorescent core-shell nanoprobes. Carbohydr Polym 2016;146:455e66. [63] Tang CR, Su ZH, Lin BG, Huang HW, Zeng YL, Li S, Huang H, Wang YJ, Li CX, Shen GL, Yu RQ. A novel method for iodate determination using cadmium sulfide quantum dots as fluorescence probes. Anal Chim Acta 2010;678:203e7. [64] Chen L, Lai C, Marchewka R, Berry RM, Tam KC. Use of CdS quantum dot-functionalized cellulose nanocrystal films for anti-counterfeiting applications. Nanoscale 2016;8:13288e96. [65] Chan WCW, Nie SM. Quantum dot bioconjugates for ultrasensitive nonisotopic detection. Science 1998;281:2016e8. [66] Das R, Bandyopadhyay R, Pramanik P. Carbon quantum dots from natural resource: a review. Materials Today Chemistry 2018;8:96e109. [67] Toda R, Kawazu K, Oyabu M, Miyazaki T, Kiuchi Y. Comparison of drug permeabilities across the blooderetinal barrier, bloodeaqueous humor barrier, and bloodebrain barrier. J Pharm Sci 2011;100:3904e11. [68] Raviola G. The structural basis of the blood-ocular barriers. Exp Eye Res 1977;25:27e63. [69] Garnett BD. The contact lens manual: a practical fitting guide. Hum Immunol 1993;63:S78. [70] Creech JL, Chauhan A, Radke CJ. Dispersive mixing in the posterior tear film under a soft contact lens. Ind Eng Chem Res 2001;40(14):3015e26. [71] Helsen JA, Breme HJ. Metals as biomaterials. 1998. p. 30e40. Chichester, New York. [72] Neoh KG, Hu X, Zheng D, Kang ET. Balancing osteoblast functions andbacterial adhesion on functionalized titanium surfaces. Biomaterials 2012;33:2813e22. [73] Li M, Liu X, Xu Z, Yeung KWK, Wu S. Dopamine modified organic-inorganic hybrid coating for antimicrobial and osteogenesis. ACS Appl Mater Interfaces 2016;8(49):33972e81. [74] Darouiche RO. Treatment of infections associated with surgical implants. N Engl J Med 2004;350:1422e9.

54 Chapter 2 [75] Liu X, Li M, Zhu Y, Yeung KWK, Chu PK, Wu S. The modulation of stem cell behaviors by functionalized nanoceramic coatings on Ti-based implants. Bioact Mater 2016;1:65e76. [76] Wu S, Liu X, Yeung KWK, Liu C, Yang X. Biomimetic porous scaffolds for bone tissue engineering. Mater Sci Eng R 2014;80:1e36. [77] Xu Z, Man L, Xia L, Liu X, Fei M, Wu S, Yeung KWK, Han Y, Chu PK. Antibacterial activity of silver doped titanate nanowires on Ti implants. ACS Appl Mater Interfaces 2016;8:16584e94. [78] Zhou T, Zhu Y, Li X, Liu X, Yeung KWK, Wu S, Wang X, Cui Z, Yang X, Chu PK. Surface functionalization of biomaterials by radical polymerization. Prog Mater Sci 2016;83:191e235. [79] Rinaudo M. Chitin and chitosan: properties and applications. Prog Polym Sci 2006;31:603e32. [80] Rajendran R, Radhai R, Kotresh TM, Csiszar E. Development of antimicrobial cotton fabrics using herb loaded nanoparticles. Carbohydr Polym 2013;91. 613-317. [81] Ul-Ialam S, Shahid M, Mohammad F. Green chemistry approaches to develop antimicrobial textiles based on sustainable biopolymers-a review. Ind Eng Chem Res 2013;52:5245e60. [82] Mahato K, Srivastava A, Chandra P. Paper based diagnostics for personalized health care: emerging technologies and commercial aspects. Biosens Bioelectron 2017;15:246e59. [83] Reshetnyak EA, Ostrovskaya VM, Goloviznina KV, Kamneva NN. Influence of tetraalkylammonium halides on analytical properties of universal acid-base indicator paper. J Mol Liq 2017;248:610e5. [84] Schoonover DV, Gibson HW. Facile removal of tosyl chloride from tosylates using cellulosic materials, e.g., filter paper. Tetrahedron Lett 2017;58:242e4. [85] Rastogi VK, Samyn P. Bio-based coatings for paper applications. Coatings 2015;5:887e930. [86] Piselli A, Garbagnoli P, Alfieri I, Lorenzi A, Curto BD. Natural-based coatings for food paper packaging. Int J Des Sci Technol 2014;20:55e78. [87] Khwaldia K, Arab-Tehrany E, Desobry S. Biopolymer coatings on paper packaging materials. Compr Rev Food Sci F 2010;9:82e91. [88] Ma Y, Liu P, Si C, Liu Z. Chitosan nanoparticles: preparation and application in antibacterial paper. J Macromol Sci B 2010;49:994e1001. [89] Gottesman R, Shukla S, Perkas N, Solovyov LA, Nitzan Y, Gedanken A. Sonochemical coating of paper by microbiocidal silver nanoparticles. Langmuir 2011;27:720e6. [90] Brobbey KJ, Haapanen J, Gunell M, Ma¨kela¨ JM, Eerola E, Toivakka M, Saarinen JJ. One-step flame synthesis of silver nanoparticles for roll-to-roll production of antibacterial paper. Appl Surf Sci 2017;420:558e65. [91] Mlalila N, Hilonga A, Swai H, Devlieghere F, Ragaert P. Antimicrobial packaging based on starch, poly(3-hydroxybutyrate) and poly(lactic-co-glycolide) materials and application challenges. Trends Food Sci Technol 2018;74:1e11. [92] Appendini P, Hotchkiss JH. Review of antimicrobial food packaging. Innov Food Sci Emerg Technol 2002;3:113e26. [93] Quintavalla S, Vicini L. Antimicrobial food packaging in meat industry. Meat Sci 2002;62:373e80. [94] Sung S-Y, Sin LT, Tee T-T, Bee S-T, Rahmat AR, Rahman WAWA, Tan AC, Vikhraman M. Antimicrobial agents for food packaging applications. Trends Food Sci Technol 2013;33:110e23. [95] Wang J-Z, Ding Z-Q, Zhang F, Ye W-B. Recent development in cell encapsulations and their therapeutic applications. Mater Sci Eng C 2017;77:1247e60. [96] Orive G, Santos E, Pedraz J, Herna´ndez R. Application of cell encapsulation for controlled delivery of biological therapeutics. Adv Drug Deliv Rev 2014;67:3e14. ˚ , Blennow K, [97] Ferreira D, Westman E, Eyjolfsdottir H, Almqvist P, Lind G, Linderoth B, Seiger A Karami A, Darreh-Shori T. Brain changes in Alzheimer’s disease patients with implanted encapsulated cells releasing nerve growth factor. J Alzheimers Dis 2015;43:1059e72. [98] Vegas AJ, Veiseh O, Gu¨rtler M, Millman JR, Pagliuca FW, Bader AR, Doloff JC, Li J, Chen M, Olejnik K. Long-term glycemic control using polymer-encapsulated human stem cell-derived beta cells in immune-competent mice. Nat Med 2016;22(3):306e11.

Pharmaceutical applications of natural polysaccharides 55 [99] Dong D, Hao T, Wang C, Zhang Y, Qin Z, Yang B, Fang W, Ye L, Yao F, Li J. Zwitterionic starchbased hydrogel for the expansion and “stemness” maintenance of brown adipose derived stem cells. Biomaterials 2018;157:149e60. [100] Cacciotti I, Ceci C, Bianco A, Pistritto G. Neuro-differentiated Ntera2 cancer stem cells encapsulated in alginate beads: first evidence of biological functionality. Mater Sci Eng C 2017;81:32e8. [101] Swioklo S, Ding P, Pacek AW, Connon CJ. Process parameters for the high-scale production of alginateencapsulated stem cells for storage and distribution throughout the cell therapy supply chain. Process Biochem 2017;59:289e96. [102] Raveendran S, Palaninathan V, Nagaoka Y, Fukuda T, Iwai S, Higashi T, Mizuki T, Sakamoto Y, Mohanan PV, Maekawa T, Kumar DS. Extremophilic polysaccharide nanoparticles for cancer nanotherapyand evaluation of antioxidant properties. Int J Biol Macromol 2015;76:310e9. [103] Pooja D, Panyaram S, Kulhari H, Reddy B, Rachamalla SS, Sistla R. Natural polysaccharide functionalized gold nanoparticles asbiocompatible drug delivery carrier. Int J Biol Macromol 2015;80:48e56. [104] Deore UV, Mahajan HS. Isolation and characterization of natural polysaccharide from Cassia Obtustifolia seed mucilage as film forming material for drug delivery. Int J Biol Macromol 2018;115:1071e8. [105] Anderson W. Human gene therapy. Science 1992;256:808e13. [106] Oliveira AV, Rosa da Costa AM, Silva GA. Non-viral strategies for ocular gene delivery. Mater Sci Eng C 2017;77:1275e89. [107] Rosenberg SA, Aebersold P, Cornetta K, Kasid A, Morgan RA, Moen R, Karson EM, Lotze MT, Yang JC, Topalian SL, Merino MJ, Culver K, Miller AD, Blaese RM, Anderson WF. Gene transfer into humansdimmunotherapy of patients with advanced melanoma, using tumor-infiltrating lymphocytes modified by retroviral gene transduction. N Engl J Med 1990;323:570e8. [108] Leiden JM. Gene therapy-promise, pitfalls, and prognosis. N Engl J Med 1995;333:871e3. [109] Cotrim AP, Baum BJ. Gene therapy: some history, applications, problems, and prospects. Toxicol Pathol 2008;36:97e103. [110] Han J, Jia C, Borjihan W, Ganbold T, Tariq M. Rana, Huricha Baigude, Preparation of novel curdlan nanoparticles for intracellular siRNA delivery, Carbohydr. Polymers 2015;117:324e30. [111] Fernandez-Pin˜eiro I, Pensado A, Badiola I, Sanchez A. Development and characterisation of chondroitin sulfate- and hyaluronic acid-incorporated sorbitan ester nanoparticles as gene delivery systems. Eur J Pharm Biopharm 2018;125:85e94. [112] Wang R, Tian Z, Chen L. Nano-encapsulations liberated from barley protein microparticles for oral delivery of bioactive compounds. Int J Pharm 2011;406:153e62. [113] Dickinson E. Hydrocolloids as emulsifiers and emulsion stabilizers. Food Hydrocolloids 2009;23:1473e82. [114] Dai L, Wei Y, Sun C, Mao L, McClements DJ, Gao Y. Development of protein-polysaccharide-surfactant ternary complex particles as delivery vehicles for curcumin. Food Hydrocolloids 2018;85:75e85. [115] Hu B, Chen Q, Cai Q, Fan Y, Wilde PJ, Rong Z, Zeng X. Gelation of soybean protein and polysaccharides delays digestion. Food Chem 2017;221:1598e605. [116] Wang Y, Xu S, Xiong W, Pei Y, Li B, Chen Y. Nanogels fabricated from bovine serum albumin and chitosan viaself-assembly for delivery of anticancer drug. Colloids Surfaces B Biointerfaces 2016;146:107e13. [117] Zhang H, Chen J, Cen Y. Burn wound healing potential of a polysaccharide from Sanguisorba officinalis L. in mice. Int J Biol Macromol 2018;112:862e7. [118] Selig HF, Lumenta DB, Giretzlehner M, Jeschke MG, Upton D, Kamolz LP. The properties of an "ideal" burn wound dressingdwhat do we need in daily clinical practice? Results of a worldwide online survey among burn care specialists. Burns 2012;38:960e6. [119] Waghmare VS, Wadke PR, Dyawanapelly S, Deshpande A, Jain R, Dandekar P. Starch based nanofibrous scaffolds for wound healing applications. Bioactive Mater 2018;3:255e66.

56 Chapter 2 [120] Basu A, Kunduru KR, Abtew E, Domb AJ. Polysaccharide-based conjugates for biomedical applications. Bioconjug Chem 2015;26:1396e412. [121] Poonguzhali R, Khaleel Basha S, Sugantha Kumari V. Fabrication of asymmetric nanostarch reinforced Chitosan/PVP membrane and its evaluation as an antibacterial patch for in vivo wound healing application. Int J Biol Macromol 2018;114:204e13. [122] Ktari N, Trabelsi I, Bardaa S, Triki M, Bkhairia I, Slama-Ben Salem RB, Nasri M, Salah RB. Antioxidant and hemolytic activities, and effects in rat cutaneous wound healing of a novel polysaccharide from fenugreek (Trigonellafoenum-graecum) seeds. Int J Biol Macromol 2017;95:625e34. [123] Singh S, Gupta A, Sharma D, Gupta B. Dextran based herbal nanobiocomposite membranes for scar free wound healing. Int J Biol Macromol 2018;113:227e39. [124] Chi N-H, Yang M-C, Chung T-W, Chou N-K, Wang S-S. Cardiac repair using chitosan-hyaluronan/silk fibroin patches in a rat heart model with myocardial infarction. Carbohydr Polym 2013;92:591e7. [125] Silvestri A, Boffito M, Sartori S, Ciardelli G. Biomimetic materials and scaffolds for myocardial tissue regeneration. Macromol Biosci 2013;13:984e1019. [126] Hollister SJ. Porous scaffold design for tissue engineering. Nat Mater 2005;4:518e24. [127] De Mulder EL, Buma P, Hannink G. Anisotropic porous biodegradable scaffolds for musculoskeletal tissue engineering. Materials 2009;2:1674e96. [128] Saludas L, Pascual-Gil S, Pro´sper F, Garbayo E, Blanco-Prieto M. Hydrogel based approaches for cardiac tissue engineering. Int J Pharm 2017;523:454e75. [129] Sokolsky-Papkov M, Agashi K, Olaye A, Shakesheff K, Domb AJ. Polymer carriers for drug delivery in tissue engineering. Adv Drug Deliv Rev 2007;59:187e206. [130] Liu Y, Wang S, Zhang R. Composite poly(lactic acid)/chitosan nanofibrous scaffolds for cardiac tissue engineering. Int J Biol Macromol 2017;103:1130e7. [131] Nelson DM, Ma Z, Fujimoto KL, Hashizume R, Wagner WR. Intra-myocardial biomaterial injection therapy in the treatment of heart failure: materials, outcomes and challenges. Acta Biomater 2011;7:1e15. [132] Place ES, Evans ND, Stevens MM. Complexity in biomaterials fortissue engineering. Nat Mater 2009;8:457e70. [133] Shakir M, Zia I, Rehman A, Ullah R. Fabrication and characterization of nanoengineered biocompatible n-HA/chitosan-tamarind seed polysaccharide: bio-inspired nanocomposites for bone tissue engineering. Int J Biol Macromol 2018;111:903e16. [134] Choudhury P, Kumar S, Singh A, Kumar A, Kaur N, Sanyasi S, Chawla S, Goswami C, Goswami L. Hydroxyethyl methacrylate grafted carboxy methyl tamarind (CMT-g-HEMA) polysaccharide based matrix as a suitable scaffold for skin tissue engineering. Carbohydr Polym 2018;189:87e98. [135] Madhusudana Rao K, Kumar A, Han SS. Polysaccharide-based magnetically responsive polyelectrolyte hydrogels for tissue engineering applications. J Mater Sci Technol 2018;34:1371e7. [136] Alhaique F, Matricardi P, Meo CD, Coviello T, Montanari E. Polysaccharide-based self-assembling nanohydrogels: an overview on 25-years research on pullulan. J Drug Deliv Sci Technol 2015;30:300e9. [137] Mansur AAP, de Carvalho FG, Mansur RL, Carvalho SM, de Oliveira LC, Mansur HS. Carboxymethylcellulose/ZnCdS fluorescent quantum dot nanoconjugates for cancer cell bioimaging. Int J Biol Macromol 2017;96:675e86. [138] Chintakunta R, Buaron N, Kahn N, Moriah A, Lifshiz R, Goldbart R, Traitel T, Tyler B, Brem H, Kost J. Synthesis, characterization, and self-assembly with plasmid DNA of a quaternary ammonium derivative of pectic galactan and its fluorescent labeling for bioimaging applications, Carbohydr. Polymers 2016;150:308e18. [139] Zhao Q, Wang S, Yang Y, Li X, Di D, Zhang C, Jiang T, Wang S. Hyaluronic acid and carbon dotsgated hollow mesoporous silica for redox and enzyme-triggered targeted drug delivery and bioimaging. Mater Sci Eng C 2017;78:475e84. [140] Xu J, Xue Y, Hu G, Lin T, Gou J, Yin T, He H, Zhang Y, Tang X. A comprehensive review on contact lens for ophthalmic drug delivery. J Control Release 2018;281:97e118.

Pharmaceutical applications of natural polysaccharides 57 [141] Silva D, Pinto LFV, Bozukova D, Santos LF, Serro AP, Saramago B. Chitosan/alginate based multilayers to control drug release from ophthalmic lens. Colloids Surfaces B Biointerfaces 2016;147:81e9. [142] Anirudhan TS, Nair AS, Parvathy J. Extended wear therapeutic contact lens fabricated from timolol imprinted carboxymethyl chitosan-g-hydroxy ethyl methacrylate-g-polyacrylamide as a onetime medication for glaucoma. Eur J Pharm Biopharm 2016;109:61e71. [143] Hu X, Tan H, Hao L. Functional hydrogel contact lens for drug delivery in the application of oculopathy therapy. Int J Mech Behav Biomed Mater 2016;64:43e52. [144] Blum MM, Ovaert TC. Investigation of friction and surface degradation of innovative boundary lubricant functionalized hydrogel material for use as artificial articular cartilage. Wear 2013;301:201e9. [145] Kobayashi M, Chang Y-S, Oka M. A two year in vivo study of polyvinyl alcohol hydrogel (PVA-H) artificial meniscus. Biomaterials 2005;26:3243e8. [146] Lin P, Zhang R, Wang X, Cai M, Yang J, Yu B, Zhou F. Articular cartilage inspired bilayer tough hydrogel prepared by interfacial modulated polymerization showing excellent combination of high loadbearing and low friction performance. ACS Macro Lett 2016;5:1191e5. [147] Arjmandi M, Ramezani M, Nand A, Neitzert T. Experimental study on friction and wear properties of interpenetrating polymer network alginate-polyacrylamide hydrogels for use in minimally invasive joint implants. Wear 2018;406e407:194e204. [148] Divakar DD, Jastaniyah NT, Altamimi HG, Alnakhli YO, Muzaheed, Alkheraif AA, Haleem S. Enhanced antimicrobial activity of naturally derived bioactivemolecule chitosan conjugated silver nanoparticle against dental implant pathogens. Int J Biol Macromol 2018;108:790e7. [149] Nawrotek K, Tylman M, Rudnicka K, Balcerzak J, Kami nski K. Chitosan-based hydrogel implants enriched with calcium ions intended for peripheral nervous tissue regeneration. Carbohydr Polym 2016;136:764e71. [150] Maryan AS, Montazer M, Harifi T. Synthesis of nano silver on cellulosic denim fabric producing yellow colored garment with antibacterial properties, Carbohydr. Polymers 2015;115:568e74. [151] Shariatinia Z, Shekarriz S, Mirhosseini Mousavi HS, Maghsoudi N, Nikfar Z. Disperse dyeing and antibacterial properties of nylon and wool fibers using two novel nanosized copper(II) complexes bearing phosphoramide ligands. Arab J Chem 2017;10:944e55. [152] Shariatinia Z, Javeri N, Shekarriz S. Flame retardant cotton fibers produced using novel synthesized halogen-free phosphoramide nanoparticles. Carbohydr Polym 2015;118:183e98. [153] Revathi T, Thambidurai S. Synthesis of chitosan incorporated neem seed extract (Azadirachtaindica) for medical textiles. Int J Biol Macromol 2017;104:1890e6. [154] Ranjbar-Mohammadi M. Production of cotton fabrics with durable antibacterial property by using gum tragacanth and silver. Int J Biol Macromol 2018;109:476e82. [155] Jung J, Kasi G, Seo J. Development of functional antimicrobial papers using chitosan/starch-silver nanoparticles. Int J Biol Macromol 2018;112:530e6. [156] Gyawali R, Ibrahim SA. Natural products as antimicrobial agents. Food Control 2014;46:412e29. [157] Meira SMM, Zehetmeyer G, Werner JO, Brandelli A. A novel active packaging material based on starch-halloysite nanocomposites incorporating antimicrobial peptides. Food Hydrocolloids 2017;63:561e70. [158] Wang D, Lv R, Ma X, Zou M, Wang W, Yan L, Ding T, Ye X, Liu D. Lysozyme immobilization on the calcium alginate film under sonication: development of an antimicrobial film. Food Hydrocolloids 2018;83:1e8. [159] Krivorotova T, Staneviciene R, Luksa J, Serviene E, Sereikaite J. Preparation and characterization of nisin-loaded pectin-inulin particles as antimicrobials. LWT - Food Sci Technol 2016;72:518e24.

CHAPTER 3

Sodium alginate in drug delivery and biomedical areas Kiran Chaturvedi1, Kuntal Ganguly1, Uttam A. More2, Kakarla Raghava Reddy3, Tanavi Dugge4, Balaram Naik4, Tejraj M. Aminabhavi1, Malleshappa N. Noolvi2 1

Department of Pharmaceutical Engineering and Polymer Science, SET’s College of Pharmacy, Dharwad, India; 2Department of Pharmaceutical Chemistry, Shree Dhanvantary Pharmacy College, Kim, Surat, India; 3School of Chemical and Biomolecular Engineering, The University of Sydney, Sydney, NSW, Australia; 4Shri Dharmasthala Manjunath Dental College, Dharwad, India

Chapter Outline 1. Introduction 60 2. Physicochemical properties 2.1 2.2 2.3 2.4 2.5 2.6 2.7

3. Regulatory status 68 4. Applications in drug delivery 4.1 4.2 4.3 4.4 4.5 4.6 4.7 4.8 4.9 4.10 4.11

62

Molecular structure and weight 62 Solubility 62 Viscosity 63 Cross-linking and gel formation 63 pH sensitivity 65 Mucoadhesivity 66 Biocompatibility 67

68

Oral delivery 69 Injectable delivery 77 Ocular delivery 81 Nasal delivery 81 Tackling obesity and weight management Wound dressings 82 Cell delivery and implants 83 Protein delivery 84 Vaccine delivery 89 Tissue engineering 89 Miscellaneous 93

5. Concluding remarks and future directions References 94

82

94

Natural Polysaccharides in Drug Delivery and Biomedical Applications. https://doi.org/10.1016/B978-0-12-817055-7.00003-0 Copyright © 2019 Elsevier Inc. All rights reserved.

59

60 Chapter 3

1. Introduction Research in the field of naturally occurring polymers has witnessed a considerable attention over the past few decades worldwide mainly because, apart from being biodegradable, these polymers are renewable if used wisely and, in most cases, available in abundance. Advances in the field of chemical technology and advanced analytical techniques have enabled better understanding of the physicochemical nature of these polymers, which in turn has hastened even more research and utilization of these natural polymers in the field of drug delivery and other biomedical applications. Alginates have emerged as one of the most extensively studied biopolymers because of their unique properties and versatility. It is cheap, nature abundant, biocompatible, water soluble, and mucoadhesive and shows the solegel transition [1,2]. Alginates are natural linear polysaccharides obtained mainly from brown algae, namely, Laminaria hyperborean, Macrocystis pyrifera, and Ascophyllum nodosum. The alginate content on dry weight basis obtained from different algal sources is Cu2þ > Cd2þ > Ba2þ > Sr2þ > Ca2þ > Co2þ, Ni2þ, Zn2þ > Mn2þ. Thus, calcium has a relatively weaker binding affinity but is still preferred because of its presence and acceptability by the human body. Apart from this, Ba2þ and Sr2þ are also considered, but Pb2þ, Cu2þ, and Cd2þ being toxic are not favored [8,9]. Jay et al. attempted for modulation of release of vascular endothelial growth factor (VEGF) from alginate

64 Chapter 3

Figure 3.2 The egg-box model for calcium alginate. Reprinted from Braccini I, Pe´rez S. Molecular basis of Ca2þinduced gelation in alginates and pectins: the egg-box model revisited. Biomacromolecules 2001;2(4):1089e96, Copyright © 2001 with permission from American Chemical Society & Lee KY, Mooney DJ. Alginate: properties and biomedical applications. Prog Polym Sci 2012;37:106e26, Copyright © 2011 with permission from Elsevier.

microparticle (80%), better ex vivo mucoadhesion, and slower drug release rate with 65.47  0.59% risperidone release after 8 h [52]. Alginate gelecoated floating beads of risperidone (EE of 73.53  1.04%) showing slow release (Q8h 75.43  1.34% vs. 73%e89% for uncoated beads) were prepared by coating olive oileentrapped calcium alginateetamarind gumemagnesium stearate composite buoyant beads using SA. The coating imparted better buoyancy to the beads (floating duration >8 h, floating lag time 0.5% chitosan coat > uncoated [59]. A multicomponent system comprising vitamin B5 liposomes encapsulated within alginate or alginate/pectin microparticles showed an EE of w60% for alginate microparticles, whereas liposomes as such could show higher EE of w75%. The decreased EE of alginate microparticles was due to loss of vitamin B5 during the particle hardening process. Moreover, as the percentage alginate content in microparticles increased (1.5%, 2%, 3% solution), the percentage of vitamin B5 release in 24 h also increased at pH 4.0 (citric acid solution) [60]. The destabilizing effect on alginate on the liposome membrane [61] is one of the probable reasons for this. Furthermore, an increase in percentage of pectin in the

Sodium alginate in drug delivery and biomedical areas 75

1.0

0.8 0.6 0.4 Colloidosome Pure alginate microsphere Bare insulin crystal

0.2

Commulative release fraction

Commulative release fraction

1.0

0.8 0.6 0.4 Colloidosome Pure alginate microsphere Bare insulin crystal

0.2 0.0

0.0 0

2 Time (h)

4

0

20

40

60 80 Time (h)

100

120

Figure 3.7 Schematic illustration of the processes for preparing colloidosome, gelation of SA, polar optical microscopy photos of colloidosomes, release curve at pH 1.2 and 7.4. Reprinted from Liu H, Wang C, Gao Q, Chen J, Ren B, Liu X, et al. Facile fabrication of well-defined hydrogel beads with magnetic nanocomposite shells. Int J Pharm 2009;376:92e8, Copyright © 2009 with permission from Elsevier.

76 Chapter 3 alginate/pectin microparticles also resulted in an increase of vitamin B5 release. Here, the cross-linking ion seems to play an important role. The electrical conductivity of alginate solution is almost double than that of pectin solution. The interaction between Ca2þ ions and alginate is strong and rapid, which is not the case with pectin where more pectin chains are free to interact with phospholipid membrane of the liposome to cause a destabilizing effect and a greater drug leakage [61]. Alginate microparticles showed diffusion controlled Higuchi kinetics, whereas alginate/pectin microparticles and vitamin B5 liposomes showed the first-order release kinetics [60]. Mucoadhesive films of SAemagnesium aluminum silicate dispersion loaded with nicotine (NCT) were prepared by adsorption of nicotine on magnesium aluminum silicate. The drug polymer dispersion prepared at pH 5 had the greatest nicotine concentration (48.4  2.1%). The particle size was 13.5  0.1 mm and zeta potential was 70.0  3.6 mV. In aqueous medium, the layered structure of magnesium aluminum silicate (MAS) gets separated into a weakly positively charged edges and negatively charged faces. At neutral and acidic pH, protonated NCT interacts with the negatively charged MAS and/or SA via an electrostatic force, leading to the formation of NCTeMAS flocculates. These flocculates that function as microreservoirs within the film causing decrease in the nicotine evaporation during film drying help sustain in vitro NCT release and permeation across the porcine esophageal mucosa. The drug release rate and permeation from these films was much slower than NCT-loaded SA film because of the difficulty in exchange of NCT molecules within MAS layers with cations in the medium. A matrix diffusionecontrolled mechanism was stated for in vitro release and permeation. A change in film crystallinity and thermal properties was observed for films prepared at different pH (i.e., 5, 7, and 10). Mucoadhesivity of these films was lower than nicotine-loaded SA films, yet it was sufficient for use in buccal delivery [62]. Moreover, in a separate study, alginate beads containing propranolol HCl intercalated with MAS showed greater drug entrapment efficiency, reduced burst release of propranolol HCl, and could modify the drug release when compared with propranolol-only alginate beads. Additionally, an increase in MAS quantity resulted in higher EE, thermal stability, and mechanical strength of the beads [63]. In yet another study, SA and cellulose nanocrystals derived from banana stem were used to develop alginateecellulose nanocrystal hybrid nanoparticles for rifampicin delivery. A slightly modified ionotropic gelation method was used to produce nanoparticles, and honey was used as a surfactant/stabilizing agent in the procedure. The optimized formulation had a particle size of w70 nm, EE of 69.73%, and a negative zeta potential of 20. The nanoparticles showed 15% release of rifampicin at pH 1.2 followed by sustained release of >90% at neutral and alkaline conditions (pH 6.8 followed by pH 7.4) up to 12 h [64].

Sodium alginate in drug delivery and biomedical areas 77 Microencapsulation of bioactive compounds helps in maintaining product stability, efficiency, and ease of usage as ingredients in a variety of food and beverages. It also helps to eliminate undesirable effects, while at the same time provide desired characteristics to the material [65]. Vitamin C was immobilized in hydrated zinc oxide layers by coprecipitation method followed by encapsulation within SA beads. These alginate beads showed sustained release (w90% in 6 h) of vitamin C, better storage stability (>95% after 4 weeks), and comparable biological activity for neat vitamin C [66]. Microencapsulation of polyphenolic extracts from medicinal plants, namely, nettle (Urtica dioica L.), hawthorn (Crataegus laevigata), raspberry leaf (Rubus idaeus L.), olive leaf (Olea europaea L.), yarrow (Achillea millefolium L.), and (Glechoma hederacea L.) was tried using alginateechitosan system. The authors used ascorbic acid for solubilizing chitosan and electrostatic extrusion technique to form microbeads containing the bioactive. These microbeads showed substantial polyphenol content and antioxidant activity. However, encapsulation efficiency and size of the beads depend on the elemental content of the bioactive. Nonetheless, these microbeads denature during storage under refrigerated condition within a month because of ascorbic acid degradation [67]. SA-grafted polyglycidyl methacrylate hydrogels (PGMA-g-SA) showed pH-sensitive release of vitamin B2 for up to 4 days in simulated intestinal fluid (pH 7.5) and almost 3 days in simulated gastric fluid (SGF pH 1.2). The scheme for graft preparation is illustrated in Fig. 3.8. The graft hydrogel (5 mm cube size) showed higher encapsulation efficiency (87.4%e99.7%) than vitamin B2eloaded calcium alginate beads (diameter, 1.4  0.05 mm). This was due to high bead porosity and its method of preparation. A striking difference in drug release was observed between the two formulations with the calcium alginate bead showing 70% vitamin B2 release in SGF and 100% vitamin B2 release in SIF after 24 h as against 18% vitamin B2 release in SGF and 30% release in SIF shown by the hydrogel in the same duration [68]. Encapsulation of vitamin B2 in chitosan/ alginate nanoparticles (diameter, 104.0  67.2 nm, PDI 0.319  0.068) helped maintain the vitamins stability up to at least 5 months. Vitamin B2 is positively charged in low pH solution, and its inclusion within the nanoparticles led to a reduction in zeta potential from 30.9  0.5 to 29.6  0.1 mV. These nanoparticles showed EE of almost 55% and loading content of 2% [69].

4.2 Injectable delivery Nanoparticle-based delivery of drugs and diagnostics in cancer therapy has proven to be superior for antitumor efficacy, reduced systemic toxicity, and undesirable side effects [70,71]. Methylene blueeloaded AOT and SA nanoparticles prepared by a multipleemulsion cross-linking process showed enhanced photodynamic efficacy. The encapsulated

78 Chapter 3

Figure 3.8 Grafting reaction mechanism of the GMA onto SA. Reprinted from El-Ghaffar MAA, Hashem MS, El-Awady MK, Rabie AM. pH-sensitive sodium alginate hydrogels for riboflavin controlled release. Carbohydr Polym 2012;89:667e75, Copyright © 2012 with permission from Elsevier.

methylene blue here acted as a photosensitizer. A photosensitizer generates cytotoxic singlet oxygen species (1O2) and reactive oxygen species (ROS) when exposed to light of a particular wavelength and hence can selectively kill tumor cells using photodynamic therapy. The nanoparticle preparation process consisted of emulsifying an aqueous solution of methylene blue and SA in AOT solution with methylene chloride. The w/o emulsion was further emulsified in polyvinyl alcohol solution, and aqueous calcium chloride solution was added to cross-link the formed methylene chloride that was evaporated to obtain nanoparticles. The nanoparticles were hypothesized to have a calcium-cross-linked alginate inner core structure with AOT head groups surrounded by one or more layers of

Sodium alginate in drug delivery and biomedical areas 79 AOT tails. Majority of encapsulated methylene blue was expected to be within the core with some amount on the particle surface as well. Furthermore, cell line studies carried out on two cancer cell lines, MCF-7 and 4T1, indicated higher accumulation of the encapsulated methylene blue mostly in the nucleus and resulted in an increase of production of reactive oxygen species [72]. Another approach for developing surface-modified nanoparticle is the layer-by-layer method of oppositely charged polymer immobilization on the surface of base via electrostatic interactions to form functional multilayers. This technique was used by Lei et al. [73] to develop nanocarrier of functionalized graphene oxide with chitosan and SA (GO-CS/SA). The modification and preparation process increased the solubility of graphene oxide and the nanocomposites. The nanocomposites were taken up internally by MCF-7 cells, and encapsulated doxorubicin showed excellent cytotoxicity to tumor cells. These nanocomposites showed 70% doxorubicin loading. On the other hand, nanocomposites of graphene oxide, protamine sulfate, and SA prepared by the same process showed higher doxorubicin loading capacity of 130% [74]. The nanocomposites showed good dispersibility and stability in physiological pH. In vitro doxorubicin release of 32% and 52% was observed in case of GO-CS/SA nanocomposites in 5 days at pH 7.4 and pH 5.0, respectively. In contrast to this, GO-PRM/SA nanocomposites showed respective doxorubicin release of 27% and 50% in 7 days at the same pH values [74]. In both the studies, nonspecific protein adhesion was reduced. The nonspecific adsorption of proteins by nanocarriers leads to internalization of nanocarriers by macrophages, resulting in inflammatory reactions. Studies carried out using BSA as a model protein for up to 24 h showed adsorption value of 15% w/w for GO-CS/SA nanocomposites compared with 152% for free GO. On the other hand, GO-PRM/SA nanocomposites showed BSA adsorption of 46% and 8% compared with 141% w/w for free GO at the end of 24 h [73,74]. Artemisinin nanocapsules coated with multilayers of chitosan/alginate (CS/ALG)6 or gelatin/chitosan (GEL/ALG)4 prepared by layer-by-layer technique showed good hydrophilicity and dispersibility in aqueous solution. The encapsulation made the artemisinin crystal hydrophilic, and increased polymer layers caused an increase in drug release [75]. The pH-triggered release of doxorubicin was observed from single-wall carbon nanotubes (SWCNTs) that were derivatized with carboxylate groups and coated with SA and chitosan. Both electrostatic and pep interactions helped in the attachment of anionic alginate and cationic chitosan onto SWCNT surface. Folic acid was attached to polysaccharide-modified SWCNTs by carbodiimide linkage, and furthermore, doxorubicin could bind at the physiological pH (pH 7.4) because of pep as well as electrostatic interactions with doxorubicin loading efficiency being highest in case of ALG-SWCNTs and lowest in CS-SWCNTs. Fig. 3.9 shows the mechanism of SWCNT modification with

Figure 3.9 Preparation of modified SWCNTs. (A) Modification of SWCNTs (derivatized with eCO2H groups) with ALG, CHI and DOX, (B) UV-Vis absorption spectra of DOX and DOX loaded SWCNTs TEM images of modified SWCNTs. (C) Cut SWCNTs, (D) ALG-SWCNTs, (E) CHI-SWCNTs, (F) CHI/ALGSWCNTs, (G) DOX-SWCNTs, (H) DOX-ALG-SWCNTs, (I) DOX-CHI-SWCNTs and (J) DOX-CHI/ALGSWCNTs, fluorescence images of cells incubated with (K) DOX-FA-CHI/ALG-SWCNTs (20 mg/mL), (L) DOX-CHI/ALG-SWCNTs (20 mg/mL), (M) FA for 2 h followed by DOX-FA-CHI/ALG-SWCNTs (20 mg/ mL), and (N) free DOX (50 mg/mL) at 37 C for 1 h. Reprinted from Zhang X, Meng L, Lu Q, Fei Z, Dyson PL. Targeted delivery and controlled release of doxorubicin to cancer cells using modified single wall carbon nanotubes. Biomaterials 2009;30:6041e7, Copyright © 2009 with permission from Elsevier.

Sodium alginate in drug delivery and biomedical areas 81 alginate, chitosan, and doxorubicin. The Dox-FA-CS/ALG-SWCNT has shown to be highly cytotoxic at the low dose administered at which even pure drug and unloaded nanotubes did not have appreciable toxicity. They showed excellent stability under physiological conditions and selectively delivered DOX into lysosomes (acidic environment) of human cervical carcinoma HeLa cells with quite greater efficiency than free DOX [76].

4.3 Ocular delivery Biodegradable ocular inserts (3 mm diameter and 0.12e0.63 mm thickness) containing brimonidine tartrate were prepared from low-molecular-weight SA. These showed sustained drug release of 99% brimonidine in 6 h. Furthermore, one-sided or both-sided coating of ethylcellulose or Eudragit RSPO was given to effectively sustain brimonidine release. Studies indicate that ocular inserts composed of 7% PVP K-90, 1.5% lowmolecular-weight SA, 5% propylene glycol, and ethylcellulose coating showed the best results with better pharmacodynamics activity, good tolerability, and decrease in intraocular pressure compared with brimonidine eye drops [77]. Mucoadhesive nanoparticles of thiolated chitosan and SA were formed as a result of interaction between the negatively charged groups of SA and the positively charged amino groups of thiolated CS. Cellular uptake studies carried out on human corneal epithelium indicated higher drug transport via these nanoparticles. This was further substantiated by in vivo experiments on rat cornea [78].

4.4 Nasal delivery Nasal delivery of drugs is convenient, painless, and noninvasive and they avoid the hepatic first-pass metabolism [79]. This route has the potential to be used as an alternative to injectable delivery by usage of suitable bioadhesive polymers and nontoxic absorption enhancers [80]. Drugs that are prone to degradation in GIT or those that are rapidly metabolized by the first-pass effect can be delivered by nasal route. A wide variety of drugs ranging from highly lipophilic to hydrophilic compounds such as proteins and peptides have been investigated by nasal route of delivery. The nasal cavity is highly vascularized and has large surface area, which permits better drug absorption either for local or for systemic use [28]. Direct nose to brain delivery of venlaflaxin using venlaflaxin-loaded alginate nanoparticles in albino Wistar rats showed higher levels of venlaflaxin in brain with greater brain:blood ratio of the nanoparticles compared with intranasal and intravenous venlaflaxin solution [81]. Mucoadhesive alginate microspheres were developed and optimized for nasal delivery of carvedilol, a nonselective b-adrenergic antagonist that showed extensive hepatic first-pass effect. The microspheres produced were in the size range of 27e54 mm with an

82 Chapter 3 encapsulation efficiency of 37%e57% [28]. The CS-TPP-ALG nanoparticles were formed by ionic gelation of CS hydrochloride with pentasodium tripolyphosphate (TPP) and simultaneous electrostatic complexation of opposite charges of alginate and CS. The nanoparticles were loaded with insulin by directly dissolving the peptide in TPP solution before nanoparticle formation. An increase in nanoparticle size was observed with the addition of alginate of molecular weight 4e74 kDa. The molecular weight of alginate had no effect on association efficiency of insulin, which ranged from w41% to w52%. Moreover, these nanoparticles showed hypoglycemic effect in in vivo studies on rabbits; the authors claimed that duration of the hypoglycemic response was affected by molecular weight of alginate used [82].

4.5 Tackling obesity and weight management A number of studies suggest that intake of SA along with calcium salt (tri- or dicalcium phosphate) either as a beverage or as a nutritional bar resulted in attenuation of postprandial glycemic response. This effect was claimed to be due to viscosification of gastric contents that resulted in decreased nutrient absorption and gastric emptying [83,84]. A randomized controlled two-way crossover study was carried out on 68 healthy volunteers (both males and females) by Paxman et al. [10], demonstrating the effect of daily ingestion of SA in reducing the energy intake. A significantly reduced energy intake was observed in the treatment group that consumed a vanilla-flavored 100 mL reconstituted beverage for 7 days prebreakfast or preevening food majorly containing 1.5 g SA (65%e75% guluronate content) along with calcium carbonate, glucono-delta-lactone apart from other taste, flavor, and palatability improving ingredients. The control group received SlimFast Simply Vanilla Milkhake Powder (Unilever, UK) to be reconstituted to 100 mL with water. In water, gluconic acid was formed due to hydrolysis of glucono-d-lactone that caused a controlled pH reduction and allowed the dissolution of calcium carbonate in a controlled manner and hence, the controlled gelation rate. This enhanced the intragastric gelation in SA-based beverage that was independent of endogenous acid secretion and resulted in a strong gel (around 30 N as per in vitro measurements). About 174 kcal reduction in energy intake, independent of gender, was observed for the treatment group compared with control group, indicating its potential in weight management [10].

4.6 Wound dressings There has been a growing interest in alginate-based materials for the production of wound dressing materials, specifically the so-called “moist healing” products such as gels, foams, and fibrous nonwoven dressing materials. The hydrophilic property of SA under normal physiological conditions in addition to its biocompatible and biodegradable nature enables

Sodium alginate in drug delivery and biomedical areas 83 the creation of physiologically moist local environment by interaction of the polymer with the wound surface [6,85]. This encouraged the formation of granulation tissues and caused healing. The migration of epithelial cells from wound periphery to the wound is faster in moist conditions in contrast to dry state. However, wet conditions at wound surface are not preferred. The unique property of high water absorbency makes alginates the material of choice for preparation of highly absorbent wound dressings. Once applied on the wound, it absorbs the wound exudate and forms a gelatinous coat on the wound surface, thus keeping the wound moist, thereby limiting the bacterial activity. This makes them useful for highly exuding wounds. Calcium alginate fiber can also act as hemostatic agent where Ca2þ ions of the fiber are exchanged with Na2þ of the body fluid at the wound site [6]. Curcumin-loaded cross-linked CS and SA sponge were prepared by lyophilization method. Sponge samples with 1:1 ratio of CS and alginate showed sustained curcumin release with 50% curcumin being released in 20 days. A decrease in % swelling was observed by decreasing chitosan quantity. Better wound healing effect was observed for these sponges (either with or without curcumin) in comparison with normal gauze-treated group. However, no significant change in wound defect area was observed between the drugloaded or drug-unloaded sponge. Nonetheless, better collagen arrangement and increased collagen content were observed in curcumin-loaded sponges [85]. Lyophilization technique was also used to produce porous wafers of SA by Boateng et al. for use on wound surface [86]. Polymers used for topical wound healing require both swelling capacity to absorb exudate and permeability to oxygen [87,88]. A better physiological environment for wound epithelialization was observed in case of PVANaAlg composite nanofiber wound dressing compared with PVA nanofiber patch in in vivo studies. The greater the hydroxyproline content of the wound, the greater the rate of wound healing [89]. Alginate has been used in a number of commercially available wound dressing materials. Some of these are Algicell (Derma Sciences), AlgiSite M (Smith and Nephew), Comfeel Plus (Coloplast), Kaltostat (ConvaTec), Sorbsan (UDL Laboratories), Tegagen (3M Healthcare), etc.

4.7 Cell delivery and implants SA hydrogels have numerous applications in the field of medicine such as diagnostics, tissue engineering and regenerative medicine, cell immobilization, etc. Apart from their systemic use, they can be used as depot systems, which slowly release the drug load, thereby maintaining a high drug concentration in the surrounding tissues for a prolonged time [90]. The success of a hydrogel implant system depends in part on the stability of polymer and its ability to resist degradation after implantation. The fact that SA forms hydrogel under mild conditions that very well maintain the cell viability and protein activity makes it suitable for encapsulating such biological moieties.

84 Chapter 3 Cells can be immobilized within the alginate network by suspending them in alginate solution, which is further cross-linked in the presence of suitable cations to form hydrogel network [91]. However, to be useful as an implant, alginate-based systems should have sufficient mechanical strength so that they can remain in place for a long time. Insufficient mechanical integrity of alginate capsule has been linked with immunological reactions [92]. Alginate microbeads were used to deliver islet cells for the treatment of type I diabetes [93]. To provide necessary mechanical strength, dual cross-linked microbeads of alginate were prepared. Methacrylation of SA using 2-aminoethyl methacrylate hydrochloride helped to introduce groups on exposure to a photoinitiator (0.05% (w/v), Irgacure 1173) that helped for covalent cross-linking in addition to cross-linking with CaCl2 [92]. The synthetic scheme is shown in Fig. 3.10. These beads were stable under inflammatory challenge created by local injection of 100 mL mg/mL of lipopolysaccharide (50 mg/mL). These beads were stable in rat’s omentumpounch for 3 weeks in contrast to unmodified alginate beads that remained stable hardly for 1 week. Calcium alginate microcapsules containing immobilized human embryonic stem cells (hESCs) could cause the differentiation of hESCs into definitive endoderm in 3D [94]. Cyropreserved islet cells within chitosanealginate microcapsules maintained exceptional cell viability with 95.4  1.3% cell recovery postthawing that was comparable with freshly isolated islet cells (97.5  0.8%). The unencapsulated cells showed only 69.4  3.5% cell viability. These microcapsules supported islet cell membrane, reduced peripheral cell loss, and maintained islet cell integrity. These cells were morphologically and functionally intact postthawing and showed significant rise in basal as well as glucose-stimulated insulin release comparable with freshly isolated islets [95].

4.8 Protein delivery Previous studies have already shown that coated or uncoated alginate beads and microcapsules can be used for delivery of proteins such as heparin, hemoglobin, melatonin, etc. Moreover, coated beads and microspheres were considered to be better for oral administration [13]. Approximately 350 nm nanoparticles of SA were prepared by interfacial cross-linking method using microemulsion-based reactor. Microemulsion of SA, DOSS, and IPM was mixed with microemulsion of aqueous calcium chloride, DOSS, and IPM, which led to spontaneous nanoparticle production. These nanoparticles showed 40% BSA loading efficiency with a burst release followed by sustained release of BSA up to 16 h [96]. Injectable IPN based on methacrylated alginate showed promising results for sustained protein delivery. The hydrogel was formed as a result of copolymerization of methacrylated alginate, poly(ethylene glycol) methacrylate, and N-isopropylacrylamide at 37 C. The N,N-methylene bis(acrylamide) was used as a cross-linker, whereas ammonium

Sodium alginate in drug delivery and biomedical areas 85

Figure 3.10 Synthetic scheme for methacrylation of alginate and photocross-linking of methacrylated alginate. Reprinted from Somo SI, Langert K, Yang C-Y, Vaicik MK, Ibarra V, Appel AA, et al. Synthesis and evaluation of dual crosslinked alginate microbeads. Acta Biomater 2018;65:53e65, Copyright © 2018 with permission from Elsevier.

86 Chapter 3 persulfate and N,N,N,N-tetramethylethylenediamine were used as redox initiators. Synthetic scheme is shown in Fig. 3.11. In vitro release showed AA, they are called proteins. Owing to their natural existence in the human body, both proteins and peptides have low toxicity, well-known pharmacology (distribution, metabolization, and elimination), and biological abundance, generally being usually biologically safe [12]. Overall, peptides are designed and synthesized to bind and modulate proteins and their interactions involved with the carcinogenic process, and proteins are commonly involved in the immune system as antibodies, cytokines, or interferons [101]. Recently, Marqus et al. [101] reviewed therapeutic peptides for cancer treatment and classified them into four categories according to their mechanism of action: targeting signal transduction pathways, targeting the cell cycle inducing cell death, targeting tumor suppressor protein, and targeting transcription factors. Peptides have high specificity of targeting when compared with proteins, as well as easy cell penetration and interaction with cell receptors, and also are more accessible to synthesize (Fig. 19.1). Some peptides are already available to cancer therapy clinical use [102]. To enumerate, somatostatin and its analogs as growth inhibitors [103], endostatin as an antiangiogenic drug [104], and well-known proteins as monoclonal antibodies [105] are used in cancer therapeutics, leading to cell apoptosis through direct tumor cell killing, targeting and delivering drug into the cells, or recruiting the immune system. For example, Herceptin, a widely used drug that targets HER-2 receptors overexpressed in some types of breast cancer [106]; Rituxan, a humanized antibody is effective against non-Hodgkin lymphoma once it targets the B-cell-specific antigen CD20 [107]; L-asparaginase, an important component of multiagent CT schemes for the treatment of acute lymphoblastic leukemia,

Natural polysaccharides 457

Figure 19.1 Mechanism of therapeutic peptides for cancer treatment. According to Marqus et al. [101].

is an enzyme that prevents tumor development by breaking down asparagine, a peptide that specific cancer cells require in higher amounts when compared to healthy cells [108]. Recently, derivatives of the human immunodeficiency virus (HIV) trans-activator of transcription protein (TAT) were patented for use as an anticancer targeting agent (US 207/0274,040 A1), as well as some recombinant cytokines in combined treatments to amplify the NK-mediated antitumor response [109]. In spite of all those benefits and already proved activity, the use of pep/pro in cancer therapeutics remains a challenge due to some drawbacks summarized in Table 19.3. Macromolecules as proteins display low stability in the organismdthis instability is majorly related to the presence of various functional fractions susceptible to chemical degradation and their high hydrophilic character [110]. Given these points, their adequate delivery to target tissues has been considerably limited in vivo because these molecules can hardly pass through various hydrophobic biological barriers, and on the other hand, peptides are more manageable to synthesize and have better cell penetrability. In the past decades, nanosized systems have been studied in function of improving pep/pro therapeutics efficacy in reaching the disease site exploiting the EPR characteristic of pathological angiogenic vasculature in cancer [6,12,19,104,110]. To summarize, studies

Table 19.3: Peptide and proteins use drawbacks in cancer therapeutics. Drawbacks Chemical degradation in the organism owing to enzymatic degradation and pH instability [101] Accumulation in nontargeted organs and tissues [102] Rapid elimination owing to renal clearance [102] Production and manufacturing challenges [101] Low permeability of cell membranes [102] Poor oral bioavailability in function of gastric proteases degradation [101]

458 Chapter 19 involving protein delivery by a PSC-derived matrix tested in cancer cell lines in vitro and in vivo are exposed in Table 19.4. Bovine lactoferrin (bLF) is a well-known protein for its anticancer and antiinflammatory properties [115]. The effects of lactoferrin-loaded NPs on breast cancer [111] and colon cancer [115] were evaluated in alginate-enclosed CSecalcium phosphate iron-loaded bovine lactoferrin nanocapsules (AE-CS-CP-Fe-bLf), which proved to enhance bioavailability and activity of this protein, with gastric protection by the alginate gel encapsulation and nanosizing with the CS. AE-CS-CP-Fe-bLf showed better blood half-life and increased antitumor activity against MDA-MB-231 cells in vitro and in vivo, orally administered. Additionally, it not only killed cancer cells but also downregulated cancer stem cells [111]. Another study demonstrated AE-CS-CP-Fe-bLf activity against Caco-2 colon cancer cells and cancer stem cells, in vitro and in vivo, with a remarkable reduction in angiogenesis markers [115]. Both studies cited above concluded that when the NPs were uptaken, they modified the expression of specific miRNAs which intensified their uptake by the cells and improved their effectiveness, with an increase in body iron and calcium levels. This feature can be valuable for cancer patients and perform as a supporting therapy, lowering conventional therapy doses [111,115]. Moreover, some pep/pro complexed with polysaccharides as nanocarriers were evaluated, aiming to improve cell targeting [114,116e118]. Antoniraj et al. [117] applied atrial natriuretic peptide (ANP, a cell-specific ligand) conjugated with CS-hydrazone-methoxy PEG copolymer for intracellular delivery of prednisone. It could be observed that conjugation with ANP enhanced the cellular uptake of the polymeric NPs by the A549 cells, with selective delivery based on the pH cleavability of the polymer in function of the acid-cleavable hydrazine linkage. On a complex strategy, Chen et al. [114] applied the dual-targeted concept to kill SKOV-3 ovarian cancer cells and MDA-MB-231 cells with granzyme B (GrB)-loaded and peptide-targeted nanogel with an HA matrix and GE-11 outer signalization. GrB is a protease secreted by NK cells and T cytotoxic cells to eliminate infected or cancerous cells, and it has multiple uses in anticancer therapy. To add, HA has a known property to target CD44þ cells [119], and GE-11 peptide has the ability to connect EGFRþ cells. These two combined enhanced cellular uptake of the complex and cytoplasmic release of GrB henceforth stimulated caspase cascade activation and cell death by apoptosis. The authors tested both in vitro and in vivo separately cytochrome C (CC) and GrB loading in nanogel, but GrB displayed better antitumor activity probably in function of overexpression of Bax and gtBid proteins, which combined together can generate CC and therefore improve cancer therapy.

Table 19.4: Polysaccharide polymers encapsulating proteins/peptides tested for anticancer activity in vitro and in vivo. PSC Alginate and chitosan

System Nanocapsules

Pep/Pro Fe-bLf

Structure PDS AEC-CP-Fe-bLf

Evaluation model MDA-MB-231 cells and C57 Balb/C xenograft mice

Caco-2 cells and xenograft mice

Nanoparticles complex

TRAIL

CMCS-FA-PEI-BSATRAIL-GA

MCF-7 (FRþ) and A549 (FR-) cells and Balb/C mice

Hyaluronic acid

Nanogel

GrB

HA-GE11-GrB

SKOV-3 and mda-mb231 cells and xenograft nude mice

Active orally as prevention, treatment after tumor growth (4.8-fold decrease in tumor size), and inhibited tumor recurrence Ability to induce apoptosis in cancer cells and cancer stem cells, when given orally in diet pH-dependent and surface chargeswitchable NPs loading GA and TRAIL achieved a precise release in specific sites, which resulted in improved anticancer efficacy and reduced undesirable side effects Tumor cell death by apoptosis owing to CD44 and EGFR-specific internalization of granzyme B and improved anticancer activity than cytochrome C

References [111]

[112]

[113]

[114]

Natural polysaccharides 459

Chitosan

Outcome

460 Chapter 19 Recently, Ding et al. [120] have exploited HA-based nanogel activity against overexpressing MCF-7 cells in vitro with a pH-dependant coiled-coil E3 and K3 peptide cross-linked structure loading saporin as a therapeutic protein model or CC as a protein release model. The peptides provided an a-helicoidal configuration that enabled better structural stability in blood with neutral conditions and caused unfolding in acidic pH-dependent manner, which in fusion with endosomal membrane helped proteins to escape from endosomal entrapment. Saporin, a highly potent protein, which is able to inactivate ribosomes and nonpenetrable into the cell membrane, has demonstrated better antitumor efficacy than CC, confirming the formulation as a good option for cationic protein delivery. Another approach for pep/pro use in polymeric nanocarriers is to improve the system stability [12,121,122] carrying proteins or chemicals. Zhang et al. [113] developed TRAIL and GA coloaded BSA-PEI NPs with CMCS-FA-based outer shell encapsulating the NPs. They tested this NP complex in MCF-7 (FRþ) and A549 (FR-) cells analyzing the synergistic mechanism of action of TRAIL interacting with cell membrane receptors, leading to a caspase enzyme cascade, and GA being released intracellular via proton sponge effect reaching the nucleus, both inducing cell death and apoptosis. The CS-derived enabled targeted release of NPs because of their pH-responsive behavior, considering that tumor tissues are more acidic than healthy tissues. Moreover, Liu et al. [123] explored the well-known self-assembly property of CMC and BSA, focusing on the effective radionuclide 131I and the CT drug camptothecin codelivery, to achieve combined chemoradioisotope synergistic therapy of cancer. Surprisingly, the CMCeBSA complex presented a pH-dependent drug release profile and high drug loading capacity, and following LLC cells, the combined therapy was significantly superior to single therapy. Protein delivery by PDS matrix is also being evaluated as a model vaccine delivery platform, using the protein antigen for presentation and the polysaccharide polymer, providing low cytotoxicity, good antigen-loading capacity, and targeted and sustained release. Its mechanism can improve cell or humoral immunity, through induced T cell proliferation and cytokine secretion by a pH-dependent release [124] and MHC I/II response, promoting intracellular processing of the antigen based on polymer bioreduction [125]. Furthermore, the P-PDS combination can make up theragnostic (tumor targeting and magnetic resonance imaging) applications in brain tumor 9L-glioma cells when complexed with heparin-coated magnetic NPs. In detail, the protein can perform as a helper for tissue targeting as B-galactosidase [126] and even as a cationic model to prove the binding capacity of the heparin-functionalized complex, as protamine [127].

Natural polysaccharides 461 Some authors studied the liberation kinetics of pep/pro encapsulated or complexed in nanocarriers [128] based in carrageenan/CS and BSA [129], starch and BMP-4 [130], mannan and bFGF [131], glucomannan/CS and bFGF [132], fucoidan/CS and BSA [133], pullulan and BoHc/A [134], heparin and bFGF [135]. Despite the promising protein delivery results, in vitro or in vivo antitumor evaluations were not conducted; therefore, they need more deep and straightforward studies to be considered.

4. Clinical trials of PDS Although several PDSs have been studied to develop an effective anticancer treatment, only a small amount of studies reached clinical trials (Table 19.5). Overall, the reasons that can explain this issue are mostly given to the impurity of some developed products that can cause systemic toxicity. Therefore, future studies using PDS should include systemic delivery evaluations [7]. In addition, codeliveries and smart designed PDS seem to be the most promising approaches in this field of research.

Table 19.5: Clinical trials with PDS. Polysaccharide Dextran

Product DE-310

Formulation

Chitosan

Milican

Exatecan mesylate, carboxymethyl dextran Camptothecin (T-2513), carboxymethyl dextran Holmium-166, CS

Hyaluronic acid Cyclodextrin

ONCOFID-R-B

PTX and HA

CALAA-01

siRNA (RRM2), cisplatin, AD-PEG, hTf b-Cyclodextrin and PEG copolymere camptothecin

Delimotecan (MEN 4901T/0128)

CRLX101/ IT-101

Cancer type

Development stage/phase References

Advanced solid tumors

I

[136]

Solid tumors

I

[137,138]

Small hepatocellular carcinoma Bladder cancer

II

[139]

I, II

[140]

Solid tumors

I

[141]

Ovarian/tubal/ peritoneal cancer Rectal cancer Advanced solid tumors Lung cancer

I, II

[142]

I, II I

[143] [144]

I, II

[145]

462 Chapter 19

Figure 19.2 A schematic representation of polysaccharides-based drug delivery systems and their biofunctionalization.

5. Conclusion There are several strategies being developed to improve therapeutics biological stability, blood half-life, and specificity to target cancer cells, with attention given to incorporation and targeting with polymers. Polysaccharide polymers represent a promising approach to promote targeted delivery and overcome these molecular deficiencies because of low toxicity, natural abundance, and biodegradability. In addition, PDSs exhibit the ability to mimic the natural extracellular matrix, self-assembling properties, and also easy ionic manipulation, which allows various possibilities of drug targeting and release (Fig. 19.2). However, further studies are necessary to improve production methods, considering synthesis issues such as stability, product variability, and impurity. Additionally, more immunogenic evaluations are needed to prove that these bioproducts are safe for systematic use.

Natural polysaccharides 463

References [1] Ferlay J, Soerjomataram I, Dikshit R, Eser S, Mathers C, Rebelo M, Parkin DM, Forman D, Bray F. Cancer incidence and mortality worldwide: sources, methods and major patterns in GLOBOCAN 2012. Int J Cancer March 2015;136(5):E359e86. https://doi.org/10.1002/ijc.29210. [2] National Institutes of Health (US). Biological sciences curriculum study. In: NIH curriculum supplement series. Bethesda (MD): National Institutes of Health (US); 2007. Available from: https://www.ncbi.nlm. nih.gov/books/NBK20364/. [3] Masood F. Polymeric nanoparticles for targeted drug delivery system for cancer therapy. Mater Sci Eng C Mater Biol Appl March 2016;60:569e78. https://doi.org/10.1016/j.msec.2015.11.067. Epub 2015 Nov 27. [4] Aftab S, Shah A, Nadhman A, Kurbanoglu S, Aysıl Ozkan S, Dionysiou DD, et al. Nanomedicine: an effective tool in cancer therapy. Int J Pharm 2018;540(1e2):132e49. https://doi.org/10.1016/ j.ijpharm.2018.02.007. [5] Cho K, Wang X, Nie S, Chen ZG, Shin DM. Therapeutic nanoparticles for drug delivery in cancer. Clin Cancer Res 2008;14(5):1310e6. https://doi.org/10.1158/1078-0432.CCR-07-1441. [6] Ranjbari J, Mokhtarzadeh A, Alibakhshi A, Tabarzad M, Hejazi M, Ramezani M. Anti-cancer drug delivery using carbohydrate-based polymers. Curr Pharmaceut Des 2018;23(39):6019e32. https://doi.org/ 10.2174/1381612823666170505124927. [7] Miao T, Wang J, Zeng Y, Liu G, Chen X. Polysaccharide-based controlled release systems for therapeutics delivery and tissue engineering: from bench to bedside. Adv Sci 2018;5(4): 1700513. [8] Dumitriu S. Polysaccharides: structural diversity and functional versatility. 2nd ed. USA: CRC Press; 2004. [9] Lee JW, Park JH, Robin JR. Bioadhesive-based dosage forms: the next generation. J Pharm Sci 2000;89(7):850e66. [10] Jung B, Shim MK, Park MJ, Jang EH, Yoon HY, Kim K, et al. Hydrophobically modified polysaccharide-based on polysialic acid nanoparticles as carriers for anticancer drugs. Int J Pharm 2017;520(1):111e8. [11] Sithole MN, Choonara YE, du Toit LC, Kumar P, Pillay V. A review of semi-synthetic biopolymer complexes: modified polysaccharide nano-carriers for enhancement of oral drug bioavailability. Pharmaceut Dev Technol 2017;22(2):283e95. [12] Zhang L, Pan J, Dong S, Li Z. The application of polysaccharide-based nanogels in peptides/proteins and anticancer drugs delivery. J Drug Target 2017;25:673e84. [13] Gomes B, Moreira I, Rocha S, Coelho M, Pereira MDC. Polysaccharide-based nanoparticles for cancer therapy. J Nanopharm Drug Deliv 2013;1:335e54. https://doi.org/10.1166/jnd.2013.1039. [14] Zhang Y, Chan JW, Moretti A, Uhrich KE. Designing polymers with sugar-based advantages for bioactive delivery applications. J Control Release 2015;2019:355e68. [15] Zhang N, Wardwell PR, Bader RA. Polysaccharide-based micelles for drug delivery. Pharmaceutics 2013;5(2):329e52. [16] Alvarez-Lorenzo C, Blanco-Fernandez B, Puga AM, Concheiro A. Crosslinked ionic polysaccharides for stimuli-sensitive drug delivery. Adv Drug Deliv Rev 2013;65(9):1148e71. [17] Huo M, Zou A, Yao C, Zhang Y, Zhou J, Wang J, et al. Somatostatin receptor-mediated tumor-targeting drug delivery using octreotide-peg-deoxycholic acid conjugate-modified n-deoxycholic acid-o, n-hydroxyethylation chitosan micelles. Biomaterials 2012;33(27):6393e407. [18] Bae YH, Park K. Targeted drug delivery to tumors: myths, reality and possibility. J Control Release 2011;153(3):198e205. [19] Posocco B, Dreussi E, de Santa J, Toffoli G, Abrami M, Musiani F, et al. Polysaccharides for the delivery of antitumor drugs. Materials 2015;8(5):2569e615. https://doi.org/10.3390/ma8052569.

464 Chapter 19 [20] Zhao Q, Han B, Wang Z, Gao C, Peng C, Shen J. Hollow chitosan-alginate multilayer microcapsules as drug delivery vehicle : doxorubicin loading and in vitro and in vivo studies. Nanomedicine 2007;3:63e74. [21] Cafaggi S, Russo E, Stefani R, Leardi R, Caviglioli G, Parodi B, et al. Preparation and evaluation of nanoparticles made of chitosan or N -trimethyl chitosan and a cisplatin e alginate complex. J Drug Deliv Sci Technol 2007;121:110e23. [22] Bisht S, Maitra A. Dextran-doxorubicin/chitosan nanoparticles for solid tumor therapy. Wiley Interdiscip Rev Nanomed Nanobiotechnol 2009;1(4):415e25. https://doi.org/10.1002/wnan.43. [23] Lee E, Lee J, Lee IH, Yu M, Kim H, Chae SY, et al. Conjugated chitosan as a novel platform for oral delivery of paclitaxel. J Med Chem 2008;51(20):6442e9. [24] Lee E, Kim H, Lee IH, Jon S. In vivo antitumor effects of chitosan-conjugated docetaxel after oral administration. J Control Release 2009;140(2):79e85. [25] Kim K, Kim JH, Park H, Kim YS, Park K, Nam H, et al. Tumor-homing multifunctional nanoparticles for cancer theragnosis: simultaneous diagnosis, drug delivery, and therapeutic monitoring. J Control Release 2010;146:219e27. [26] Chen KJ, Chiu YL, Chen YM, Ho YC, Sung HW. Intracellularly monitoring/imaging the release of doxorubicin from pH-responsive nanoparticles using Fo¨rster resonance energy transfer. Biomaterials 2011;32:2586e92. [27] Lee SJ, Koo H, Jeong H, Huh MS, Choi Y, Jeong SY, et al. Comparative study of photosensitizer loaded and conjugated glycol chitosan nanoparticles for cancer therapy. J Control Release 2011;152:21e9. [28] Liang Y, Zhao X, Ma PX, Guo B, Du Y, Han X. pH-responsive injectable hydrogels with mucosal adhesiveness based on chitosan-grafted-dihydrocaffeic acid and oxidized pullulan for localized drug delivery. J Colloid Interface Sci 2019;536:224e34. Available from: https://doi.org/10.1016/j.jcis.2018.10. 056. [29] Abazari R, Mahjoub AR, Ataei F, Morsali A, Carpenter-Warren CL, Mehdizadeh K, et al. Chitosan immobilization on bio-MOF nanostructures: a biocompatible pH-responsive nanocarrier for doxorubicin release on MCF-7 cell lines of human breast cancer. Inorg Chem 2018;57(21):13364e79. https://doi.org/ 10.1021/acs.inorgchem.8b01955. [30] Sahu P, Kashaw SK, Sau S, Kushwah V, Jain S. Colloids and surfaces B: biointerfaces pH responsive 5fluorouracil loaded biocompatible nanogels for topical chemotherapy of aggressive melanoma. Colloids Surfaces B Biointerfaces 2019;174:232e45. [31] Chen J, Yang X, Huang L, Lai H, Gan C, Luo X. Development of dual-drug-loaded stealth nanocarriers for targeted and synergistic anti-lung cancer efficacy. Drug Deliv 2018;25(1):1932e42. https://doi.org/ 10.1080/10717544.2018.1477856. [32] Zhang X, Li L, Liu Q, Wang Y, Yang J, Qiu T, Zhou G. Co-delivery of rose bengal and doxorubicin nanoparticles for combination photodynamic and chemo-therapy. J Biomed Nanotechnol 2019;15(1):184e95. https://doi.org/10.1166/jbn.2019.2674. [33] Choi KY, Chung H, Min KH, Yoon HY, Kim K, Park JH, et al. Self-assembled hyaluronic acid nanoparticles for active tumor targeting. Biomaterials 2010;31(1):106e14. [34] Jeong YI, Kim ST, Jin SG, Ryu HH, Jin YH, Jung TY, et al. Cisplatin-incorporated hyaluronic acid nanoparticles based on ion-complex formation. J Pharm Sci 2008;97:1268e76. [35] Ganesh S, Iyer AK, Gattacceca F, Morrissey DV, Amiji MM. In vivo biodistribution of siRNA and cisplatin administered using CD44-targeted hyaluronic acid nanoparticles. J Control Release 2013;172(3):699e706. [36] Chen Z, Li Z, Lin Y, Yin M, Ren J, Qu X. Bioresponsive hyaluronic acid-capped mesoporous silica nanoparticles for targeted drug delivery. Chem Eur J 2013;19:1778e83. [37] Cai S, Thati S, Bagby TR, Diab HM, Davies NM, Cohen MS, et al. Localized doxorubicin chemotherapy with a biopolymeric nanocarrier improves survival and reduces toxicity in xenografts of human breast cancer. J Control Release 2010;146:212e8.

Natural polysaccharides 465 [38] Xie Y, Aillon KL, Cai S, Christian JM, Davies NM, Berkland CJ, et al. Pulmonary delivery of cisplatinhyaluronan conjugates via endotracheal instillation for the treatment of lung cancer. Int J Pharm 2010;392:156e63. [39] Thomas RG, Moon M, Lee S, Jeong YY. Paclitaxel loaded hyaluronic acid nanoparticles for targeted cancer therapy: in vitro and in vivo analysis. Int J Biol Macromol 2015;72:510e8. [40] Li N, Chen Y, Zhang YM, Yang Y, Su Y, Chen JT, et al. Polysaccharide-gold nanocluster supramolecular conjugates as a versatile platform for the targeted delivery of anticancer drugs. Sci Rep 2014;4:4164. https://doi.org/10.1038/srep04164. [41] Gaio E, Conte C, Esposito D, Miotto G, Quaglia F, Moret F, et al. Co-delivery of docetaxel and disulfonate tetraphenyl chlorin in one nanoparticle produces strong synergism between chemo- and photodynamic therapy in drug-sensitive and -resistant cancer cells. Mol Pharm 2018;15(10):4599e611. https://doi.org/10.1021/acs.molpharmaceut.8b00597. Epub 2018 Sep. 7. [42] Zhao J, Wan Z, Zhou C, Yang Q, Dong J, Song X, et al. Hyaluronic acid layer-by-layer (LbL) nanoparticles for synergistic chemo-phototherapy. Pharm Res 2018;35(10):196. https://doi.org/10.1007/ s11095-018-2480-8. [43] Wang Y, Yang M, Qian J, Xu W, Wang J, Hou G, et al. Sequentially self-assembled polysaccharidebased nanocomplexes for combined chemotherapy and photodynamic therapy of breast cancer. Carbohydr Polym 2019;203:203e13. [44] Luo GF, Xu XD, Zhang J, Yang J, Gong YH, Lei Q, et al. Encapsulation of an adamantane-doxorubicin prodrug in pH-responsive polysaccharide capsules for controlled release. ACS Appl Mater Interfaces 2012;4(10):5317e24. [45] Pramod PS, Shah R, Chaphekar S, Balasubramanian N, Jayakannan M. Polysaccharide nano-vesicular multidrug carriers for synergistic killing of cancer cells. Nanoscale 2014;6(20):11841e55. [46] Su H, Zhang W, Wu Y, Han X, Liu G, Jia Q, et al. Schiff base-containing dextran nanogel as pH-sensitive drug delivery system of doxorubicin: synthesis and characterization. J Biomater Appl 2018;33(2):170e81. https://doi.org/10.1177/0885328218783969. [47] Fang Y, Wang H, Dou HJ, Fan X, Fei XC, Wang L, et al. Doxorubicin-loaded dextran-based nanocarriers for highly efficient inhibition of lymphoma cell growth and synchronous reduction of cardiac toxicity. Int J Nanomed 2018;13:5673e83. [48] Lee SJ, Hong GY, Jeong YI, Kang MS, Oh JS, Song CE, et al. Paclitaxel-incorporated nanoparticles of hydrophobized polysaccharide and their antitumor activity. Int J Pharm 2012;433:121e8. [49] Wang D, Zhang S, Zhang T, Wan G, Chen B, Xiong Q, et al. Pullulan-coated phospholipid and Pluronic F68 complex nanoparticles for carrying IR780 and paclitaxel to treat hepatocellular carcinoma by combining photothermal therapy/photodynamic therapy and chemotherapy. Int J Nanomed 2017;12:8649e70. https://doi.org/10.2147/IJN.S147591. [50] Laksee S, Puthong S, Kongkavitoon P, Palaga T, Muangsin N. Facile and green synthesis of pullulan derivative-stabilized Au nanoparticles as drug carriers for enhancing anticancer activity. Carbohydr Polym 2018;198:495e508. https://doi.org/10.1016/j.carbpol.2018.06.119. [51] Metaxa AF, Efthimiadou EK, Boukos N, Fragogeorgi EA, Loudos G, Kordas G. Hollow microspheres based ondfolic acid modifieddhydroxypropyl cellulose and synthetic multi-responsive bio-copolymer for targeted cancer therapy: controlled release of daunorubicin in vitro and in vivo studies. J Colloid Interface Sci 2014;435:171e81. [52] Thipapun Plyduang LL. Carboxymethylcellulose-tetrahydrocurcumin conjugates for colon-specific delivery of a novel anti-cancer agent, 4-amino tetrahydrocurcumin. Eur J Pharm Biopharm 2014;88:351e60. [53] Wu W, Aielli M, Zhou T, Berliner A, Banerjee P, Zhou S. In-situ immobilizaton of quantum dots in polysaccharide-based nanogels for integraion of optical pH-sensing, tumor cell imaging and drug delivery. Biomaterials 2010;31:3023e31. [54] Capanema NSV, Mansur AAP, Carvalho SM, Carvalho IC, Chagas P, de Oliveira LCA, Mansur HS. Bioengineered carboxymethyl cellulose-doxorubicin prodrug hydrogels for topical chemotherapy of

466 Chapter 19

[55]

[56] [57]

[58]

[59]

[60]

[61]

[62] [63] [64] [65] [66] [67] [68] [69]

[70]

[71] [72] [73]

melanoma skin cancer. Carbohydr Polym 2018;195:401e12. https://doi.org/10.1016/ j.carbpol.2018.04.105. Asabuwa Ngwabebhoh F, Ilkar Erdagi S, Yildiz U. Pickering emulsions stabilized nanocellulosic-based nanoparticles for coumarin and curcumin nanoencapsulations: in vitro release, anticancer and antimicrobial activities. Carbohydr Polym 2018;201:317e28. https://doi.org/10.1016/ j.carbpol.2018.08.079. Zhang C, Wang W, Liu T, Wu Y, Guo H, Wang P, et al. Biomaterials doxorubicin-loaded glycyrrhetinic acid-modified alginate nanoparticles for liver tumor chemotherapy. Biomaterials 2012;33(7):2187e96. Ahn DG, Lee J, Park SY, Kwark YJ, Lee KY. Doxorubicin-loaded alginate-g-poly(Nisopropylacrylamide) micelles for cancer imaging and therapy. ACS Appl Mater Interfaces 2014;6(24):22069e77. Wang Y, Zhou J, Qiu L, Wang X, Chen L. Biomaterials Cisplatin e alginate conjugate liposomes for targeted delivery to EGFR-positive ovarian cancer cells. Biomaterials 2014;35(14):4297e309. Available from: https://doi.org/10.1016/j.biomaterials.2014.01.035. Manatunga DC, de Silva RM, de Silva KMN, Wijeratne DT, Malavige GN, Williams G. Fabrication of 6-gingerol, doxorubicin and alginate hydroxyapatite into a bio-compatible formulation: enhanced anti-proliferative effect on breast and liver cancer cells. Chem Cent J 2018;12(1):119. https://doi.org/ 10.1186/s13065-018-0482-6. Raveendran S, Poulose AC, Yoshida Y, Maekawa T, Kumar DS. Bacterial exopolysaccharide based nanoparticles for sustained drug delivery, cancer chemotherapy and bioimaging. Carbohydr Polym 2013;91:22e32. Pinhassi RI, Assaraf YG, Farber S, Stark M, Ickowicz D, Drori S, et al. Arabinogalactan-folic acid-drug conjugate for targeted delivery and target-activated release of anticancer drugs to folate receptoroverexpressing cells. Biomacromolecules 2010;11(1):294e303. Park JH, Saravanakumar G, Kim K, Kwon IC. Targeted delivery of low molecular drugs using chitosan and its derivatives. Adv Drug Deliv Rev 2010;62(1):28e41. Aruffo A, Stamenkovic I, Melnick M, Underhill C, Seed B. CD44 is the principal cell surface receptor for hyaluronate. Cell 1990;61(7):1303e13. https://doi.org/10.1016/0092-8674(90)90694-A. Lostale´-Seijo I, Montenegro J. Synthetic materials at the forefront of gene delivery. Nat Rev Chem 2018;2:258e77. Kaur K, Rath G, Chandra S, Singh R, Goyal AK. Chemotherapy with si-RNA and anti-cancer drugs. Curr Drug Deliv 2018;15(3):300e11. Pai SI, Lin YY, Macaes B, Meneshian A, Hung CF, Wu TC. Prospects of RNA interference therapy for cancer. Gene Ther 2006;13(6):464e77. Jackson AL, Linsley PS. Noise amidst the silence: off-target effects of siRNAs? Trends Genet 2004;20:521e4. Choi KY, Silvestre OF, Huang X, Hida N, Liu G, Ho DN, et al. A nanoparticle formula for delivering siRNA or miRNAs to tumor cells in cell culture and in vivo. Nat Protoc 2014;9(8):1900e15. Yang X. Cluster of differentiation 44 targeted hyaluronic acid based nanoparticles for MDR1 siRNA delivery to overcome drug resistance in ovarian cancer. Pharm Res 2015;32(6):2097e109. https://doi.org/ 10.1007/s11095-014-1602-1. Talekar M, Ouyang Q, Goldberg MS, Amiji MM. Co-silencing of PKM-2 and MDR-1 sensitizes multidrug-resistant ovarian cancer cells to paclitaxel in a murine model of ovarian cancer. Mol Canc Therapeut 2015;14(7):1521e31. Lin WJ, Lee WC. Polysaccharide-modified nanoparticles with intelligent CD44 receptor targeting ability for gene delivery. Int J Nanomed 2018;13:3989e4002. https://doi.org/10.2147/IJN.S163149. Zhao Y, Wang W, Guo S, Wang Y, Miao L, Xiong Y, et al. PolyMetfomin combines carrier and anticancer activities for in vivo siRNA delivery. Nat Commun 2016;7:11822. Kim GH, Won JE, Byeon Y, Kim MG, Wi TI, Lee JM, et al. Selective delivery of PLXDC1 small interfering RNA to endothelial cells for anti-angiogenesis tumor therapy using CD44-targeted chitosan

Natural polysaccharides 467

[74] [75]

[76]

[77]

[78]

[79]

[80]

[81]

[82]

[83]

[84] [85] [86]

[87]

[88] [89] [90] [91]

nanoparticles for epithelial ovarian cancer. Drug Deliv 2018;25(1):1394e402. https://doi.org/10.1080/ 10717544.2018.1480672. Hyun EJ, Hasan MN, Kang SH, Cho S, Lee YK. Oral siRNA delivery using dual transporting systems to efficiently treat colorectal liver metastasis. Int J Pharm 2018;555:250e8. Talekar M, Trivedi M, Shah P, Ouyang Q, Oka A, Gandham S, et al. Combination wt-p53 and MicroRNA-125b transfection in a genetically engineered lung cancer model using dual CD44/EGFRtargeting nanoparticles. Mol Ther 2016;24(4):759e69. Li YF, Zhang HT, Xin L. Hyaluronic acid-modified polyamidoamine dendrimer G5-entrapped gold nanoparticles delivering METase gene inhibits gastric tumor growth via targeting CD44þ gastric cancer cells. J Cancer Res Clin Oncol 2018;144(8):1463e73. https://doi.org/10.1007/s00432-018-2678-5. Cosco D, Cilurzo F, Maiuolo J, Federico C, Di Martino MT, Cristiano MC, et al. Delivery of miR-34a by chitosan/PLGA nanoplexes for the anticancer treatment of multiple myeloma. Sci Rep 2015;5:17579. https://doi.org/10.1038/srep17579. Gaur S, Wen Y, Song JH, Parikh NU, Mangala LS, Blessing AM, et al. Chitosan nanoparticle-mediated delivery of miRNA-34a decreases prostate tumor growth in the bone and its expression induces non-canonical autophagy. Oncotarget 2015;6(30):29161e77. https://doi.org/10.18632/oncotarget.4971. Sadio A, Gustafsson JK, Pereira B, Gomes CP, Hansson GC, David L, et al. Modified-chitosan/siRNA nanoparticles downregulate cellular CDX2 expression and cross the gastric mucus barrier. PLoS One 2014;9(6):e99449. https://doi.org/10.1371/journal.pone.0099449. Fan L, Yang Q, Tan J, Qiao Y, Wang Q, He J, et al. Dual loading miR-218 mimics and Temozolomide using AuCOOH@FA-CS drug delivery system: promising targeted anti-tumor drug delivery system with sequential release functions. J Exp Clin Cancer Res 2015;34:106. https://doi.org/10.1186/s13046-015-0216-8. Wang F, Pang JD, Huang LL, Wang R, Li D, Sun K, et al. Nanoscale polysaccharide derivative as an AEG-1 siRNA carrier for effective osteosarcoma therapy. Int J Nanomed 2018;13:857e75. https://doi.org/10.2147/IJN.S147747. Wang K, Kievit F, Sham JG, Jeon M, Stephen ZR, Bakthavatsalam A, et al. Iron-oxide-based nanovector for tumor target siRNA delivery in an orthotopic hepatocellular carcinoma xenograft mouse model. Small 2016;12:477e87. https://doi.org/10.1002/smll.201501985. Van Woensel M, Wauthoz N, Rosie`re R, Mathieu V, Kiss R, Lefranc F, et al. Development of siRNA-loaded chitosan nanoparticles targeting Galectin-1 for the treatment of glioblastoma multiforme via intranasal administration. J Control Release 2016;227:71e81. https://doi.org/10.1016/ j.jconrel.2016.02.032. Zhao F, Yin H, Li J. Supramolecular self-assembly forming a multifunctional synergisti system for targeted co-delivery of gene and drug. Biomaterials 2014;35(3):1050e62. Liu T, Xue W, Xie MQ, Ma D. Star-shaped cyclodextrin-poly(L-lysine) derivative co-delivering docetaxel and MMP-9 siRNA plasmid in cancer therapy. Biomaterials 2014;35:3865e72. Liu T, Wu X, Wang Y, Zhag T, Wu T, Liu F, et al. Folate-targeted star-shaped cationic copolymer co-delivering docetaxel and MMP-9 siRNA for nasopharyngeal carcinoma therapy. Oncotarget 2016;7(27):42017e30. Shen J, Kim HC, Su H, Wang F, Wolfram J, Kirui D, et al. Cyclodextrin and polyethylenimine functionalized mesoporous silica nanoparticles for delivery of siRNA cancer therapeutics. Theranostics 2014;4(5):487e97. https://doi.org/10.7150/thno.8263. Davis ME. The first targeted delivery of siRNA in human via a self-assembling, cyclodextrin polymer-based nanoparticle: from concept to clinic. Mol Pharm 2009;6(3):659e68. Ribas A, Tolcher AW, Yen Y. Safety study of CALAA-01 to treat solid tumor cancers. Calando pharmaceuticals. 2018. Available in: https://clinicaltrials.gov/ct2/show/NCT00689065. Underhill C. CD44: the hyaluronan receptor. J Cell Sci 1992;103(Pt 2):293e8. Senbanjo LT, Chellaiah MA. CD44: a multifunctional cell surface adhesion receptor is a regulator of progression and metastasis of cancer cells. Front Cell Dev Biol 2017;5:18. https://doi.org/10.3389/ fcell.2017.00018.

468 Chapter 19 [92] Taniguchi D, Saeki H, Nakashima Y, Kudou K, Nakanishi R, Kubo N, et al. CD44v9 is associated with epithelial-mesenchymal transition and poor outcomes in esophageal squamous cell carcinoma. Cancer Med 2018. https://doi.org/10.1002/cam4.1874. [93] Misra S, Heldin P, Hascall VC, Karamanos NK, Skandalis SS, Markwald RR, et al. Hyaluronan-CD44 interactions as potential targets for cancer therapy. FEBS J 2011;278(9):1429e43. [94] Kato Y, Ozawa S, Miyamoto C, Maehata Y, Suzuki A, et al. Acidic extracellular microenvironment and cancer. Cancer Cell Int 2013;13(1):89. https://doi.org/10.1186/1475-2867-13-89. [95] Huang G, Huang H. Application of hyaluronic acid as carriers in drug delivery. Drug Deliv 2018;25(1):766e72. [96] Raja MA, Katas H, Jing Wen T. Stability, intracellular delivery, and release of siRNA from chitosan nanoparticles using different cross-linkers. PLoS One 2015;10(6):e0128963. https://doi.org/10.1371/ journal.pone.0128963. [97] Martin DT, Steinbach JM, Liu J, Shimizu S, Kaimakliotis HZ, Wheeler MA, et al. Surface-modified nanoparticles enhance transurothelial penetration and delivery of survivin siRNA in treating bladder cancer. Mol Canc Therapeut 2013;13(1):71e81. [98] Li JM, Zhang W, Su H, Wang YY, Tan CP, Ji LN, et al. Reversal of multidrug resistance in MCF-7/Adr cells by codelivery of doxorubicin and BCL2 siRNA using a foic acid-conjugated polyethylenimine hydroxypropyl-b-cyclodextrin nanocarrier. Int J Nanomed 2015;10:3147e62. https://doi.org/10.2147/ IJN.S67146. [99] Ceborska M. Folate appended cyclodextrins for drug, DNA, and siRNA delivery. Eur J Pharm Biopharm 2017;120:133e45. https://doi.org/10.1016/j.ejpb.2017.09.005. Epub 2017 Sep. 9. [100] Ma X, et al. Redox-responsive mesoporous silica nanoparticles: a physiologically sensitive codelivery vehicle for siRNA and doxorubicin. Antioxid Redox Signal 2014;21(5):707e22. https://doi.org/10.1089/ ars.2012.5076. Epub 2013 Sep 28. [101] Marqus S, Pirogova E, Piva TJ. Evaluation of the use of therapeutic peptides for cancer treatment. J Biomed Sci 2017;24(1):1e15. [102] Torchilin VP, Lukyanov AN. Peptide and protein drug delivery to and into tumors: challenges and solutions. Drug Discov Today 2003;8(6):259e66. [103] Pyronnet S, Bousquet C, Najib S, Azar R, Laklai H, Susini C. Antitumor effects of somatostatin. Mol Cell Endocrinol 2008;286(1e2):230e7. [104] Yu Y, Wang YY, Wang YQ, Wang X, Liu YY, Wang JT, et al. Antiangiogenic therapy using endostatin increases the number of ALDHþ lung cancer stem cells by generating intratumor hypoxia. Sci Rep 2016;6:34239. https://doi.org/10.1038/srep34239. [105] Wold ED, Smider VV, Felding BH. Antibody therapeutics in oncology. Immunotherapy 2016;2(1):1e18. [106] Bryant J, inventor, Genentech Inc, assignee. Herceptin adjuvant therapy. Patent US 2006/0275305. 2006. Available from: https://patents.google.com/patent/US20060275305A1/en. [107] Grillo-Lopez A, Wihite CA, inventors, Biogen Inc, assignee. Chimeric anti-CD20 antibody, rituxan, for use in the treatment of chronic lymphocytic leukemia. A1European patent EP 1 616 572 B1. 1999. Available from: https://patents.google.com/patent/EP1616572B1/en. [108] Salzer W, Bostrom B, Messinger Y, Perissinotti AJ, Marini B. Asparaginase activity levels and monitoring in patients with acute lymphoblastic leukemia. Leuk Lymphoma 2018;59(8):1797e806. Available from: https://doi.org/10.1080/10428194.2017.1386305. [109] Ardolino M, Hsu J, Raulet DH. Cytokine treatment in cancer immunotherapy. Oncotarget 2015;6(23):19346e7. [110] dos Santos MA, Grenha A. Polysaccharide nanoparticles for protein and Peptide delivery: exploring less-known materials. In: Advances in protein chemistry and structural biology. 1st ed., 98. Elsevier Inc.; 2015. p. 223e61 Available from: https://doi.org/10.1016/bs.apcsb.2014.11.003. [111] Mahidhara G, Kanwar RK, Roy K, Kanwar JR. Oral administration of iron-saturated bovine lactoferrineloaded ceramic nanocapsules for breast cancer therapy and influence on iron and calcium metabolism. Int J Nanomed 2015;10:4081e98.

Natural polysaccharides 469 [112] Kanwar JR, Mahidhara G, Roy K, Sasidharan S, Krishnakumar S, Prasad N, et al. Fe-bLf nanoformulation targets survivin to kill colon cancer stem cells and maintains absorption of iron, calcium and zinc. Nanomedicine 2015;10(1):35e55. [113] Zhang Y, Tan X, Ren T, Jia C, Yang Z, Sun H. Folate-modified carboxymethyl-chitosan/ polyethylenimine/bovine serum albumin based complexes for tumor site-specific drug delivery. Carbohydr Polym 2018;198(54):76e85. Available from: https://doi.org/10.1016/j.carbpol.2018.06.055. [114] Chen J, Ouyang J, Chen Q, Deng C, Meng F, Zhang J, et al. EGFR and CD44 dual-targeted multifunctional hyaluronic acid nanogels boost protein delivery to ovarian and breast cancers in vitro and in vivo. ACS Appl Mater Interfaces 2017;9(28):24140e7. [115] Kanwar JR, Mahidhara G, Kanwar RK. Research Article: novel alginate-enclosed chitosan e calcium nanocarriers for oral delivery in colon cancer therapy. Nanomedicine 2012;7(10):1521e50. [116] Shimoda A, Sawada S, ichi, Akiyoshi K. Cell specific peptide-conjugated polysaccharide nanogels for protein delivery. Macromol Biosci 2011;11(7):882e8. [117] Gover Antoniraj M, Senthil Kumar C, Henry LJK, Natesan S, Kandasamy R. Atrial natriuretic peptide-conjugated chitosan-hydrazone-mPEG copolymer nanoparticles as pH-responsive carriers for intracellular delivery of prednisone. Carbohydr Polym 2017;157:1677e86. [118] Dissanayake S, Denny WA, Gamage S, Sarojini V. Recent developments in anticancer drug delivery using cell penetrating and tumor targeting peptides. J Control Release 2017;250:62e76. [119] Liang K, Ng S, Lee F, Lim J, Chung JE, Lee SS, et al. Targeted intracellular protein delivery based on hyaluronic acid-green tea catechin nanogels. Acta Biomater 2016;33:142e52. [120] Ding L, Jiang Y, Zhang J, Klok HA, Zhong Z. PH-sensitive coiled-coil peptide-cross-linked hyaluronic acid nanogels: synthesis and targeted intracellular protein delivery to CD44 positive cancer cells. Biomacromolecules 2018;19(2):555e62. [121] Wu DY, Ma Y, Hou XS, Zhang WJ, Wang P, Chen H, et al. Co-delivery of antineoplastic and protein drugs by chitosan nanocapsules for a collaborative tumor treatment. Carbohydr Polym 2017;157: 1470e8. [122] Dionı´sio M, Braz L, Corvo M, Lourenc¸o JP, Grenha A, Rosa da Costa AM. Charged pullulan derivatives for the development of nanocarriers by polyelectrolyte complexation. Int J Biol Macromol 2016;86:129e38. [123] Liu K, Zheng D, Zhao J, Tao Y, Wang Y, He J, et al. PH-Sensitive nanogels based on the electrostatic self-assembly of radionuclide131I labeled albumin and carboxymethyl cellulose for synergistic combined chemo-radioisotope therapy of cancer. J Mater Chem B Royal Soc Chem 2018;6(29):4738e46. [124] Zhang C, Shi G, Zhang J, Song H, Niu J, Shi S, et al. Targeted antigen delivery to dendritic cell via functionalized alginate nanoparticles for cancer immunotherapy. J Control Release 2017;256(April):170e81. [125] Li P, Luo Z, Liu P, Gao N, Zhang Y, Pan H, et al. Bioreducible alginate-poly(ethylenimine) nanogels as an antigen-delivery system robustly enhance vaccine-elicited humoral and cellular immune responses. J Control Release 2013;168(3):271e9. [126] Chertok B, David AE, Yang VC. Magnetically-enabled and MR-monitored selective brain tumor protein delivery in rats via magnetic nanocarriers. Biomaterials 2011;32(26):6245e53. [127] Zhang J, Shin MC, David AE, Zhou J, Lee K, He H, et al. Long-circulating heparin-functionalized magnetic nanoparticles for potential application as a protein drug delivery platform. Mol Pharm 2013;10(10):3892e902. [128] Koshy ST, Zhang DKY, Grolman JM, Stafford AG, Mooney DJ. Injectable nanocomposite cryogels for versatile protein drug delivery. Acta Biomater 2018;65:36e43. [129] Rodrigues S, Cordeiro C, Seijo B, Remun˜a´n-Lo´pez C, Grenha A. Hybrid nanosystems based on natural polymers as protein carriers for respiratory delivery: stability and toxicological evaluation. Carbohydr Polym 2015;123:369e80.

470 Chapter 19 [130] Huang Y, Liu M, Gao C, Yang J, Zhang X, Zhang X, et al. Ultra-small and innocuous cationic starch nanospheres: preparation, characterization and drug delivery study. Int J Biol Macromol 2013;58: 231e9. [131] Gou ML, Dai M, Li XY, Yang L, Huang MJ, Wang YS, et al. Preparation of mannan modified anionic PCL-PEG-PCL nanoparticles at one-step for bFGF antigen delivery to improve humoral immunity. Colloids Surfaces B Biointerfaces 2008;64(1):135e9. [132] Cuna M, Alonso-Sandel M, Remunan-Lopez C, Pivel JP, Alonso-Lebrero JL, Alonso MJ. Development of phosphorylated glucomannan-coated chitosan nanoparticles as nanocarriers for protein delivery. J Nanosci Nanotechnol 2006;6(9e10):2887e95. [133] Liu Y, Yao W, Wang S, Geng D, Zheng Q, Chen A. Preparation and characterization of fucoidan-chitosan nanospheres by the sonification method. J Nanosci Nanotechnol 2014;14(5):3844e9. [134] Nochi T, Yuki Y, Takahashi H, Sawada SI, Mejima M, Kohda T, et al. Nanogel antigenic protein-delivery system for adjuvant-free intranasal vaccines. Nat Mater Nat Publ Group 2010;9(7):572e8. Available from: https://doi.org/10.1038/nmat2784. [135] Liang Y, Kiick KL. Heparin-functionalized polymeric biomaterials in tissue engineering and drug delivery applications. Acta Biomater 2014;10(4):1588e600. [136] Soepenberg O, de Jonge MJ, Sparreboom A, de Bruin P, Eskens FA, de Heus G, et al. Phase I and pharmacokinetic study of DE-310 in patients with advanced solid tumors. Clin Cancer Res 2005;11:703e11. [137] Veltkamp SA, Witteveen EO, Capriati A, Crea A, Animati F, Voogel-Fuchs M, et al. Clinical and pharmacologic study of the novel prodrug delimotecan (MEN 4901/T-0128) in patients with solid tumors. Clin Cancer Res November 15, 2008;14(22):7535e44. https://doi.org/10.1158/1078-0432.CCR08-0438. [138] Bigioni M, Parlani M, Bressan A, Bellarosa D, Rivoltini L, Animati F, et al. Antitumor activity of delimotecan against human metastatic melanoma: pharmacokinetics and molecular determinants. Int J Cancer 2009;125(10):2456e64. https://doi.org/10.1002/ijc.24661. [139] Kim JK, Han KH, Lee JT, Pail YH, Ahn SH, Lee JD, et al. Long-term clinical outcome of phase IIb clinical trial of percutaneous injection with holmium-166/chitosan complex (Milican) for the treatment of small hepatocellular carcinoma. Clin Cancer Res 2006;12:543e8. https://doi.org/10.1158/10780432.CCR-05-1730. [140] Bassi PF, Volpe A, D’Agostino D, Palermo G, Renier D, Franchini S, et al. Paclitaxel-hyaluronic acid for intravesical therapy of bacillus Calmette-Gue´rin refractory carcinoma in situ of the bladder: results of a phase I study. Urol Times 2011;185(2):445e9. [141] CRLX101 in combination with bevacizumab for recurrent ovarian/tubal/peritoneal cncer. 2018. Available from: https://clinicaltrials.gov/ct2/show/study/NCT01652079?term¼CRLX101&recr¼Recruiting&rank¼3. [142] Neoadjuvant chemoradiotherapy with CRLX-101 and capecitabine for rectal cancer. 2018. Available from: https://clinicaltrials.gov/ct2/show/study/NCT02010567?term¼CRLX101&recr¼Recruiting&rank¼4. [143] Alternative dosing for CRLX101 alone and with avastin in advanced solid tumors. 2018. Available from: Https://clinicaltrials.gov/ct2/show/study/NCT02648711?term¼CRLX101&recr¼Recruiting&rank¼2. [144] Trial of CRLX101, a nanoparticle camptothecin with olaparib in people with relapsed/refractory small cell lung cancer. 2018. Available from: ttps://clinicaltrials.gov/ct2/show/study/NCT02769962? term¼CRLX101&recr¼Recruiting&rank¼1. [145] Trial of CRLX101 (Formerly Named IT-101) in the treatment of Advanced Solid Tumors. Available from: https://clinicaltrials.gov/ct2/show/NCT00333502.

C H A P T E R 20

Organic nanocomposites for the delivery of bioactive molecules Pedro M. Castro1, 2, Bruno Sarmento2, 3, 4, Ana Raquel Madureira1, Manuela E. Pintado1 CBQF e Centro de Biotecnologia e Quı´mica Fina e Laborato´rio Associado, Escola Superior de Biotecnologia, Universidade Cato´lica Portuguesa/Porto, Porto, Portugal; 2CESPU, Instituto de Investigacaeo e Formacaeo Avancada em Cieˆncias e Tecnologias da Saude, Gandra-PRD, Portugal; 3 i3S - Instituto de Investigac¸a˜o e Inovac¸a˜o em Sau´de, Universidade do Porto, Porto, Portugal; 4 INEB e Instituto de Engenharia Biome´dica, Universidade do Porto, Porto, Portugal 1

Chapter Outline 1. Introduction 471 2. Advanced uses of composite nanoparticles 473 2.1 2.2 2.3 2.4 2.5

Drug delivery 473 Diagnosis (600) 477 Biomedical engineering 478 Food safety, enrichment, and shelf-life extension Phytopharmacy and ecology 483

480

3. Toxicity 485 4. Conclusions, future perspectives, and legal concerns Acknowledgments 488 References 488

487

1. Introduction A great number of polymer matrices present several attractive functionalities for a wide range of areas of interest such as low weight, biocompatibility, low price, and flexibility. However, some characteristics can be improved (e.g., mechanical, optical, barrier, rheological, electrical) by the inclusion of organic or inorganic nanoparticles, turning them suitable for being used in several areas such as drug therapy, theragnostics, phytopharmacy, food packaging, and food additives [1,2]. Nanocomposites can be characterized as multiphase solid nanostructured particles formed by overlapping of two or more layers of different materials with the condition that at least one phase has one, two, Natural Polysaccharides in Drug Delivery and Biomedical Applications. https://doi.org/10.1016/B978-0-12-817055-7.00020-0 Copyright © 2019 Elsevier Inc. All rights reserved.

471

472 Chapter 20

Figure 20.1 Scheme of nanocomposite where a matricial structure in the micrometer range (A) entraps smaller particles in the nanometric range (B).

or three dimensions at the nanosize range, that is, between 1 and 999 nm [3]. A scheme of the possible configuration of a nanocomposite is outlined in Fig. 20.1. Based on structure, there is a wide array of nanocomposites: scaffolds, nanofibers, nanotubes, nanoparticles. The fact that nanostructured particles have several layers offers some improvements over conventional nanoparticles, as the possibility of a more controlled release of carried molecules, better stability (e.g., chemical, mechanical and/or thermal stability), and potential shelf-life enhancement. Moreover, layers can be composed of distinct materials that confer different and unique properties to the nanocomposites. Effectively, for instance, mechanical properties of biopolymers produced for several purposes (e.g., food packaging, drug delivery, phytochemicals, replacement/improvement of faulty or damaged anatomical structures) present a very positive enhancement regarding mechanical properties [4]. Indeed, a representative portion of mechanical (e.g., tensile strength, Young’s modulus) but also block properties (e.g., permeability to O2 and/or CO2) seem to be mainly dependent on the filler material, and the materials composing the composite blend offer improved characteristics over to the isolate materials or to conventional nanoparticles alone. Effectively, the concept of nanocomposites includes the presence of nanostructured materials (e.g., clays, metal oxides, silica nanoparticles). The fact that nanocomposites present, by definition, at least one layer at the nanoscale presents toxicity concerns that are somehow shared with nanoparticles. Indeed, small-sized structures may permeate organs, tissues, cells, or even organelles, potentially causing a chemical and/or physical disturbance to physiological activities. Also, a great number of nanocomposites include inorganic ions that expose cellular viability, for instance, to oxidative stress. Also, when nanocomposites are used to produce packaging for food products, migration may occur and compromise the quality of the packed product. Indeed,

Organic nanocomposites for the delivery of bioactive molecules 473 oxidation, off-flavors, color changing, or significant alteration of the nutritional composition of the product may occur if there is a faulty nanocomposite-based packaging system. Extreme conditions of pH and enzymatic activity may compromise the stability of nutrients when crossing the stomach. Association with nanoparticles may lead to an increased stability and, potentially, bioavailability, by avoiding direct contact of carried molecules with gastric juices. Also, since nanoparticles are capable of hampering digestive enzymes from reaching site-specific parts of carried molecules, protection is therefore enhanced [5]. Indeed, mucoadhesive polymeric nanoparticles are especially effective in the protection of proteins and peptides from the harsh gastric environment [6]. Poly(lactic acid) (PLA), poly(lactic-co-glycolic acid) (PLGA), poly(sebacic acid) (PSA), and poly(acrylic acid) (PAA) are good examples of polymers that can adhere to mucous on the walls of gastrointestinal tracts by one or by the combination of the following factors: hydrophobic interactions, entrapment within mucin matrices, hydrogen bonds. Effectiveness and safety of nanocomposites produced for bioengineering purposes, especially for tissue replacement or healing, depends on multiple factors such as porosity achieved by the ceramic/glass-nanoparticle interface, degradation times of used polymers, among several other factors that are thoroughly discussed in this chapter. A brief discussion on the legislation of nanocomposites as nanoparticles is also addressed in this chapter.

2. Advanced uses of composite nanoparticles 2.1 Drug delivery A nanocomposite for drug delivery is a nanosized structure that differs from conventional nanoparticles for necessarily being composed by, at least, two components other than a nanosized structure and one (or more) active pharmaceutical ingredient. Nanocomposites may be used as delivery systems that can improve major key features when compared to conventional delivery systems. Indeed, nanocomposites can offer controlled release of drugs, enhancing efficacy of the drug and reducing the frequency of drug administration, with consequent improvement of patient compliance to therapies [3]. Nanocomposites can also be used for targeted delivery, minimizing toxicity on healthy tissues/organs. A brief summary of common uses and production methods of nanocomposites is outlined in (Table 20.1). Core-shell nanocomposites have been widely used in drug delivery for the treatment of a wide array of health issues. Chitosan-based nanocomposites were tested as drug carriers for cancer therapy [7]. Doxorubicin was incorporated into nanocomposites of Ag2Schitosan with an oleoyl cap. Chitosan offered pH-sensitive release (preferential release at acidic pH, characteristic of tumor cells, targeting the nuclei) oleoyl conjugation increased affinity of nanocarriers to doxorubicin by hydrophobic interaction, greatly enhancing association efficiency.

474 Chapter 20 Table 20.1: Examples of nanocomposites as drug delivery systems and their production methods. Composite

Production method

Ag2S-Chitosan-Oleoyol

Conjugation and hydrophobic incorporation

Magnetic-graphite-alginate Chitosan-Ag

Coprecipitation method Cross-linking and ionic gelation Water/oil/water double emulsion with solvent evaporation Solution blending method

PLGA-montmorillonite

Jatropha Curcas oil-based alkyd-epoxy-graphene oxide Magnetite-PLGA Pectin-coated chitosan-LDH

Solvent evaporationsupercritical fluid extraction Coprecipitation

Uses Targeted delivery of doxorubicin to the nuclei of cancer cells Ibuprofen drug delivery Antibacterial drug delivery system Oral drug delivery with controlled release

Ref. [7]

[8] [9] [10]

Potential drug delivery

[11]

Targeted drug delivery

[12]

Colon-targeted drug delivery or 5-aminosalicylic acid

[13]

Core-shell poly(lactic-co-glycolic acid) (PLGA) nanocomposites are also produced to increase circulating times, stimuli-sensitivity, targeting specificity, and bioadhesion over conventional PLGA nanoparticles [3]. Indeed, targeted delivery of PLGA nanoparticles was assured by adding the cyclo(1,12)PenITDGEATDSGC to the surface, producing a functionalized nanocomposite able to preferentially target cells with upregulated intercellular cell-adhesion molecule-1 (ICAM-1) expression on the cell surface. ICAM-1 is upregulated in several inflammatory responses or in certain cancers and is, therefore, an important target for functionalized nanocomposites [14e16]. PLGA is also commonly used for the preparation of nanocomposites intended for drug delivery [3]. Venlafaxine hydrochloride has been incorporated into montmorillonite-PLGA nanocomposites aiming oral extended drug delivery [17]. Steady state elimination half-life of venlafaxine hydrochloride is short (4e5 h) and, therefore, administrations must be performed 2e3 times a day in order to assure efficacy of the treatment. Thus, a drug delivery form that reduces the frequency of administration of venlafaxine could greatly contribute to compliance to treatment. Venlafaxine hydrochloride association efficiency to montmorillonite-PLGA was high (85%) and release was characterized by an initial burst release with 17% of the drug being delivered in the first 0.5 h followed by a slower release with 100% of the drug being released after 12 h, in PBS. Results indicated that montmorillonite-PLGA nanocomposites offer a slower release of venlafaxine when compared with pure drug (42% released after 0.5 h and 100% after 3.5 h) or nanoparticles produced only with PLGA (25% released after 0.5 h and 100% after 7.5 h).

Organic nanocomposites for the delivery of bioactive molecules 475 PLGA was also combined with methacrylate (Eudragit E100) to produce nanocomposites for gene therapy. A plasmid encoding interleukin-10 used for the prevention of autoimmune diseases was incorporated into nanocomposites produced by W/O/W double emulsion/solvent evaporation, using PLGA and E100 as polymers (50% w/w of each) and cetyltrimethylammonium bromide (0.5% w/v), a cationic surfactant, was used to increase z-potential [18]. Both cellular uptake of nanocomposites and expression of interleukin-10 in vitro were tested on Human Embryonic Kidney (HEK293) cells. Endosomal internalization of the plasmid was similar with PLGA nanoparticles but effectiveness of the treatment, that is, the amount of DNA that reached the nuclei of the cells and thus promoted protein expression, was significantly higher for PLGA-E100 nanocomposites. Effectively, expression of interleukin-10 was 4.7-fold and 13.8-fold superior, at 24 and 72 h, respectively, for plasmids delivered by PLGA-E100 when compared with PLGA nanoparticles. This difference may to be due to the Proton Sponge (PS) effect. Indeed, E100 copolymer seems to induce the osmotic swelling and membrane disruption, leading to the delivery of DNA on the cytoplasm and greater transfection efficiency, both in vitro and in vivo [18,19]. A nanocomposite (biohydrid scaffold) was also produced using PLGA, calcium phosphate (CaP), and silk fibroin (SF) as components, for the delivery of the angiogenic factor vascular endothelial growth factor (VEGF) targeted for bone regeneration [20]. VEGF was reported as being a promoter of bone overall metabolism (e.g., mineralization, bone turnover, osteoblast migration) by promoting vascularization [21e24]. PLGA forms a barrier enclosing previously freeze-dried silk fibroin-calcium phosphate (SF-CaP) loaded with VEGF. Electrospinning technique was chosen for preparation of the scaffold for being a faster and more robust method than conventional techniques. Both in vitro and in vivo assays were performed to assess the biocompatibility and effectiveness of the biohybrid scaffold silk fibroin-calcium phosphate-PLGA (SFeCaP-PLGA). In vivo assays were performed in nine New Zealand rabbits. Bone sockets (negative control), SF-CaPPLGA scaffolds (placebo) and SF-CaP-PLGA scaffolds loaded with PLGA were implanted onto a cranial previously drilled orifice of three rabbits, respectively. After 10 days of treatment, rabbits treated with bone sockets and placebo SF-CaP-PLGA scaffolds demonstrated the formation of a thin fibrous connective tissue sheet in the damaged area, with a more pronounced effect on the rabbits treated with SF-CaP-PLGA scaffolds, as expected based on previously described data in the literature, namely due to the fact that SF-CaP biohybrid scaffold offers physical support for osteoblasts along with a porosity profile (70%e75%) that fits in the range that is considered to promote the enhancement of nutrients and elimination of waste resulting from bone metabolism [25e27]. Moreover, the effects of SF-CaP-PLGA loaded with VEGF were greatly improved, when compared with placebo or control. Indeed, SF-CaP-PLGA loaded with VEGF induced a pronounced bone growth with significantly inferior formation of fibrous tissue when compared to

476 Chapter 20 placebo or control. Moreover, the formation of new capillary structures was evident inside the bone island, most probably due to the incorporation of VEGF loaded by SF-CaPPLGA biohybrid scaffolds. The fact that osteogenesis occurred is likely related with the formation of new vascular structures [28,29]. In vitro cell viability assay was verified by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium (MTT) reduction assay using highly purified osteoblast for control, placebo, and VEGF-containing formulations. Cell viability was not compromised in day 3 and, after 7 and 14 days of incubation times, osteoblast cell number had highly increased, as predicted based on previously published literature [30,31]. Indeed, primary human osteoblasts growth was significantly (P < .05) higher for SF-CaP-PLGA biohybrid scaffolds containing VEGF when compared with negative control or placebo formulations. It has been previously suggested that the proliferation of primary human osteoblasts was mediated by VEGF receptor-1 [23]. Also, highly purified osteoblast cells increased production of alkaline phosphatase (ALP) when in contact with SF-CaP or SF-CaP-PLGA biohybrid scaffolds containing VEGF, despite the fact that significant differences were only observed 7 days after contact with osteoblast cells. Moreover, in accordance to what was verified for the MTT assay, SF-CaP-PLGA biohybrid scaffolds containing VEGF induced a significantly higher (P < .05) osteoblast response than the placebo. Results regarding ALP activity were also in accordance with information predicted in the literature [32]. Bacterial cellulose, produced by the bacterium Acetobacter xylinum, has been used as polymer for the formation of carriers for drug delivery [33,34]. Recently, bacterial cellulose has been associated with graphene oxide for the production of nanocomposites as carriers for ibuprofen [35]. Indeed, graphene oxide is reported to offer high drug loading efficiency and, due to the numerous accessible functional groups, is also easily functionalized especially on the surface. Also, the fact that graphene oxide presents a sheetlike shape grants a large surface area and a great amount of available functional groups that can be easily modified. Also, even though it can cause aggregation, the fact that graphene oxide tends to aggregate to proteins can be advantageous in the right circumstances (e.g., mucoadhesion) [36]. On the other hand, bacterial cellulose (BC) from A. xylinum presents very promising morphologic characteristics (e.g., good porosity profile, high crystallinity, tensile strength, and elastic modulus) along with great capacity to retain water [37,38]. BC-GO nanocomposite was used as carrier of ibuprofen (drug model) due to short biological half-life, well-studied bioactivity, and suitable molecular size and shape to be adsorbed to both BC and GO. BC-GO demonstrated a pH-dependent release of ibuprofen. Ibuprofen release from BC-GO nanocomposites was faster near physiological pH and slower at more acidic pH values. Release profile was associated with different swelling behavior of BC at different pH values. Also, release of ibuprofen from BC-GO nanocomposite follows a Korsmeyer-Peppas model indicating that release profile depends on the structural and geometric dosage form.

Organic nanocomposites for the delivery of bioactive molecules 477 In another study, a nanocomposite-based delivery system with magnetic targeting to specific tissues and temperature-induced release was developed by producing nanocomposites of poly(N-isopropylacrylamide) with y-Fe2O3 and mesoporous silica (SBA-15), using gentamicin as model drug [39]. Even though the gentamicin was only modestly loaded in the nanocomposites (28%), release in response to temperature increase was successfully achieved. Indeed, release assay performed at 10 C reached a plateau of about 10% gentamicin release after 1 h. But, for the release assay performed at 40 C, a closer temperature to physiological human temperature, a burst release occurred till 2 h and a plateau was reached till the end of the assay with accumulated release of gentamicin reaching about 70%. Moreover, the y-Fe2O3 enclosed in the SBA-15 porous structure offered magnetism to the particles, therefore allowing targeted release of the drug content.

2.2 Diagnosis (600) Nanocomposites are an important improvement over nanoparticles to be used as sensors with great performance and, generally, low prices when compared with reference sensors. Nanocomposite sensors are easily customizable and can be tailored, among many other uses, to indicate small changes of the pH of an environment or indicate the presence (and, in some cases, the concentration) of indicators of the occurrence of particular diseases [40]. As happens with the administration of drugs, invasive routes used for diagnosis, control, or management of diseases may reduce compliance with consequent aggravation of the health problems of the patients for lack of premature intervention. Besides the invasive insulin treatment, control of plasmatic glycemia by the patients with diabetes mellitus represents a paradigmatic case, since current procedures involve finger pricking or phlebotomizing which causes pain and increases the possibility of infections and lesions on the sampling anatomic area. In that instance, several glucose nanocomposite sensors have been developed, which are simple, cheap, sensitive, and highly selective [41]. 5,10,15,20tetrakis(4-carboxyphenyl)-porphyrin-Fe3O4 nanocomposites have been developed with peroxidaselike activity, leading to the oxidation of H2O2 with the appearance of blue color. Peroxidase activity was significantly higher than for Fe3O4 nanoparticles alone. Sensitive assays to perform an early diagnosis of cancer are also of paramount importance in order to enhance the odds of remission success. Therefore, aiming to obtain sensitive, robust, electrochemical immunosensor for the detection of a-fetoprotein, an oncofetal glycoprotein expressed in hepatocellular carcinoma, a metal-organic immunosensor based on a nanocomposite of poly(methylene blue)-Au modified with graphene was developed [42]. The immunosensor revealed to be sensitive (current correlated linearly with a-fetoprotein concentration) and selective to a-fetoprotein and demonstrated to be stable

478 Chapter 20 as well, even after being stored at 4 C for about 2 weeks, not significantly compromising electrochemical response. poly(methylene blue)-Au-graphene also demonstrated good accuracy when compared to ELISA for the analysis of clinical serum samples containing a-fetoprotein, with acceptable relative errors from 8.00% to 8.96%. Repeatability was also assured by performing successive, intraday and interday determinations of a-fetoprotein, for five consecutive days with relative standard deviations of 3.21% and 4.86%, respectively.

2.3 Biomedical engineering It is conventionally accepted that the higher the complexity of the anatomical structure to repair/replace, the more intricate it is to fix damaged tissues [43]. Nanocomposite structures for biomedical engineering are mainly used for tissue repairing or replacement. Other than biocompatibility, bioactivity, ease of manipulation, and cost, mechanical characteristics (namely Young’s modulus and tensile strength) are of paramount importance. Nanocomposites represent relatively straightforward tools to replace or repair tissues, especially bone and cartilage tissues. Effectively, bone itself is nanocompositebased, being composed of a composite of collagen type I fibers and nanohydroxyapatite crystals as matrix [44]. The organization of tendons and articular cartilage ligaments is also micro- and nanostructured [45]. This chapter and the chapter on nanocomposites are based totally or partially on organic compounds. Therefore, with regard to tissue bioengineering, mainly polymer-matrix nanocomposites will be addressed. Polymer-matrix nanocomposites can be labeled as biodegradable (e.g., polylactide e PLA e poly(glycolides) e PGA e and poly-ε-caprolactone) or nonbiodegradable (e.g., polypropylene, polyethylene, polytetrafluoroethylene, polyamide) depending on the polymer chosen for the dispersion of the nanoparticles. Even though conventional glass-ceramic A-W can be used for the replacement of a wide array of bone fillers and structures (e.g., maxillofacial bone, iliac crest, intervertebral discs), it is not a suitable replacement for structures that are subjected to very high loads such as tibial or femoral bones [46e48]. On the other hand, HAPEX, a hydroxyapatiteparticle-reinforced high-density polyethylene (HDPE) composite, was improved by forming a composite with glass-ceramic A-W thus improving bioactivity of the material, to better mimic the natural bone tissue. Nevertheless, the inclusion of glass-ceramic A-W also led to a decrease of mechanical resistance of the composite, being unusable for the bones usually subjected to great stress [49,50]. Attempting to offer HDPE more characteristics that resemble human healthy bone, a nanocomposite was formed by adding TiO2 nanoparticles to the hydroxyapatite matrix. Indeed, Young’s modulus values greatly increased for the new-formed nanocomposite, but it also resulted in a decline of bioactivity and biocompatibility.

Organic nanocomposites for the delivery of bioactive molecules 479 Nanophase forsterite (Mg2SiO4) was also used as a bioceramic alternative to hydroxyapatite, aiming to produce a nanocomposite-based bone tissue repairing product [51]. Nanoscaled (exclusively) forsterite presents several advantages such as good cellular interactions with osteoclasts and osteoblasts, being reported as possessing very promising apatite formation capability when tested in vitro [52,53]. Also, forsterite has improved mechanical characteristics over conventional ceramics, glasses, or glass-ceramics [54,55]. Forsterite nanoparticles were combined with polycaprolactone (PCL) to obtain bonerepairing nanocomposites (scaffolds). PCL was chosen due to higher and more durable mechanical strength along with a degradation time that correlates very well with bone regeneration period [56]. The obtained nanocomposite was demonstrated to have the ideal porosity profile for migration and proliferation of osteoblast cells. Nevertheless, as the relative amount of forsterite nanoparticles in the nanocomposite formulation was increased, degradation of the porosity profile occurred (due to a decrease of average pore size with consequent limitation of osteoblast action), impairing bone tissue healing. Attempting to improve previous work, forsterite was esterified with dodecyl alcohol in order to increase interfacial adhesion and dispersion of nanosized forsterite onto/within the ceramic polymer matrix (PCL) [51]. However, esterification with dodecyl alcohol also decreases hydrophilicity, therefore compromising the interaction with bone matrix. To overcome the solubility problem, the esterification reaction was reversed after modified forsterite was dispersed onto/within the PCL polymeric matrix, therefore reobtaining bioactivity and the potential to perform cellular interactions. One of the most used polymer for bioengineering of tissue (bone) repair is poly(methyl methacrylate) but, when added alone, it does not have the capability to positively interact with damaged bone [57]. Indeed, poly(methyl methacrylate) does not chemically or physically adhere to bone tissue and may release debris over time, caused by the motion, potentially damaging the surrounding tissues and causing implant loss [58,59]. In an attempt to achieve osteoconductivity, calcium ions and silica nanoparticles were used to produce nanocomposites with poly(methyl methacrylate) with the formation of hydroxy carbonated apatite. Resulting hydroxy carbonated apatite was tested in simulated body fluid resulting in the nucleation of silanol SieO- groups and, thus, a higher bioactivity with enhanced, long lasting bone matrix repairing activity. Also in an attempt to formulate nanocomposites that are osteocompatible but also osteoconductive and prevent implant loosening, PCL was associated with Bioglass or with calcium phosphate [60]. Bioglass is reported as being extremely biocompatible, osteoconductive, and osteoinductive, and has been used as bone filler or even as adjuvant in bone grafts [61,62]. Joining nanostructured Bioglass to an organic polymeric matrix may increase adsorption of proteins as fibronectin or vitronectin that are, in turn, related with enhanced cell adhesion to the nanocomposite structure [63,64]. Bioglass-polymer nanocomposites can also lead to an improvement of physical obstruction from bacterial

480 Chapter 20 deposition and also reduce bone resorption [65]. In comparison with PCL alone or PCLcalcium phosphate, PCL-Bioglass nanocomposite presented enhanced in vitro bioactivity (i.e., induced greater formation of cellular bridges with sheep bone marrow stromal cells and promoted greater cell growth). On the other hand, PCL-Bioglass composite had lower compressive Young’s modulus when compared to PCL-calcium phosphate or PCL alone, indicating a textural handicap. On the other hand, PCL-calcium phosphate provides a balanced outcome of both compressive Young’s modulus and cell proliferation. Nevertheless, results may vary depending on the animal and anatomical region from where the cell line used for in vitro growth and compatibility tests was obtained [66,67]. Therefore, further in vivo along with in vitro studies are regarded to determine the regenerating capacity of the developed nanocomposites. The loss of structures that support tooth may result in the unwanted migration of epithelial tissue to the damaged area, leading to the hindrance of periodontal regeneration [68]. Conventional intervention includes a membrane that prevents epithelial overgrowth and wound closure but does not significantly induce the regeneration of periodontal tissues [69,70]. In the attempt to improve periodontal tissue regeneration, Bioglass nanostructures were used to form nanocomposites with an alginate hydrogel [43]. The inclusion of Bioglass into the alginate scaffold resulted in a formulation with great resilience to degradation along with swelling capability. Moreover, Bioglass-alginate nanocomposite structure not only was not toxic to MG-63 (human bone fibroblasts obtained from osteosarcoma) or hPDLF (human periodontal ligament fibroblasts) cells but induced cell attachment and proliferation, leading to the conclusion that Bioglass-alginate nanocomposites can potentially accelerate in vivo bone regeneration when compared to standard procedures. Also, hPDLF cells increased alkaline phosphatase activity, suggesting an osteoblastlike behavior, that is, favoring bone growth. The authors concluded that the composite formed by alginate and Bioglass could represent a good bioactive matrix for periodontal tissue regeneration.

2.4 Food safety, enrichment, and shelf-life extension Biopolymers can be extremely useful for the agro-food industry and have been mainly developed as food-preserving and food-safety matrices [71]. Moreover, conversion of neat biopolymers into nanocomposites offers a wide array of advantages such as higher potential to enhance protection to products (e.g., by the incorporation of antimicrobial agents, protecting from post-production contamination, thereby extending shelf-life of the product). Biodegradable packaging nanocomposites present better chemical, mechanical, and thermal resistance than unprocessed biopolymers. Packaging nanocomposites would ideally present high capacity to prevent flavor compounds, water vapor, oxygen, and carbon dioxide diffusion [72].

Organic nanocomposites for the delivery of bioactive molecules 481 There is a wide array of biopolymers that can be used for the production of nanocomposite-based food packaging and can be divided into natural (e.g., starch, cellulose, pullulan, chitosan, curdlan, alginate) and synthetic (e.g., poly(L-lactide) (PLA), poly(lactic-co-glycolic acid) (PLGA)) biomaterials [73]. Natural or synthetic (or semisynthetic) biomaterials usually present good biocompatibility and are capable of offering reasonable mechanical, thermal, and chemical resilience, gas and water vapor permeability. However, the formation of hybrid nanocomposites with materials (clays are reported as being the most used) offers improvements on overall physicochemical characteristics of nanocomposite and biodegradable food packaging systems [72,74]. The formation of biopolymer-clay nanocomposite may be performed by being exfoliated or intercalated with biopolymers [75,76]. Biopolymers intercalated with clay materials occur when there is moderate biopolymer-clay affinity. Biopolymer penetrates the basal space of clay without changing the shape of the structure formed by the clay. On the other hand, formation of biopolymer-clay nanocomposite by exfoliation occurs when there is great affinity between clay material and biopolymer. Clay becomes totally and homogeneously dispersed in the polymer matrix [75,77,78]. A third way of blending biopolymers with clay compounds, tactoid, does not lead to the formation of nanocomposites and therefore will not be further explored in this review. With the formation of nanocomposite-based biodegradable food packaging, different categories can be enlisted and are outlined in Table 20.2. Chitosan, cellulose, starch, polylactic acid (PLA), polyhydroxybutyrate (PHB), polycaprolactone (PCL), poly-(butene succinate), and other biocompatible, biodegradable polymers have been studied to replace packaging materials made from fossil fuels [82]. Nanocomposites produced exclusively with cellulose, obtained from byproduct of the Table 20.2: Nanocomposite-based types of packaging systems for food products. Packaging type Degradable

Production

Smart

• Incorporation of inorganic nanofillers to the organic matrix. • Incorporation of biosensors.

Improved

• Incorporation of nanofillers.

Active

• Incorporation of nanofillers with antioxidant and antimicrobial activities.

Purposes • Mechanical improvement of packaging, increasing shelf-life of food products. • Monitorization of food quality for consumption. • Improvement of the quality of packaged food by block of water/gas exchanges with the surrounding environment; • Improvement of food appearance. • Improvement of quality and shelf-life of packaged food product.

Ref. [79,80]

[79] [79]

[79,81]

482 Chapter 20 processing of sugarcane bagasse, presented appreciable tensile strength (140 MPa) and represented an interesting low-cost raw material to produce protective films for food packaging. Due to water-sensitivity that leads to deterioration of mechanical strength, biopolymers were combined with nanosized layered silicates with the formation of nanocomposite films [75,83]. The formation of organic-inorganic nanocomposite films for food packaging also improved thermal stability and resistance to solvents and decreased permeability of gases and liquids, without compromising biodegradability or biocompatibility [79,84]. Chitosan-ZnO nanocomposites have been used to coat polyethylene films for antibacterial protection, maintain food colors, and improve mechanical and barrier properties of the package [85]. Indeed, an effective antimicrobial activity against Escherichia coli, Staphylococcus aureus, and Staphylococcus enterica were obtained when combining chitosan and ZnO nanoparticles as nanocomposites for coating polyethylene films, leading to an improved shelf-life and overall safety for the consumer [71]. It was reported that chitosan-ZnO nanocomposites increase antimicrobial activity 10- to 20-fold when compared to chitosan alone [86]. Some nanocomposite-based packages are already being extensively used in food industry. Imperm nylon-silicate composites are used to coat food/beverage products, hindering both the loss of CO2 and the entrance of O2 into the system. Imperm is of paramount importance to maintain organoleptic conditions of gaseous (carbonated) beverages, simultaneously impeding the contact with unwanted outer oxygen that could degrade the food product. A composite formed of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHA) and zinc oxide (ZnO) was studied as biodegradable food packaging [87]. Even though PHA is a biodegradable, biocompatible polymer suitable for production of active food packaging, low mechanical robustness and thermal resistance implied using ZnO nanoparticles to overcome formulation problems but also to offer antimicrobial activity. The antimicrobial effect of the PHA-ZnO observed was variable according to the shape of ZnO particles. Other than size, ZnO nanoparticles used for the production of antibacterial biomaterials also presented distinct shapes after incorporation within PHA lattice. Indeed, PHA-P-ZnO nanocomposite showed much more effective antibacterial activity against Listeria monocytogenes and presented similar antibacterial activity against Salmonella enterica when compared with PHA-BZnO, PHA-RZnO, or PHA-SZnO composites. Moreover, BZnO, RZnO, and SZnO induced an inhibition of growing of both S. enterica and L. monocytogenes during 24 h of incubation, that is, induced a bacteriostatic effect. On the other hand, even though PZnO also induced a bacteriostatic effect on S. enterica, a reduction of >6 log units of L. monocytogenes was obtained when using PZnO. Differences in antibacterial effect seem to be related with the higher surface area of

Organic nanocomposites for the delivery of bioactive molecules 483 PZnO (74% of the surface area with accessible zinc atoms, surface area of 34 m2 g 1, and a prismatic morphology) than BZnO (67% of the surface area with accessible zinc atoms, surface area of 14 m2 g 1, and starlike morphology), RZnO (83% of total surface area with accessible zinc atoms, surface area of 14 m2 g 1, and a rodlike morphology) or SZnO (33% of total surface area with accessible zinc atoms, 17 m2 g 1, balllike morphology). Thus, packaging systems based on PHA-ZnO can be good alternatives to conventional packaging systems, either as degradable, active, or improved food packaging.

2.5 Phytopharmacy and ecology Toxicity caused by exposure to herbicides, either occupational or by intake of contaminated food products, is of paramount importance for public health of workers and consumers. A herbicide is a compound that, ideally, effectively eliminates weeds that compete with crops for space, light, nutrients, and water but does not present a residence time that is long enough that environmental, crop, or consumer health is somehow injured [88,89]. Besides the residence time, specificity of the herbicide is another characteristic of paramount importance [90]. Finally, as happens, for instance, for medicine formulation, dosage of pesticides must be just enough to be effective when the target is attained without compromising the physiological conditions of the system. Environment conditions (e.g., sunlight, wind, rain, animals) may compromise the effectiveness of pesticides by transformation or removal from the site of action. Indeed, photolysis (either direct or indirect) is a very frequent degradation inducer of pesticides [91]. Photolysis can, therefore, lead not only to loss of effectiveness of the pesticide but to unexpected toxicity coming from the photodegradation products [90]. The photoprotection of pesticides susceptible to photodegradation has been subject to intense study and the incorporation of pesticides into biopolymers could be an interesting idea, since the pesticide is protected and the environment is not seriously affected if biodegradable, nontoxic, polymers are used [92e95]. But, biodegradable polymers are fragile to environment factors and would demand a very high amount of material to encapsulate the pesticide. Therefore, the idea of producing a nanocomposite based on biopolymers (blocks radiation but suffering degradation instead of the incorporated pesticide) and inorganic phase that was capable of absorbing ultraviolet radiation [96]. In order to decrease photodegradation of ametryne, a selective herbicide for the control of grass weeds and broadleaf, a composite of starch and montmorillonite was produced. Starch is abundant, cheap, biodegradable, renewable, and easy to manipulate. Moreover, starch is a hydrophilic biopolymer and montmorillonite is reported to exfoliate in aqueous medium, improving the chances of compatibilization between the polymer and the clay without further use of stabilizers or

484 Chapter 20 other components [97e99]. To determine the photoprotection and degradation times of ametryne, samples were artificially irradiated with eight parallel tube lamps (16 W each) that mimic UV-C radiation, for 8 consecutive days [100]. Ametryne photodegradation was determined by weight loss, using an analytical scale. As a result, pure ametryne suffered higher weight loss, that is, higher photodegradation. Also, as predicted, starch-ametryne (1:1) blend also experienced a faster weight loss than required after 7 days. Nevertheless, ametryne UV-C photodegradation when encapsulated in starch was effective, despite starch degradation, indicating that starch photodegradation shielded the degradation of the herbicide. Starch UV degradation was reported as occurring by breaking C2eC3 bond of the glucopyranose ring [101]. Starch-montmorillonite-ametryne composite offered very good control of weight loss, without significant alteration after 7 days. Protection of ametryne when incorporated in starch-montmorillonite composite was dependent on the amount of clay used in the formulation. Nevertheless, the amount of montmorillonite used in the composite formulation with starch could not be indiscriminately increased due to phase segregation that occurred in the formulation, leading to partial decrease of protection of ametryne from photodegradation. Starch-montmorillonite-ametryne composite not only protected ametryne from photodegradation but also from thermal degradation. Finally, the nanocomposite offered slow release of the herbicide, a potential third advantage of incorporating herbicides into nanocomposites. Production of nanocomposites is also important for the detection of herbicides. Most commonly used herbicides have legislation to control the maximum amount that can be present in soil or food products. Acetochlor (2-chloro-N-(ethoxymethyl)-N-(2-ethyl-6methylphenyl)-acetamide), as ametryne, is a herbicide used to eliminate grasses and broadleaf weeds, commonly used in corn crops. However, acetochlor was characterized as a B-2 carcinogen by the United States Environmental Protection Agency (US EPA) and therefore monitoring of the amounts of acetochlor and transformation byproducts (ethanesulfonic acid is the most representative) is of paramount importance [102,103]. A photoelectrochemical sensor for indirect detection of acetochlor was produced using poly(3-hexylthiophene) (P3HT) as organic photoelectric material, a functionizer of TiO2 that works as a catalyst and is capable of absorbing ultraviolet light [104]. However, TiO2 only performs as a catalyst under ultraviolet light that corresponds to a very little portion (3%e5%) of the incident solar light and, therefore, had to be functionalized with P3HT [105]. The composite resulting from nanosized TiO2 with P3HT had already been successfully tested for the detection of organophosphorus pesticide chlorpyrifos [106]. For the indirect detection of acetochlor, P3HT-TiO2 was dispersed in ionic liquid (IL) 1-butyl-3-methylimidazolium hexafluorophosphate, commonly used due to thermal stability, great electrochemical characteristics, avoiding the use of supporting electrolytes, being an interesting medium for photochemical reactions [107]. The P3HTTiO2-IL nanocomposite uses a glassy carbon electrode (GCE) as the working electrode.

Organic nanocomposites for the delivery of bioactive molecules 485 Effectively, acetochlor photolysis reaction products are able to be accurately detected by P3HT-TiO2-IL nanocomposite, in water samples. Another way to manage human-caused environmental pollution is, for instance, to attempt to convert the pollutant compounds into nonpollutant, biodegradable, ecofriendly molecules or to isolate and remove the pollutants from the environment. Nanocomposites can be functionalized in order to promote degradation of toxic molecules into nontoxic compounds. In that instance, a graphitelike carbon nitride (g-C3N4) semiconductor material was associated with TiO2 in order to induce the photodegradation of 2,4,6trichlorophenol (2,4,6-TCP), a phytochemical widely used in forest and woodworking industries [108]. 2,4,6-Trichlorophenol was reported as being lethal to animals and plants and to cause mutagenicity and carcinogenicity [109], and amplify the absorption wavelength of TiO2. Graphitelike carbon nitride has also been commonly used for pollution degradation purposes but has a major flaw: g-C3N4 is unrecyclable and impossible to be reused because of the high dispersibility in nature [110,111]. To recover used geC3N4eTiO2 nanocomposite, Fe3O4 nanoparticles were added to confer magnetism to the nanocomposite. It was previously reported that the combination of TiO2 and Fe3O4 facilitated the manipulation of the composite (agglomeration prevention and increase of durability) but also allowed to separate the magnetic composite from a solution just by applying a magnetic field. Indeed, not only the geC3N4eTiO2eFe3O4 nanocomposite was very effective in the photodegradation of 2,4,6-TCP (95%,7% degraded after 100 min under 500 W of sunlike radiation) but the composite was also easily recovered and reused (92.0% of degradation occurred after 5 cycles of degradation).

3. Toxicity Nanocomposites have been extensively reported as safe, mainly recurring to the fact that, if biodegradable and biocompatible materials are used for the formulations, no toxicity is expected. Nonetheless, the fact that there is a lack of information regarding the toxicity profile of nanocomposites still persists and is commonly acknowledged by researchers [112,113]. The necessity of controlling the toxicity of nanosized systems as nanocomposites is of the greatest importance since a wide array of products already marketed claim to contain nanoparticles as components. Indeed, it has been forecast that the market value of nanoparticle-based products by 2024 will exceed the mark of US$125 billion [114]. It is also of great importance to acknowledge that the presence of nanoparticle-based products is highly relevant in cosmetic, biomedical, food and agriculture industries, therefore having a direct contact with the consumer. Organic nanocomposites are usually produced with materials with well-known biocompatibility. Nevertheless, safety of chronic nanocomposite exposition and the effects of

486 Chapter 20 bioaccumulation are yet to be properly assessed [115]. Moreover, it has been demonstrated that, due to specific in vivo immunomodulatory effects (along with specific uptake methods) of nanoparticles administered through parenteric route, expected toxicity of nanocomposites is not necessarily circumscribed to the toxicity profile of the individual components of the formulations [116,117]. Thus, even though of high importance, the establishment of a correlation between nanocomposite composition and biological responses in vitro is rather simplistic. Indeed, even in vivo toxicity studies using animal models according to the preclinical toxicity studies applied today might not be sufficient for nanocomposites as chronical administration of nanoparticle-based products might lead to different toxicity profiles over extended periods of contact [118]. One of the major characteristics that leads to different toxicity profiles of nanocomposites and the bulk materials used to produce them is the reduced size. Indeed, as the size of a particle decreases, the surface-to-mass ratio increases considerably, leading to a significant change in the interaction with biological structures [119]. Moreover, surface structure and nanocomposite shape are also important factors to characterize and control due to potential alterations regarding biological fate. Indeed, a higher surface area induces a higher exposure of functional groups of the excipients, therefore leading to a higher reactivity of the nanocomposites when compared with the bulk materials. Thus, besides from the cell models, commonly used to mimic different biologic tissues for the assessment of in vitro toxicity of nanocomposites, immunotoxicity has also been assessed [120]. Immunotoxicity is dependent on the dosage, number of specific cell receptors, surface functionalization, z-potential, cellular uptake and particle size of the nanocomposites. It has been reported that the intravenous administration of nanocarriers is associated with the activation of the complement, that is, with a response of the innate immune system. For instance, the cascade of inflammasomes, responsible for beginning acute inflammation processes that can lead to an upstart of autoimmune disorders, was reported to be activated by nanocarriers, leading to an unwanted activation of macrophages and inherent oxidative stress along with premature elimination of the particles [121,122]. Besides from the activation of the complement, cytokine storms were also reported to occur with the administration of nanocarriers, either due to endotoxin contamination of the formulation or over activation of innate immune cells and usually lead to the occurrence of fever, hypotension and severe inflammatory symptoms and can even be life-threatening [123]. In brief, for nanocomposites to be characterized regarding toxicological profile, besides from the commonly performed in vitro tests (e.g., MTT and lactate dehydrogenase releasedLDH) usually performed using cancer cells as cell models, the assessment of innate immune system response (e.g., by monitoring the activity of the complement and

Organic nanocomposites for the delivery of bioactive molecules 487 lymphocytes of the innate immune system), the determination of the overproduction reactive oxygen species along with tests to assess potential mutagenic effects caused by the nanocomposites and protein/gene expression must be performed [118].

4. Conclusions, future perspectives, and legal concerns Future of nanocomposites, as a somehow improved version of nanoparticles, strongly depends on consumer/patient/user acceptance. Indeed, this is a long way to be taken in order to fully understand the impact that nanoparticles or nanocomposites may induce on the application environment, either a human body or an ecosystem. Therefore, legislation based on solid scientific evidence must be elaborated in order to offer assurances to the users of nanocomposites. The facts that consumers are still fearful of the concept of food products functionalized with nanoparticles and that there is not yet sufficient legislation that, for instance, properly defines the term nanomaterials or any specific steps for the elaboration of a food product containing nanoparticles, from production to labeling, are strongly hindering industries from stepping forward into the production of food products enhanced with the inclusion of nanoparticles [124]. Indeed, the concept that knowledge of the potential hazard that can be induced by the intake of nanoparticles can be extrapolated from existing information (and regulation) regarding the single components that constitute the nanoparticle is still generally accepted [125]. Also, it is claimed by the Organization for Economic Co-operation and Development (OECD) that compounds can be grouped in chemical categories so that the characterization of a product as safe is overall simplified [126]. Instead, once a compound is attributed to a chemical category, hazard estimation would be the one attributed to the overall category and not to the single compound itself. This approach does not seem entirely suitable for the risk assessment of nanoparticles as food ingredients. Indeed, besides the potential creation of new chemical entities during production of nanoparticles, particle size represents a characteristic of paramount importance that is not necessarily related with the chemical composition of nanoparticles. Effectively, size is a main characteristic to consider when toxicity issues are concerned, since crossing of biological barriers along with tissue and cellular distribution are highly dependent on the dimensions of the particle [125]. Also, particle numbers, charge, shape, and surface area are considered to contribute to different toxicity profiles of nanoparticles, when compared with larger particles formulated with the same components [127]. Thus, grouping compounds into chemical categories is rather insufficient for the risk assessment for the intake of nanoparticles in food. The European parliament and the council of the European Union regulated that consumers must be informed of the presence of engineered nanomaterials, suggesting that labels must include this information [128]. But, even though engineered nanomaterials are defined by the EU regulation, no clear boundaries on the size of particles are defined.

488 Chapter 20 Regarding biosensors, nanocomposites can really represent breakthrough tools, being accurate, reproducible, easy to produce, and cheaper than conventional sensors and are surely going to get even greater attention of researchers in a very proximate future.

Acknowledgments This article is a result of the project NORTE-01-0145-FEDER-000012, supported by Norte Portugal Regional Operational Program (NORTE 2020), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF). Pedro M. Castro would like to thank Comissa˜o de Coordenac¸a˜o e Desenvolvimento Regional do Norte (CCDRN), Portugal, for his Ph.D. grant (NORTE-08-5369-FSE-000007). The authors acknowledge the support granted by national funds from FCT through project PTDC/BBB-NAN/ 3249/2014. This work was also financed by FEDER - Fundo Europeu de Desenvolvimento Regional funds through the COMPETE 2020 - Operacional Program for Competitiveness and Internationalisation (POCI), Portugal 2020, and by Portuguese funds through FCT - Fundac¸a˜o para a Cieˆncia e a Tecnologia/Ministe´rio da Cieˆncia, Tecnologia e Ensino Superior in the framework of the project “Institute for Research and Innovation in Health Sciences” (POCI-01-0145-FEDER-007274).

References [1] Breiner JM, Mark JE. Preparation, structure, growth mechanisms and properties of siloxane composites containing silica, titania or mixed silica-titania phases. Polymer 1998;39(22):5483e93. [2] Rong MZ, et al. Improvement of tensile properties of nano-SiO2/PP composites in relation to percolation mechanism. Polymer 2001;42(7):3301e4. [3] Shen S, et al. Nanocomposites for drug delivery. In: Nanotechnologies for the life Sciences. Wiley-VCH Verlag GmbH & Co. KGaA; 2007. [4] Lee SR, et al. Microstructure, tensile properties, and biodegradability of aliphatic polyester/clay nanocomposites. Polymer 2002;43(8):2495e500. [5] Teng Z, et al. Cationic beta-lactoglobulin nanoparticles as a bioavailability enhancer: effect of surface properties and size on the transport and delivery in vitro. Food Chem 2016;204:391e9. [6] Ensign LM, Cone R, Hanes J. Oral drug delivery with polymeric nanoparticles: the gastrointestinal mucus barriers. Adv Drug Deliv Rev 2012;64(6):557e70. [7] Tan L, et al. Chitosan-based core-shell nanomaterials for pH-triggered release of anticancer drug and near-infrared bioimaging. Carbohydr Polym 2017;157:325e34. [8] Ribeiro LN, et al. Bionanocomposites containing magnetic graphite as potential systems for drug delivery. Int J Pharm 2014;477(1e2):553e63. [9] Yadollahi M, Farhoudian S, Namazi H. One-pot synthesis of antibacterial chitosan/silver bionanocomposite hydrogel beads as drug delivery systems. Int J Biol Macromol 2015;79:37e43. [10] Jain S, Datta M. Montmorillonite-PLGA nanocomposites as an oral extended drug delivery vehicle for venlafaxine hydrochloride. Appl Clay Sci 2014;99:42e7. [11] Gogoi P, Boruah R, Dolui SK. Jatropha curcas oil based alkyd/epoxy/graphene oxide (GO) bionanocomposites: effect of GO on curing, mechanical and thermal properties. Prog Org Coating 2015;84:128e35. [12] Furlan M, et al. Preparation of biocompatible magnetiteePLGA composite nanoparticles using supercritical fluid extraction of emulsions. J Supercrit Fluids 2010;54(3):348e56. [13] Ribeiro LN, et al. Pectin-coated chitosan-LDH bionanocomposite beads as potential systems for colontargeted drug delivery. Int J Pharm 2014;463(1):1e9.

Organic nanocomposites for the delivery of bioactive molecules 489 [14] Weishaupt C, et al. T-cell distribution and adhesion receptor expression in metastatic melanoma. Clin Cancer Res 2007;13(9):2549e56. [15] Ramudo L, et al. ICAM-1 and CD11b/CD18 expression during acute pancreatitis induced by bilepancreatic duct obstruction: effect of N-acetylcysteine. Exp Biol Med 2007;232(6):737e43. [16] Eniola AO, Hammer DA. Artificial polymeric cells for targeted drug delivery. J Control Release 2003;87(1e3):15e22. [17] Jain S, Datta M. Oral extended release of dexamethasone: montmorillonite-PLGA nanocomposites as a delivery vehicle. Appl Clay Sci 2015;104:182e8. [18] Basarkar A, Singh J. Poly (lactide-co-glycolide)-polymethacrylate nanoparticles for intramuscular delivery of plasmid encoding interleukin-10 to prevent autoimmune diabetes in mice. Pharm Res 2009;26(1):72e81. [19] Benjaminsen RV, et al. The possible “proton sponge” effect of polyethylenimine (PEI) does not include change in lysosomal pH. Mol Ther 2013;21(1):149e57. [20] Farokhi M, et al. Bio-hybrid silk fibroin/calcium phosphate/PLGA nanocomposite scaffold to control the delivery of vascular endothelial growth factor. Mater Sci Eng C Mater Biol Appl 2014;35:401e10. [21] Borselli C, et al. Bioactivation of collagen matrices through sustained VEGF release from PLGA microspheres. J Biomed Mater Res A 2010;92a(1):94e102. [22] Gerber HP, et al. VEGF couples hypertrophic cartilage remodeling, ossification and angiogenesis during endochondral bone formation. Nat Med 1999;5(6):623e8. [23] Mayr-Wohlfart U, et al. Vascular endothelial growth factor stimulates chemotactic migration of primary human osteoblasts. Bone 2002;30(3):472e7. [24] Street J, et al. Vascular endothelial growth factor stimulates bone repair by promoting angiogenesis and bone turnover. Proc Natl Acad Sci USA 2002;99(15):9656e61. [25] Hulshoff JE, et al. Biological evaluation of the effect of magnetron sputtered Ca/P coatings on osteoblast-like cells in vitro. J Biomed Mater Res 1995;29(8):967e75. [26] Zhang Y, et al. The osteogenic properties of CaP/silk composite scaffolds. Biomaterials 2010;31(10):2848e56. [27] Zhang Y, et al. The effects of pore architecture in silk fibroin scaffolds on the growth and differentiation of mesenchymal stem cells expressing BMP7. Acta Biomater 2010;6(8):3021e8. [28] Borselli C, et al. Bioactivation of collagen matrices through sustained VEGF release from PLGA microspheres. J Biomed Mater Res A 2010;92(1):94e102. [29] Murphy WL, et al. Sustained release of vascular endothelial growth factor from mineralized poly(lactideco-glycolide) scaffolds for tissue engineering. Biomaterials 2000;21(24):2521e7. [30] Shokrgozar M, et al. Biocompatibility evaluation of HDPE-UHMWPE reinforced b-TCP nanocomposites using highly purified human osteoblast cells. J Biomed Mater Res A 2010;95(4):1074e83. [31] Farokhi M, et al. Porous crosslinked poly (ε-caprolactone fumarate)/nanohydroxyapatite composites for bone tissue engineering. J Biomed Mater Res A 2012;100(4):1051e60. [32] Midy V, Plouet J. Vasculotropin/vascular endothelial growth factor induces differentiation in cultured osteoblasts. Biochem Biophys Res Commun 1994;199(1):380e6. [33] Huang L, et al. Nano-cellulose 3D-networks as controlled-release drug carriers. J Mater Chem B 2013;1(23):2976e84. [34] Mohite BV, Suryawanshi RK, Patil SV. Study on the drug loading and release potential of bacterial cellulose. Cellul Chem Technol 2016;50(2):219e23. [35] Luo H, et al. Bacterial cellulose/graphene oxide nanocomposite as a novel drug delivery system. Curr Appl Phys 2017;17(2):249e54. [36] Zheng XT, Ma XQ, Li CM. Highly efficient nuclear delivery of anti-cancer drugs using a biofunctionalized reduced graphene oxide. J Colloid Interface Sci 2016;467:35e42. [37] Pertile RA, et al. Bacterial cellulose: long-term biocompatibility studies. J Biomater Sci Polym Ed 2012;23(10):1339e54.

490 Chapter 20 [38] Saska S, et al. Characterization and in vitro evaluation of bacterial cellulose membranes functionalized with osteogenic growth peptide for bone tissue engineering. J Mater Sci Mater Med 2012;23(9):2253e66. [39] Zhu YF, et al. Magnetic SBA-15/poly(N-isopropylacrylamide) composite: preparation, characterization and temperature-responsive drug release property. Microporous Mesoporous Mater 2009;123(1e3):107e12. [40] Lu W, et al. Full-color mechanical sensor based on elastic nanocomposite hydrogels encapsulated threedimensional colloidal arrays. Sensor Actuator B Chem 2016;234:527e33. [41] Liu Q, et al. Glucose-sensitive colorimetric sensor based on peroxidase mimics activity of porphyrinFe3O4 nanocomposites. Mater Sci Eng C Mater Biol Appl 2014;41:142e51. [42] Conti F, et al. Biomarkers for the early diagnosis of bacterial infection and the surveillance of hepatocellular carcinoma in cirrhosis. Biomark Med 2015;9(12):1343e51. [43] Srinivasan S, et al. Biocompatible alginate/nano bioactive glass ceramic composite scaffolds for periodontal tissue regeneration. Carbohydr Polym 2012;87(1):274e83. [44] Liu W, Webster TJ. 4 e toxicity and biocompatibility properties of nanocomposites for musculoskeletal tissue regeneration. In: Nanocomposites for musculoskeletal tissue regeneration. Oxford: Woodhead Publishing; 2016. p. 95e122. [45] Sun F, Zhou H, Lee J. Various preparation methods of highly porous hydroxyapatite/polymer nanoscale biocomposites for bone regeneration. Acta Biomater 2011;7(11):3813e28. [46] Yamamuro T, et al. Replacement of the lumbar vertebrae of sheep with ceramic prostheses. J Bone Joint Surg Br 1990;72(5):889e93. [47] Kokubo T, et al. Mechanical-properties of a new type of apatite-containing glass ceramic for prosthetic application. J Mater Sci 1985;20(6):2001e4. [48] Kozo O, et al. Quantitative study on osteoconduction of apatite-wollastonite containing glass ceramic granules, hydroxyapatite granules and alumina granules. Biomaterials 1990;11(4):265e71. [49] Juhasz JA, et al. Apatite-forming ability of glass-ceramic apatite-wollastonite - polyethylene composites: effect of filler content. J Mater Sci Mater Med 2003;14(6):489e95. [50] Juhasz JA, et al. Mechanical properties of glass-ceramic A-W-polyethylene composites: effect of filler content and particle size. Biomaterials 2004;25(6):949e55. [51] Kharaziha M, Fathi M, Edris H. Effects of surface modification on the mechanical and structural properties of nanofibrous poly (ε-caprolactone)/forsterite scaffold for tissue engineering applications. Mater Sci Eng C 2013;33(8):4512e9. [52] Kharaziha M, Fathi M. Synthesis and characterization of bioactive forsterite nanopowder. Ceram Int 2009;35(6):2449e54. [53] Kharaziha M, Fathi MH. Improvement of mechanical properties and biocompatibility of forsterite bioceramic addressed to bone tissue engineering materials. J Mech Behav Biomed Mater 2010;3(7):530e7. [54] Rezwan K, et al. Biodegradable and bioactive porous polymer/inorganic composite scaffolds for bone tissue engineering. Biomaterials 2006;27(18):3413e31. [55] Suchanek W, Yoshimura M. Processing and properties of hydroxyapatite-based biomaterials for use as hard tissue replacement implants. J Mater Res 1998;13(1):94e117. [56] Diba M, Fathi MH, Kharaziha M. Novel forsterite/polycaprolactone nanocomposite scaffold for tissue engineering applications. Mater Lett 2011;65(12):1931e4. [57] Rhee SH, Choi JY. Preparation of a bioactive poly(methyl methacrylate)/silica nanocomposite. J Am Ceram Soc 2002;85(5):1318e20. [58] Lewis G. Properties of acrylic bone cement: state of the art review. J Biomed Mater Res 1997;38(2):155e82. [59] Kango S, et al. Surface modification of inorganic nanoparticles for development of organic-inorganic nanocomposites-A review. Prog Polym Sci 2013;38(8):1232e61.

Organic nanocomposites for the delivery of bioactive molecules 491 [60] Poh PS, et al. In vitro and in vivo bone formation potential of surface calcium phosphate-coated polycaprolactone and polycaprolactone/bioactive glass composite scaffolds. Acta Biomater 2016;30:319e33. [61] Peter M, et al. Development of novel a-chitin/nanobioactive glass ceramic composite scaffolds for tissue engineering applications. Carbohydr Polym 2009;78(4):926e31. [62] Peter M, et al. Novel biodegradable chitosan-gelatin/nano-bioactive glass ceramic composite scaffolds for alveolar bone tissue engineering. Chem Eng J 2010;158(2):353e61. [63] Binulal NS, et al. Role of nanofibrous poly(caprolactone) scaffolds in human mesenchymal stem cell attachment and spreading for in vitro bone tissue engineering–response to osteogenic regulators. Tissue Eng 2010;16(2):393e404. [64] Kumar PS, et al. b-Chitin hydrogel/nano hydroxyapatite composite scaffolds for tissue engineering applications. Carbohydr Polym 2011;85(3):584e91. [65] Misra SK, et al. Effect of nanoparticulate bioactive glass particles on bioactivity and cytocompatibility of poly(3-hydroxybutyrate) composites. J R Soc Interface 2010;7(44):453e65. [66] Yang ZJ, et al. Osteogenesis in extraskeletally implanted porous calcium phosphate ceramics: variability among different kinds of animals. Biomaterials 1996;17(22):2131e7. [67] Ripamonti U. Osteoinduction in porous hydroxyapatite implanted in heterotopic sites of different animal models. Biomaterials 1996;17(1):31e5. [68] Nyman S, et al. New attachment following surgical treatment of human periodontal disease. J Clin Periodontol 1982;9(4):290e6. [69] Ivanovski S. Periodontal regeneration. Aust Dent J 2009;54(Suppl. 1):S118e28. [70] Reynolds MA, Aichelmann-Reidy ME, Branch-Mays GL. Regeneration of periodontal tissue: bone replacement grafts. Dent Clin N Am 2010;54(1):55e71. [71] Al-Naamani L, Dobretsov S, Dutta J. Chitosan-zinc oxide nanoparticle composite coating for active food packaging applications. Innov Food Sci Emerg Technol 2016;38:231e7. [72] Rhim JW, Park HM, Ha CS. Bio-nanocomposites for food packaging applications. Prog Polym Sci 2013;38(10e11):1629e52. [73] Sorrentino A, Gorrasi G, Vittoria V. Potential perspectives of bio-nanocomposites for food packaging applications. Trends Food Sci Technol 2007;18(2):84e95. [74] Arora A, Padua GW. Review: nanocomposites in food packaging. J Food Sci 2010;75(1):R43e9. [75] Ray SS, Okamoto M. Polymer/layered silicate nanocomposites: a review from preparation to processing. Prog Polym Sci 2003;28(11):1539e641. [76] Carrado KA. Synthetic organo- and polymer-clays: preparation, characterization, and materials applications. Appl Clay Sci 2000;17(1e2):1e23. [77] Torre L, et al. Processing and characterization of epoxy-anhydride-based intercalated nanocomposites. J Appl Polym Sci 2003;90(9):2532e9. [78] Sorrentino A, et al. Barrier properties of polymer/clay nanocomposites. In: Polymer nanocomposites; 2006. p. 273. [79] Yu YH, et al. Preparation and properties of poly(vinyl alcohol)-clay nanocomposite materials. Polymer 2003;44(12):3553e60. [80] Huang JY, Li X, Zhou WB. Safety assessment of nanocomposite for food packaging application. Trends Food Sci Technol 2015;45(2):187e99. [81] Cushen M, et al. Nanotechnologies in the food industry - recent developments, risks and regulation. Trends Food Sci Technol 2012;24(1):30e46. [82] Ghaderi M, et al. All-cellulose nanocomposite film made from bagasse cellulose nanofibers for food packaging application. Carbohydr Polym 2014;104:59e65. [83] Alexandre M, Dubois P. Polymer-layered silicate nanocomposites: preparation, properties and uses of a new class of materials. Mater Sci Eng R Rep 2000;28(1e2):1e63. [84] Schmidt D, Shah D, Giannelis EP. New advances in polymer/layered silicate nanocomposites. Curr Opin Solid State Mater Sci 2002;6(3):205e12.

492 Chapter 20 [85] Shi LE, et al. Synthesis, antibacterial activity, antibacterial mechanism and food applications of ZnO nanoparticles: a review. Food Addit Contam Part A Chem Anal Control Expo Risk Assess 2014;31(2):173e86. [86] Kong M, et al. Antimicrobial properties of chitosan and mode of action: a state of the art review. Int J Food Microbiol 2010;144(1):51e63. [87] Castro-Mayorga JL, et al. The impact of zinc oxide particle morphology as an antimicrobial and when incorporated in poly(3-hydroxybutyrate-co-3-hydroxyvalerate) films for food packaging and food contact surfaces applications. Food Bioprod Process 2017;101:32e44. [88] Singh B, et al. Controlled release of the fungicide thiram from starchealginateeclay based formulation. Appl Clay Sci 2009;45(1e2):76e82. [89] Fernandez-Perez M, et al. Use of activated bentonites in controlled-release formulations of atrazine. J Agric Food Chem 2004;52(12):3888e93. [90] Giroto AS, et al. Photoprotective effect of starch/montmorillonite composites on ultraviolet-induced degradation of herbicides. React Funct Polym 2015;93:156e62. [91] Liu P, et al. Photodegradation mechanism of deltamethrin and fenvalerate. J Environ Sci (China) 2010;22(7):1123e8. [92] Carr M, Wing R, Doane W. Encapsulation of atrazine within a starch matrix by extrusion processing. Cereal Chem 1991;68(3):262e6. [93] Grillo R, et al. Poly(epsilon-caprolactone)nanocapsules as carrier systems for herbicides: physicochemical characterization and genotoxicity evaluation. J Hazard Mater 2012;231e232:1e9. [94] Guan H, et al. Encapsulated ecdysone by internal gelation of alginate microspheres for controlling its release and photostability. Chem Eng J 2011;168(1):94e101. [95] Jerobin J, et al. Biodegradable polymer based encapsulation of neem oil nanoemulsion for controlled release of Aza-A. Carbohydr Polym 2012;90(4):1750e6. [96] Muro-Sun˜e´ N, et al. Predictive property models for use in design of controlled release of pesticides. Fluid Phase Equilib 2005;228e229:127e33. [97] Shogren RL, et al. Structure and morphology of baked starch foams. Polymer 1998;39(25):6649e55. [98] El-Nahhal Y, et al. Reduction of photodegradation and volatilization of herbicides in organo-clay formulations. Appl Clay Sci 1999;14(1e3):105e19. [99] Chiou BS, et al. Extruded starchenanoclay nanocomposites: effects of glycerol and nanoclay concentration. Polym Eng Sci 2007;47(11):1898e904. [100] Campos A, et al. The influence of UV-C irradiation on the properties of thermoplastic starch and polycaprolactone biocomposite with sisal bleached fibers. Polym Degrad Stabil 2012;97(10):1948e55. [101] Bertolini AC, et al. Photodegradation of cassava and corn starches. J Agric Food Chem 2001;49(2):675e82. [102] Aga D, et al. Identification of a new sulfonic acid metabolite of metolachlor in soil. Environ Sci Technol 1996;30(2):592e7. [103] Graham WH, et al. Metolachlor and alachlor breakdown product formation patterns in aquatic field mesocosms. Environ Sci Technol 1999;33(24):4471e6. [104] Jin D, et al. A derivative photoelectrochemical sensing platform for herbicide acetochlor based on TiO(2)-poly (3-hexylthiophene)-ionic liquid nanocomposite film modified electrodes. Talanta 2014;127:169e74. [105] Linsebigler AL, Lu GQ, Yates JT. Photocatalysis on Tio2 surfaces - principles, mechanisms, and selected results. Chem Rev 1995;95(3):735e58. [106] Li HB, et al. Poly(3-hexylthiophene)/TiO2 nanoparticle-functionalized electrodes for visible light and low potential photoelectrochemical sensing of organophosphorus pesticide chlopyrifos. Anal Chem 2011;83(24):9681e6. ´ lvaro M, et al. Screening of an ionic liquid as medium for photochemical reactions. Chem Phys Lett [107] A 2002;362(5e6):435e40.

Organic nanocomposites for the delivery of bioactive molecules 493 [108] Yang J, et al. Synthesis of Fe3O4/g-C3N4 nanocomposites and their application in the photodegradation of 2,4,6-trichlorophenol under visible light. Mater Lett 2016;164:183e9. [109] Rengaraj S, Li XZ. Enhanced photocatalytic activity of TiO2 by doping with Ag for degradation of 2,4,6-trichlorophenol in aqueous suspension. J Mol Catal A Chem 2006;243(1):60e7. [110] Jiang D, et al. Highly efficient heterojunction photocatalyst based on nanoporous g-C3N4 sheets modified by Ag3PO4 nanoparticles: synthesis and enhanced photocatalytic activity. J Colloid Interface Sci 2014;417:115e20. [111] Cao S, et al. Polymeric photocatalysts based on graphitic carbon nitride. Adv Mater 2015;27(13):2150e76. [112] Ali A, Ahmed S. A review on chitosan and its nanocomposites in drug delivery. Int J Biol Macromol 2018;109:273e86. [113] Elgadir MA, et al. Impact of chitosan composites and chitosan nanoparticle composites on various drug delivery systems: a review. J Food Drug Anal 2015;23(4):619e29. [114] Wood L. Global nanotechnology market 2018-2024: market is expected to exceed US$ 125 billion. 2018. Available from: https://www.prnewswire.com/news-releases/global-nanotechnology-market-20182024-market-is-expected-to-exceed-us-125-billion-300641054.html. [115] Lopez-Cuesta JM, Longuet C, Chivas-Joly C. 13 e thermal degradation, flammability, and potential toxicity of polymer nanocomposites. In: Health and environmental safety of nanomaterials. Woodhead Publishing; 2014. p. 278e310. [116] Environmental N. Health and safety research needs for engineered nanoscale materials. USA: National Nanotechnology Initiative (NNI); 2006. [117] Nohynek G, Dufour E, Roberts MS. Nanotechnology, cosmetics and the skin: is there a health risk? Skin Pharmacol Physiol 2008;21(3):136e49. [118] Jones CF, Grainger DW. In vitro assessments of nanomaterial toxicity. Adv Drug Deliv Rev 2009;61(6):438e56. [119] Grainger DW, Castner DG. Nanobiomaterials and nanoanalysis: opportunities for improving the science to benefit biomedical technologies. Adv Mater 2008;20(5):867e77. [120] Pandey RK, Prajapati VK. Molecular and immunological toxic effects of nanoparticles. Int J Biol Macromol 2018;107(Pt A):1278e93. [121] Li H, et al. Cutting edge: inflammasome activation by alum and alum’s adjuvant effect are mediated by NLRP3. J Immunol 2008;181(1):17e21. [122] Kuroda E, et al. Silica crystals and aluminum salts regulate the production of prostaglandin in macrophages via NALP3 inflammasome-independent mechanisms. Immunity 2011;34(4):514e26. [123] Vallhov H, et al. The importance of an endotoxin-free environment during the production of nanoparticles used in medical applications. Nano Letters 2006;6(8):1682e6. [124] Amenta V, et al. Regulatory aspects of nanotechnology in the agri/feed/food sector in EU and non-EU countries. Regul Toxicol Pharmacol 2015;73(1):463e76. [125] Arts JH, et al. A critical appraisal of existing concepts for the grouping of nanomaterials. Regul Toxicol Pharmacol 2014;70(2):492e506. [126] OECD. Guidance on grouping of chemicals. Series on testing and assessment. 2014. Available from: http://www.oecd.org/chemicalsafety/risk-assessment/groupingofchemicalschemicalcategoriesandreadacross.htm. [127] Hagens WI, et al. What do we (need to) know about the kinetic properties of nanoparticles in the body? Regul Toxicol Pharmacol 2007;49(3):217e29. [128] Parliament E. Regulation (EU) 1169/2011 of the European Parliament and of the council of 25 October 2011 on the provision of food information to consumers. Off J Eur Communities L 2011;304:18.

C H A P T E R 21

Natural polysaccharides for growth factors delivery Sneha S. Rao1, P.D. Rekha1, Sukumaran Anil2, Baboucarr Lowe3, Jayachandran Venkatesan1 1

Yenepoya Research Centre, Yenepoya (Deemed to be University), Mangalore, Karnataka, India; Department of Dentistry, Hamad Medical Corporation, Doha, Qatar; 3School of Dentistry, The University of Queensland, Brisbane, QLD, Australia

2

Chapter Outline List of abbreviations 495 1. Introduction 496 2. Structure and properties of polysaccharides 496 3. Classification of polysaccharides 497 4. Role of polysaccharides in drug delivery 498 5. Growth factors and their importance 498 6. Important polysaccharides for growth factors delivery 499 6.1 6.2 6.3 6.4 6.5 6.6 6.7

Alginate 499 Chitin/chitosan 499 Hyaluronic acid 501 Cellulose 503 Fucoidan 503 Carrageenan 505 Ulvan 507

7. Conclusions 508 Acknowledgments 508 References 508

List of abbreviations bFGF BMP-2 ECM EGF IL-1 PDGF

Basic fibroblast growth factor Bone morphogenic protein-2 Extracellular matrix Epidermal growth factor Interleukin-1 Platelet derived growth factor

Natural Polysaccharides in Drug Delivery and Biomedical Applications. https://doi.org/10.1016/B978-0-12-817055-7.00021-2 Copyright © 2019 Elsevier Inc. All rights reserved.

495

496 Chapter 21 PDLLA PLGA PRP SDF-1 SEM TGF-b VEGF

Poly DL-lactic acid poly (D,L-lactic acid-co-glycolic acid) Platelets rich plasma Stromal cell derived factor-1 Scanning electron microscopy Transforming growth factor-b Vascular endothelial growth factor

1. Introduction Natural polysaccharides are polymeric materials which have been in use through the centuries owing to their exceptional properties. They are biological macromolecules composed of one or more monosaccharides connected together to yield a linear or a branched polysaccharide. They are biocompatible, biodegradable, and possess excellent mechanical strength and drug releasing capacity making them accessible for an extensive array of applications in biomedical, pharmaceutical, and cosmetic industries [1]. They are capable of interacting with different cellular components and can bring about cellular migration, proliferation, and cellular attachment [2]. Current day studies show that these polysaccharides also hold antitumor [3], antioxidant [4], antiinflammatory [5], antiHIV [6], and chelating properties [7]. They are obtained by biosynthesis by plants, animals, and microorganisms. Some of the most popular polysaccharides include alginate, chitin, chitosan, cellulose, hyaluronic acid, fucoidan, carrageenan, dextran, etc. These polysaccharides can be easily isolated from the resources in economical ways and find numerous potential biomedical uses including tissue engineering, drug delivery, and biosensors [8].

2. Structure and properties of polysaccharides Polysaccharides are macromolecules comprising of a large number of monosaccharide residues linked to each other through glycosidic linkages. They are termed as homopolysaccharides or homoglucans if they are composed of only one type of the monosaccharides and heteropolysaccharides or heteroglucans if they are composed of more than one type of the monosaccharide units. Fig. 21.1. shows the structure of alginate and carrageenan. Chemical component and substitute of the polysaccharide mainly depends on geographical area and climate condition. The following are the properties of most of the polysaccharide materials: • • • •

Biocompatible Biodegradable Scaffold and film forming ability Nanoparticle formation

Natural polysaccharides for growth factors delivery

497

Figure 21.1 (A) Chemical structure of alginate and (B) different forms of carrageenan (kappa, iota, and lambda). Figure adapted with permission from de Jesus Raposo M, de Morais A, de Morais R. Marine polysaccharides from algae with potential biomedical applications. Mar Drugs 2015;13(5):2967e3028.

• • • •

Less toxic to normal cells Drug and growth factor holding capacity Inexpensive, abundant, and easily available Act as ECM for tissue development.

3. Classification of polysaccharides The polysaccharides are mainly categorized based on their source, structure, and charge. Based on the source of origin they are classified as of plant, animal, microbial, and marine origin [9,10]. The polysaccharides like starch, cellulose, pectin, amylose, and gum are from plant sources while, chitosan, hyaluronic acid, chitin, and chondroitin sulfate are of animal origin; xanthan, dextran, and pullulan are of microbial origin and marine origin polysaccharides include fucoidan, carrageenan, and alginate. Based on their structure they are classified into linear polysaccharides like amylose, cellulose, and pectin and branched like amylopectin, xanthan gum, etc. Based on charge they are classified into neutral like amylose, pectin, cellulose, etc., and anionic polysaccharides like alginates, carrageenan, xanthan, etc.

498 Chapter 21

4. Role of polysaccharides in drug delivery Noteworthy research has been carried out on naturally derived polysaccharides as a drug delivery system [11e14]. These polysaccharides can be converted into scaffolds, hydrogels, nanoparticles, films using different methods such as covalent cross-linking, ionic gelation method, electrospinning, and self-assembly to get a desirable shape [11]. These polysaccharides are widely utilized as a drug delivery system in cancer drug delivery [15], protein delivery [16], growth factor delivery [17,18], antibiotic delivery [19], and insulin delivery [20]. Polysaccharides have advantages in delivering proteins and growth factors in a sustainable way and hold the protein and growth factor for several days.

5. Growth factors and their importance Growth factors are protein molecules which can mediate in cell proliferation, differentiation, migration, etc., resulting in the complete growth of the cell [21]. They are required for the occurrence of different body functions. Platelets are an abundant source of growth factors and they coordinate events like immune response, angiogenesis, and wound healing. The growth factors are encapsulated within the a-granules of the platelets. On activation the growth factors, contained within the a-granules of the platelets are released, to bring about cellular proliferation, migration, cell signaling and adhesion, wound debridement, and angiogenesis [22]. Cell-based therapies have shown promising outcomes; use of Platelets Rich Plasma (PRP) has overcome the hitches associated with rejections, has exhibited enhanced wound closure, and is economic. PRP is a rich source of growth factors as it contains concentrated platelet numbers, usually six-fold more than baselines [23]. The growth factors include PDGF, VEGF, EGF, VEGF, FGF, TGF-b, and IL-1 [24]. Randomized control trial screened the efficacy of the PRP gel against standard care. The wound closure was faster in the group receiving the PRP gel compared to the standard treatments. Recent treatment strategies aim to supply the growth factors into the wound site. The efficiency of the growth factors is enhanced using natural biopolymers or polysaccharides. Different studies show that biopolymers can regulate the growth of cells leading to three-dimensional tissue regeneration. These properties of the polysaccharides and the growth factors have received great attention and their successful in combination with different polysaccharides have been applied in various tissue engineering applications. Rossi et al. [25] used freeze-dried chitosan glutamate and sodium hyaluronate for supply of platelets to chronic tissue wounds. This exhibited a sustained release of growth factors from the platelets.

Natural polysaccharides for growth factors delivery

499

6. Important polysaccharides for growth factors delivery 6.1 Alginate Alginate is an anionic polysaccharide and mostly isolated from the cell walls of brown algae (Laminaria, Macrocystis, Ascophyllum, Ecklonia, Sargassum, etc.). Alginic acid is a linear anionic polysaccharide substance and mainly composed of mannuronic acid and guluronic acid units. The chemical structure of alginates is simplified into two monomer units M-block and G-block, which increases its demand for commercial purposes [26]. They are widely used in cosmetic, drug delivery, tissue engineering, pharmaceutical, and food industries owing to properties such as highly biocompatibility, nontoxicity, ability to retain water to form gels, and easy interaction with cellular components [27]. Pawar et al. in 2011, studied the properties of the alginate-based hydrogels which were suitable for the delivery of bioactive molecules like nucleic acids and proteins [28]. Alginates are comprehensively being used as additives [26]. Alginates are being used to treat bone defects; Kolambkar et al. [29] utilized alginatebased scaffolds for the delivering of osteogenic signal molecules. Here they introduced the peptide-modified alginate hydrogel into an electrospun nanofiber mesh tube which showed continuous growth factor release and exhibited bone regeneration as shown in (Fig. 21.2). Gu et al. [30] used alginate beads for the delivery of VEGF to promote angiogenesis. The calcium alginate beads were encapsulated with VEGF by external gelation method and were developed for the treatment of limb ischemia and myocardial ischemia. In another study, Wang et al. [31] used chemically cross-linked and highly porous alginate-based scaffolds for the growth factors delivery. The fabricated alginate scaffolds were biodegradable and used for minimally invasive surgical applications. Rabbany et al. [32] used an alginate-based hydrogel for delivering the naturally occurring cytokine SDF-1 for treating pressure ulcers. Study outcomes showed significant wound closure in the alginatebased hydrogel containing SDF-1.

6.2 Chitin/chitosan Chitin is predominantly found in the exoskeletons of the marine crustaceans like shrimps and lobsters and in exoskeletons of different insects. It is the most abundant material next to cellulose. Chitosan is a derivative of chitin and it is a cationic polysaccharide mainly composed of two units, namely glucosamine and N-acetyl glucosamine with [1e4] glycosidic bonds [33]. They also possess chemotactic, wound healing, proliferative, antimicrobial, antitumor activities [34e36]. They are biocompatible, biodegradable, and

Figure 21.2 (I) (A) SEM images of alginate hydrogel and nanofiber mesh tubes for surgery. (B) Hollow tubular implant without perforation (C) with perforation (D) segmental bone defect for implantation for alginate hydrogels, with and without BMP-2 (E) defect picture after implantation and perforated mesh tube (F) after 1-week period (G) alginate release kinetics 21 days at in vitro condition and sustainable release of rhBMP-2. (II) (A) mCT analysis of bone regeneration at 4 and 12 weeks (B) Quantification of regenerated bone volume and (C) Local density of regenerated bone. Adapted with permission from Kolambkar YM, Dupont KM, Boerckel JD, Huebsch N, Mooney DJ, Hutmacher DW, et al. An alginate-based hybrid system for growth factor delivery in the functional repair of large bone defects. Biomaterials 2011;32(1):65e74.

Natural polysaccharides for growth factors delivery

501

less toxic and can be easily utilized to produce scaffolds, gels, nanoparticles, and nanofibers, therefore, making it an important material in biomedical applications [37]. Murakami et al. [38] prepared an alginateechitin/chitosanefucoidanebased hydrogel. These were used for retaining the moisture in the wound bed and showed better healing outcomes in impaired wounds. Chitin/chitosan is widely being used in different bone healing applications. Yokoyama et al. [39] developed cement composed of calcium phosphate using chitin and citric acid, and the cement exhibited good mechanical and biocompatible properties and thus could be used as a prospective bone substitute material. BMP-2 growth factor plays an vital role in bone tissue repair and regeneration [40,41]. Kim et al. [42] developed a porous chitosan scaffold holding microspheres containing TGF-b1. These scaffolds showed continual release of the growth factors stimulating the proliferation and differentiation of chondrocytes, making it potent for treating cartilage regenerative problems as highlighted in (Fig. 21.3). Alemdaroglu et al. [43] developed a chitosan gel blended with epidermal growth factors. They were used for treating second degree burns in rats and results show better proliferation in treatment groups when compared to control groups. The utilization of polysaccharides in growth factor delivery helps in sustainable release and maintenance of growth factor bioactivity by protection from degradation. Mizuno et al. [44] used chitosan films for the sustained release of bFGF for enhancing the growth and differentiation of different cell types in the wound bed. Therefore the chitosan film containing the growth factors fasten healing rates in wounds in diabetic mice. Lee et al. [45] evaluated release of PDGF from a combination of chitosan and tricalcium phosphate sponge system and shows the sustainable release of PDGF and also exhibited osteogenic effect on bone restoration at in-vivo condition.

6.3 Hyaluronic acid Hyaluronic acid (HA) is a linear polysaccharide, and mainly composed of N-acetyl-Dglucosamine and glucuronic acid unit. It possesses good viscoelastic properties and high molecular mass. It is naturally occurring in the biological fluids and is fundamental for many cellular and tissue processes [46]. It can be used to develop viscoelastic gels, soft/ stiff hydrogels, porous sponges, etc. The cross-linking of these polysaccharides ensures the encapsulation of the cells with sustained release. Kurisawa et al. synthesized biodegradable and injectable hydrogels consisting of HA-tyramine conjugate for drug delivery applications [47]. Ekaputra et al. [48] prepared the scaffold with a combination of HA and poly (ε-caprolactone)/collagen fibers to form a hydrogel. These electrospun fibers exhibit characteristics similar to the ECM and facilitate differentiation of different cell typesdfibroblasts, keratinocytes, and osteoblasts.

502 Chapter 21

Figure 21.3 (A) SEM images of chitosan microspheres developed in the absence of FITC-BSA and (B) presence of FITC-BSA as a reference protein. (C) SEM images of chitosan scaffold with chondrocytes containing TGF-b1 after 4 days. (D) High magnified view (E) after 7 days. Adapted with permission from Kim SE, Park JH, Cho YW, Chung H, Jeong SY, Lee EB, et al. Porous chitosan scaffold containing microspheres loaded with transforming growth factor-b1: implications for cartilage tissue engineering. J Control Release 2003;91(3):365e74.

HA-based materials like scaffolds and gels have been designed to deliver growth factor for various tissue engineering purposes. Lisignoli et al. [49] synthesized a nonwoven hyaluronic acidebased scaffold containing bone marrow stromal cells, and in the presence of bFGF they observed that differentiation and the mineralization of the rat bone marrow stromal cells were increased. This study shows the biocompatibility of HA with the bone marrow stromal cells and its ability to promote differentiation taking up the bFGF from the media. A study by Yoo et al. [50] confirmed the role of HA in enhancing adhesion, proliferation, and differentiation of chondrocytes for cartilage healing, and they have developed a macroporous biodegradable scaffold PLGA on which the HA was immobilized. The outcome of the study shows the HA-PLGA scaffolds augmented the

Natural polysaccharides for growth factors delivery

503

formation of collagen type II, therefore allowing the cartilage tissue development. Liu et al. [51] designed hyaluronate-heparin conjugate gels. The recombinant bFGFe2 bound to the heparin regions of the gel and was released in vitro from the gels. The functional analysis of the gel showed increased stability and growth factor release from the gels. In Fig. 21.4, the combination of hydrogel with BMP-2 yields better bone regeneration as compared to control and absence of BMP-2 protein.

6.4 Cellulose Cellulose is the most ample polysaccharide which is isolated from the secondary cell walls of the plant cell, accounting for up to 20%e30% of its dry weight. It is known for its long-term energy storing abilities. Cellulose is obtained by extraction from the cell wall using chelating and alkali agents and mainly composed of a linear b-4-linked D-glucan [53]. Cellulose can also be isolated from bacterial source [54]. The semisynthetic derivatives of cellulose are extensively used in the cosmetic and pharmaceutical fields, and they are exclusively used as coating materials, in extended release of growth factors, as matrix materials, thickeners, and adhesives [55]. Cellulose derivatives are extensively being used as biomaterials for several applications like dialysis membranes and biosensors. Entcheva et al. [56] showed scaffolds composed of cellulose acetate (CA) and regenerated cellulose (RC) could be used to make cardiac cell constructs in vitro, and these scaffolds stimulated cell growth and increased cell connectivity. Cellulose has been extensively used as a growth factor drug delivery system for tissue engineering applications. Santa-Comba et al. [57] used cellulose-based carriers for the delivery of bone morphogenic proteins (rhBMP-2) to induce osteogenesis. The use of rhBMP-2 in combination with carboxymethyl cellulose considerably increased the alkaline phosphatase (ALP) activity indicating it to be a potential absorbent and bring about release of rhBMP-2 in vitro. Li et al. [58] designed a collagen/cellulose nanocrystal scaffold loaded with bFGF. The scaffold was biocompatible, ensured the sustained release of the bFGF, enhancing the cell proliferation both in in vitro and in vivo conditions.

6.5 Fucoidan Fucoidan, a sulfated polysaccharide, is commonly observed in brown algae at minimum level and mainly composed of fucose and sulfate forms [59]. Hong et al. [60] observed antioxidant effect of fucoidandshowed protective effect against the acetaminopheninduced lung damage by increasing the production of glutathione, superoxide dismutase, and glutathione peroxide. They can easily interact with different macromolecules in the biological system and bring about cellular adhesion, proliferation, and migration. This adds to the therapeutic benefits and can be utilized to develop effective wound care products [61].

504 Chapter 21

Figure 21.4 Implantation of hydrogels in muscle of the rat hind limb at in vivo condition (A) A photograph shows the surgical insertion of a PCL-TCP tube containing BMP-2 (B) after 8 weeks post implantation (X-ray image) shows the bone formation in the presence of BMP-2 (dotted lines). (C) Corresponding digital and X-ray images of hydrogels and histological observations. Figure adapted with permission from Bhakta G, Rai B, Lim ZX, Hui JH, Stein GS, van Wijnen AJ, et al. Hyaluronic acidbased hydrogels functionalized with heparin that support controlled release of bioactive BMP-2. Biomaterials 2012;33(26):6113e22.

Natural polysaccharides for growth factors delivery

505

Aisa et al. [62] shows the inhibition in proliferation and induced apoptosis through the mitochondrial pathway of human lymphoma HS-sultan cell lines using fucoidan. This reflected the anticancerous properties possessed by fucoidan. Synytsya et al. [63] isolated and characterized the fucoidan from Undaria pinnatifida and showed the antitumor activity against HeLa, PC-3, A549, HepG2 cells. Venkatesan et al. [64] developed a polymer-based scaffold consisting of chitosan-alginate with fucoidan for bone tissue repair and regeneration applications. The scaffold shows excellent water uptake and retention abilities and high mineralization abilities which can be potentially used as bone graft substitute. Fucoidan is popularly known for different properties with biological importance, and it has been used in the field of tissue engineering due to its ability to encapsulate growth factors and induce proliferation and migration. Huang et al. [65] prepared the combination of chitosan, tripolyphosphate, and fucoidan in the form of nanoparticles for SDF-1 delivery applications. The study outcomes saw the migration and enhanced the expression of PI3K under the influence of the released SDF-1. Purnama et al. [66] prepared a fucoidan-based 3D scaffold combined with VEGF. This promoted neovascularization in mice and could be a prospective agent to treat ischemia by enhancing angiogenesis as shown in (Fig. 21.5). Certain studies show that the interaction of growth factors with the polysaccharides increases their half-life. Nakamura et al. [67] developed a chitosan/fucoidan-based hydrogel; FGF-2 on interaction with the hydrogel had prolonged biological half-life and results indicated the slow release of FGF-2 from the hydrogel and could be useful in treating ischemic conditions.

6.6 Carrageenan Carrageenan is a linear sulfated polysaccharide found in edible red seaweeds and it is water soluble. It is a linear polysaccharide with repeating structures of altering galactose and 3,6 anhydrogalactose units. It is said to be nontoxic, biodegradable, biocompatible and so has gained huge attention in the field of medicine, pharmaceuticals, cosmetics, and the food industries [68]. Due to their strong negative charge, controlled drug release, and gel forming abilities, they are extensively used for drug delivery [69]. Carrageenan is extensively being used for growth factor delivery. Kurtz et al. [70] developed a blend of carrageenan with insulinlike growth factor. The results showed that the growth factors from the blends were sustainable. The rats recovered from the Achilles tendon injury faster on treatment with the blend. Studies have shown that the carrageenanbased hydrogels show persistent release of growth factors. Santo et al. developed the hydrogel with carrageenan with the combination of PDGF-BB and beads show the

Figure 21.5 After 2-week implantation of microporous scaffolds control, medium molecular weight fucoidan (MMWF), VEGF, and MMWF þ VEGF and histological examination was done on hematoxylin/ eosin-stained sections at different magnifications. Results were expressed as neovessels area (mm2) and density (N/mm2). Figure adapted with permission from Purnama A, Aid-Launais R, Haddad O, Maire M, Mantovani D, Letourneur D, et al. Fucoidan in a 3D scaffold interacts with vascular endothelial growth factor and promotes neovascularization in mice. Drug Deliv Transl Res 2015;5(2):187e97.

Natural polysaccharides for growth factors delivery

507

Figure 21.6 Beads containing 2% carrageenan with 0.2% OVA-FITC in 5% KCl 60 min (fluorescence stereomicroscopy images): (A) raw beads before release of growth factor (B) after release of growth factor in 2 days. Figure adapted with permission from Santo VE, Frias AM, Carida M, Cancedda R, Gomes ME, Mano JF, et al. Carrageenan-based hydrogels for the controlled delivery of PDGF-BB in bone tissue engineering applications. Biomacromolecules 2009;10(6):1392e401.

controlled release of growth factors for the development of vascular network [71] (Fig. 21.6). Hydrogel based on carrageenan have been utilized for the delivery of injectable thermoresponsive formulations for cartilage healing. Rocha et al. [72] developed a carrageenan-based hydrogel with adipose derived stem cells and TGF-b-1 encapsulated within. The study outcomes showed the differentiation of human adipose derived stem cells to cartilage lineage which makes the development of cartilage.

6.7 Ulvan Ulvan are sulfated polysaccharides that are mainly obtained from green algae Ulva species and Enteromorpha [73]. It is extracted by water solutions using ammonium oxalate as a divalent cation at 80e90 C temperature [74], and it is mainly composed of sulfate, rhamnose, xylose, and glucuronic acid units. They are water soluble and have good rheological and biochemical properties making them a novel compound with potential applications [75]. It is a fairly studied biomaterial at the moment in tissue engineering application, and is an excellent biocompatible and biologically active substance. Dash et al. [76] developed a UV-cross-linked ulvan scaffold which was intended for the enzyme mediated apatite mineral formation. These scaffolds enhanced the mineralization in MC3T3 cells. In another study they developed a novel ulvan-based 3D scaffold enriched with PDLLA. The study results showed that they have excellent physiochemical and cytocompatible properties which can be applied in bone tissue repair and regeneration applications [77]. Alves et al. [78] designed a 3D degradable, porous ulvan structure. These structures had good mechanical strength and water uptake ability. They also showed enhanced cell proliferation abilities making them suitable for several biological and biomedical applications.

508 Chapter 21

7. Conclusions The development of combination of biocompatible and biodegradable polysaccharide, where bioactive molecules like growth factors can be accommodated for sustained release is gaining considerable attention. These combinations will provide greater advantages in tissue regeneration and also meet several requirements such as sustainable release and protect the growth factor bioactivity for longer time, which will enable better cell proliferation, migration, and adhesion of the cells. The polysaccharides are easy to isolate and inexpensive and combine with the growth factors in an effective and economic way. Different studies show that the combination of polysaccharides and growth factors shows better results in terms of stem cell differentiation toward osteogenic, chondrogenic lineages. The results from various literature ensure better tissue construction and healing capacity which can be potentially useful in tissue engineering, cosmetics, and other biomedical applications.

Acknowledgments Ms. Sneha Rao and Dr. J Venkatesan thanks to YRC, Yenepoya (Deemed to be University) for support to write this book chapter. Baboucarr Lowe acknowledges funding received from the Australian Government Research Training Program, The University of Queensland.

References [1] Rinaudo M. Main properties and current applications of some polysaccharides as biomaterials. Polym Int 2008;57(3):397e430. [2] Mann BK, West JL. Cell adhesion peptides alter smooth muscle cell adhesion, proliferation, migration, and matrix protein synthesis on modified surfaces and in polymer scaffolds. J Biomed Mater Res 2002;60(1):86e93. [3] Wasser S. Medicinal mushrooms as a source of antitumor and immunomodulating polysaccharides. Appl Microbiol Biotechnol 2002;60(3):258e74. [4] Ping CX, Yan C, Bing LS, Guo CY, Yun LJ, Ping LL. Free radical scavenging of Ganoderma lucidum polysaccharides and its effect on antioxidant enzymes and immunity activities in cervical carcinoma rats. Carbohydr Polym 2009;77(2):389e93. [5] Matsui MS, Muizzuddin N, Arad S, Marenus K. Sulfated polysaccharides from red microalgae have antiinflammatory properties in vitro and in vivo. Appl Biochem Biotechnol 2003;104(1):13e22. [6] Be´ress A, Wassermann O, Bruhn T, Be´ress L, Kraiselburd EN, Gonzalez LV, et al. A new procedure for the isolation of anti-HIV compounds (polysaccharides and polyphenols) from the marine alga Fucus vesiculosus. J Nat Prod 1993;56(4):478e88. [7] Crini G. Recent developments in polysaccharide-based materials used as adsorbents in wastewater treatment. Prog Polym Sci 2005;30(1):38e70. [8] de Jesus Raposo M, de Morais A, de Morais R. Marine polysaccharides from algae with potential biomedical applications. Mar Drugs 2015;13(5):2967e3028. [9] Prajapati VD, Jani GK, Zala BS, Khutliwala TA. An insight into the emerging exopolysaccharide gellan gum as a novel polymer. Carbohydr Polym 2013;93(2):670e8.

Natural polysaccharides for growth factors delivery

509

[10] Ruocco N, Costantini S, Guariniello S, Costantini M. Polysaccharides from the marine environment with pharmacological, cosmeceutical and nutraceutical potential. Molecules 2016;21(5):551. [11] Liu Z, Jiao Y, Wang Y, Zhou C, Zhang Z. Polysaccharides-based nanoparticles as drug delivery systems. Adv Drug Deliv Rev 2008;60(15):1650e62. [12] Marchessault RH, Ravenelle F, Zhu XX. Polysaccharides for drug delivery and pharmaceutical applications. Amer Chemical Society; 2006. [13] Luo Y, Wang Q. Recent development of chitosan-based polyelectrolyte complexes with natural polysaccharides for drug delivery. Int J Biol Macromol 2014;64:353e67. [14] Sinha VR, Kumria R. Polysaccharides in colon-specific drug delivery. Int J Pharm 2001;224(1e2):19e38. [15] Na K, Lee TB, Park K-H, Shin E-K, Lee Y-B, Choi H-K. Self-assembled nanoparticles of hydrophobically-modified polysaccharide bearing vitamin H as a targeted anti-cancer drug delivery system. Eur J Pharm Sci 2003;18(2):165e73. [16] Vermonden T, Censi R, Hennink WE. Hydrogels for protein delivery. Chem Rev 2012;112(5):2853e88. [17] Zhang L, Furst EM, Kiick KL. Manipulation of hydrogel assembly and growth factor delivery via the use of peptideepolysaccharide interactions. J Control Release 2006;114(2):130e42. [18] Lee K, Silva EA, Mooney DJ. Growth factor delivery-based tissue engineering: general approaches and a review of recent developments. J R Soc Interface 2010;8(55):153e70. [19] Zhao Y, Zhang X, Wang Y, Wu Z, An J, Lu Z, et al. In situ cross-linked polysaccharide hydrogel as extracellular matrix mimics for antibiotics delivery. Carbohydr Polym 2014;105:63e9. [20] Sonia TA, Sharma CP. An overview of natural polymers for oral insulin delivery. Drug Discov Today 2012;17(13e14):784e92. [21] Barrientos S, Stojadinovic O, Golinko MS, Brem H, Tomic-Canic M. Growth factors and cytokines in wound healing. Wound Repair Regen 2008;16(5):585e601. [22] Spruit MA, Singh SJ, Garvey C, ZuWallack R, Nici L, Rochester C, et al. An official American Thoracic Society/European Respiratory Society statement: key concepts and advances in pulmonary rehabilitation. Am J Respir Crit Care Med 2013;188(8):e13e64. [23] Everts PA, Knape JT, Weibrich G, Scho¨nberger JP, Hoffmann J, Overdevest EP, et al. Platelet-rich plasma and platelet gel: a review. J Extra-Corporeal Technol 2006;38(2):174. [24] Savi P, Pereillo J, Uzabiaga M, Combalbert J, Picard C, Maffrand J, et al. Identification and biological activity of the active metabolite of clopidogrel. Thromb Haemostasis 2000;84(05):891e6. [25] Rossi S, Faccendini A, Bonferoni M, Ferrari F, Sandri G, Del Fante C, et al. “Sponge-like” dressings based on biopolymers for the delivery of platelet lysate to skin chronic wounds. Int J Pharm 2013;440(2):207e15. [26] Nalamothu N, Potluri A, Muppalla MB. Review on marine alginates and its applications. J Pharm Res 2014;4(10). [27] Lee KY, Mooney DJ. Alginate: properties and biomedical applications. Prog Polym Sci 2012;37(1):106e26. [28] Pawar SN, Edgar KJ. Alginate derivatization: a review of chemistry, properties and applications. Biomaterials 2012;33(11):3279e305. [29] Kolambkar YM, Dupont KM, Boerckel JD, Huebsch N, Mooney DJ, Hutmacher DW, et al. An alginatebased hybrid system for growth factor delivery in the functional repair of large bone defects. Biomaterials 2011;32(1):65e74. [30] Gu F, Amsden B, Neufeld R. Sustained delivery of vascular endothelial growth factor with alginate beads. J Control Release 2004;96(3):463e72. [31] Wang L, Shansky J, Borselli C, Mooney D, Vandenburgh H. Design and fabrication of a biodegradable, covalently crosslinked shape-memory alginate scaffold for cell and growth factor delivery. Tissue Eng 2012;18(19e20):2000e7. [32] Rabbany SY, Pastore J, Yamamoto M, Miller T, Rafii S, Aras R, et al. Continuous delivery of stromal cell-derived factor-1 from alginate scaffolds accelerates wound healing. Cell Transplant 2010;19(4):399e408.

510 Chapter 21 [33] [34] [35] [36] [37] [38]

[39] [40] [41]

[42]

[43]

[44]

[45]

[46] [47]

[48]

[49]

[50] [51] [52]

[53]

Koide S. Chitin-chitosan: properties, benefits and risks. Nutr Res (NY) 1998;18(6):1091e101. Rinaudo M. Chitin and chitosan: properties and applications. Prog Polym Sci 2006;31(7):603e32. Kumar MNVR. A review of chitin and chitosan applications. React Funct Polym 2000;46(1):1e27. Dutta PK, Dutta J, Tripathi V. Chitin and chitosan: chemistry, properties and applications. J Sci Ind Res 2004;63:20e31. Anitha A, Sowmya S, Kumar PS, Deepthi S, Chennazhi K, Ehrlich H, et al. Chitin and chitosan in selected biomedical applications. Prog Polym Sci 2014;39(9):1644e67. Murakami K, Aoki H, Nakamura S, Nakamura SI, Takikawa M, Hanzawa M, et al. Hydrogel blends of chitin/chitosan, fucoidan and alginate as healing-impaired wound dressings. Biomaterials 2010;31(1):83e90. Yokoyama A, Yamamoto S, Kawasaki T, Kohgo T, Nakasu M. Development of calcium phosphate cement using chitosan and citric acid for bone substitute materials. Biomaterials 2002;23(4):1091e101. Li C, Vepari C, Jin H-J, Kim HJ, Kaplan DL. Electrospun silk-BMP-2 scaffolds for bone tissue engineering. Biomaterials 2006;27(16):3115e24. Rickard D, Sullivan T, Shenker B, Leboy P, Kazhdan I. Induction of rapid osteoblast differentiation in rat bone marrow stromal cell cultures by dexamethasone and BMP-2. Dev Biol 1994;161(1):218e28. Kim SE, Park JH, Cho YW, Chung H, Jeong SY, Lee EB, et al. Porous chitosan scaffold containing microspheres loaded with transforming growth factor-b1: implications for cartilage tissue engineering. J Control Release 2003;91(3):365e74. ¨ ztu¨rk S, Erdo Alemdaroglu C, Degim Z, C¸elebi N, Zor F, O gan D. An investigation on burn wound healing in rats with chitosan gel formulation containing epidermal growth factor. Burns 2006;32(3):319e27. Mizuno K, Yamamura K, Yano K, Osada T, Saeki S, Takimoto N, et al. Effect of chitosan film containing basic fibroblast growth factor on wound healing in genetically diabetic mice. J Biomed Mater Res Part A 2003;64(1):177e81. Lee Y-M, Park Y-J, Lee S-J, Ku Y, Han S-B, Klokkevold PR, et al. The bone regenerative effect of platelet-derived growth factor-BB delivered with a chitosan/tricalcium phosphate sponge carrier. J Periodontol 2000;71(3):418e24.  ´ s L, Stern R, Gemeiner P. Hyaluronic acid: a natural biopolymer with a broad range of Kogan G, Solte biomedical and industrial applications. Biotechnol Lett 2007;29(1):17e25. Kurisawa M, Chung JE, Yang YY, Gao SJ, Uyama H. Injectable biodegradable hydrogels composed of hyaluronic acidetyramine conjugates for drug delivery and tissue engineering. Chem Commun 2005;(34):4312e4. Ekaputra AK, Prestwich GD, Cool SM, Hutmacher DW. The three-dimensional vascularization of growth factor-releasing hybrid scaffold of poly (3-caprolactone)/collagen fibers and hyaluronic acid hydrogel. Biomaterials 2011;32(32):8108e17. Lisignoli G, Zini N, Remiddi G, Piacentini A, Puggioli A, Trimarchi C, et al. Basic fibroblast growth factor enhances in vitro mineralization of rat bone marrow stromal cells grown on non-woven hyaluronic acid based polymer scaffold. Biomaterials 2001;22(15):2095e105. Yoo HS, Lee EA, Yoon JJ, Park TG. Hyaluronic acid modified biodegradable scaffolds for cartilage tissue engineering. Biomaterials 2005;26(14):1925e33. Liu LS, Ng CK, Thompson AY, Poser JW, Spiro RC. Hyaluronate-heparin conjugate gels for the delivery of basic fibroblast growth factor (FGF-2). J Biomed Mater Res 2002;62(1):128e35. Bhakta G, Rai B, Lim ZX, Hui JH, Stein GS, van Wijnen AJ, et al. Hyaluronic acid-based hydrogels functionalized with heparin that support controlled release of bioactive BMP-2. Biomaterials 2012;33(26):6113e22. McNeil M, Darvill AG, Fry SC, Albersheim P. Structure and function of the primary cell walls of plants. Annu Rev Biochem 1984;53(1):625e63.

Natural polysaccharides for growth factors delivery

511

[54] Helenius G, Ba¨ckdahl H, Bodin A, Nannmark U, Gatenholm P, Risberg B. In vivo biocompatibility of bacterial cellulose. J Biomed Mater Res Part A 2006;76(2):431e8. [55] Shokri J, Adibkia K. Application of cellulose and cellulose derivatives in pharmaceutical industries. Cellulose-medical, pharmaceutical and electronic applications. InTech; 2013. [56] Entcheva E, Bien H, Yin L, Chung C-Y, Farrell M, Kostov Y. Functional cardiac cell constructs on cellulose-based scaffolding. Biomaterials 2004;25(26):5753e62. [57] Santa-Comba A, Pereira A, Lemos R, Santos D, Amarante J, Pinto M, et al. Evaluation of carboxymethylcellulose, hydroxypropylmethylcellulose, and aluminum hydroxide as potential carriers for rhBMP-2. J Biomed Mater Res 2001;55(3):396e400. [58] Li W, Lan Y, Guo R, Zhang Y, Xue W, Zhang Y. In vitro and in vivo evaluation of a novel collagen/ cellulose nanocrystals scaffold for achieving the sustained release of basic fibroblast growth factor. J Biomater Appl 2015;29(6):882e93. [59] Li B, Lu F, Wei X, Zhao R. Fucoidan: structure and bioactivity. Molecules 2008;13(8):1671e95. [60] Hong S-W, Lee H-S, Jung KH, Lee H, Hong S-S. Protective effect of fucoidan against acetaminopheninduced liver injury. Arch Pharm Res (Seoul) 2012;35(6):1099e105. [61] Wijesinghe W, Jeon Y-J. Biological activities and potential industrial applications of fucose rich sulfated polysaccharides and fucoidans isolated from brown seaweeds: a review. Carbohydr Polym 2012;88(1):13e20. [62] Aisa Y, Miyakawa Y, Nakazato T, Shibata H, Saito K, Ikeda Y, et al. Fucoidan induces apoptosis of human HS-sultan cells accompanied by activation of caspase-3 and down-regulation of ERK pathways. Am J Hematol 2005;78(1):7e14. [63] Synytsya A, Kim W-J, Kim S-M, Pohl R, Synytsya A, Kvasnicka F, et al. Structure and antitumour activity of fucoidan isolated from sporophyll of Korean brown seaweed Undaria pinnatifida. Carbohydr Polym 2010;81(1):41e8. [64] Venkatesan J, Bhatnagar I, Kim S-K. Chitosan-alginate biocomposite containing fucoidan for bone tissue engineering. Mar Drugs 2014;12(1):300e16. [65] Huang Y-C, Liu T-J. Mobilization of mesenchymal stem cells by stromal cell-derived factor-1 released from chitosan/tripolyphosphate/fucoidan nanoparticles. Acta Biomater 2012;8(3):1048e56. [66] Purnama A, Aid-Launais R, Haddad O, Maire M, Mantovani D, Letourneur D, et al. Fucoidan in a 3D scaffold interacts with vascular endothelial growth factor and promotes neovascularization in mice. Drug Deliv Transl Res 2015;5(2):187e97. [67] Nakamura S, Nambu M, Ishizuka T, Hattori H, Kanatani Y, Takase B, et al. Effect of controlled release of fibroblast growth factor-2 from chitosan/fucoidan micro complex-hydrogel on in vitro and in vivo vascularization. J Biomed Mater Res Part A 2008;85(3):619e27. [68] Prajapati VD, Maheriya PM, Jani GK, Solanki HK. Carrageenan: a natural seaweed polysaccharide and its applications. Carbohydr Polym 2014;105:97e112. [69] Li L, Ni R, Shao Y, Mao S. Carrageenan and its applications in drug delivery. Carbohydr Polym 2014;103:1e11. [70] Kurtz CA, Loebig TG, Anderson DD, DeMeo PJ, Campbell PG. Insulin-like growth factor I accelerates functional recovery from Achilles tendon injury in a rat model. Am J Sports Med 1999;27(3):363e9. [71] Santo VE, Frias AM, Carida M, Cancedda R, Gomes ME, Mano JF, et al. Carrageenan-based hydrogels for the controlled delivery of PDGF-BB in bone tissue engineering applications. Biomacromolecules 2009;10(6):1392e401. [72] Rocha PM, Santo VE, Gomes ME, Reis RL, Mano JF. Encapsulation of adipose-derived stem cells and transforming growth factor-b1 in carrageenan-based hydrogels for cartilage tissue engineering. J Bioact Compat Polym 2011;26(5):493e507. [73] Silva TH, Alves A, Popa EG, Reys LL, Gomes ME, Sousa RA, et al. Marine algae sulfated polysaccharides for tissue engineering and drug delivery approaches. Biomatter 2012;2(4):278e89.

512 Chapter 21 [74] Yamamoto M. Physicochemical studies on sulfated polysaccharides extracted from seaweeds at various temperatures. Agric Biol Chem 1980;44(3):589e93. [75] Lahaye M, Robic A. Structure and functional properties of ulvan, a polysaccharide from green seaweeds. Biomacromolecules 2007;8(6):1765e74. [76] Dash M, Samal SK, Bartoli C, Morelli A, Smet PF, Dubruel P, et al. Biofunctionalization of ulvan scaffolds for bone tissue engineering. ACS Appl Mater Interfaces 2014;6(5):3211e8. [77] Alves A, Duarte ARC, Mano JF, Sousa RA, Reis RL. PDLLA enriched with ulvan particles as a novel 3D porous scaffold targeted for bone engineering. J Supercrit Fluids 2012;65:32e8. [78] Alves A, Sousa RA, Reis RL. Processing of degradable ulvan 3D porous structures for biomedical applications. J Biomed Mater Res Part A 2013;101(4):998e1006.

C H A P T E R 22

Marine polysaccharides for drug delivery in tissue engineering Manoj Kumar Sarangi1, M.E. Bhanoji Rao2, Versha Parcha3, Dong Kee Yi4, Sitansu Sekhar Nanda4 1

School of Pharmaceutical Sciences & Technology, Sardar Bhagwan Singh University, Dehradun, Uttarakhand, India; 2Calcutta Institute of Pharmaceutical Technology & Allied Health Sciences, Kolkata, West Bengal, India; 3Dolphin (PG) Institute of Biomedical & Natural Sciences, Dehradun, Uttarakhand, India; 4Department of Chemistry, Myongji University, Yongin, South Korea

Chapter Outline 1. Introduction 514 2. Polysaccharides from marine algae 2.1 2.2 2.3 2.4

514

Alginates 514 Carrageenans 516 Fucoidans 516 Ulvans 516

3. Polysaccharides from marine animals 3.1 Chitosans 518 3.2 Hyaluronans 518 3.3 Chondroitin sulfates

517

520

4. Emerging glycosaminoglycanlike polysaccharides from marine origin 4.1 4.2 4.3 4.4 4.5 4.6

520

Dermatan sulfate 520 Heparan sulfates 520 Keratan sulfates 521 Agarose 521 Laminaran 522 Ascophyllan 522

5. Conclusions 522 References 523 Further reading 530

Natural Polysaccharides in Drug Delivery and Biomedical Applications. https://doi.org/10.1016/B978-0-12-817055-7.00022-4 Copyright © 2019 Elsevier Inc. All rights reserved.

513

514 Chapter 22

1. Introduction Marine organisms, with diverse biological properties, are a vast source of different compounds. Many scientific areas are concentrating on marine compounds in order to justify their simplicity, biodiversity, purification and extraction processes [1,2]. Biocompatibility, biodegradability, and noncytotoxic characteristics were attributed as major properties of marine biomaterials in their applications for biomedicine. These applications have allowed the discovery of a basic cornerstone in pharmaceutical industry [1,3,4]. Marine biomaterials showed antitumoral properties for cancer treatment [5e7], like polypeptides extracted from sponges [8] and tunicates [9]. Marine biomaterials are already applied in clinical trials, such as Ecteinascidin 743 [10]and Aplidin [11]. Food and cosmetic industries use marine polysaccharides, but their bioactive property attracted pharmaceutical sciences also [12]. Algae, cartilaginous fish tissue, and skeletons of crustaceans are the primary source for marine polysaccharides [13]. It has dissimilar biological properties of different macromolecules [14,15]. Marine polysaccharides exhibit dissimilar biological properties and structures, ability to form hydrogels with their biodegradability, biocompatibility, and adhesive properties. Sulfated polysaccharides [16] showed anticoagulant [17], anticancer [18], antioxidant [19], antiadhesive, antiangiogenic, antiinflammatory [20], antiviral [21], antiallergic [22] actions. Marine polysaccharides studied systematically for chemical modification and drug delivery systems (DDSs) exhibited affinity to specific drugs and improved biological activities. Marine polysaccharides loading lower drug dosages, led to reduced adverse drug effects. DDSs are used as diagnostic instruments and target a specific location [23,24]. Limitations of health risk associated with viral vectors have been achieved through gene therapy [25] (Fig. 22.1).

2. Polysaccharides from marine algae Primary source of marine polysaccharides, algae, are extracted from marine prokaryotes like microalgae, grown in bioreactors under controlled conditions. Most used sources of polysaccharides are red macro algae. There is possibility to obtain polysaccharides from green or brown macro algae. Multicellular marine algae were obtained from seaweeds, a vital source of polysaccharides. They are described as green, red, and brown algae. In coastal regions, marine algae deposits are used to produce pharmaceutical products.

2.1 Alginates Marine polysaccharide, extracted from brown seaweeds like Ascophyllum nodosum, Laminaria hyperborea, Laminaria digitata, Laminaria japonica, and Macrocystis pyrifera [26] are known as alginates. Alginates are comprised of a sequence of b-D-mannuronate

Marine polysaccharides for drug delivery in tissue engineering 515

Figure 22.1 This figure depicts role of cell wall and its general organization in brown algae. Cell polarity (Fucus serratus zygotes) with its defense responses, tissue integrity (protoplasts derived from Ectocarpus siliculosus), cell adhesion (cortical cells within a stipe of Saccharina latissima), cell differentiation, cell elongation, and osmotic adjustment. This figure was adapted from Deniaud-Boue¨t E, Hardouin K, Potin P, Kloareg B, Herve´ C. A review about brown algal cell walls and fucose-containing sulfated polysaccharides: cell wall context, biomedical properties and key research challenges. Carbohydr Polym 2017;175:395e408. with permission.

˜ 4)-linkeda-L-guluronate(G). M and G blocks proportion: (M) monomers and two (1N Ascophyllum nodosum and Laminaria digitata, extracted alginate, with M/G ratios of 1.82 and 1.16, respectively [27]. It is available with the widest biomedical application. It acts as a stabilizer agent and as an excipient in DDSs in pharmaceutical formulations [28]. Carboxyl groups of alginate with greater than 3e4 pH values make the alginates soluble in alkaline and neutral environments [29]. This kind of pH sensitivity serves as drug protection with suitable absorption in the intestinal tract (where the pH is alkaline) [30]. So, pH sensitivity and solubility make alginate a suitable biomaterial for DDSs [31]. High bioavailability, low toxicity, lower extraction and purification cost, biocompatibility are major properties of alginate as compared with other biopolymers. It is used to process beads, particles, and hydrogel matrices [32e34].

516 Chapter 22

2.2 Carrageenans A sulfated polysaccharide available in red algae, carrageenan comprises of galactose ˜ 4) (unit B) and a-(1N ˜ 3) residues. These residues are connected by alternating b-(1N (unit A) glycosidic bonds. D-configuration is observed in unit A and D- or L-configuration in unit B. Negative charges occur due to the sulfated groups, and this categorizes carrageenans as polyanions [35]. Degree of sulfation decides carrageenans classification. They are classified as lambda (l), iota (i), and kappa (k), with three, two, or one sulfate groups, respectively. i and k types are isolated from algae of the Eucheuma and Kappaphycus genera. l type is isolated from algae of Gigantinaceae family. Gelation capability is influenced by sulfated groups. In the presence of cations, carrageenans i and k can form gels. Gelation is prevented with high sulfation degree of l carrageenan. Food industries use stabilizer and emulsifier properties of carrageenans, cosmetic and pharmaceutical industries use their gelation capability [36]. Carrageenan has a protective activity against fungi, virus, and bacteria [37,38]. Physicochemical properties of carrageenan decide its role as in oral delivery or in the manufacture of devices as excipient. Carrageenan is generally used in cell therapies, cell capsules, cartilage regeneration applications [39,40], and for cell encapsulation [41,42].

2.3 Fucoidans ˜ 3)-linked Sulfated polysaccharide, present in brown algae, is a polymer chain of (1N a-L-fucopyranosyl residues known as fucoidan. Although it is possible to find alternating ˜ 4)-linked a-L-fucopyranosyl and (1N ˜ 3) residues, structure and composition of fucoidan (1N depends on the extraction and isolation source and algae type. Fucus vesiculosus is rich in sulfate and fucose serves as isolation source for fucoidan, extracted from Sargassum stenophyllum. Fucoidans contain glucuronic acid, mannose, galactose, xylose, and glucose [43]. Dependent on its degree of sulfation, fucoidan has antitumor activity and can inhibit cancer cell growth and proliferation [44,45]. Fucoidan showed antiinflammatory, anticoagulant, antiviral, and antiadhesive properties [46,47]. Fucoidan is used as a raw material for DDSs to make fucospheres by electrostatic interactions with chitosan [48], suggested for burn treatments (Table 22.1).

2.4 Ulvans Sulfated polysaccharide present in green algae, ulvan, is extracted from Enteromorpha and Ulva genera. Ulvan is a polymer chain of residues like xylose, glucose, iduronic acid, ˜ 4) linkages. Ulvan may exhibit variations glucuronic acid, and rhamnose with a-and b-(1N in the charge distribution, electronic density, and molecular weight, due to the large number of sugars. Instead of resorting to chemical synthesis, depolymerization is used for isolation of ulvan: a simple extraction process of adding an organic solvent to the feed stock [49].

Marine polysaccharides for drug delivery in tissue engineering 517 Table 22.1: Important properties of algae are showed in this table. Polysaccharide

Seaweed type

Gelation

Degradation

Cell interaction

Alginate

Brown seaweed

Ionic (Ca2þ)

Ion exchange; others

Low

Agarose

Red seaweed

Thermal

Nondegradable

Low

Carrageenan

Red seaweed

Thermal and ionic (Ca2þ/Kþ)

Ion exchange

Low

Ulvan

Green seaweed

Ionic (boric acid and divalent cations)

Enzymatic degradation

n.d.

This table was adapted from Popa EG, Reis RL, Gomes ME. Seaweed polysaccharide-based hydrogels used for the regeneration of articular cartilage. Crit Rev Biotechnol 2015;35(3):410e24; with permission.

Biological properties, such as antiviral, sex habiting, antitumor, antioxidant, antihyperlipidemic, immune system enhancing, anticoagulant activities, are attributed to ulvan. Wide range of applications of ulvan due to its low cytotoxicity include[50] in the cosmetic and food industries, development of new DDSs; its biological properties are use in pharmacological formulations [51], in the treatment for heavy metal poisoning as a chelating agent [52], in wound healing treatments for regenerative medicine and tissue engineering [53, 54]. When chemically modified, ulvan synthesizes thermostable hydrogels. These hydrogels used in the construction of membranes [55], which are a polymeric component of bone cement [56].

3. Polysaccharides from marine animals Not only the microorganisms as well as algae from the marine sources but also different marine animals are considered to be a rich source of polysaccharides. Mostly, two major categories of polymers namely glycosaminoglycans (GAGs) and chitinderived polymers are obtained from marine animals.

518 Chapter 22

3.1 Chitosans A linear polysaccharide obtained from chitin is known as chitosan [57]. It is prepared by chitin deacetylation, comprised of N-acetyl-D-glucosamine (acetylated unit) and D-glucosamine residues (deacetylated unit) [58,59]. The enzymes such as lysozyme and chitinase are responsible for degradation of both chitosan as well as chitin [60]. Chitin is found to be an essential constituent of arthropod exoskeleton as well as crustaceans like crabs, lobsters, and shrimps; it is also obtained from some nematodes and fungi. It is practically insoluble in water, commonly converted into carboxymethyl chitosan (soluble in alkaline and acidic solutions), chitosan (soluble in acidic conditions). Chitosan contains amine groups which are sensitive to pH variations, hence neutral in alkaline pH and positively charged in acidic environments (pKa close to 6) [29]. The most popular marine polysaccharide chitosan is utilized in the development of beads, microcapsules, microspheres, and nanoparticles toward modified release drug delivery systems [61e64]. The antimicrobial property of chitosan is highly utilized in development of films for wound healing [65,66]. It also exhibts antitumor and antiinflammatory activity [67,68]. Hence considering all such biological properties, chitosan is believed to be a supreme element for developing devices which can have contact with biological environments, and can be an excipient for DDSs [69,70]. The chemical modification of chitosan is related to the efficiency of release and boost-up of other activities like drug stabilization and protection [59]. The reaction between propylene epoxide in presence of alkali produces hydroxy propyl chitosan (HPCH), grafted with carboxymethyl b-cyclodextrin mediated via a water-soluble 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) [71]. Because of its presence, hydrophobic drugs can be encapsulated within it. Free amine groups present in chitosan gets protonated at lower pH thus making it pH-responsive. The in vitro release data of ketoprofen proved that the chitosan derivate has potential for developing biodegradable pH responsive modified release delivery system [72,73] (Fig. 22.2).

3.2 Hyaluronans Hyaluronan, a GAG, is a negatively charged, linear and heteropolysaccharide containing repeated units of uronic acid and N-acetylated hexosamine [74]. This polysaccharide comprises of disaccharide units of N-acetyl-D-glucosamine and D-glucuronic acid which ˜ 4) and b-(1N ˜ 3) glycosidic bonds [75]. Hyaluronan is present in the are linked by b-(1N vitreous humor, cartilage tissue, and synovial fluid. Hyaluronan is used for biomedical applications such as in cell proliferation, differentiation, and migration, as well as in tissue regeneration [76]. Hyaluronan is used as a biological marker for rheumatoid arthritis owing to its presence in the synovial fluid of different joints [77]. It has also been suggested for its applications in development of several wound healing structures

Marine polysaccharides for drug delivery in tissue engineering 519

Figure 22.2 This figure shows the macroscopic design of hydrogel with dissimilar delivery systems. This figure was adapted from Moreira CD, Carvalho SM, Mansur HS, Pereira MM. Thermogelling chitosanecollagene bioactive glass nanoparticle hybrids as potential injectable systems for tissue engineering. Mater Sci Eng C 2016;58:1207e16; Li J, Mooney DJ. Designing hydrogels for controlled drug delivery. Nat Rev Mater 2016;1(12):16071.

because of its biocompatibility and biodegradability [78] and also as a supplement for arthritic patients [76]. Currently, the production of hyaluronan is carried out by bacterial fermentation on a large-scale basis [79e81]. However, it can be isolated directly from several marine animal sources like from vitreous humor and cartilages of different species of fishes [82]. The biodegradation of hyaluronan is usually carried out by hyaluronidase, which causes the cleavage of the glycosidic bond between two residues [83]. Hyaluronan is used as a biomarker for monitoring self-movement in biological fluids in human body [84,85]. The hydrogels of hyaluronan with dual stimuliresponsiveness can be developed by altering the pH and temperature conditions. Hydrogels of hyaluronan and PNIPAAm were obtained by using TEMED as a crosslinking agent [86]. Just like several polyanions, hyaluronan can develop microspheres as well as nanoparticles by complexing with chitosan [87,88]. The adsorption of hyaluronan and chitosan can be altered by development of cross-linked chitosan spheres [89], utilized as a diagnostic tool and as reservoirs for several bioactive agents usually obtained via various construction methods [35]. The gold nanoparticles of hyaluronan are widely used as markers for cancer and rheumatoid arthritis because of their potential to emit fluorescence [85,90e92].

520 Chapter 22

3.3 Chondroitin sulfates Chondroitin, a sulfated GAG is made up units of N-acetyl galactosamine (GalNAc) linked ˜ 3) and glucuronic acid (GlcA). Usually chondroitin gets extracted not only from by b-(1N some marine animals, such as the shark and whale, but also from the cartilage of porcine cattle and bovine. There are nevertheless sources, like the ray, salmon fish, some cnidarians, mollusks, and the sea cucumber [39]. Chondroitin sulfate possesses anticoagulant properties and thus can be used as a natural alternative for heparin, one of the most popular as well as widely used anticoagulants [93,94]. The polysaccharide is used as an active component along with the drugs having anticoagulant properties in several pharmaceutical industries and also as a supplement for preventing arthritis [95], as hydrogels for regeneration of cartilage tissue [96,97].

4. Emerging glycosaminoglycanlike polysaccharides from marine origin With diverse biological properties and dissimilar types of glycosaminoglycans, unlike chondroitin sulfate and hyaluronan, they possess low bioavailability; hence are difficult to extract as well as to produce, for which they are least useful in pharmaceutical sciences. Thus, they are used as essential component in supplements. Sulfated dermatan sulfate, glycosaminoglycans, keratin sulfate, nonsulfated agarose, and heparin sulfate are the examples.

4.1 Dermatan sulfate Glycosaminoglycan dermatan sulfate comprises of a linear disaccharide chain containing ˜ 4) or (1N ˜ 3) and units of N-acetyl-galactosamine or glucuronic acid linked by b-(1N hexosamine. The compound possesses residues of L-iduronic acid, which is usually considered as the main difference between chondroitin sulfate and dermatan sulfate. Dermatan sulfate is utilized as a stabilizer for growth factors of cytokines. From the studies, it has been observed that dermatan sulfate possesses a very good anticoagulant activity, and thus can be used for treatments made with heparin [98e100]. The anticoagulant character of dermatan sulfate inhibits thrombin and does not show any effect on clotting cascade (factor X) and does not show any interaction in platelet function. Hence, it is considered to be a very good alternative as well as a substitute for heparin [93,101].

4.2 Heparan sulfates Glycosaminoglycan heparan sulfate has a structure analogous with heparin. It contains a linear chain of alternating residues of D-glucosamine as well as D-glucuronic acid or iduronic acid, which could be acetylated or sulfated. However, the sulfated residues of heparan are associated with some biological properties which could be influenced by their

Marine polysaccharides for drug delivery in tissue engineering 521 affinity toward several proteins [102]. For example, heparan sulfate shows its response by inhibiting the activity of DNA topoisomerase in cell nucleus [103], and thereby controls the cell cycle as well as their proliferation. The processes of oncogenesis may be developed with increased cellular proliferation due to increased heparin sulfate/cell complexation. Hence, heparan sulfate is significantly responsible for development of cancer, associated with cancer cells differentiation, angiogenesis in tumors, and metastasis followed by increased cellular proliferation [104]. The outcome of heparan sulfate on tumor cells is not only based on the structure of glycosaminoglycan, but also on the type of tumor microenvironment and tumor cell [105]. However the enzymatic action of heparanase causes the biodegradability of the sulfated polysaccharide [106]. Because of the presence of sulfated groups, the polysaccharide is able to bind to several proteins and thereby regulates biological processes like regulation and coagulation. It also binds to several polypeptides like growth factors as well as cellular receptor complexes [107], and is used in the development of some new antineoplastic drugs along with novel drug delivery systems, as well as some new diagnostic approaches [108].

4.3 Keratan sulfates Glycosaminoglycan keratan sulfate is lacking of the uronic acid unit. The disaccharide ˜ 4) linkages and galactose unit comprises of N-acetyl glucosamine bonded by b-(1N residues. Keratan sulfate is classified into three different classes which are differentiated from each other on the basis of their protein binding efficiency. Class I of the keratin sulfate is commonly found in small cartilages as well as in cornea. In case of Class II, which is also available in small cartilages, the protein binding occurs in between the O- of N-acetyl glucosamine and either with a threonine or a serine. In case of Class III (first ever isolated from nervous tissue), the protein binding takes place in between the O- of the mannose residue to either a threonine or serine [109]. The keratan sulfate is responsible for maintaining moisture content of the corneal tissue, thus dealing with its level of transparency. The cellular level studies conducted for keratan sulfate revealed that it has antiadhesive properties. Keratan sulfate in nervous tissues is highly useful for preventing the growth of axons and decreases the immune response in diseases like osteoarthritis in cartilage [110]. However, in case of nerve injury keratan sulfate shows an inhibitory response toward the regeneration of nerves [111,112].

4.4 Agarose Agarose, one of the fabulous biomaterial from marine sources, has a structure analogous to carrageenan, an important component found in red algae cell wall. Agarose contains monosaccharide as its structural components, which are connected alternatively in a conformation (AB)n. The units comprise of galactose residues which are linked by

522 Chapter 22 ˜ 3) (unitA) and b-(1N ˜ 4) (unit B) linkages. The carrageenan is differenced from agar a-(1N by the respect of conformation of unit A, that is, unit A of carrageenan is always in the D-conformation, whereas the unit A in agar is only in the L-conformation [113]. Agarose is connected with various biomedical applications specifically as hydrogels for taking benefit of its ability to jellify, release of bioactive agents, biocompatibility, and biodegradability [114,115].

4.5 Laminaran Glucans, laminarans, present in either soluble or insoluble form [116,117]. D glucose with b-(1.3) linkages, with b-(1.6) intrachain branching are the composition of this polysaccharide [118e120]. It is known as leucosin or laminarin, isolated by Schmiedeberg from the Laminariaceae (Phaeophyceae) [121]. It is a food reserve of brown algae and is located in vacuoles. It is found in the fronds of Saccharina and Laminaria species [122]. (1.3)-b-D-glucan is known as laminaran and comprised of low molecular weight storage b-glucans [123]. It consist of some b-(1.6)-intrachain links and (1.3)-b-D-glucopyranose residues [124]. Its molecular weight 5 kDa is dependent upon the polymerization with 20e25 glucose moieties [125,126]. The molecular weight of laminaran from Saccharina longicruris is 2.89e3.32 kDa depending on the extraction conditions [127], including the used solvent type [128]. Laminarans are comprised of different biological properties [120,128]. Laminarins are characterized by solubility in cold water, and the insoluble form dissolves only in hot-water. Moreover, water-soluble laminarin, a carbohydrate reserve of brown algae with degree of polymerization between 20-25 glucose units [127,129].

4.6 Ascophyllan Brown algae ascophyllan, Ascophyllum nodosum (Phaeophyceae), is a fucose-containing sulfated polysaccharide. It has a chemical structure similar to fucoidan and distinct characteristic monosaccharide composition [121]. Ascophyllan (xylofucoglycuronan) has similar but obviously distinct composition characteristics from those of fucoidans isolated from A. nodosum and Fucus vesiculosus [130,131], with fucose and xylose in about equimolecular proportion. A. nodosum (ascophyllan) and F. vesiculosus (fucoidans) sulfate levels were 9.6%, 19.4%, and 22.6%, respectively [132]. Molecular mass of ascophyllan was about 390 kDa higher than fucoidan [133].

5. Conclusions Marine polysaccharides were quite engaged in synthesizing DDSs as they are biocompatible, often biodegradable, nontoxic, and stimuli-responsive; hence, they are

Marine polysaccharides for drug delivery in tissue engineering 523 considered to be suitable raw materials for designing as well as developing most suitable complex devices meant for controlled release. It has been observed that such complex devices can be designed as well as constructed by several methods and can be utilized in developing several dosage forms like hydrogels, particles, and capsules, along with the ability of protecting nuclei acids and proteins. Every polymer unveils different biochemical properties, in order to make the marine biomaterials and their derivatives an extraordinary material toward the construction of loading devices as well as a versatile excipient for pharmaceutical formulations and food supplements. The biomaterials from natural origin permit for incorporating drugs, nucleic acids and proteins, which might be toxic for the body, which may not be possible with many synthetic materials. Value-added DDS designs and efficient control over the release rate can be used in innovative therapies. By looking into response of the polysaccharides along with interactions between drugs, polymer, and native biological tissues, it seems to be possible.

References [1] Cardoso MJ, Costa RR, Mano JF. Marine origin polysaccharides in drug delivery systems. Mar Drugs 2016;14(2):34. [2] Wijesekara I, Pangestuti R, Kim SK. Biological activities and potential health benefits of sulfated polysaccharides derived from marine algae. Carbohydr Polym 2011;84(1):14e21. [3] Faulkner DJ. Marine natural products. Nat Prod Rep 2001;18(1):1Re49R. [4] Arumugam V, Venkatesan M, Ramachandran S, Sundaresan U. Bioactive peptides from marine ascidians and future drug development e A review. Int J Pept Res Ther 2018;24(1):13e8. [5] Zheng L, Xu Y, Lin X, Yuan Z, Liu M, Cao S, et al. Recent progress of marine polypeptides as anticancer agents. Recent Pat Anti-cancer Drug Discov 2018;13(4):445e54. [6] Cabral C, Efferth T, Pires IM, Severino P, Lemos MF. Natural products as a source for new leads in cancer research and treatment. Evid Based Complement Altern Med 2018;2018. [7] Gomes AR, Freitas AC, Duarte AC, AP T. Clinical trials for deriving bioactive compounds from marine invertebrates. Nat Prod Clin Trials 2018;1(1):1. [8] Giordano D, Costantini M, Coppola D, Lauritano C, Pons LN, Ruocco N, et al. Biotechnological applications of bioactive peptides from marine sources. Adv Microb Physiol 2018;73:171e220. [9] Alves C, Silva J, Pinteus S, Gaspar H, Alpoim MC, Botana LM, Pedrosa R. From marine origin to therapeutics: the antitumor potential of marine algae-derived compounds. Front Pharmacol 2018;9. [10] Miller JH, Field JJ, Kanakkanthara A, Owen JG, Singh AJ, Northcote PT. Marine invertebrate natural products that target microtubules. J Nat Prod 2018;81(3):691e702. [11] Pereira RB, Dasari R, Lefranc F, Kornienko A, Kiss R, Gomes NG. Targeting enzymatic pathways with marine-derived clinical agents. In: Natural products targeting clinically relevant enzymes; 2017. p. 255e75. [12] Guzma´n EA, Xu Q, Pitts TP, Mitsuhashi KO, Baker C, Linley PA, et al. Leiodermatolide, a novel marine natural product, has potent cytotoxic and antimitotic activity against cancer cells, appears to affect microtubule dynamics, and exhibits antitumor activity. Int J Cancer 2016;139(9):2116e26. [13] Ruocco N, Costantini S, Guariniello S, Costantini M. Polysaccharides from the marine environment with pharmacological, cosmeceutical and nutraceutical potential. Molecules 2016;21(5):551.

524 Chapter 22 [14] Abrahamse H, Kumar SSD, Houreld NN. The potential role of photobiomodulation and polysaccharidebased biomaterials in wound healing applications. In: Wound healing: stem cells repair and restorations. Basic and Clinical Aspects; 2017. p. 211. [15] Deniaud-Boue¨t E, Hardouin K, Potin P, Kloareg B, Herve´ C. A review about brown algal cell walls and fucose-containing sulfated polysaccharides: cell wall context, biomedical properties and key research challenges. Carbohydr Polym 2017;175:395e408. [16] Faggio C, Pagano M, Dottore A, Genovese G, Morabito M. Evaluation of anticoagulant activity of two algal polysaccharides. Nat Prod Res 2016;30(17):1934e7. [17] Peasura N, Laohakunjit N, Kerdchoechuen O, Wanlapa S. Characteristics and antioxidant of Ulva intestinalis sulphated polysaccharides extracted with different solvents. Int J Biol Macromol 2015;81:912e9. [18] Arata PX, Genoud V, Lauricella AM, Ciancia M, Quintana I. Alterations of fibrin networks mediated by sulfated polysaccharides from green seaweeds. Thromb Res 2017;159:1e4. [19] Ahmadi A, Zorofchian Moghadamtousi S, Abubakar S, Zandi K. Antiviral potential of algae polysaccharides isolated from marine sources: a review. BioMed Research International; 2015. [20] Fernando IS, Nah JW, Jeon YJ. Potential anti-inflammatory natural products from marine algae. Environ Toxicol Pharmacol 2016;48:22e30. [21] Newman DJ, Cragg GM. Natural products as sources of new drugs from 1981 to 2014. J Nat Prod 2016;79(3):629e61. [22] Abdelahad N, Barbato F, O’Heir S, Fratini F, Valletta A, Ninivaggi L, Alfinito S. Reproduction of Sphaerococcus coronopifolius (Gigartinales, Rhodophyta) in natural populations of the Lazio coasts (central Italy) and in culture. Algologie: Cryptogamie; 2016. [23] Ma J, Porter AL, Aminabhavi TM, Zhu D. Nano-enabled drug delivery systems for brain cancer and Alzheimer’s disease: research patterns and opportunities. Nanomed Nanotechnol Biol Med 2015;11(7):1763e71. [24] Venkatesan J, Lowe B, Anil S, Manivasagan P, Kheraif AAA, Kang KH, Kim SK. Seaweed polysaccharides and their potential biomedical applications. Starch Staerke 2015;67(5e6):381e90. [25] Masood F. Polymeric nanoparticles for targeted drug delivery system for cancer therapy. Mater Sci Eng C 2016;60:569e78. [26] Ramamoorth M, Narvekar A. Non viral vectors in gene therapy-an overview. J Clin Diagn Res 2015;9(1):GE01. [27] Martins M, Barros AA, Quraishi S, Gurikov P, Raman SP, Smirnova I, et al. Preparation of macroporous alginate-based aerogels for biomedical applications. J Supercrit Fluids 2015;106:152e9. [28] Wang QC, Zhao X, Pu JH, Luan XH. Influences of acidic reaction and hydrolytic conditions on monosaccharide composition analysis of acidic, neutral and basic polysaccharides. Carbohydr Polym 2016;143:296e300. [29] Lee S, Kim YC, Park JH. Zein-alginate based oral drug delivery systems: protection and release of therapeutic proteins. Int J Pharm 2016;515(1e2):300e6. [30] Blum AP, Kammeyer JK, Rush AM, Callmann CE, Hahn ME, Gianneschi NC. Stimuli-responsive nanomaterials for biomedical applications. J Am Chem Soc 2015;137(6):2140e54. [31] Farhadian A, Dounighi NM, Avadi M. Enteric trimethyl chitosan nanoparticles containing hepatitis B surface antigen for oral delivery. Hum Vaccines Immunother 2015;11(12):2811e8. [32] Mogos‚anu GD, Grumezescu AM. Pharmaceutical natural polymers: structure and chemistry. In: Handbook of polymers for pharmaceutical technologies, structure and chemistry. 1; 2015. p. 477. [33] Zhang Z, Zhang R, Zou L, McClements DJ. Protein encapsulation in alginate hydrogel beads: effect of pH on microgel stability, protein retention and protein release. Food Hydrocoll 2016;58:308e15.

Marine polysaccharides for drug delivery in tissue engineering 525 [34] Yu S, Zhang X, Tan G, Tian L, Liu D, Liu Y, et al. A novel pH-induced thermosensitive hydrogel composed of carboxymethyl chitosan and poloxamer cross-linked by glutaraldehyde for ophthalmic drug delivery. Carbohydr Polym 2017;155:208e17. [35] Xu J, Strandman S, Zhu JX, Barralet J, Cerruti M. Genipin-crosslinked catechol-chitosan mucoadhesive hydrogels for buccal drug delivery. Biomaterials 2015;37:395e404. [36] Ahmadi F, Oveisi Z, Samani SM, Amoozgar Z. Chitosan based hydrogels: characteristics and pharmaceutical applications. Res Pharm Sci 2015;10(1):1. [37] Diogo JV, Novo SG, Gonza´lez MJ, Ciancia M, Bratanich AC. Antiviral activity of lambda-carrageenan prepared from red seaweed (Gigartina skottsbergii) against BoHV-1 and SuHV-1. Res Vet Sci 2015;98:142e4. [38] Song F, Li X, Wang Q, Liao L, Zhang C. Nanocomposite hydrogels and their applications in drug delivery and tissue engineering. J Biomed Nanotechnol 2015;11(1):40e52. [39] Liu J, Zhan X, Wan J, Wang Y, Wang C. Review for carrageenan-based pharmaceutical biomaterials: favourable physical features versus adverse biological effects. Carbohydr Polym 2015;121:27e36. [40] Inic-Kanada A, Stein E, Stojanovic M, Schuerer N, Ghasemian E, Filipovic A, et al. Effects of iota-carrageenan on ocular Chlamydia trachomatis infection in vitro and in vivo. J Appl Phycol 2018:1e10. [41] Popa EG, Reis RL, Gomes ME. Seaweed polysaccharide-based hydrogels used for the regeneration of articular cartilage. Crit Rev Biotechnol 2015;35(3):410e24. [42] Tziveleka LA, Pippa N, Georgantea P, Ioannou E, Demetzos C, Roussis V. Marine sulfated polysaccharides as versatile polyelectrolytes for the development of drug delivery nanoplatforms: complexation of ulvan with lysozyme. Int J Biol Macromol Oct 15, 2018;118(Pt A):69e75. [43] Pina S, Oliveira JM, Reis RL. Natural-based nanocomposites for bone tissue engineering and regenerative medicine: a review. Adv Mater 2015;27(7):1143e69. [44] Puvaneswary S, Talebian S, Raghavendran HB, Murali MR, Mehrali M, Afifi AM, et al. Fabrication and in vitro biological activity of bTCP-Chitosan-Fucoidan composite for bone tissue engineering. Carbohydr Polym 2015;134:799e807. [45] Olabisi RM. Cell microencapsulation with synthetic polymers. J Biomed Mater Res A 2015;103(2):846e59. [46] Anastyuk SD, Shevchenko NM, Usoltseva RV, Silchenko AS, Zadorozhny PA, Dmitrenok PS, Ermakova SP. Structural features and anticancer activity in vitro of fucoidan derivatives from brown alga Saccharina cichorioides. Carbohydr Polym 2017;157:1503e10. [47] Isnansetyo A, Lutfia FNL, Nursid M, Susidarti RA. Cytotoxicity of fucoidan from three tropical Brown algae against breast and colon cancer cell lines. Phcog J 2017;9(1). [48] Kim SK. Marine cosmeceuticals: trends and prospects. CRC Press; 2016. [49] Sridhar R, Lakshminarayanan R, Madhaiyan K, Barathi VA, Lim KHC, Ramakrishna S. Electrosprayed nanoparticles and electrospun nanofibers based on natural materials: applications in tissue regeneration, drug delivery and pharmaceuticals. Chem Soc Rev 2015;44(3):790e814. ¨ ner ET, Akbu [50] Sezer AD, Kazak Sarılmıs‚er H, Rayaman E, C ¸ evikbas‚ A, O ga J. Development and characterization of vancomycin-loaded levan-based microparticular system for drug delivery. Pharmaceut Dev Technol 2017;22(5):627e34. [51] Ulaganathan T, Boniecki MT, Foran E, Buravenkov V, Mizrachi N, Banin E, et al. New ulvan-degrading polysaccharide lyase family: structure and catalytic mechanism suggests convergent evolution of active site architecture. ACS Chem Biol 2017;12(5):1269e80. [52] Thanh TTT, Quach TMT, Nguyen TN, Luong DV, Bui ML, Van Tran TT. Structure and cytotoxic activity of ulvan extracted from green seaweed Ulva lactuca. Int J Biol Macromol 2016;93:695e702. [53] Abd-Ellatef GEF, Ahmed OM, Abdel-Reheim ES, Abdel-Hamid AHZ. Ulva lactuca polysaccharides prevent Wistar rat breast carcinogenesis through the augmentation of apoptosis, enhancement of antioxidant defense system, and suppression of inflammation. Breast Cancer Target Ther 2017;9:67.

526 Chapter 22 [54] Reis RP, de Carvalho Junior AA, Facchinei AP, dos Santos Calheiros AC, Castelar B. Direct effects of ulvan and a flour produced from the green alga Ulva fasciata Delile on the fungus Stemphylium solani Weber. Algal Res 2018;30:23e7. [55] Dhivya S, Selvamurugan N. Biocomposite scaffolds derived from renewable resources for bone tissue repair. In: Handbook of composites from renewable materials, biodegradable materials. 5; 2017. p. 439. [56] Melcher RL, Neumann M, Werner JPF, Gro¨hn F, Moerschbacher BM. Revised domain structure of ulvan lyase and characterization of the first ulvan binding domain. Sci Rep 2017;7:44115. [57] Yu-Qing T, Mahmood K, Shehzadi R, Ashraf MF. Ulva lactuca and its polysaccharides: food and biomedical aspects. J Biol Agric Healthc 2016;6(1):140e51. [58] Li M, Mitra D, Kang ET, Lau T, Chiong E, Neoh KG. Thiol-ol chemistry for grafting of natural polymers to form highly stable and efficacious antibacterial coatings. ACS Appl Mater Interfaces 2017;9(2):1847e57. [59] Elgadir MA, Uddin MS, Ferdosh S, Adam A, Chowdhury AJK, Sarker MZI. Impact of chitosan composites and chitosan nanoparticle composites on various drug delivery systems: a review. J Food Drug Anal 2015;23(4):619e29. [60] Younes I, Rinaudo M. Chitin and chitosan preparation from marine sources. Structure, properties and applications. Mar Drugs 2015;13(3):1133e74. [61] Boamah PO, Huang Y, Hua M, Zhang Q, Wu J, Onumah J, et al. Sorption of heavy metal ions onto carboxylate chitosan derivativesda mini-review. Ecotoxicol Environ Saf 2015;116:113e20. [62] Poth N, Seiffart V, Gross G, Menzel H, Dempwolf W. Biodegradable chitosan nanoparticle coatings on titanium for the delivery of BMP-2. Biomolecules 2015;5(1):3e19. [63] Reis CP, Neufeld RJ, Veiga F. Preparation of drug-loaded polymeric nanoparticles. In: Nanomedicine in cancer. Pan Stanford; 2017. p. 197e240. [64] Ahmed TA, Aljaeid BM. Preparation, characterization, and potential application of chitosan, chitosan derivatives, and chitosan metal nanoparticles in pharmaceutical drug delivery. Drug Des Dev Ther 2016;10:483. [65] Li J, Mooney DJ. Designing hydrogels for controlled drug delivery. Nat Rev Mater 2016;1(12):16071. [66] Moreira CD, Carvalho SM, Mansur HS, Pereira MM. Thermogelling chitosanecollagenebioactive glass nanoparticle hybrids as potential injectable systems for tissue engineering. Mater Sci Eng C 2016;58:1207e16. [67] Ma Z, Garrido-Maestu A, Jeong KC. Application, mode of action, and in vivo activity of chitosan and its micro-and nanoparticles as antimicrobial agents: a review. Carbohydr Polym 2017;176:257e65. [68] LogithKumar R, KeshavNarayan A, Dhivya S, Chawla A, Saravanan S, Selvamurugan N. A review of chitosan and its derivatives in bone tissue engineering. Carbohydr Polym 2016;151:172e88. [69] Zou P, Yang X, Wang J, Li Y, Yu H, Zhang Y, Liu G. Advances in characterisation and biological activities of chitosan and chitosan oligosaccharides. Food Chem 2016;190:1174e81. [70] Folkerts J, Stadhouders R, Redegeld FA, Tam SY, Hendriks RW, Galli SJ, Maurer M. Effect of dietary fiber and metabolites on mast cell activation and mast cell-associated diseases. Front Immunol 2018;9. [71] Bharate SS, Bharate SB, Bajaj AN. Interactions and incompatibilities of pharmaceutical excipients with active pharmaceutical ingredients: a comprehensive review. J Excipients Food Chem 2016;1(3):1131. [72] Raafat D, Leib N, Wilmes M, Franc¸ois P, Schrenzel J, Sahl HG. Development of in vitro resistance to chitosan is related to changes in cell envelope structure of Staphylococcus aureus. Carbohydr Polym 2017;157:146e55. [73] Karakasyan C, Mathos J, Lack S, Davy J, Marquis M, Renard D. Microfluidics-assisted generation of stimuli-responsive hydrogels based on alginates incorporated with thermo-responsive and amphiphilic polymers as novel biomaterials. Colloids Surfaces B Biointerfaces 2015;135:619e29.

Marine polysaccharides for drug delivery in tissue engineering 527 [74] Pan Q, Lv Y, Williams GR, Tao L, Yang H, Li H, Zhu L. Lactobionic acid and carboxymethyl chitosan functionalized graphene oxide nanocomposites as targeted anticancer drug delivery systems. Carbohydr Polym 2016;151:812e20. [75] Chen Y, Huang Y, Qin D, Liu W, Song C, Lou K, et al. b-cyclodextrin-based inclusion complexation bridged biodegradable self-assembly macromolecular micelle for the delivery of paclitaxel. PLoS One 2016;11(3):e0150877. [76] Clausen TM, Pereira MA, Al Nakouzi N, Oo HZ, Agerbæk MO, Lee S, et al. Oncofetal chondroitin sulfate glycosaminoglycans are key players in integrin signaling and tumor cell motility. Mol Canc Res Dec 2016;14(12):1288e99. molcanres-0103. [77] Highley CB, Prestwich GD, Burdick JA. Recent advances in hyaluronic acid hydrogels for biomedical applications. Curr Opin Biotechnol 2016;40:35e40. [78] Browne S, Jha AK, Ameri K, Marcus SG, Yeghiazarians Y, Healy KE. TGF-b1/CD105 signaling controls vascular network formation within growth factor sequestering hyaluronic acid hydrogels. PLoS One 2018;13(3):e0194679. [79] Rao NV, Yoon HY, Han HS, Ko H, Son S, Lee M, et al. Recent developments in hyaluronic based nanomedicine for targeted cancer treatment. Expert Opin Drug Deliv 2016;13(2):239e52. [80] Tracy LE, Minasian RA, Caterson EJ. Extracellular matrix and dermal fibroblast function in the healing wound. Adv Wound Care 2016;5(3):119e36. [81] Va´zquez JA, Pastrana L, Pin˜eiro C, Teixeira JA, Pe´rez-Martı´n RI, Amado IR. Production of hyaluronic acid by Streptococcus zooepidemicus on protein substrates obtained from Scyliorhinus canicula discards. Mar Drugs 2015;13(10):6537e49. [82] Amado IR, Va´zquez JA, Pastrana L, Teixeira JA. Cheese whey: a cost-effective alternative for hyaluronic acid production by Streptococcus zooepidemicus. Food Chem 2016;198:54e61. [83] Hamad G, Taha T, Hafez E, El Sohaimy S. Physicochemical, molecular and functional characteristics of hyaluronic acid as a functional food. Am J Food Technol 2017;12(2):72e85. [84] Va´zquez JA, Blanco M, Fraguas J, Pastrana L, Pe´rez-Martı´n R. Optimisation of the extraction and purification of chondroitin sulphate from head by-products of Prionace glauca by environmental friendly processes. Food Chem 2016;198:28e35. [85] Sakai S, Ueda K, Taya M. Peritoneal adhesion prevention by a biodegradable hyaluronic acid-based hydrogel formed in situ through a cascade enzyme reaction initiated by contact with body fluid on tissue surfaces. Acta Biomater 2015;24:152e8. [86] Nykopp TK, Pasonen-Seppa¨nen S, Tammi MI, Tammi RH, Kosma VM, Anttila M, Sironen R. Decreased hyaluronidase 1 expression is associated with early disease recurrence in human endometrial cancer. Gynecol Oncol 2015;137(1):152e9. [87] Tarquini C, Mattera R, Mastrangeli F, Agostinelli S, Ferlosio A, Bei R, et al. Comparison of tissue transglutaminase 2 and bone biological markers osteocalcin, osteopontin and sclerostin expression in human osteoporosis and osteoarthritis. Amino Acids 2017;49(3):683e93. [88] Wei W, Hu X, Qi X, Yu H, Liu Y, Li J, et al. A novel thermo-responsive hydrogel based on salecan and poly (N-isopropylacrylamide): synthesis and characterization. Colloids Surfaces B Biointerfaces 2015;125:1e11. [89] Sacco P, Furlani F, de Marzo G, Marsich E, Paoletti S, Donati I. Concepts for developing physical gels of chitosan and of chitosan derivatives. Gels 2018;4(3):67. [90] Aston R, Wimalaratne M, Brock A, Lawrie G, Grøndahl L. Interactions between chitosan and alginate dialdehyde biopolymers and their layer-by-layer assemblies. Biomacromolecules 2015;16(6):1807e17. [91] Reddy N, Reddy R, Jiang Q. Crosslinking biopolymers for biomedical applications. Trends Biotechnol 2015;33(6):362e9.

528 Chapter 22 [92] Cai Z, Zhang H, Wei Y, Cong F. Hyaluronan-inorganic nanohybrid materials for biomedical applications. Biomacromolecules 2017;18(6):1677e96. [93] Jordan AR, Lokeshwar SD, Lopez LE, Hennig M, Chipollini J, Yates T, et al. Antitumor activity of sulfated hyaluronic acid fragments in pre-clinical models of bladder cancer. Oncotarget 2017;8(15):24262. [94] Lidbury JA, Hoffmann AR, Fry JK, Suchodolski JS, Steiner JM. Evaluation of hyaluronic acid, procollagen type III N-terminal peptide, and tissue inhibitor of matrix metalloproteinase-1 as serum markers of canine hepatic fibrosis. Can J Vet Res 2016;80(4):302e8. [95] Bougatef H, Krichen F, Capitani F, Amor IB, Maccari F, Mantovani V, et al. Chondroitin sulfate/ dermatan sulfate from corb (Sciaena umbra) skin: purification, structural analysis and anticoagulant effect. Carbohydr Polym 2018;196:272e8. [96] Casu B, Naggi A, Torri G. Re-visiting the structure of heparin. Carbohydr Res 2015;403:60e8. [97] Hochberg MC, Martel-Pelletier J, Monfort J, Mo¨ller I, Castillo JR, Arden N, et al. Combined chondroitin sulfate and glucosamine for painful knee osteoarthritis: a multicentre, randomised, doubleblind, non-inferiority trial versus celecoxib. Ann Rheum Dis 2016;75(1):37e44. [98] Chen F, Yu S, Liu B, Ni Y, Yu C, Su Y, et al. An injectable enzymatically crosslinked carboxymethylated pullulan/chondroitin sulfate hydrogel for cartilage tissue engineering. Sci Rep 2016;6:20014. [99] Liu M, Zeng X, Ma C, Yi H, Ali Z, Mou X, et al. Injectable hydrogels for cartilage and bone tissue engineering. Bone research 2017;5:17014. [100] Claure-Del Granado R, Bouchard J, Mehta RL. Principles of anticoagulation in extracorporeal circuits. In: Critical care nephrology; 2019. p. 860e6. [101] Suleria HAR, Masci PP, Zhao KN, Addepalli R, Chen W, Osborne SA, Gobe GC. Anti-coagulant and anti-thrombotic properties of blacklip abalone (Haliotis rubra): in vitro and animal studies. Mar Drugs 2017;15(8):240. [102] Sethi SK, Bansal SB, Khare A, Dhaliwal M, Raghunathan V, Wadhwani N, et al. Heparin free dialysis in critically sick children using sustained low efficiency dialysis (SLEDD-f): a new hybrid therapy for dialysis in developing world. PLoS One 2018;13(4):e0195536. [103] Mizumoto S, Kosho T, Yamada S, Sugahara K. Pathophysiological significance of dermatan sulfate proteoglycans revealed by human genetic disorders. Pharmaceuticals 2017;10(2):34. [104] Meneghetti MC, Hughes AJ, Rudd TR, Nader HB, Powell AK, Yates EA, Lima MA. Heparan sulfate and heparin interactions with proteins. J R Soc Interface 2015;12(110):20150589. [105] Harisi R, Jeney A. Extracellular matrix as target for antitumor therapy. OncoTargets Ther 2015;8:1387. [106] Kuhnast B, El Hadri A, Boisgard R, Hinnen F, Richard S, Caravano A, et al. Synthesis, radiolabeling with fluorine-18 and preliminary in vivo evaluation of a heparan sulphate mimetic as potent angiogenesis and heparanase inhibitor for cancer applications. Org Biomol Chem 2016;14(6):1915e20. ˚ , Mani K. Nucleolin is a nuclear target of heparan sulfate derived from [107] Cheng F, Belting M, Fransson LA glypican-1. Exp Cell Res 2017;354(1):31e9. [108] Weissmann M, Arvatz G, Horowitz N, Feld S, Naroditsky I, Zhang Y, et al. Heparanase-neutralizing antibodies attenuate lymphoma tumor growth and metastasis. Proc Natl Acad Sci USA 2016;113(3):704e9. [109] Langereis EJ, van Vlies N, Church HJ, Geskus RB, Hollak CE, Jones SA, et al. Biomarker responses correlate with antibody status in mucopolysaccharidosis type I patients on long-term enzyme replacement therapy. Mol Genet Metabol 2015;114(2):129e37. [110] Kumar AV, Katakam S, Kiesel L, Go¨tte M. The heparan sulfate sulfotransferase HS2ST1 modulates breast cancer cell invasiveness via the EGFR and MAPK signaling pathways. Exp Clin Endocrinol Diabetes 2015;123(03):P03_26. [111] Uchimura K. Keratan sulfate: biosynthesis, structures, and biological functions. In: Glycosaminoglycans. New York, NY: Humana Press; 2015. p. 389e400.

Marine polysaccharides for drug delivery in tissue engineering 529 [112] Hodgson K, Supanich K. Toward understanding keratan sulfate biosynthesis in the chick embryonic cornea. 2016. [113] Melrose J. Keratan sulfate (KS)-Proteoglycans and neuronal regulation in health and disease: the importance of KS-glycodynamics and interactive capability with neuroregulatory ligands. J Neurochem Apr 2019;149(2):170e94. [114] Ueno R, Miyamoto K, Tanaka N, Moriguchi K, Kadomatsu K, Kusunoki S. Keratan sulfate exacerbates experimental autoimmune encephalomyelitis. J Neurosci Res 2015;93(12):1874e80. [115] Restrepo-Espinosa DC, Roma´n Y, Colorado-Rı´os J, de Santana-Filho AP, Sassaki GL, Cipriani TR, et al. Structural analysis of a sulfated galactan from the tunic of the ascidian Microcosmus exasperatus and its inhibitory effect of the intrinsic coagulation pathway. Int J Biol Macromol 2017;105:1391e400. [116] Merino S, Martin C, Kostarelos K, Prato M, Vazquez E. Nanocomposite hydrogels: 3D polymerenanoparticle synergies for on-demand drug delivery. ACS Nano 2015;9(5):4686e97. [117] Sharma K, Kumar V, Swart-Pistor C, Chaudhary B, Swart HC. Synthesis, characterization, and antimicrobial activity of superabsorbents based on agarepoly (methacrylic acid-glycine). J Bioact Compat Polym 2017;32(1):74e91. [118] McConville MJ. Chemical composition and biochemistry of sea ice microalgae. In: Sea ice biota. CRC Press; 2017. p. 105e29. [119] Matos J, Cardoso C, Bandarra NM, Afonso C. Microalgae as healthy ingredients for functional food: a review. Food funct 2017;8(8):2672e85. [120] El Atouani S, Bentiss F, Reani A, Zrid R, Belattmania Z, Pereira L, et al. The invasive brown seaweed Sargassum muticum as new resource for alginate in Morocco: spectroscopic and rheological characterization. Phycol Res 2016;64(3):185e93. [121] Kadam SU, Tiwari BK, O’donnell CP. Extraction, structure and biofunctional activities of laminarin from brown algae. Int J Food Sci Technol 2015;50(1):24e31. [122] Kadam SU, O’Donnell CP, Rai DK, Hossain MB, Burgess CM, Walsh D, Tiwari BK. Laminarin from Irish brown seaweeds Ascophyllum nodosum and Laminaria hyperborea: ultrasound assisted extraction, characterization and bioactivity. Mar Drugs 2015;13(7):4270e80. [123] Schmuhl HW. Die Gesellschaft Deutscher Neurologen und Psychiater im Nationalsozialismus. SpringerVerlag; 2015. [124] Kawai H, Henry EC, Phaeophyta I, Archibald J, Simpson A, Slamovits C. Handbook of the protists. Cham: Springer; 2017. [125] Jeong SC, Jeong YT, Lee SM, Kim JH. Immune-modulating activities of polysaccharides extracted from brown algae Hizikia fusiforme. Biosci Biotechnol Biochem 2015;79(8):1362e5. [126] Wang D, Kim DH, Yun EJ, Park YC, Seo JH, Kim KH. The first bacterial b-1, 6-endoglucanase from Saccharophagus degradans 2-40 T for the hydrolysis of pustulan and laminarin. Appl Microbiol Biotechnol 2017;101(1):197e204. [127] Graiff A, Ruth W, Karsten U. Identification and quantification of laminarins in brown algae, vol. 11; 2017 [Chapter]. [128] Becker S, Scheffel A, Polz MF, Hehemann JH. Accurate quantification of laminarin in marine organic matter with enzymes from marine microbes. Appl Environ Microbiol Apr 17, 2017;83(9). AEM-03389. e03389e16. [129] Ayoub A, Pereira JM, Rioux LE, Turgeon SL, Beaulieu M, Moulin VJ. Role of seaweed laminaran from Saccharina longicruris on matrix deposition during dermal tissue-engineered production. Int J Biol Macromol 2015;75:13e20. [130] Sanjeewa KKA, Lee JS, Kim WS, Eon YJ. The potential of brown algae polysaccharides for the development of anticancer agents: an update on anticancer effects reported for fucoidan and laminaran. Carbohydr Polym 2017;177:451459. [131] Abu R, Jiang Z, Ueno M, Isaka S, Nakazono S, Okimura T, et al. Anti-metastatic effects of the sulfated polysaccharide ascophyllan isolated from Ascophyllum nodosum on B16 melanoma. Biochem Biophys Res Commun 2015;458(4):727e32.

530 Chapter 22 [132] Priyan Shanura Fernando I, Kim KN, Kim D, Jeon YJ. Algal polysaccharides: potential bioactive substances for cosmeceutical applications. Crit Rev Biotechnol 2019;39(1):99e113. [133] Skriptsova AV. Fucoidans of brown algae: biosynthesis, localization, and physiological role in thallus. Russ J Mar Biol 2015;41(3):145e56.

Further reading [1] Castro LSEPW, de Sousa Pinheiro T, Castro AJG, Santos MDSN, Soriano EM, Leite EL. Potential antiangiogenic, antiproliferative, antioxidant, and anticoagulant activity of anionic polysaccharides, fucans, extracted from brown algae Lobophora variegata. J Appl Phycol 2015;27(3):1315e25. [2] Yuan Y, Macquarrie D. Microwave assisted extraction of sulfated polysaccharides (fucoidan) from Ascophyllum nodosum and its antioxidant activity. Carbohydr Polym 2015;129:101e7. [3] Kim YS, Kim EK, Nawarathna WPAS, Dong X, Shin WB, Park JS, et al. Immune-stimulatory effects of althaea rosea flower extracts through the MAPK signaling pathway in RAW264. 7 cells. Molecules 2017;22(5):679.

C H A P T E R 23

Natural polysaccharides in tissue engineering applications Amit Kumar Nayak1, Syed Anees Ahmed2, Mohammad Tabish3, Md Saquib Hasnain4 1

Department of Pharmaceutics, Seemanta Institute of Pharmaceutical Sciences, Mayurbhanj, Odisha, India; 2Department of Pharmacology, Hygia Institute of Pharmaceutical Application and Research, Lucknow, India; 3Department of Pharmacology, College of Medicine, Saqra University, Kingdom of Saudi Arabia; 4Department of Pharmacy, Shri Venkateshwara University, Gajraula, India

Chapter Outline 1. 2. 3. 4. 5.

Introduction 531 Sources of natural polysaccharides 534 Functionalization of polysaccharides 534 Tissue engineering 535 Role of natural polysaccharides in tissue engineering 5.1 5.2 5.3 5.4 5.5 5.6 5.7

Alginates 536 Chitosan 537 Cellulose 539 Starch 539 Pectins 540 Dextran and pullulan 540 Gellan gum and xanthan gum

536

540

6. Conclusion 541 References 541

1. Introduction Polymers are macromolecules made up of repeated structural units, which are chemically linked with covalent bond. The word “polymer” is originated from a Greek word “polus” meaning “much/many” and “meros” meaning “part.” Thus, polymers are the structural molecules made up of several repeated units with relatively higher molecular weight [1,2]. The polymeric units are made up of molecules having relatively lower molecular weight. The word “polymer” was coined by J.J. Berzelius in 1833 [1]. Natural Polysaccharides in Drug Delivery and Biomedical Applications. https://doi.org/10.1016/B978-0-12-817055-7.00023-6 Copyright © 2019 Elsevier Inc. All rights reserved.

531

532 Chapter 23 During past few decades, various natural polymers (polysaccharides and proteins) are being exploited for the use in different important biomedical applications [2e6]. Even, these are being exploited to develop various useful strategies for the use in tissue engineering regenerative medicines (TERM) (Fig. 23.1). Polysaccharides are composed of numerous polymeric materials of natural origin, such as algae, plants, and animals having a linkage of glycosidic monosaccharides [2,7,8]. Thus, on the basis of nature of monosaccharide units, polysaccharides are divided into two groups: 1. Linear chain polysaccharides, and 2. Branched chain polysaccharides Polysaccharides have numerous reactive functional groups such as amino, hydroxyl, carboxylic acid, etc., groups. There are possibilities of chemical modification because of different functional groups attached to the compounds [9]. The molecular mass of polysaccharides occurring naturally ranges from 100 to 1000 Da [10]. The naturally occurring polysaccharides are biological macromolecules consisting of one or more monosaccharides connected with each other in a chain of sequence [11]. The linkings of monosaccharides are arranged in numerous ways and form either linear or branched polysaccharides.

Figure 23.1 Classification of natural polymers with their important properties, biomedical applications, and strategies for tissue engineering regenerative medicine (TERM).

Natural polysaccharides in tissue engineering applications 533 Natural polysaccharides are naturally formed by the process of capturing carbon, known as photosynthesis, and by the biosynthetic modifications. These are found in abundance in nature and have its own industrial importance in respect to the medicinal aspect and some are used as food. Polysaccharides are the carbohydrate molecules having long repeated monosaccharide units linked together by glycosidic bonds [12,13]. These comprise of plentiful biomolecules in nature and play vital role in numerous processes. Polysaccharides are intended to be used as raw materials in several kinds of textiles, cosmetics, foods, paints, papers, adhesives, and biopharmaceutical organizations [14e21]. Number of hydrogels containing polysaccharides has been reviewed for modified release formulations [22]. These have now become the essential part for drug delivery; these may be conventional or novel drug delivery systems. The polysaccharides are particularly the one that is swellable and are promising versatile carriers [23e25]. The use of polysaccharides is advantageous as these have a wide range of properties such as solubility, permeability, and diffusivity, which are essential for the controlled drug delivery [26,27]. Therefore, a variety of natural polysaccharides have gained much interest because of the immense pharmaceutical/biopharmaceutical usages such as thickening, binding, gelling, suspending, emulsifying, stabilizing, film forming, matrix forming, disintegrating, and controlled release matrix forming [21,22]. The natural polysaccharides are preferred to use over semisynthetic and synthetic excipients and described in Fig. 23.2. There are certain limitations that need to be resolved such as predisposition to microbial attack, decreased drug loading capacity, and possibility of variations from batch to batch.

Figure 23.2 Advantages of natural polysaccharides.

534 Chapter 23

2. Sources of natural polysaccharides Polysaccharides are found abundantly in nature and usually found in many higher plants, seaweeds, fungi, plants, and animals [17,18]. These are also obtained from the microbial sources, where these carry out numerous structural and metabolic functions. Kingdom Plantae provides a huge quantity of polysaccharides [2]. These comprise a structurally diverse class of biological macromolecules having a wider range of physicochemical properties, used in numerous applications in pharmaceuticals and medicines [28e30]. The different types of polysaccharides are enlisted in the table given below (Table 23.1).

3. Functionalization of polysaccharides Natural polysaccharides contain number of functional groups such as carboxyl and hydroxyl groups to provide attachment of other functional groups for modification in their chemical nature [31]. Recently, the natural polysaccharides are being modified in different ways to design formulation for the delivery of drugs and are being explored for the use in place of synthetic excipients or polymers [32,33]. The synthetic polymers have already been replaced by the use of functionalized biopolymers [34]. The functionalizations of polysaccharides are made to prevent the interaction in between the polymers as well as in between drugs and polymers to increase the drug loading capability, drug release profile, solubility, and to alter the bioadhesive or mucoadhesive property [32,34]. It has become essential in the fields of drug delivery and biopharmaceutical/pharmaceutical technology. The importance of studying the effect of chemical modifications on the functional and structural properties of polysaccharides lies in the design of new molecules with altered physicochemical properties [34,35]. The functionalization of polysaccharides as reported is done by the process of oxidation, reduction, ethylation, carboxymethylation, quaternization, etherification, thiolation, cross-linking, grafting, acrylation, PEGylation, and so on [17,21,32,34e36]. Among these, cross-linking and grafting are being commonly used for the functionalization of biopolysaccharides [35].

Table 23.1: Natural polysaccharides obtained from various sources. Sources

Examples

Microbial origin (bacteria and fungi) Marine origin (algal or seaweed gums) Animal origin Plant origin

Dextran, xanthan, schizophyllan, curdian, pullulan, Baker’s yeast glycan, scleroglucan, zanflo, emulsan, krestin, lentinan, etc. Laminarin, carrageenans, agar, alginic acid, etc. Chitin, chitosan, chondroitin sulfate, hyaluronic acid, etc. Starch, gum karaya, pectin, albizia gums, cellulose, gum Arabica, gum ghatti, gum tragacanth, khaya gum, guar gum, tamarind gum, locust bean gum, amylose, fenugreek gum, potato starch, psyllium, etc.

Natural polysaccharides in tissue engineering applications 535

4. Tissue engineering Tissue engineering is an interdisciplinary field that implies the application of principles and methods of engineering and biological sciences in understanding the structure function relationship in tissues of living organism as well as formation of biological substitutes to restore the normal functioning [8,29]. From both therapeutic as well as economic standpoint, the potential impact of tissue engineering is enormous [37]. The strategy for tissue engineering is shown below (Fig. 23.3). Generally, there are multiple properties of polysaccharides due to which these are used as a starting material and have numerous biomaterial applications. The presence of multiple eOH groups on particular molecule enables at different sites for the attachment of various side groups. These functional groups attached to the molecules give specificity in function and feature for the recognition of biological and even alter the biological or mechanical properties of the parent molecules. The polysaccharides have the capability to be processed as hydrogels of numerous densities because of its hydrophilic nature [8,29]. The polysaccharidic molecules containing charges have additional property that may be useful in the development of materials. The natural polysaccharides and these monomers are degraded easily, and these items are normally nontoxic or least toxic [38,39]. The generalized combined method from the material sciences and life sciences utilized to regenerate the artificially developed constructs consisting of matrices (scaffolds) along with living cells is called as “tissue engineering” [40]. To meet the above requirement, an interdisciplinary field has been emerged from past few decades that include the methods and concepts of engineering, medicine, and biology. Recent years, various kinds

Figure 23.3 Strategy for tissue engineering.

536 Chapter 23 of nanocomposite materials are used for tissue engineerings of bone, dental and for prosthetics and cell therapies are quite different from the tissue engineering, one need to focus upon tissue differentiation processes [41e47]. There is an increasing attention towards the polysaccharide-based scaffold materials, nowadays, and are mainly because of: 1. Deep knowledge about the growing body and the role of saccharide entity in cell signaling and in immune recognition. 2. Formulations of potent and cost-effective techniques for potent biological oligosaccharide syntheses permitting decode and developing the signaling of oligosaccharides. 3. Its need in tissue engineering for newer scaffold materials with precise, accurate, and controlled biological activity in combination with varied degradation and resorption kinetics.

5. Role of natural polysaccharides in tissue engineering 5.1 Alginates Alginates are anionic polymer found naturally and chemically, a copolymer of a-(1, 4)-linked L-guluronic acid and b-(1, 4) D-mannuronic acid [48,49]. It belongs to the family of linear polysaccharide. It is produced or obtained from brown seaweeds and bacterial species such as A. chroococcum, Azotobacter, and Pseudomonas. In comparison with other polysaccharides, alginate molecules do not have repeated monomer sequence units along the chain of polymer. Therefore, there is an immense heterogeneity in composition among the family of alginates isolated from various organisms and in samples obtained from the various parts of the same organisms [50]. It is widely used and has biomedical applications because of its least toxicity, biocompatibility, gelatin property by the addition of cations (divalent or trivalent), cost-effectiveness, etc [51e55]. Day-by-day applications of alginates in biopharmaceuticals and in pharmaceuticals are increasing, which have led to carry out major researches aiming to understand the major metabolic pathways, biological/physiological functions, and production of microbes, its optimization, and quality maintenance [56e63]. Alginate is an approved polymer by the US Food and Drug Administration (USFDA). It is the most widely used biomaterial in the field of regenerative medicine, semipermeable separation, and nutritional supplements [64]. Alginates are traditionally used in food industry. It is used as emulsifying, stabilizing agent, and gelling agent. To use alginates in acidic conditions, it is modified chemically by propylene glycol and used in milled food stuffs [65,66]. It is used in paper and textile printing industry because of its shear-thinning agent and its print levelness and higher color yield [66]. In pharmaceuticals, it is used as drug excipients [66e68], dressing of wounds [69], impression material for dental applications [70], etc.

Natural polysaccharides in tissue engineering applications 537 Alginates, as the biomaterial, have numerous advantages that include nonimmunogenicity, biodegradability, and biocompatibility [71]. Its strong gelling nature permits the encapsulation of numerous substances with negligible trauma [72]. Various chemically modified alginates are being clinically used at present as carrier matrices for drug delivery of proteins that enhances regeneration of mineralized tissues [73] and used as carriers for the cell transplantations [74]. In the tissue engineering field, alginates are being used in formulation and development of beads, microparticles, nanoparticles, hydrogels, composites, nanocomposites, scaffolds, etc [21,54,64]. It has numerous applications such as cell encapsulation, delivery of protein, and wound healing [75]. The hydrogels of alginates possess wide range of application. These have been used in numerous studies for the delivery of chondrocytes [74]. These also have been given by subcutaneous route at the defective sites [76,77]. Recently, the concept of fabrication of alginates was developed by Bonassar et al., by the use of silicone-based molding technology [78]. For the differentiation of mesenchymal stem cells to chondrocytes, the alginate gel was used as matrices [79]. For the fabrication of cartilage in nude mouse model, Paige et al. used alginates for the delivery of isolated chondrocytes [76]. According to Yang et al., alginate hydrogel polymerization may be controlled by injecting chondrocytes to produce cartilage in rabbit at the subcutaneously dorsal site [80]. Alginate hydrogels are being widely used for the localized and sustained delivery of drugs [21]. The drugs used has traditionally low molecular weight for tissue regeneration applications [8,29]. The release kinetics of low molecular weight drugs from the alginate gels may be controlled by the drugealginate interactions. In the tissue engineering field, alginate gels are used as vehicle for cell delivery [81]. These are permitted for the transplanted cell localization and also, to control the fate [78,81]. Alginate gels are tremendously versatile and flexible biomaterials. These are useful in the extensively diverse medical applications. These materials have been employed as the supportive matrices, as cell delivery vehicles to be used in tissue engineering, as model extracellular matrix for in vitro experiments related to cells, and as depots for different drugs. In tissue engineering, alginate-based hydrogels and microsphere combinations have been used for the controlled growth factor delivery [82e84].

5.2 Chitosan Chitin is a long chain of N-acetylglucosamine, a derivative of glucose [85]. It is found in the exoskeleton of insects and crustaceans [86]. It is a mucopolysaccharide occurring naturally and most abundant polysaccharide on earth. In terms of abundance, it ranks second to cellulose [87]. It is cellulose derivative having amine group (in place of hydroxyl group) and is polycationic. Chitosan is the major derivative of chitin obtained by

538 Chapter 23 the process of alkaline deacetylation of chitin [88,89]. It is a linear polysaccharide, composed of semicrystalline polysaccharide (1,4)-2-amino-2-deoxy-b-D-glucan (D-glucosamine) and (1,4)-2-acetamido-2-deoxy-b-D-glucan (N-acetyl D-glucosamine) [90]. Owing to least biodegradability and toxicity, chitosan has significant pharmaceutical applications. It has been reported that chitosan has numerous useful pharmacological activities such as hypocholesterolemic [91], antiulcer [92], wound healing [93], antitumor [94], hemostatic [95], and spermicidal [96]. Recently, chitosan has been used in tissue engineering applications and as matrix materials for the delivery of drugs [88,89,97]. In the pharmaceutical industry, chitosan is being widely used. It is used as excipients in the formulations of tablets and gels [98,99], for enhancement of absorption [100,101], controlled release dosage forms [102,103], drug dissolution [104], formulation and development of nanoparticles and microparticles [105], and in wound healing items [93,106]. Chitosan-based nanoparticles and microparticles are being widely used as devices for the delivery of numerous drug molecules [107]. The two important features of chitosan responsible for its use as matrix materials in drug delivery are its molecular weight and degree of acetylation. It affects its hydrophobicity and aqueous solubility, and thus, it is capable of altering drug encapsulations [108]. In neutral and basic conditions, chitosan is sparingly soluble in aqueous solvents. In acidic conditions, owing to protonation of the amino group, the solubility of chitosan increases [109]. Another significant feature of chitosan is its use as transiently opening of tight epithelial junctions and as mucoadhesive agent in bioadhesive drug delivery. Because of this property of chitosan, it is widely used in the delivery of drug molecules across the epithelial cells of lungs [110], intestinal mucosa [111,112], nasal mucosa [113,114], buccal mucosa [115], and eye [116]. Because of these reasons, chitosan has been used for coating of numerous nanoparticle-based formulations [117]. For the synthesis of chitosan nanoparticles, the commonly used methods are ionic gelation technique, reverse micellar technique, precipitation/coacervation-based techniques, and self-assembly technique [107]. Thus, for the synthesis of drug-encapsulated chitosan nanoparticles, these techniques are widely used and have been reported [107,118]. The use of chitosan is restricted as carrier material for delivery of drug molecules because of its low solubility in the biological samples (pH 7.4) [119]. Numerous methods have been employed for the improvement of chitosan solubility such as acetylation [120], alkylation [121], quaternization [122], generation of N-trimethyl chitosan [123], carboxymethylation [120], formation of chitosan/polyol salt combinations [124], conjugation of polyethylene oxide [125], etc. Sugar-bearing chitosans were developed by Park et al. with increased solubility and broad range of pH value [119]. Among the derivatives of chitosan having enhanced hydrophilicity, glycol chitosan is a well-known drug carrier material due to its biocompatibility and water solubility [126]. Thus, the particles produced from glycol chitosan are used in drug delivery of different

Natural polysaccharides in tissue engineering applications 539 therapeutic molecules such as L-asparginase, doxorubicin, and siRNA [127]. Because of the characteristic physicochemical properties of chitosan and its derivatives, these possess numerous advantageous features like cell/tissue supporting biomaterial matrices. By using the processes such as electrospinning, lyophilization, and gas foaming, chitosan are molded to porous scaffolds [128e130]. These are prepared through blending by using different types of natural and synthetic polymers [131,132]. The development of polyelectrolyte complexes having broad range of anionic glycosaminoglycans from chitosan is due to its cationic nature. It also includes chondroitin sulfate and heparin. Thus, the activity of number of cytokines and growth factors are altered by chitosan glycosaminoglycans. It has been experimentally performed for cartilage and bone regeneration [133,134]. As the chitosan is a polymer, which is biodegradable and its rate of degradation can be controlled by altering the degree of deacytylation, it can be used for nearly all types of tissues because of its varying rates of degradation [135]. It is also used in dentistry as it possesses regenerative property, biocompatibility, and ease in treatment with chemicals. Chitosan is also used for the treatment of damaged tissues of oral cavity. Because of its regenerative and antibacterial property having enhanced biocompetency, it is used as a potential biomaterial in the field of medicine and dentistry [136].

5.3 Cellulose Cellulose is a naturally occurring polymer and most abundant in nature [137]. It is made up of b-(1,4) linked-D-glucopyranose units, discovered in 1838, by Payen, in the green plants [138]. The chemical structure of cellulose is (C6H10O5)n. It is water-insoluble, stable, and fibrous polysaccharide. The plant-based cellulose has wide range of applications in biomedical and in tissue engineering [8,29]. The oxidized cellulose is broadly utilized in wound healing materials having good features such as antibacterial, antiviral, antiadhesive, etc., properties. It has increased absorbability as nontoxic in nature [139,140]. As cellulose has the capability to increase the coagulation of blood at the site, the oxidized cellulose has been used as a hemostatic material [141]. It is also used as a promising carrier for controlled drug delivery [142]. In tissue engineering field, oxidized dialdehyde cellulose has been used as biodegradable scaffold, such as lamina propria [139e141].

5.4 Starch Starch is occurred in plants as a storage polysaccharide [15]. It is in form of branched as amylopectin and linear as amylase [15,56]. Pure starch microfibrils prepared by electrospinning technique have been investigated for the use in tissue engineering applications [143]. In combinations, starch has been used with various synthetic or natural polymers to accomplish enhanced bioactivity and mechanical properties of the

540 Chapter 23 scaffolds [8,29]. Collagen I nanofibers in combination with starch microfibers were made to produce extracellular matrix equivalent for bone tissue engineering applications [144]. Polylactide or polycaprolactone, a synthetic polymer, is blended with starch for the purpose of bone tissue engineering [145]. The advantage of combination or blending with starch improves the differentiation and growth of articular chondrocytes on polycaprolactone scaffolds [146].

5.5 Pectins Pectins are polysaccharides that are complex in nature, having residues of a-(1,4)-linkedD-galactosyl uronic acid [12,13]. It is mostly found in nonwoody part of terrestrial plants and primarily in cell walls. As pectin is cytocompatible, it has numerous biomedical applications such as in tissue engineering, gene delivery, drug delivery, and wound healing applications [147]. The deposition of nanostructured pectin films over glass, polystyrene, and titanium substrates enhances the growth, adhesion, and differentiation of osteogenic cells of primary rat osteoblasts and murine preosteoblastic MC3T3-E1 cells [148]. Pectins have been used in combinations for the potential bone tissue engineering. For this purpose, it is blended with chitosan (a natural polymer) [149] and polylactide (a synthetic polymer) [150].

5.6 Dextran and pullulan Dextran is a naturally occurring polysaccharide made up of repeated units of glucose having chain of different lengths ranging from 3 to 2000 kDa [29]. It is synthesized by specific lactic acid bacteria such as Lactobacillus brevis, Streptococcus mutans, and Leuconostoc mesenteroides. Medically, it is used as antithrombotic. In combination with pullulan, it has potential applications in tissue engineering. Pullulan is another naturally occurring polysaccharide having units of maltotriose formed by Aureobasidium pullulans (fungi). The combination of both dextran and pullulan has been used in vascular and bone tissue engineering [151].

5.7 Gellan gum and xanthan gum Gellan gum and xanthan gum are two natural-occurring polysaccharides [1,2]. These have been proved and shown to be superior carriers for matrices, the releasing of drugs, proteins, cells, growth factors, etc [29]. These have been employed for numerous applications in tissue engineering. The combination of gellan gum and xanthan gum forms gels within seconds and retains huge amount of water providing same requirement for environmental conditions like that for extracellular matrices [152,153]. The derivatives of xanthan gum are employed for the encapsulation of drug and chondrocyte delivery in cartilage tissue engineering [154].

Natural polysaccharides in tissue engineering applications 541

6. Conclusion Tissue engineering has put an immense impact in preparation of various types of formulations by using natural polysaccharides. It is mainly multidisciplinary and interdisciplinary areas of research mounting exponentially with time. Fabrication and scaffold materials have an essential role in tissue engineering applications. The carrier properties such as specific cell type targeting of biomacromolecules and selective tissue distribution must be investigated. The tissue engineering process also allows the simple manufacturing and fabrication of high-quality polysaccharide-based carriers. A continuous research progress is going on for the design and development of various multifunctional scaffolds made of natural polysaccharides for the use in tissue engineering applications. Still, more researches are required for further progress to overcome several limitations in using natural polysaccharides in tissue engineering applications.

References [1] Bashir A, Warsi MH, Sharma PK. An overview of natural gums as pharmaceutical excipient: their chemical modification. World J Pharm Pharmaceut Sci 2016;5:2025e39. [2] Shukla RK, Tiwari A. Carbohydrate polymers: applications and recent advances in delivering drugs to the colon. Carbohydr Polym 2012;88:399e416. [3] Ray S, Sinha P, Laha B, Maiti S, Bhattacharyya UK, Nayak AK. Polysorbate 80 coated crosslinked chitosan nanoparticles of ropinirole hydrochloride for brain targeting. J Drug Deliv Sci Technol 2018;48:21e9. [4] Jena AK, Nayak AK, De A, Mitra D, Samanta A. Development of lamivudine containing multiple emulsio stabilized by gum odina. Future J Pharm Sci 2018;4:71e9. [5] Nayak AK, Pal D. Tamarind seed polysaccharide: an emerging excipient for pharmaceutical use. Indian J Pharm Educ Res 2017;51:S136e46. [6] Verma A, Dubey J, Verma N, Nayak AK. Chitosan-hydroxypropyl methylcellulose matrices as carriers for hydrodynamically balanced capsules of moxifloxacin HCl. Curr Drug Deliv 2017;14:83e90. [7] Nayak AK, Pal D, Santra K. Swelling and drug release behavior of metformin HCl-loaded tamarind seed polysaccharide-alginate beads. Int J Biol Macromol 2016;82:1023e7. [8] Nayak AK, Pal D. Natural polysaccharides for drug delivery in tissue engineering. Everyman’s Sci 2012;XLVI:347e52. [9] Liu Z, Jiao Y, Wang Y, Zhou C, Zhang Z. Polysaccharides-based nanoparticles as drug delivery systems. Adv Drug Deliv Rev 2008;60:1650e62. [10] Saravanakumar G, Jo D-G, Park J. Polysaccharide-based nanoparticles: a versatile platform for drug delivery and biomedical imaging. Curr Med Chem 2012;19:3212e29. [11] Nayak AK, Pal D, Santra K. Screening of polysaccharides from tamarind, fenugreek and jackfruit seeds as pharmaceutical excipients. Int J Biol Macromol 2015;79:756e60. [12] Nayak AK, Pal D, Santra K. Development of pectinate-ispagula mucilage mucoadhesive beads of metformin HCl by central composite design. Int J Biol Macromol 2014;66:203e21. [13] Nayak AK, Pal D, Santra K. Development of calcium pectinate-tamarind seed polysaccharide mucoadhesive beads containing metformin HCl. Carbohydr Polym 2014;101:220e30. [14] Nayak AK, Bera H, Hasnain MS, Pal D. Graft-copolymerization of plant polysaccharides. In: Thakur VK, editor. Biopolymer grafting, synthesis and properties. Elsevier Inc.; 2018. p. 1e62.

542 Chapter 23 [15] Nayak AK, Pal D. Natural starches-blended ionotropically-gelled microparticles/beads for sustained drug release. In: Thakur VK, Thakur MK, Kessler MR, editors. Handbook of composites from renewable materials, volume 8, nanocomposites: advanced applications. USA: Wiley-Scrivener; 2017. p. 527e60. [16] Pal D, Nayak AK. Plant polysaccharides-blended ionotropically-gelled alginate multiple-unit systems for sustained drug release. In: Thakur VK, Thakur MK, Kessler MR, editors. Handbook of composites from Renewable materials, volume 6, polymeric composites. USA: Wiley-Scrivener; 2017. p. 399e400. [17] Nayak AK, Pal D. Sterculia gum-based hydrogels for drug delivery applications. In: Kalia S, editor. Polymeric hydrogels as smart biomaterials, Springer series on polymer and composite materials. Switzerland: Springer International Publishing; 2016. p. 105e51. [18] Nayak AK. Tamarind seed polysaccharide-based multiple-unit systems for sustained drug release. In: Kalia S, Averous L, editors. Biodegradable and bio-based polymers: environmental and biomedical applications. USA: Wiley-Scrivener; 2016. p. 471e94. [19] Nayak AK, Pal D. Plant-derived polymers: ionically gelled sustained drug release systems. In: Mishra M, editor. Encyclopedia of biomedical polymers and polymeric biomaterials, vol. VIII. New York, NY, USA: Taylor & Francis Group; 2016. p. 6002e17. [20] Nayak AK, Pal D. Chitosan-based interpenetrating polymeric network systems for sustained drug release. In: Tiwari A, Patra HK, Choi J-W, editors. Advanced theranostics materials. USA: Wiley-Scrivener; 2015. p. 183e208. [21] Pal D, Nayak AK. Alginates, blends and microspheres: controlled drug delivery. In: Mishra M, editor. Encyclopedia of biomedical polymers and polymeric biomaterials, vol. I. New York, NY, USA: Taylor & Francis Group; 2015. p. 89e98. [22] Coviello T, Matricardi P, Marianecci C, Alhaique F. Polysaccharide hydrogels for modified release formulations. J Control Release 2007;2007:5e24. [23] Pal D, Nayak AK. Interpenetrating polymer networks (IPNs): natural polymeric blends for drug delivery. In: Mishra M, editor. Encyclopedia of biomedical polymers and polymeric biomaterials, vol. VI. New York, NY, USA: Taylor & Francis Group; 2015. p. 4120e30. [24] Sinha P, Ubaidulla U, Nayak AK. Okra (Hibiscus esculentus) gum-alginate blend mucoadhesive beads for controlled glibenclamide release. Int J Biol Macromol 2015;72:1069e75. [25] Nayak AK, Pal D, Das S. Calcium pectinate-fenugreek seed mucilage mucoadhesive beads for controlled delivery of metformin HCl. Carbohydr Polym 2013;96:349e57. [26] Das B, Dutta S, Nayak AK, Nanda U. Zinc alginate-carboxymethyl cashew gum microbeads for prolonged drug release: development and optimization. Int J Biol Macromol 2014;70:505e15. [27] Nayak AK, Pal D, Santra K. Tamarind seed polysaccharide-gellan mucoadhesive beads for controlled release of metformin HCl. Carbohydr Polym 2014;103:154e63. [28] Prajapati VD, Jani GK, Moradiya NG, Randeria NP. Pharmaceutical applications of various natural gums, mucilages and their modified forms. Carbohydr Polym 2013;92:1685e99. [29] Hasnain M, Nayak AK, Singh R, Ahmad F. Emerging trends of natural-based polymeric systems for drug delivery in tissue engineering applications. Sci J UBU 2010;1:1e13. [30] Nayak AK, Ahmad SA, Beg S, Ara TJ, Hasnain MS. Drug delivery: present, past and future of medicine. In: Inamuddin, Asiri AM, Mohammad A, editors. Applications of nanocomposite materials in drug delivery, a volume in Woodhead Publishing Series in biomaterials. Elsevier Inc.; 2018. p. 255e82. [31] Quignard F, Di Renzo F, Guibal E. From natural polysaccharides to materials for catalysis, adsorption, and remediation. Carbohydrates in sustainable development I. Springer; 2010. p. 165e97. [32] Pal D, Nayak AK, Saha S. Interpenetrating polymer network hydrogels of chitosan: applications in controlling drug release. In: Mondal IH, editor. Cellulose-based superabsorbent hydrogels, polymers and polymeric composites: a reference series. Springer; 2018. p. 1e41. [33] Bera H, Boddupalli S, Nayak AK. Mucoadhesive-floating zinc-pectinate-sterculia gum interpenetrating polymer network beads encapsulating ziprasidone HCl. Carbohydr Polym 2015;131:108e18. [34] Nayak AK, Pal D. Functionalization of tamarind gum for drug delivery. In: Thakur VK, Thakur MK, editors. Functional biopolymers. Switzerland: Springer International Publishing; 2018. p. 35e56.

Natural polysaccharides in tissue engineering applications 543 [35] Cumpstey I. Chemical modification of polysaccharides. ISRN Org Chem 2013:2013. [36] Jana S, Saha A, Nayak AK, Sen KK, Basu SK. Aceclofenac-loaded chitosan-tamarind seed polysaccharide interpenetrating polymeric network microparticles. Colloids Surf B Biointerf 2013;105:303e9. [37] Vacanti JP, Langer R. Tissue engineering: the design and fabrication of living replacement devices for surgical reconstruction and transplantation. Lancet 1999;354:S32e4. [38] Matthew HW. Polymers for tissue engineering scaffolds. Polymeric biomaterials, revised and expanded. CRC Press; 2001. p. 181e200. [39] Suh J-KF, Matthew HW. Application of chitosan-based polysaccharide biomaterials in cartilage tissue engineering: a review. Biomaterials 2000;21:2589e98. [40] Pa`mies P. Tissue engineering: paper scaffolds. Nat Mater 2015;15:3. [41] Hasnain MS, Nayak AK. Nanocomposites for improved orthopedic and bone tissue engineering applications. In: Inamuddin, Asiri AM, Mohammad A, editors. Applications of nanocomposite materials in orthopedics, a volume in Woodhead Publishing Series in biomaterials. Elsevier Inc.; 2019. p. 145e77. [42] Hasnain MS, Ahmad SA, Chaudhary N, Hoda MN, Nayak AK. Biodegradable polymer matrix nanocomposites for bone tissue engineering. In: Inamuddin, Asiri AM, Mohammad A, editors. Applications of nanocomposite materials in orthopedics, a volume in Woodhead Publishing Series in biomaterials. Elsevier Inc.; 2019. p. 1e37. [43] Hasnain MS, Ahmad SA, Minhaj MA, Ara TJ, Nayak AK. Nanocomposite materials for prosthetic devices. In: Inamuddin, Asiri AM, Mohammad A, editors. Applications of nanocomposite materials in orthopedics, a volume in Woodhead Publishing Series in biomaterials. Elsevier Inc.; 2019. p. 127e44. [44] Nayak AK, Mazumder S, Ara TJ, Ansari MT, Hasnain MS. Calcium fluoride-based dental nanocomposites. In: Inamuddin, Asiri AM, Mohammad A, editors. Applications of nanocomposite materials in dentistry, a volume in Woodhead Publishing Series in biomaterials. Elsevier Inc.; 2019. p. 27e45. [45] Rani P, Pal D, Hoda MN, Ara TJ, Beg S, Hasnain MS, Nayak AK. Dental pulp capping nanocomposites. In: Inamuddin, Asiri AM, Mohammad A, editors. Applications of nanocomposite materials in dentistry, a volume in Woodhead Publishing Series in biomaterials. Elsevier Inc.; 2019. p. 65e91. [46] Hasnain MS, Ahmad SA, Chaudhary N, Minhaj MA, Nayak AK. Degradation and failure of dental composite materials. In: Inamuddin, Asiri AM, Mohammad A, editors. Applications of nanocomposite materials in dentistry, a volume in Woodhead Publishing Series in biomaterials. Elsevier Inc.; 2019. p. 108e21. [47] Mazumder S, Nayak AK, Ara TJ, Hasnain MS. Hydroxyapatite composites for dentistry. In: Inamuddin, Asiri AM, Mohammad A, editors. Applications of nanocomposite materials in dentistry, a volume in Woodhead Publishing Series in biomaterials. Elsevier Inc.; 2019. p. 108e21. [48] Malakar J, Das K, Nayak AK. In situ cross-linked matrix tablets for sustained salbutamol sulfate release - formulation development by statistical optzation. Polym Med 2014;44:221e30. [49] Hasnain MS, Rishishwar P, Rishishwar S, Ali S, Nayak AK. Isolation and characterization of Linum usitatisimum polysaccharide to prepare mucoadhesive beads of diclofenac sodium. Int J Biol Macromol 2018;116:162e72. [50] Smidsrod O, Draget K. Chemistry and physical properties of alginates. 1996. [51] Nayak AK, Pal D. Development of pH-sensitive tamarind seed polysaccharide-alginate composite beads for controlled diclofenac sodium delivery using response surface methodology. Int J Biol Macromol 2011;49:784e93. [52] Nayak AK, Khatua S, Hasnain MS, Sen KK. Development of alginate-PVP K 30 microbeads for controlled diclofenac sodium delivery using central composite design. DARU J Pharm Sci 2011;19:356e66. [53] Malakar J, Nayak AK, Pal D. Development of cloxacillin loaded multiple-unit alginate-based floating system by emulsionegelation method. Int J Biol Macromol 2012;50(1):138e47.

544 Chapter 23 [54] Hasnain MS, Nayak AK. Alginate-inorganic composite particles as sustained drug delivery matrices. In: Inamuddin, Asiri AM, Mohammad A, editors. Applications of nanocomposite materials in drug delivery, a volume in Woodhead Publishing Series in biomaterials. Elsevier Inc.; 2018. p. 39e74. [55] Nayak AK, Ara TJ, Hasnain MS, Hoda N. Okra gum-alginate composites for controlled releasing drug delivery. In: Inamuddin, Asiri AM, Mohammad A, editors. Applications of nanocomposite materials in drug delivery, a volume in Woodhead Publishing Series in biomaterials. Elsevier Inc.; 2018. p. 761e85. [56] Malakar J, Nayak AK, Jana P, Pal D. Potato starch-blended alginate beads for prolonged release of tolbutamide: development by statistical optimization and in vitro characterization. Asian J Pharm 2013;7:43e51. [57] Nayak AK, Pal D, Santra K. Plantago ovata F. Mucilage-alginate mucoadhesive beads for controlled release of glibenclamide: development, optimization, and in vitro-in vivo evaluation. J Pharm 2013;2013. Article ID 151035. [58] Nayak AK, Pal D, Hasnain MS. Development and optimization of jackfruit seed starch-alginate beads containing pioglitazone. Curr Drug Deliv 2013;10:608e19. [59] Hasnain MS, Nayak AK, Singh M, Tabish M, Ansari MT, Ara TJ. Alginate-based bipolymericnanobioceramic composite matrices for sustained drug release. Int J Biol Macromol 2016;83:71e7. [60] Pal D, Nayak AK. Development, optimization and anti-diabetic activity of gliclazide-loaded alginate-methyl cellulose mucoadhesive microcapsules. AAPS PharmSciTech 2011;12:1431e41. [61] Nayak AK, Das B, Maji R. Calcium alginate/gum Arabic beads containing glibenclamide: development and in vitro characterization. Int J Biol Macromol 2012;51:1070e8. [62] Nayak AK, Pal D, Pradhan J, Hasnain MS. Fenugreek seed mucilage-alginate mucoadhesive beads of metformin HCl: design, optimization and evaluation. Int J Biol Macromol 2013;54:144e54. [63] Nayak AK, Beg S, Hasnain MS, Malakar J, Pal D. Soluble starch-blended Ca2þ-Zn2þ-alginate composites-based microparticles of aceclofenac: formulation development and in vitro characterization. Future J PharmSci 2018;4:63e70. [64] Bouhadir KH, Lee KY, Alsberg E, Damm KL, Anderson KW, Mooney DJ. Degradation of partially oxidized alginate and its potential application for tissue engineering. Biotechnol Prog 2001;17:945e50. [65] Rowley JA, Madlambayan G, Mooney DJ. Alginate hydrogels as synthetic extracellular matrix materials. Biomaterials 1999;20:45e53. [66] Glicksman M. Utilization of seaweed hydrocolloids in the food industry. Hydrobiol (Sofia) 1987;151:31e47. [67] Sinha P, Ubaidulla U, Hasnain MS, Nayak AK, Rama B. Alginate-okra gum blend beads of diclofenac sodium from aqueous template using ZnSO4 as a cross-linker. Int J Biol Macromol 2015;79:555e63. [68] Bera H, Kandukuri SG, Nayak AK, Boddupalli S. Alginate-sterculia gum gel-coated oil-entrapped alginate beads for gastroretentive risperidone delivery. Carbohydr Polym 2015;120:74e84. [69] Matthew IR, Browne RM, Frame JW, Millar BG. Subperiosteal behaviour of alginate and cellulose wound dressing materials. Biomaterials 1995;16:275e8. [70] Ashley M, McCullagh A, Sweet C. Making a good impression: (a‘how to’paper on dental alginate). Dent Update 2005;32:169e70. [71] Shapiro L, Cohen S. Novel alginate sponges for cell culture and transplantation. Biomaterials 1997;18:583e90. [72] Klo¨ck G, Pfeffermann A, Ryser C, Gro¨hn P, Kuttler B, Hahn H-J, et al. Biocompatibility of mannuronic acid-rich alginates. Biomaterials 1997;18:707e13. [73] Bratthall G, Lindberg P, Havemose-Poulsen A, Holmstrup P, Bay L, So¨derholm G, et al. Comparison of ready-to-use EMDOGAIN®-gel and EMDOGAIN® in patients with chronic adult periodontitis: a multicenter clinical study. J Clin Periodontol 2001;28:923e9. [74] Bent AE, Tutrone RT, McLennan MT, Lloyd K, Kennelly MJ, Badlani G. Treatment of intrinsic sphincter deficiency using autologous ear chondrocytes as a bulking agent. Neurourol Urodyn 2001;20:157e65.

Natural polysaccharides in tissue engineering applications 545 [75] Yang J, Han S, Zheng H, Dong H, Liu J. Preparation and application of micro/nanoparticles based on natural polysaccharides. Carbohydr Polym 2015;123:53e66. [76] Paige KT, Cima LG, Yaremchuk MJ, Vacanti JP, Vacanti CA. Injectable cartilage. Plast Reconstr Surg 1995;96:1390e8. [77] Atala A, Kim W, Paige KT, Vacanti CA, Retik AB. Endoscopic treatment of vesicoureteral reflux with a chondrocyte-alginate suspension. J Urol 1994;152:641e3. [78] Chang SC, Rowley JA, Tobias G, Genes NG, Roy AK, Mooney DJ, et al. Injection molding of chondrocyte/alginate constructs in the shape of facial implants. J Biomed Mater Res 2001;55:503e11. [79] Kavalkovich KW, Boynton RE, Murphy JM, Barry F. Chondrogenic differentiation of human mesenchymal stem cells within an alginate layer culture system. In Vitro Cell Dev Biol Anim 2002;38:457e66. [80] Yang W, Chen S, Mao T, Chen F, Lei D, Tao K, et al. A study of injectable tissue-engineered autologous cartilage. Chin J Dental Res 2000;3:10e5. [81] Atala A, Koh C. Applications of tissue engineering in the genitourinary tract. Expert Rev Med Devices 2005;2:119e26. [82] Chen R, Curran SJ, Curran JM, Hunt JA. The use of poly (l-lactide) and RGD modified microspheres as cell carriers in a flow intermittency bioreactor for tissue engineering cartilage. Biomaterials 2006;27:4453e60. [83] Ma K, Titan AL, Stafford M, Zheng C, Levenston ME. Variations in chondrogenesis of human bone marrow-derived mesenchymal stem cells in fibrin/alginate blended hydrogels. Acta Biomater 2012;8:3754e64. [84] Ghahramanpoor MK, Najafabadi SAH, Abdouss M, Bagheri F, Eslaminejad MB. A hydrophobicallymodified alginate gel system: utility in the repair of articular cartilage defects. J Mater Sci 2011;22:2365. [85] Hasnain MS, Nayak AK. Chitosan as responsive polymer for drug delivery applications. In: Makhlouf ASH, Abu-Thabit NY, editors. Stimuli responsive polymeric nanocarriers for drug delivery applications, volume 1, types and triggers. Woodhead Publishing Series in Biomaterials, Elsevier Ltd.; 2018. p. 581e605. [86] Aranaz I, Harris R, Heras A. Chitosan amphiphilic derivatives. Chemistry and applications. Curr Org Chem 2010;14:308e30. [87] Roberts GA. Structure of chitin and chitosan. Chitin chemistry. Springer; 1992. p. 1e53. [88] Jana S, Manna S, Nayak AK, Sen KK, Basu SK. Carbopol gel containing chitosan-egg albumin nanoparticles for transdermal aceclofenac delivery. Colloids Surf B Biointerf 2014;114:36e44. [89] Jana S, Maji N, Nayak AK, Sen KK, Basu SK. Development of chitosan-based nanoparticles through inter-polymeric complexation for oral drug delivery. Carbohydr Polym 2013;98:870e6. [90] Rinaudo M. Chitin and chitosan: properties and applications. Prog Polym Sci 2006;31:603e32. [91] Sugano M, Watanabe S, Kishi A, Izume M, Ohtakara A. Hypocholesterolemic action of chitosans with different viscosity in rats. Lipids 1988;23:187e91. [92] Hillyard IW, Doczi J, Kiernan PB. Antacid and antiulcer properties of the polysaccharide chitosan in the rat. Proc Soc Exp Biol Med 1964;115:1108e12. [93] Kozen BG, Kircher SJ, Henao J, Godinez FS, Johnson AS. An alternative hemostatic dressing: comparison of CELOX, HemCon, and QuikClot. Acad Emerg Med 2008;15:74e81. [94] Suzuki K, Mikami T, Okawa Y, Tokoro A, Suzuki S, Suzuki M. Antitumor effect of hexa-Nacetylchitohexaose and chitohexaose. Carbohydr Res 1986;151:403e8. [95] Millner RW, Lockhart AS, Bird H, Alexiou C. A new hemostatic agent: initial life-saving experience with Celox (chitosan) in cardiothoracic surgery. Ann Thorac Surg 2009;87:e13e4. [96] Felse PA, Panda T. Studies on applications of chitin and its derivatives. Bioprocess Eng 1999;20:505e12. [97] Felt O, Buri P, Gurny R. Chitosan: a unique polysaccharide for drug delivery. Drug Dev Ind Pharm 1998;24:979e93.

546 Chapter 23 [98] Sawayanagi Y, Nambu N, Nagai T. Use of chitosan for sustained-release preparations of water-soluble drugs. Chem Pharm Bull 1982;30:4213e5. [99] Sawayanagi Y, Nambu N, Nagai T. Directly compressed tablets containing chitin or chitosan in addition to lactose or potato starch. Chem Pharm Bull 1982;30:2935e40. [100] Thanou M, Verhoef J, Junginger H. Oral drug absorption enhancement by chitosan and its derivatives. Adv Drug Deliv Rev 2001;52:117e26. [101] Schipper NG, Va˚rum KM, Stenberg P, Ocklind G, Lennerna¨s H, Artursson P. Chitosans as absorption enhancers of poorly absorbable drugs: 3: Influence of mucus on absorption enhancement. Eur J Pharm Sci 1999;8:335e43. [102] Nagai T, Sawayanagi Y, Nambu N. Application of chitin and chitosan to pharmaceutical preparations. Chitin, chitosan, and related enzymes. Elsevier; 1984. p. 21e39. [103] Nigalaye AG, Adusumilli P, Bolton S. Investigation of prolonged drug release from matrix formulations of chitosan. Drug Dev Ind Pharm 1990;16:449e67. [104] Hou W, Miyazaki S, Takada M, Komai T. Sustained release of indomethacin from chitosan granules. Chem Pharm Bull 1985;33:3986e92. [105] Ilium L. Chitosan and its use as a pharmaceutical excipient. Pharm Res 1998;15:1326e31. [106] Ueno H, Mori T, Fujinaga T. Topical formulations and wound healing applications of chitosan. Adv Drug Deliv Rev 2001;52:105e15. [107] Agnihotri SA, Mallikarjuna NN, Aminabhavi TM. Recent advances on chitosan-based micro-and nanoparticles in drug delivery. J Control Release 2004;100:5e28. [108] Sannan T, Kurita K, Iwakura Y. Studies on chitin, 2. Effect of deacetylation on solubility. Makromol Chem 1976;177:3589e600. [109] Kumar MR, Muzzarelli RA, Muzzarelli C, Sashiwa H, Domb A. Chitosan chemistry and pharmaceutical perspectives. Chem Rev 2004;104:6017e84. [110] Al-Qadi S, Grenha A, Carrio´n-Recio D, Seijo B, Remun˜a´n-Lo´pez C. Microencapsulated chitosan nanoparticles for pulmonary protein delivery: in vivo evaluation of insulin-loaded formulations. J Control Release 2012;157:383e90. [111] Artursson P, Lindmark T, Davis SS, Illum L. Effect of chitosan on the permeability of monolayers of intestinal epithelial cells (Caco-2). Pharm Res 1994;11:1358e61. [112] Prego C, Garcia M, Torres D, Alonso M. Transmucosal macromolecular drug delivery. J Control Release 2005;101:151e62. [113] Ferna´ndez-Urrusuno R, Calvo P, Remun˜a´n-Lo´pez C, Vila-Jato JL, Alonso MJ. Enhancement of nasal absorption of insulin using chitosan nanoparticles. Pharm Res 1999;16:1576e81. [114] Vila A, Sa´nchez A, Janes K, Behrens I, Kissel T, Jato JLV, et al. Low molecular weight chitosan nanoparticles as new carriers for nasal vaccine delivery in mice. Eur J Pharm Biopharm 2004;57:123e31. [115] Portero A, Remunan-Lopez C, Criado M, Alonso M. Reacetylated chitosan microspheres for controlled delivery of anti-microbial agents to the gastric mucosa. J Microencapsul 2002;19:797e809. [116] De Campos AM, Sa´nchez A, Alonso Ma J. Chitosan nanoparticles: a new vehicle for the improvement of the delivery of drugs to the ocular surface. Application to cyclosporin A. Int J Pharm 2001;224:159e68. [117] Rodrigues S, Dionı´sio M, Lo´pez CR, Grenha A. Biocompatibility of chitosan carriers with application in drug delivery. J Funct Biomater 2012;3:615e41. [118] Liu H, He J. Simultaneous release of hydrophilic and hydrophobic drugs from modified chitosan nanoparticles. Mater Lett 2015;161:415e8. [119] Park JH, Cho YW, Chung H, Kwon IC, Jeong SY. Synthesis and characterization of sugar-bearing chitosan derivatives: aqueous solubility and biodegradability. Biomacromolecules 2003;4:1087e91. [120] Muzzarelli RA, Tanfani F, Emanuelli M, Mariotti S. N-(carboxymethylidene) chitosans and N-(carboxymethyl) chitosans: novel chelating polyampholytes obtained from chitosan glyoxylate. Carbohydr Res 1982;107:199e214.

Natural polysaccharides in tissue engineering applications 547 [121] Sashiwa H, Shigemasa Y. Chemical modification of chitin and chitosan 2: preparation and water soluble property of N-acylated or N-alkylated partially deacetylated chitins. Carbohydr Polym 1999;39:127e38. [122] Murata J-I, Ohya Y, Ouchi T. Possibility of application of quaternary chitosan having pendant galactose residues as gene delivery tool. Carbohydr Polym 1996;29:69e74. [123] Sieval A, Thanou M, Kotze A, Verhoef J, Brussee J, Junginger H. Preparation and NMR characterization of highly substituted N-trimethyl chitosan chloride. Carbohydr Polym 1998;36:157e65. [124] Chenite A, Chaput C, Wang D, Combes C, Buschmann M, Hoemann C, et al. Novel injectable neutral solutions of chitosan form biodegradable gels in situ. Biomaterials 2000;21:2155e61. [125] Sugimoto M, Morimoto M, Sashiwa H, Saimoto H, Shigemasa Y. Preparation and characterization of water-soluble chitin and chitosan derivatives. Carbohydr Polym 1998;36:49e59. [126] Kim K, Kwon S, Park JH, Chung H, Jeong SY, Kwon IC, et al. Physicochemical characterizations of self-assembled nanoparticles of glycol chitosandeoxycholic acid conjugates. Biomacromolecules 2005;6:1154e8. [127] Yoon HY, Son S, Lee SJ, You DG, Yhee JY, Park JH, et al. Glycol chitosan nanoparticles as specialized cancer therapeutic vehicles: sequential delivery of doxorubicin and Bcl-2 siRNA. Sci Rep 2014;4:6878. [128] Wu X, Black L, Santacana-Laffitte G, Patrick Jr CW. Preparation and assessment of glutaraldehydecrosslinked collagenechitosan hydrogels for adipose tissue engineering. J Biomed Mater Res Part A 2007;81:59e65. [129] Lee J-Y, Tan B, Cooper AI. CO2-in-water emulsion-templated poly (vinyl alcohol) hydrogels using poly (vinyl acetate)-based surfactants. Macromolecules 2007;40:1955e61. [130] Kievit FM, Cooper A, Jana S, Leung MC, Wang K, Edmondson D, et al. Aligned chitosanpolycaprolactone polyblend nanofibers promote the migration of glioblastoma cells. Adv Health Mater 2013;2:1651e9. [131] Sarasam A, Madihally SV. Characterization of chitosanepolycaprolactone blends for tissue engineering applications. Biomaterials 2005;26:5500e8. [132] Huang Y, Onyeri S, Siewe M, Moshfeghian A, Madihally SV. In vitro characterization of chitosanegelatin scaffolds for tissue engineering. Biomaterials 2005;26:7616e27. [133] Kavya K, Dixit R, Jayakumar R, Nair SV, Chennazhi KP. Synthesis and characterization of chitosan/ chondroitin sulfate/nano-SiO2 composite scaffold for bone tissue engineering. J Biomed Nanotechnol 2012;8:149e60. [134] Chen Y-L, Chen H-C, Lee H-P, Chan H-Y, Hu Y-C. Rational development of GAG-augmented chitosan membranes by fractional factorial design methodology. Biomaterials 2006;27:2222e32. [135] Lee KY, Ha WS, Park WH. Blood compatibility and biodegradability of partially N-acylated chitosan derivatives. Biomaterials 1995;16:1211e6. [136] Wieckiewicz M, Boening K, Grychowska N, Paradowska-Stolarz A. Clinical application of chitosan in dental specialities. Mini Rev Med Chem 2017;17:401e9. [137] Klemm D, Heublein B, Fink HP, Bohn A. Cellulose: fascinating biopolymer and sustainable raw material. Angew Chem Int Ed 2005;44:3358e93. [138] Payen A. Me´moire sur la composition du tissu propre des plantes et du ligneux. C R Chim 1838;7:1052e6. [139] Jeschke MG, Sandmann G, Schubert T, Klein D. Effect of oxidized regenerated cellulose/collagen matrix on dermal and epidermal healing and growth factors in an acute wound. Wound Repair Regen 2005;13:324e31. [140] Bassetto F, Vindigni V, Scarpa C, Botti C, Botti G. Use of oxidized regenerated cellulose to stop bleeding after a facelift procedure. Aesth Plast Surg 2008;32:807e9. [141] Schonauer C, Tessitore E, Moraci A, Barbagallo G, Albanese V. The use of local agents: bone wax, gelatin, collagen, oxidized cellulose. Haemostasis in spine surgery. Springer; 2005. p. 89e96. [142] Zhu L, Kumar V, Banker GS. Examination of oxidized cellulose as a macromolecular prodrug carrier: preparation and characterization of an oxidized cellulose-phenylpropanolamine conjugate. Int J Pharm 2001;223:35e47.

548 Chapter 23 [143] Kong L, Ziegler GR. Role of molecular entanglements in starch fiber formation by electrospinning. Biomacromolecules 2012;13:2247e53. [144] Tuzlakoglu K, Santos MI, Neves N, Reis RL. Design of nano-and microfiber combined scaffolds by electrospinning of collagen onto starch-based fiber meshes: a man-made equivalent of natural extracellular matrix. Tissue Eng Part A 2010;17:463e73. [145] Gomes ME, Azevedo HS, Moreira A, Ella¨ V, Kelloma¨ki M, Reis R. Starchepoly (ε-caprolactone) and starchepoly (lactic acid) fibre-mesh scaffolds for bone tissue engineering applications: structure, mechanical properties and degradation behaviour. J Tissue Eng Regen Med 2008;2:243e52. [146] Oliveira JT, Crawford A, Mundy J, Moreira A, Gomes ME, Hatton PV, et al. A cartilage tissue engineering approach combining starch-polycaprolactone fibre mesh scaffolds with bovine articular chondrocytes. J Mater Sci: Mater Med 2007;18:295e302. [147] Munarin F, Tanzi MC, Petrini P. Advances in biomedical applications of pectin gels. Int J Biol Macromol 2012;51:681e9. [148] Kokkonen HE, Ilvesaro JM, Morra M, Schols HA, Tuukkanen J. Effect of modified pectin molecules on the growth of bone cells. Biomacromolecules 2007;8:509e15. [149] Coimbra P, Ferreira P, De Sousa H, Batista P, Rodrigues M, Correia IJ, et al. Preparation and chemical and biological characterization of a pectin/chitosan polyelectrolyte complex scaffold for possible bone tissue engineering applications. Int J Biol Macromol 2011;48:112e8. [150] Liu L, Won YJ, Cooke PH, Coffin DR, Fishman ML, Hicks KB, et al. Pectin/poly (lactide-co-glycolide) composite matrices for biomedical applications. Biomaterials 2004;25:3201e10. [151] Shi L, Aid R, Le Visage C, Chew SY. Biomimicking polysaccharide nanofibers promote vascular phenotypes: a potential application for vascular tissue engineering. Macromol Biosci 2012;12:395e401. [152] Khang G, Lee S, Kim H, Silva-Correia J, Gomes ME, Viegas C, et al. Biological evaluation of intervertebral disc cells in different formulations of gellan gum-based hydrogels. J Tissue Eng Regen Med 2015;9:265e75. [153] Dyondi D, Webster TJ, Banerjee R. A nanoparticulate injectable hydrogel as a tissue engineering scaffold for multiple growth factor delivery for bone regeneration. Int J Nanomed 2013;8:47. [154] Mendes AC, Baran ET, Pereira RC, Azevedo HS, Reis RL. Encapsulation and survival of a chondrocyte cell line within xanthan gum derivative. Macromol Biosci 2012;12:350e9.

C H A P T E R 24

Natural polysaccharides in wound dressing applications Patrı´cia Hissae Yassue-Cordeiro1, Patrı´cia Severino2, Eliana Maria Souto3, 4, Cristiana Maria Pedroso Yoshida5, Classius Ferreira da Silva5 1

Universidade Tecnolo´gica Federal do Parana´, Londrina, Brazil; 2Universidade Tiradentes, Aracaju´, Brazil; 3Faculty of Pharmacy, University of Coimbra, Coimbra, Portugal; 4Centre of Biological Engineering (CEB), University of Minho, Braga, Portugal; 5Instituto de Cieˆncias Ambientais, Quı´micas e Farmaceˆuticas, Universidade Federal de Sa˜o Paulo, Diadema, Brazil

Chapter Outline 1. Introduction 549 2. Flat systems: concepts 2.1 Wound dressings

550

552

3. Properties of dressings 557 3.1 Fluid-handling capacity 3.2 Healing activity 558

558

4. Research on natural polysaccharide dressings: overview 558 5. Mathematical models for releasing film active compounds 561 6. Conclusions 564 References 564

1. Introduction This chapter discusses a new class of dosage forms, the so-called two-dimensional systems or flat systems. Such systems comprise one of the innovative pharmaceutical forms that have emerged in recent years. They can be subdivided into films or membranes, or even fabrics (woven). Many of these systems are composed of multilayers of different materials to promote the best microenvironment for the application at the desired site in human body. The main applications of these systems are in the development of controlled release formulations, artificial organs, and in tissue engineering. This chapter focuses on these systems applied as wound dressings for healing. A brief conceptualization and introduction

Natural Polysaccharides in Drug Delivery and Biomedical Applications. https://doi.org/10.1016/B978-0-12-817055-7.00024-8 Copyright © 2019 Elsevier Inc. All rights reserved.

549

550 Chapter 24 on the main characteristics of these systems are presented in Section 2, as well as an overview of the products available in the world market and the methods of preparation of the dressings. The materials used to produce these systems may be of synthetic origin such as Nylon or polyurethane, or of natural sources, such as carboxymethylcellulose, collagen, or alginate. In this chapter, only natural polysaccharides or semisynthetic polysaccharides will be addressed. Section 3 discusses two of the main properties of dressings: fluid-handling capacity and wound healing. Section 4 presents an overview of publications on polysaccharides-based dressings, especially those of natural sources. The main mathematical models for the release of active compounds from flat systems are presented in Section 5. Finally, the last section, Section 6, summarizes the main conclusions.

2. Flat systems: concepts Flat systems are divided into film (or membranes) and fabrics. The terms films and membranes are commonly used to describe the same device. Professionals from the exact sciences or engineering usually employ the term “film,” whereas those from biological and health sciences usually use the term “membrane.” Strictly speaking, the distinction can be made regarding the moisture content of the device, the films are dried membranes, or similarly, the membranes are hydrated films. The concept also varies greatly from author to author. According to Mulder [1], a membrane is defined as a selective barrier between two phases. The membrane may be thin or thick, its structure may be homogeneous or heterogeneous, it may be used in active or passive transport phenomena (under pressure gradients, concentration, or temperature), and it may even be charged or neutral. We chose the term “films” to use herein. Membrane technology is of great importance in human medicine. Membranes have a set of applications, such as in artificial organs (e.g., oxygenators, pancreas, kidneys, liver, and others), as controlled drug release systems, diagnostic devices, tissue regeneration, as coating medical devices, in bioseparation process, among others. The medical membrane market has been growing due to the demand for new methods of treatments for several pathologies [2]. Within the pharmaceutical field, films or membranes are flat devices used to separate and isolate a surface from the medium in which it is present. These devices shall be capable of allowing selective mass transfer between the surrounding means. Fig. 24.1 depicts a scheme of the existing possibilities of mass transfer when a film/membrane is applied on a “living” surface, such as mucosa or the skin for dressing. The first mass transport expected in the development of a flat system is the transport of molecule “A” present in the system (Fig. 24.1). This molecule is the drug or the active

Natural polysaccharides in wound dressing applications 551

Figure 24.1 Schematic representation of the mass transfer processes that occur in flat systems.

compound(s) of the dressing in question. The surface here, called the “living” surface, is the surface where the dressing is placed and may be, for example, the skin or mucosa. When the drug’s action target is on the living surface, molecule “A” must be transported to this surface and then exhibit the desired pharmacological effect, for example, active dressings containing bioactive substances capable of promoting healing or fight against an infection. These dressings should avoid transporting the bioactive substance to the external environment and, for this reason, they may be provided with an outer layer which has the purpose of occluding it to protect the bioactive substance either from evaporation or loss to the medium on account of fluids in the wound. The transport of the molecule “A” to the microenvironment can also be desirable, as is the case of release films in the buccal cavity, as happens with the recent Oral Disintegration Films (ODF). Such films are used in the administration of drugs, mainly in pediatric and geriatric patients with difficulty in swallowing, nausea, and vomiting, but with the advantage of not requiring water for the administration of the drug. ODF can also be used as dressings for the oral cavity in dental treatments. Molecules present in the medium may also cross the flat system to get access to the “living” surface (molecule “B” of Fig. 24.1), as happens with some wound dressings that should allow the passage of oxygen gas (O2) from the air to the “living” surface. Selectively, dressings should also be able to prevent the passage of a pathogenic microorganism from the medium (air) into the wound bed. Unlike “B” molecules, the passage of molecules from the “living” surface into the environment is also possible through these flat systems (molecule “C” of Fig. 24.1). This happens in the case of dressings that should allow the passage of wound exudate through the dressing, for later evaporation into the medium, promoting the renewal of fluid from the middle of the wound bed. The molecule “C” can also be carbon dioxide as part of the wound breathing process. Some wound dressings should also be able to adsorb fetid odor molecules that are

552 Chapter 24 generated by the wound without being released into the environment, so that such molecules are retained in the dressing, usually in adsorbent material such as activated charcoal. The mass transfer mechanisms coexist in the same wound dressing, for example, the dressing may be capable of releasing an active ingredient into the wound to accelerate wound healing and, at the same time, allow permeation of wound exudate from the wound to the external medium. It should also allow the passage of oxygen from the external environment to the wound. The dressing should also block the entry of microorganisms into the wound and capture fetid substances released by the wound.

2.1 Wound dressings The flat systems, especially those of external use, such as dressings and transdermal patches, may be composed of several layers (Fig. 24.2), which may be: (a) coating adhesive layer; (b) controlling membrane (or film) of drug release; (c) drug reservoirs and (d) outer layer for occlusion/protection of the dressing (or patch). The drug may even be present in the film itself (layer b), or, when one of the layers is a fabric, it may be retained by impregnation in this fabric. Regarding dressings, the number and type of layers are very diverse. The design of the dressings is determined by their functionality. Generally they are structures with high porosity, with transport capacity of the nutrients to the cells, and to remove residues, they must have good mechanical properties, compatible with the tissues of the application site [3,4]. It is very common to find dressings with a layer of textile material, such as fabrics or nonwoven fabrics, and sponges (or foams). The sponges (or foams) may be of synthetic or natural materials like natural polysaccharides. Analogous to the textile materials, they may also contain the drug in their inner cavities.

Figure 24.2 Multilayer scheme of the adhesivelike flat system (transdermal patches) or wound dressings.

Natural polysaccharides in wound dressing applications 553 (A)

Non-woven fabrics fibers

(B)

Fabric fibers

Figure 24.3 Layout of the nonwoven fabric fibers (A) and fabric fibers (B).

Nonwoven fabric is a flat, flexible, and porous structure, consisting of a veil or blanket of fibers or filaments, oriented in a specific direction or in a random way (Fig. 24.3A), consolidated by mechanical and/or chemical and/or thermal process and combinations thereof. In contrast, the fabric is a structure produced by the interweaving of one set of warp yarns and another set of weft yarns at or near this 90 degrees angle (Fig. 24.3B). A wide variety of synthetic and natural materials has been applied in the development of dressings. Polymers have been widely employed in dressings for application in cardiovascular muscles or substitutes, for example, natural polymers such as collagen, fibrin, gelatin, poly(hydroxybutyrate), and polysaccharides (chitosan, starch, and alginates), or synthetic ones such as polyesters, poly(lactic acid), poly(glycolic acid), poly(propylene fumarates), and poly(anhydrides) [2]. The most common techniques used in the manufacture of dressings are summarized in Tables 24.1 and 24.2. Table 24.1: Methodologies for obtaining dressings from (bio)polymers. Methodology

Description

Casting

The dressing is produced by pouring the polymer solution into a mold. The solid phase is then separated from the solvent, either by the induction of phase separation by immersion and precipitation of the polymer phase or by solvent evaporation. The polymer is homogenized with the solvent which may be water. The continuous phase consists of the polymer-rich phase, and the solvent will be dispersed. The polymer suspension is rapidly cooled down to freeze, and the frozen suspension is then lyophilized, which consists of the phenomenon of sublimation of the solvent at very low pressures. The advantage of this methodology is the formation of large pores of a thin and homogeneous thickness [2]. In the supercritical region, inert gas (such as CO2 and N2) is used as the blowing agent to form a foam with the biopolymer under pressure. One can use the pure polymer or polymer and ceramic composites (for rigid fabrics). In this case, no solvents are used, eliminating the risk of residues remaining and not using high temperature which can cause degradation of the polymer [5,6].

Freeze-drying

Foaming

554 Chapter 24 Table 24.2: Methodologies for obtaining dressings from (bio)polymers. Methodology

Description

Particle leaching

It is applied in combination with different techniques, casting, compression molding, or foaming. In this methodology, the particles (salts, sugars or specific beads) are incorporated into the polymers. After formation of the polymeric structure, the particles are dissolved and washed with a suitable solvent, forming the porosity of the dressing. The most significant advantage is the formation of uniform pores, but it cannot be applied to all materials such as proteins [8]. The method consists of charging the polymer and ejecting through a capillary or needle under electric field in the range of 10e30 kV. The solvent is evaporated after passing through the capillary, forming a fiber in the collector. Fiber is formed with diameters in micro and nanometric scales. The advantage of this method is the formation of dressings with high flexibility and good mechanical properties, and it is still possible to align the fibers [9]. It consists of thermal treatment of the powder forming particles that adhere to each other. The dressings formed by this technique are used mainly in harder fabrics. Generally, ceramic powders are used but may be metals, glass, and specific polymers. The advantage is the possibility of creating controlled and graduated porosity. However, there may be low interconnectivity between the pores, the formation of fragile and brittle dressings [10]. It consists of mixing the polymer and the other ingredients in the dry state. This mixture is then heated and extruded into suitable equipment. Any solvents present are removed during extrusion. Although seldomly described in the literature, even by the cost of the extruder, this technique has the advantage of uniformity of drug content [11]. However, this technique encounters the problem of decomposition of the biopolymer in the heating that precedes the extrusion.

Eletrospinning

Sintering

Extrusion

Some characteristics are required to apply films or membranes to the release of drugs, such as thickness, pore size, porosity, mechanical and optical properties [7]. Such properties are strongly influenced by the method of preparation of the devices. Since earliest times, dressings have been used by humankind; however, more recently, some companies have sought innovations to enhance their healing properties, such as (1) incorporation of drugs into the dressing itself; (2) improving the permeation properties of gases and wound exudates in order to provide the best permeation for each healing step; (3) facilitate the application and removal of the dressing with the least possible trauma; (4) prevent contamination of the wound; (5) incorporate indicator substances that promote a response to contamination or need to change dressings (smart dressings). According to the Transparency Market Research report [12], the healing dressing market is segmented into two broad categories, that is, the traditional dressings and the advanced dressings. The dressing market is more favorable toward advanced dressings because of the healing speed they can offer. Traditional dressings have lower costs than advanced dressings; however, the growth rate of this market is smaller than advanced ones.

Natural polysaccharides in wound dressing applications 555 This same report estimates that the global dressing market will grow by an average of 7.8% per year. Europe and the United States represent the largest consumer market for dressings, although emerging economies such as China, Taiwan, and India also show increasing growth rates. In recent decades, interactive/bioactive dressings and antimicrobials have shown a more significant increase in the dressing market of these emerging economies. The Surgical Dressing Manufacturers Association (SDMA) exists for more than 80 years, encompassing companies that manufacture or market dressings or related products within the UK and Ireland. The associated companies range from family businesses to large multinationals, such as 3M HealthCare, ConvaTec, Coloplast, Smith and Nephew, and Mo¨lnlycke Health Care. SDMA maintains a website (www.dressings.org) that compiles information on dozens of dressings, such as composition, usage, presentation, applications, manufacturer, etc. Even though this page is not continually being updated, as companies change product names or remove from the production line, it presents handy information for those who wish to delve into this content. Table 24.3 shows some commercial dressings that have polysaccharides in their composition. Alginate and/or carboxymethylcellulose dressings are recurring in most dressing manufacturers. Some polysaccharides can be obtained directly from natural raw materials, like alginate which is extracted from seaweed. Polysaccharides obtained from natural polysaccharides (semisynthetic), such as carboxymethylcellulose and chitosan which are produced from cellulose and chitin, respectively, may also be used. It is important to note that these dressings are not always exclusively composed of these polysaccharides, but may contain synthetic polymers such as polyurethane or polyester in their structure. The dressings of chitosan have a strong military appeal on the site of their manufacturers, mainly for its hemostatic property, and have been widely used by military patients in warfare. Although the growth trend of (bio)active dressings is imminent, the range of bioactive compounds added to dressings is still very restricted. The use of active silver compounds is virtually unanimous in all major dressing manufacturers, as shown in Table 24.4. Due to its antimicrobial property, silver is one of the active ingredients most present in marketed dressings. It may be present either in the form of silver nanocrystals, silver sulfate, or silver metal. Many manufacturers report that silver is released in a sustained manner, through a mechanism of ionic exchange of the same by cations, especially sodium and calcium, present in the exudate of the wound. As the dressing is hydrated by the exudate, the silver ions are released into the wound and then its antimicrobial activity is triggered. Few companies already manufacture dressings with other antimicrobial or antiseptic substances. Table 24.4 summarizes some examples available on the international market.

556 Chapter 24 Table 24.3: Dressings containing polysaccharides and their respective commercial names and producers. Polysaccharides Alginate

Carboxymethylcellulose (CMC) Alginate/CMC

Cellulose and cellulose derivatives

Cross-linked polysaccharide Starch Esterified Hyaluronic Acid Chitin Chitosan

Commercial Name (Producer, Country) • • • • • • • • • • • • • • • • • • • • • • •

Algisite M (Smith and Nephew Medical Ltd, UK) Suprasorb A (Lohmann and Hauscher Company, Germany) MediHoney Calcium Alginate (Derma Sciences, USA) Tegaderm Alginate (3M, USA) SILVERCEL Antimicrobial Alginate Dressing (Johnson and Johnson, USA) ´dio (Curatec, Brazil) Curatec Alginato de Ca´lcio e So ActivHeal Aquafiber (ActivHeal, UK) Aquacel (ConvaTec Ltd, UK) UrgoTul and UrgoStart (Urgo Medical, France) Biostep (Smith and Nephew Medical Ltd, UK) ActivHeal Aquafiber Ag (ActivHeal, UK) UrgoSorb (Urgo Medical, France) Durafiber (Smith and Nephew Medical Ltd, UK) Suprasorb X (Lohmann and Hauscher Company, Germany) Bactigras (cotton gauze) (Smith and Nephew Medical Ltd, UK) Membracel (Vuelo Pharma, Brazil) Iodoflex (Smith and Nephew Medical Ltd, UK) Hyalomatrix (Medline Industries, Inc., USA) Beschitin W (Unitika, Ltd, Japan) Axiostat (Axio Biosolutions Private Limited, India) Hemo-bandage (CoreLeader Biotech, Taiwan) HemCon (Tricol Biomedical, Inc, USA) ChitoSAM 100 (SAM Medical, USA)

Table 24.4: Some drugs incorporated into commercial dressings containing polysaccharides. Drug Ionic silver

Ionic silver complex (Silver sodium hydrogen zirconium phosphate) Iodine Polyhexamethylene biguanide (PHMB) Silver sulfadiazine Chlorhexidine acetate NOSF (nanooligosaccharide factor) Manuka flower honey (active leptospermum honey)

Commercial Name (Producer, Country) • Aquacel Ag (ConvaTec Ltd, UK) • Suprasorb A þ Ag (Lohmann and Hauscher Company, Germany) • Biostep Ag (Smith and Nephew Medical Ltd, UK) • UrgoTul Ag/Silver (Urgo Medical, France) • ActivHeal Aquafiber Ag (ActivHeal, UK) • UrgoSorb Silver (Urgo Medical, France) • Iodoflex (Smith and Nephew Medical Ltd, UK) • Iodosorb (Smith and Nephew Medical Ltd, UK) • Suprasorb X þ PHMB (Lohmann and Hauscher Company, Germany) • UrgoTul SSD (Urgo Medical, France) • Bactigras (Smith and Nephew Medical Ltd, UK) • UrgoStart (Urgo Medical, France) • MediHoney Calcium Alginate (Derma Sciences, USA) • Algivon (Advancis Medical, UK) • MANUKAtex (ManukaMed, New Zeland)

Natural polysaccharides in wound dressing applications 557 They range from substances traditionally used in wound asepsis such as iodopovidone, to fewer common substances such as polyhexamethylene biguanide (PHMB) and iodized cadexomer, or alternative substances such as manuka flower honey. Most active dressings are composed of drugs with antimicrobial properties, since microbial infections represent a severe problem in wound healing that may even lead to patient death. However, active dressings with other features are also targeted by the dressing industries and especially in research conducted by many research groups at universities and research centers worldwide. NOSF is one of the drugs with healing properties (Table 24.4). According to the manufacturer, NOSF is a chemical compound derived from the oligosaccharide family, which is known to have antiprotease properties. NOSF binds to the damaged area where its antimetalloprotease activity (anti-MMPs) leads to accelerated wound healing. While it is not possible to list all the commercial dressings available in the international market, many manufacturers also do not provide significant information on the composition of dressings, making it difficult to identify the presence of natural polysaccharides in many available brands. Another fact worth to be mentioned is the diverse information regarding the use of dressings. Dressings are recommended for the treatment of ulcers (venous, arterial, diabetic), pressure sores, donor/graft sites, surgical and trauma wounds (dermal lesions, trauma injuries, or incisions), burns (first and second degree), cavity wounds, as well as to control the bleeding in superficial wounds. It is recommended that every reader check the catalogs and information available on the manufacturers’ websites to choose the most appropriate dressing.

3. Properties of dressings Many properties are essential in developing wound dressings. Mechanical properties are of the utmost importance since the dressing should be sufficiently capable of resisting the handling of the person responsible for applying the dressing. It should also be able to withstand efforts and movements performed by the patient in the site of application, especially in articulable members and portions. Because of this demand, many biopolymeric dressings like natural polysaccharides require reinforcements of synthetic materials such as polyethylene or polyurethane. This reinforcement is often possible by the existence of several layers in the dressing, as previously described. Moreover, dressings have also to be exchanged frequently. Only a few dressings are bioabsorbable, and therefore, they do not require an exchange, while other dressings need a frequent exchange. The exchanging processes may represent a trauma to the patient when a significant adhesion of the dressing occurs to the injured tissue. In addition to the mechanical properties, two other properties must be taken into account,

558 Chapter 24 namely (1) a physical property, that is, the fluid-handling capacity (FHC); and (2) a biological property, that is, the wound healing activity.

3.1 Fluid-handling capacity The fluid-handling capacity (FHC) is defined as the sum of the moisture vapor permeability and the absorbency. The standard Paddington cup technique, described in the European Standard BS EM 13,726e1 [13], is used to determine the FHC. Absorbency and permeability are calculated by contacting the dressing sample with an isotonic sodium/ calcium chloride solution containing 142 mmol/L of sodium ions and 2.5 mmol/L of calcium ions; these values are typically found in serum and wound fluid. The cup is placed in a chamber at 37 C (2 C) and relative humidity below 20%. Dressing samples are kept either in or not in contact with the fluid. After 24, 48, and 72 h, the cup is weighted so that the absorbency and the moisture vapor permeability can be calculated. Determining this property is very important because it defines the specific use of the dressing. Different types of wounds produce different amounts of exudate, and various stages of healing require moist or drier environments. For example, Lamke [14] reported that donor sites, third-degree burns, and unspecified granulating wounds produce between 3.4 and 5.1 g of exudate per 10 cm2 over 24 h. Although the mechanical properties of the polysaccharides are not always satisfactory, they generally exhibit high hydrophilicity and may therefore contribute to a greater or lesser degree both for moisture vapor permeability and the absorbency.

3.2 Healing activity The active dressings (described previously in Section 2) are prominent both in the dressing market and in academic research, but the healing activity that some polysaccharides can attribute to the dressing when present in its composition is a plus, regardless the presence of a drug. A large number of articles and reviews on the biological properties of these polysaccharides have been published. These polysaccharides such as chitosan [15], alginate [16], hyaluronic acid [17], dextran [18] may have healing properties, or even antimicrobial properties, such as chitosan [19].

4. Research on natural polysaccharide dressings: overview Table 24.5 shows a compilation of scientific papers on dressings of natural and semisynthetic polysaccharides, which are polysaccharides industrially produced from natural polysaccharides. A wide diversity of polysaccharides, both natural and semisynthetic, is listed. These macromolecules may be used alone or in the form of blends

Natural polysaccharides in wound dressing applications 559 Table 24.5: Scientific papers on the polysaccharide-based dressing containing bioactive compounds and their therapeutic purposes. Polysaccharides

Bioactive

Chitosan

Fucoidan from Fucus vesiculosus

Chitosan

Minocycline hydrochloride Basic fibroblast growth factor (bFGF) Silver sulfadiazine Paracetamol Ibuprofen Essential oils (elicriso italic, chamomile blue, cinnamon, lavender, tea tree, peppermint, eucalyptus, lemongrass, and lemon oils) Asiaticoside (substance from the plant Centella asiatica) Silver nanoparticles and gentamicin Simvastatin Silver nanoparticles Epidermal growth factor (EGF)

Hydroxypropylchitosan Chitosan/chondroitin Chitosan/alginate Alginate Alginate

Alginate

Alginate/starch Alginate/pectin Xanthan Hyaluronic acid

Therapeutic purpose

Reference Sezer et al. [20]

Antioxidant, antiinflammatory, anticancer Antibiotic

Aoyagi et al. [21]

Wound healing

Mizuno et al. [22]

Antibiotic Analgesics Antiinflammatory Antibiotic

Fajardo et al. [23] Lai et al. [24] Thu et al. [25] Liakos et al. [26]

Wound healing

Sikareepaisan et al. [27]

Antibiotic

Arockianathan et al. [28] Rezvanian et al. [29] Huang et al. [30] Matsumoto and Kuroyanagi [31]

Wound healing Antibiotic Wound healing

to adjust the properties of the formulation. About the active (drug), there is a predominance of allopathic drugs for the most diverse applications, but also containing natural bioactive compounds, such as essential oils. The scientific literature on dressings has grown substantially in the last 20 years. Initially, the words “dressings” and “wound” were used to search only scientific papers in journals between 1999 and 2018 (until November 2018) at Web of Science database by Clarivate Analytics. The result showed that the number of published papers increased from 96 in 1999 to 934 in 2018, that is, about 10 times within 20 years. The total number of articles in this period was 8027. This search was refined by the inclusion of one more word in the search, corresponding to the name of a polysaccharide. It was not possible to determine the exact number of scientific articles about the use of a polysaccharide alone since many papers combine more than one polysaccharide, or even present composite dressings or polysaccharide blends with other natural or synthetic polymeric materials. Polyethylene and polyurethane are two

560 Chapter 24

Figure 24.4 Most frequent polysaccharides (natural and semisynthetic) in dressings according to the search carried out in the Web of Science database.

of the most common synthetic polymers in commercial dressings. Similarly, it was also possible to find in this search that they are also the two most common synthetic polymers in scientific publications. Results are summarized in Fig. 24.4. For a better understanding of the results, for some polysaccharides, the “*” was used as a wildcard to find inflections and distinct ways of referring to the same polysaccharide. For example, by including the expression “Agar*” in article search, it was possible to obtain results for either “agarose”, “agar” or “agareagar”, which are all terms used for the same polysaccharide. Or even “Algin*”, which allows results for both “alginate” and “alginic acid”, also corresponding to the same compound. Similarly, “Hyaluron*” for “hyaluronate” or “hyaluronic” and “Chit*” for “chitosan” and “chitin”. Fig. 24.4 also demonstrates that chitosan (and/or chitin) is the polysaccharide mostly used in healing dressings, reaching almost half of the total number of publications. Chitosan is followed by alginate and cellulose. It is important to note that the healing properties of chitosan have been widely disseminated in the literature, which could be the reason for the choice of chitosan as polysaccharide for dressings. However, chitosan is a semisynthetic polysaccharide, whereas chitin, alginate, and cellulose are natural polysaccharides. The same Fig. 24.4 shows a so-called “Others” which is the fraction corresponding to other natural polysaccharides that still exhibit inexpressive amounts in applications as dressings. This fraction is plotted separately in Fig. 24.5 to show the polysaccharides that make up such inexpressive polysaccharides, pectin and dextran being highlighted.

Natural polysaccharides in wound dressing applications 561

Figure 24.5 Other polysaccharides (natural and semisynthetic), less frequent, in dressings according to the search carried out in the Web of Science database.

5. Mathematical models for releasing film active compounds In order to verify how active ingredients are released from the film or membrane to the recipient organ (burn bed, buccal cavity, etc.), the release assays are performed, and mathematic models are fitted to provide a base for the study of the main mass transport mechanisms that predominate in the system. These models make it possible to simulate the effect of the parameters delineated, provide information on which mechanism controls the release of the drug, and thus facilitate the development of new products. Several theoretical models have been proposed and the ones that best describe the release phenomenon are, in general, the Higuchi model [32], first order model [33], Korsmeyer model [34], and Peppas-Sahlin [35] (Table 24.6). The Higuchi model (Eq. 24.1) is the most popular among all those already proposed in the literature, describing the mechanism of drug release as a diffusion process based on the Fick’s Law, being dependent on the square root of time. Diffusion occurs when the drug or other active agent crosses the membrane. This diffusion occurs due to the difference in drug concentration inside the membrane (more concentrated medium) to the external environment (less concentrated medium), spontaneously until the establishment of equilibrium. Diffusion may occur through the pores of the polymer matrix (macroscopic scale) or through passage between the polymer chains of the polymer (microscopic scale).

562 Chapter 24 Table 24.6: Kinetic models for drug release. Model

Equation

Higuchi

pffiffi Qt ¼ Kh t QN

First order

  Qt ln ¼ K1 t QN

Equation number (24.1)

(24.2)

Korsmeyer

Qt ¼ K2 t n QN

(24.3)

Peppas-Sahlin

Qt ¼ K3 t m þ K4 t 2m

(24.4)

Symbols labels: Qt =QN is the fraction of drug released; Kh, K1, K2, K3 and K4, correspond to the Higuchi release constants, first order release, Korsmeyer release kinetics, Fickian and non Fickian diffusion constants respectively; n and m are exponents that characterize the release kinetics.

Although widely used, the Higuchi model has strong limitations in the interpretation of the mechanisms of controlled release, since it is better applicable in poorly soluble and/or nonswelling polymer films [32,36]. An example is its use in films made of cellulose with the incorporation of soluble drugs [37]. The First Order Model (Eq. 24.2) can be used to describe porous matrix systems containing water-soluble drugs. This model does not describe the dissolution of the active agent that is entrapped between polymer chains of natural polymers, such as those of chitosan, representing only the behavior of the drug that is on the surface of the film, being characteristic of an instant dissolution process [38]. Eq. (24.3), proposed by Korsmeyer et al. [34] is used when the drug release controlling mechanism is not known or when there is a combination of the following independently governed processes: drug transport within the matrix (Fickian diffusion) and transport mechanisms case II. In this latter mechanism, when the film comes into contact with a solvent, such as burn wound exudate or saliva, the chains of the polymer film unfold, with a transition from a semirigid to a more flexible state (associated with the absorption of fluid), resulting in the formation of a gel layer around the dry areas of the polymer matrix. The polymeric film increases its volume due to the absorption of the fluid and reorganizes its polymer chains until the establishment of a new equilibrium condition [39,40]. The values of “n” of the Korsmeyer model can be calculated from the slope of lnðMt =MNÞ versus ln t. According to Peppas [41], for this model, when n equals 0.5 the release is

Natural polysaccharides in wound dressing applications 563 controlled by diffusion. When n is equal to 1, the controlling mechanism is swelling, corresponding to zero-order kinetics. For 0.5 < n < 1, the release occurs by overlapping the two phenomena cited or anomalous transport. The data used for the estimation of parameter n must be Qt =QN < 0:6. To quantify the contributions of Fickian diffusion (first term of Eq. 24.4) and the contribution of relaxation and swelling of the polymer matrix or transport mechanism case II (second term of Eq. 24.4), Peppas and Sahlin [35] elaborated a mathematical model that leads account of these two factors. In general, zero-order models and the Higuchi model are mutually exclusive, that is, only one will be suitable for the controlled release system in question. The Korsmeyer and Peppas-Sahlin models can be used together and as a complement or verification of the mechanisms obtained by all other models. For the determination of the model that best fits the obtained experimental data, one can use the method of analysis of the correlation coefficient R2. However, according to Costa and Lobo [36], the R2 value becomes larger with the addition of more parameters, regardless the meaning of the variable added to the model (Eq. 24.5). When comparing models with different amounts of parameters, it is more appropriate to use the adjusted correlation coefficient R2adjusted . R2adjusted ¼ 1 

 ðn  1Þ  1  R2 ðn  PÞ

(24.5)

where s is the number of experimental points of the release tests and P is the number of parameters of the model. The most suitable model will be the one with the highest adjusted correlation coefficient. In addition to the correlation coefficient R2 and the adjusted correlation coefficient R2adjusted , we can also use the sum of the squares of residuals (SSR), mean square error (MMS), and the Akaike Information Criterion (AIC) and F-test to determine the applicability of the drug release models [36]. Polymeric films of chitosan with silver nitrate impregnated in zeolites were synthesized (AgY/chitosan) to be applied as dressings for the treatment of burn by Yassue-Cordeiro et al. [42]. The experimental data obtained in the silver release assays from the polymer films were adjusted to several mathematical models to analyze the kinetic behavior of the system. As the films synthesized by these authors exhibited great swelling capacity, the Higuchi kinetic model did not fit the experimental data. This result confirms the fact that the main release mechanism is not governed by the Fick’s Law of diffusion, as the value found for parameter n of the Korsmeyer model was lower than 0.5.

564 Chapter 24

6. Conclusions The use of natural and semisynthetic polysaccharides in wound dressings increases each year mainly due to the intrinsic properties of these materials such as biodegradability, toxicity, and biocompatibility. In addition to these properties, many polysaccharides show healing activity, which can offer active properties to the wound dressings, such as chitosan, which combines all these properties and has been the most studied polysaccharide in the last 20 years. The world market already has active dressings containing drugs, such as ionic silver and chlorhexidine, and virtually all significant manufacturers worldwide have dressings containing ionic silver which has antibiotic properties. In addition to these drugs, the loading of natural extracts into the dressings has also become marketable, when combined with antimicrobial, antiinflammatory, and healing activities.

References [1] Mulder J. Basic principles of membrane technology. Springer Science & Business Media; 2012. [2] Stamatialis DF, Papenburg BJ, Girone´s M, Saiful S, Bettahalli SN, Schmitmeier S, Wessling M. Medical applications of membranes: drug delivery, artificial organs and tissue engineering. J Membr Sci 2008;308(1):1e34. [3] Hutmacher DW. Scaffolds in tissue engineering bone and cartilage. Biomaterials 2000;21(24):2529e43. [4] Agrawal C, Ray RB. Biodegradable polymeric scaffolds for musculoskeletal tissue engineering. J Biomed Mater Res A 2001;55(2):141e50. [5] Harris LD, Kim BS, Mooney DJ. Open pore biodegradable matrices formed with gas foaming. J Biomed Mater Res 1998;42(3):396e402. [6] Wang X, Li W, Kumar V. A method for solvent-free fabrication of porous polymer using solid-state foaming and ultrasound for tissue engineering applications. Biomaterials 2006;27(9):1924e9. [7] Suntornnond R, An J, Yeong WY, Chua CK. Biodegradable polymeric films and membranes processing and forming for tissue engineering. Macromol Mater Eng 2015;300(9):858e77. [8] Katoh K, Tanabe T, Yamauchi K. Novel approach to fabricate keratin sponge scaffolds with controlled pore size and porosity. Biomaterials 2004;25(18):4255e62. [9] Boudriot U, Dersch R, Greiner A, Wendorff JH. Electrospinning approaches toward scaffold engineering-a brief overview. Artif Organs 2006;30(10):785e92. [10] Rezwan K, Chen Q, Blaker J, Boccaccini AR. Biodegradable and bioactive porous polymer/inorganic composite scaffolds for bone tissue engineering. Biomaterials 2006;27(18):3413e31. [11] Arya A, Chandra A, Sharma V, Pathak K. Fast dissolving oral films: an innovative drug delivery system and dosage form. Int J Chem Res 2010;2(1):576e83. [12] Transparency Market Research-TMR. Wound dressings market - global industry size, market share, trends, analysis, and forecast 2012e2018. United States of America. 2013. [13] British standards BS EN 13726-1 test methods for primary wound dressings. Part 1: aspects of absorbency, section 3.3 fluid handling capacity. 2002. [14] Lamke LO, Nilsson GE, Reithner HL. The evaporative water loss from burns and water vapour permeability of grafts and artificial membranes used in the treatment of burns. Burns 1977;3(3):159e65. [15] Singla AK, Chawla M. Chitosan: some pharmaceutical and biological aspects-an update. J Pharm Pharmacol 2001;53(8):1047e67.

Natural polysaccharides in wound dressing applications 565 [16] Lee WR, Park JH, Kim KH, Kim SJ, Park DH, Chae MH, et al. The biological effects of topical alginate treatment in an animal model of skin wound healing. Wound Repair Regen 2009;17(4):505e10. [17] Doillon CJ, Silver FH. Collagen-based wound dressing: effects of hyaluronic acid and firponectin on wound healing. Biomaterials 1986;7(1):3e8. [18] Sun G, Zhang X, Shen YI, Sebastian R, Dickinson LE, Fox-Talbot K, et al. Dextran hydrogel scaffolds enhance angiogenic responses and promote complete skin regeneration during burn wound healing. Proc Natl Acad Sci USA 2011;108(52):20976e81. [19] Raafat D, Sahl HG. Chitosan and its antimicrobial potential-a critical literature survey. Microb Biotechnol 2009;2(2):186e201. [20] Sezer AD, Hatipoglu F, Cevher E, Ogurtan Z, Bas AL, Akbu ga J. Chitosan film containing fucoidan as a wound dressing for dermal burn healing: preparation and in vitro/in vivo evaluation. AAPS PharmSciTech 2007;8(2):E94e101. [21] Aoyagi S, Onishi H, Machida Y. Novel chitosan wound dressing loaded with minocycline for the treatment of severe burn wounds. Int J Pharm 2007;330(1e2):138e45. [22] Mizuno K, Yamamura K, Yano K, Osada T, Saeki S, Takimoto N, et al. Effect of chitosan film containing basic fibroblast growth factor on wound healing in genetically diabetic mice. J Biomed Mater Res A 2003;64(1):177e81. [23] Fajardo AR, Lopes LC, Caleare AO, Britta EA, Nakamura CV, Rubira AF, et al. Silver sulfadiazine loaded chitosan/chondroitin sulfate films for a potential wound dressing application. Mater Sci Eng C Mater Biol Appl 2013;33(2):588e95. [24] Lai HL, Abu’Khalil A, Craig DQ. The preparation and characterisation of drug-loaded alginate and chitosan sponges. Int J Pharm 2003;251(1e2):175e81. [25] Thu HE, Zulfakar MH, Ng SF. Alginate based bilayer hydrocolloid films as potential slow-release modern wound dressing. Int J Pharm 2012;434(1e2):375e83. [26] Liakos I, Rizzello L, Scurr DJ, Pompa PP, Bayer IS, Athanassiou A. All-natural composite wound dressing films of essential oils encapsulated in sodium alginate with antimicrobial properties. Int J Pharm 2014;463(2):137e45. [27] Sikareepaisan P, Ruktanonchai U, Supaphol P. Preparation and characterization of asiaticoside-loaded alginate films and their potential for use as effectual wound dressings. Carbohydr Polym 2011;83(4):1457e69. [28] Arockianathan PM, Sekar S, Sankar S, Kumaran B, Sastry TP. Evaluation of biocomposite films containing alginate and sago starch impregnated with silver nano particles. Carbohydr Polym 2012;90(1):717e24. [29] Rezvanian M, Ahmad N, Amin MCIM, Ng SF. Optimization, characterization, and in vitro assessment of alginate-pectin ionic cross-linked hydrogel film for wound dressing applications. Int J Biol Macromol 2017;97(1):131e40. [30] Huang J, Ren J, Chen G, Deng Y, Wang G, Wu X. Evaluation of the xanthan-based film incorporated with silver nanoparticles for potential application in the nonhealing infectious wound. J Nanomater 2017. Article ID 6802397. [31] Matsumoto Y, Kuroyanagi Y. Development of a wound dressing composed of hyaluronic acid sponge containing arginine and epidermal growth factor. J Biomater Sci Polym Ed 2010;21(6e7):715e26. [32] Higuchi T. Mechanism of sustained-action medication. Theoretical analysis of rate of release of solid drugs dispersed in solid matrices. J Pharm Sci 1963;52(12):1145e9. [33] Wagner JG. Interpretation of percent dissolved-time plots derived from in vitro testing of conventional tablets and capsules. J Pharm Sci 1969;58(10):1253e7. [34] Korsmeyer RW, Gurny R, Doelker E, Buri P, Peppas NA. Mechanisms of solute release from porous hydrophilic polymers. Int J Pharm 1983;15(1):25e35. [35] Peppas NA, Sahlin JJ. A simple equation for the description of solute release. III. Coupling of diffusion and relaxation. Int J Pharm 1989;57(2):169e72.

566 Chapter 24 [36] Costa P, Lobo JMS. Modeling and comparison of dissolution profiles. Eur J Pharm Sci 2001;13(2):123e33. [37] Tahara K, Yamamoto K, Nishihata T. Application of model-independent and model analysis for the investigation of effect of drug solubility on its release rate from hydroxypropyl methylcellulose sustained release tablets. Int J Pharm 1996;133(1):17e27. [38] Balcerzak J, Mucha M. Analysis of model drug release kinetics from complex matrices of polylactidechitosan. Prog Chem Appl Chitin Deriv 2010;15(1):117e25. [39] Caraballo I. Critical points in the formulation of pharmaceutical swellable controlled release dosage forms-Influence of particle size. Particuology 2009;7(6):421e5. [40] Colombo P, Bettini R, Santi P, Peppas NA. Swellable matrices for controlled drug delivery: gel-layer behaviour, mechanisms and optimal performance. Pharmaceut Sci Technol Today 2000;3(6):198e204. [41] Peppas N. Analysis of Fickian and non-Fickian drug release from polymers. Pharm Acta Helv 1985;60(4):110. [42] Yassue-Cordeiro PH, Zandonai CH, Silva CF, Fernandes-Machado NRC. Desenvolvimento e caracterizac¸a˜o de filmes compo´sitos de quitosana e zeo´litas com prata. Polimeros 2015;25(5):492e502.

C H A P T E R 25

Polysaccharides from leafy vegetables: chemical, nutritional and medicinal properties Esha Bala, Siddhartha Singha, Sanjukta Patra Centre for Rural Technology, Indian Institute of Technology, Guwahati, Assam, India

Chapter Outline 1. Introduction 568 2. Polysaccharide localization in leaves 568 3. Structural illustration of some relevant polysaccharides from leaves 3.1 Fructans 571 3.2 Xylans 573 3.2.1 Arabinoxylan 573 3.2.2 Glucuronoarabinoxylan 573 3.2.3 Xyloglucan 574 3.3 Mannans 574 3.3.1 Glucomannan 574 3.3.2 Galactomannan 574 3.4 Pectin 575 3.4.1 Homogalacturonan 575 3.4.2 Rhamnogalacturonan 575 3.4.3 Xylogalacturonan 576 3.5 Arabinogalactan 576 3.6 Functional attributes of the polysaccharides from leaves 3.6.1 Dietary fiber 576 3.6.2 Immunostimulant 577 3.6.3 Antioxidant 580

569

576

4. Polysaccharide from different leafy vegetables 580 5. Effect of processing condition on polysaccharide structure and functional relationship 582 6. Negative physiological impact of plant polysaccharides: food and drug interaction 583 7. Conclusions 584 References 584 Further reading 588

Natural Polysaccharides in Drug Delivery and Biomedical Applications. https://doi.org/10.1016/B978-0-12-817055-7.00025-X Copyright © 2019 Elsevier Inc. All rights reserved.

567

568 Chapter 25

1. Introduction Leafy vegetables are important part of human diet and well recognized for their nutritional and often medicinal values. Botanically the term leafy vegetable is not well defined. The term should strictly refer to leaves which are eaten with meals in cooked form. However, in common practice across cultures, a great variety of leaves or plant parts with leaves are consumed in different processed forms and not necessarily only with meals. Sometimes they overlap with herbs, medicinal leaves, and weeds as famine food. Hence in this chapter, all such plants are included where leaves are the major part consumed in either raw or processed form. Typically, basic nutritional properties of such leaves are known but their extended functions like their therapeutic values, drug-food interactions, or other broader physiological impacts are yet to be studied. A leaf contains myriad of molecules and most of them are having some or the other effect on human health. This chapter deals with one such group of compounds called polysaccharides and their beneficial as well as detrimental effect on human health. Though polysaccharides/glycans are defined as chains consisting of more than 10 units of monosaccharides, some shorter chains or oligosaccharides (i.e., chains containing 3e10 monomer units) are also included in the scope of the chapter for the sake of complicity. Based on structure, polysaccharides are classified as homopolysaccharide that is, those consisted of only one type of sugar unit (e.g., glucans, fructans, mannans, etc.) and heteropolysaccharides, that is, those composed of two or more types of sugar units (like arabinoxylans and glucomannans). In terms of functions they can be further classified as dietary fiber, prebiotics, immunostimulant, and antioxidant polysaccharides [1]. So far, many polysaccharides from different plant sources have been studied and utilized and serve as an important component of food products, nutraceuticals, pharmaceuticals, and other industrial products like packaging film, adhesive, etc. However, very few leafy vegetables are known for their polysaccharides with significant nutritional or medicinal properties. In this communication the aim is to facilitate applications of leafy vegetable polysaccharides through understanding of their chemical, nutritional, and medicinal properties in isolated or in native form.

2. Polysaccharide localization in leaves In order to exploit the wealth of leaf-polysaccharides for various functional and medicinal properties, knowledge about their localization and metabolic role in leaves is important. Primarily three cellular substructures of leaves are pertinent with respect to polysaccharides and oligosaccharides: cell wall, vacuoles, and plastids. Cell walls of plants including leaves are made up of a principal structure and an auxiliary structure. The principal structure is consisted of cellulose, microfibrils and hemicellulose which is further

Polysaccharides from leafy vegetables: chemical, nutritional and medicinal properties

569

entrenched by pectic polysaccharides [2,3] (Fig. 25.1). This hemicellulose and pectic polysaccharides are the major source of polysaccharides in leaves. Another special source of polysaccharides and oligosaccharides (e.g., fructans) are the vacuoles of plant cells. Vacuoles are very important for plants due to their multiplicity of roles as storage compartment and lytic organelles [4]. In a plant cell, plastid is a complex organelle for synthesis and storage of many physiologically important molecules including starch. However, the starch in leaves are usually transitory in nature. It typically gets synthesized in day time subsequent to photosynthesis and degrades during the night time to meet metabolic demand of energy and C-precursor. Starch chains in leaves are quite different than their seed counterpart. For example, degree of phosphorylation is more in the starch from seed/tuber cells than that in the leaves [5]. In addition to these cell organelles some small amount of polysaccharides including complexes like glycoproteins can be identified in various other subcellular compartments.

3. Structural illustration of some relevant polysaccharides from leaves Polysaccharides are highly diverse in structure and biological functions. A wide variety of linear or branched polysaccharide structures can be seen in various leaves. Sometimes the polysaccharide chains are cross-linked to give rise to a complex substance like pectin. Often these heteropolysaccharides are quite challenging to characterize. With the advancement of sophisticated extraction techniques and analytical methods, some of these structures have been established. The ubiquitous plant polysaccharides like starch and cellulose are excluded here because there are numerous reviews available on them. Instead other polysaccharides typical but not always exclusive to leaves are listed below (Fig. 25.2). Many of them has shown beneficial effect on human body. This section explores structural and other important physicochemical aspects of such polysaccharides present in leaves, specifically, fructans, hemicelluloses, and pectins. Hemicellulose is the most common component of the cell wall in all plant tissues including leaves and represents a heterogeneous group of polysaccharides. They contain a backbone structure with either of the sugars like xylose, mannose, glucose, or galactose. Apart from the main chain, it might have side chains with other sugars or sugar derivatives like arabinose, fucose, and methyl glucuronic acid. They are obtained in the alkali-soluble fraction of the cell wall polysaccharides. Branching occurs in hemicellulose with smaller chains that are easily degradable. They show much smaller degree of polymerization (i.e., 50e200) compared to that of cellulose (i.e., 300e15,000). The hemicellulose content in dried leafy vegetables is in the range of 15e34 g/100 g on dry basis [6]. With the availability of the sophisticated separation (e.g., chromatography, electrophoresis) and analytical techniques (spectroscopy, nuclear magnetic resonance, mass spectrometry,

570 Chapter 25

Figure 25.1 Localization of major polysaccharides in plant leaves.

Polysaccharides from leafy vegetables: chemical, nutritional and medicinal properties

571

Polysaccharide Chemical Structure

Homopolysaccharide Inulin

Heteropolysaccharide Hemicellulose Xylan Arabinoxylan Glucuronoarabinoxylan Xyloglucan Arabinogalactan Galactomannan Glucomannan Pectin Galacturonan Homogalacturonan Rhamnogalacturonan Xylogalacturonan

Physiological Function

Dietary Fiber Xylan Arabinoxylan

Prebiotic

Immuno-stimulant

Xylan Arabinoxylan Glucuronoarabino- Glucuronoarabinoxylan xylan Xyloglucan Xyloglucan Arabinogalactan Arabinogalactan Galactomannan Galactomannan Glucomannan Glucomannan HomogalacturonGalacturonan Homogalacturonan an Xylogalacturonan Xylogalacturonan Inulin Inulin

Antioxidant

Glucomannan Arabinoxylan Glucuronoarabinoxylan

Glucomannan Sulfated Polysaccharide Proteoglycan

Xyloglucan Arabinogalactan Rhamnogalacturonan Galactomannan Inulin Xylan Homogalacturo- Inulin nan Xyloogalacturonan

Proteoglycan Arabinogalactan-proteins

Figure 25.2 Structure and biological function of polysaccharide.

electron microscopy, etc.) knowledge of these hemicellulose structures is evolving very rapidly. Depending on the primary chain forming monomer and side chains, hemicellulose can be categorized further.

3.1 Fructans Some plants do not store C-source as starch, instead they store it in the form of polymers of fructose called fructans. Fructans help the plant in resisting drought and cold conditions. On the basis of monomer-monomer bonding fructans can be of three types; inulins with b 2-1 bonds (Fig. 25.3A), levans with b 2-1 bonds, and graminins with both types of bonds. Out of the three, fructans and inulins are found to be prebiotic in nature. Apart from that, in many species inulin provides protection in membrane during dehydration. Membrane lipids interact more with inulin-fructans than levan-fructans [11]. However, this polysaccharide is dynamic in nature. In leaves at night, inulin is transformed to sugar (i.e., fructose) and gets transferred to other plant tissues. Fructose chain of different chains occurs during this transition. That is the reason leaves contain inulins with variety of structures starting from just 2 to 60 monomer length. The stems and stalk also contain inulins along with fructooligosaccharides or oligofructose (degree of polymerization below 10) and fructose. Generally, low molecular weight structural polysaccharides and fructans are more in stems compared to leaves. Inulin in the stems increases from aerial part to root. Long-chain inulins are found more toward the middle of the stem in lignin-containing tissues, while shorter-chain inulin molecules are more

572 Chapter 25 Figure 25.3 (A) Chemical structure of inulin [7]. (B) Chemical structure of the xylan [8]. (C) Chemical structure of pectic substances [9]. (D) Proteoglycan (example; arabinogalactan-proteins) [10].

Polysaccharides from leafy vegetables: chemical, nutritional and medicinal properties

573

prevalent toward the stem base. Chicory (Cichorium intybus L.) is one example of leafy vegetables that contain high amount of inulin which can be as high as 15% of the fresh weight and 75% of the dry weight.

3.2 Xylans Xylans are a class of polymers consisting of b-1, 4 xylopyranose linkages and act as a cross-linking polymer among cellulose, microfibrils and lignin component of plant cell wall (Fig. 25.3B) [12]. Structure, composition, and their metabolism differ widely in different plant species and even in the same species grown in dissimilar conditions. In leaves, hardly bare xylan chain is available. The xylan may have different types of branches consisting of acetyl, arabinosyl, and glucuronosyl residues which mainly depend on the origin of the compound and on the way of extraction [13]. The xylan chain containing a-D-glucopyranosyl uronate residues or more specifically 4-O-methyl-a-Dglucopyranosyl uronate residues are called glucuronoxylan. Glucuronoxylan is generally present in the secondary cell wall but their content is regulated by different environmental factors. In mulberry leaves, the polysaccharide content increases with decrease in protein and water activity. 3.2.1 Arabinoxylan Arabinoxylans are composed of branched side-chains of arabinose with the backbone of xylose chain [14]. The branching pattern is diverse in different species. Usually, the skeleton contains b-anomer of xylose and a-anomers of arabinose residues. Often the arabinose residue is bonded with glucuronic acid, ferulic acid, and acetyl groups. In a recent study, an unusual arabinoxylan has been reported in leaves. The green leaves of Litsea glutinosa contains arabinoxylan consists of /4)–d-Xylp-(1/structure, interchanged at C-2 position by -l-Araf-(1/3)–l-Araf-(1/3)–l-Araf-(1/. The isolated arabinoxylan possesses arabinose residues with both a- and b-anomers. This molecule showed strong phagocytotic and immune activation along with the activation of T- and B-lymphocytes [15]. 3.2.2 Glucuronoarabinoxylan Glucuronoarabinoxylan is a type of xylan with arabinosyl and/or glucuronyl residues. They form hydrogen bond with cellulose and therefore strengthen the cell wall [16]. Cell walls in both monocotyledon and dicotyledon plants contain the polymer. Though only 5% of the cell wall content is glucuronoarabinoxylan in the dicotyledons, cell walls in monocotyledons contain 25%. The neutral arabinoxyl- and/or xylosyl-containing side chains is a common structural feature of the primary and secondary walls of monocotyledons. Unlike glucuronoxylan in glucuronoarabinoxylan there are no uronosyl

574 Chapter 25 residues in the middle of the chain but all the residues are located at the end. In stronger leaves like Aechmea the presence of glucuronoarabinoxylan is usually more. 3.2.3 Xyloglucan Xyloglucan possesses 1,4-b-glucan structure with 1,6-a-xylosyl residues [17]. Chemically, the structure of xyloglucan is rigid and identical to cellulose, that is, straight-chain polymer of b-(1/4)-linked D-glucopyranose residues. The distribution of side-chains with the structure follows an order [18]. In dicotyledons, xyloglucan is present in middle lamella of the primary wall and its content in the dry matter is around 20%. It is also present in nongraminaceous monocotyledons and in gelatinous plants. In different plant species, the substitution of side-chain xylosyl residues with different monosaccharides, disaccharides, or trisaccharides produces a broad array of xyloglucan structures [19].

3.3 Mannans Mannans are plant polysaccharides with linear or branched chains primarily made up of D-mannose. The basic mannose chain branches with D-galactose and D-glucose and gets the nomenclature as galactomannan and glucomannan, respectively. The total mannan content and branching of mannan chains depends on the plant species. It is one of the principal components of plant hemicellulose and sometimes acts as C-source storage molecule. During metabolism both endo and exo type hydrolases cleave the mannan backbone to produce variety of oligosaccharides and fermentable sugars. 3.3.1 Glucomannan Glucomannan is abundantly available in plants (e.g., Plantago major). However, the ratio of mannose/glucose monomer in glucomannan matrix depends on the origin. The shape of the glucomannan polymer is similar to cellulose [20]. In the photosynthetically active leaves, the outer green cortex contains glucomannan but in the parenchyma cells it exists in acetylated form (called as acemannan). Glucomannan helps in maintaining cell moisture content. It gets synthesized in Golgi apparatus and travels out of the cell to form part of the cell wall [21]. 3.3.2 Galactomannan It is a group of polysaccharides with rigid hydrophilic backbone (polymannose or mannan), and grafted galactose units. In terms of its distribution in the plant world, this biopolymer represents the second largest group of storage polysaccharides. It has been identified that some of the leafy vegetables like fenugreek leaves, butterfly pea leaves contain galactomannan with antioxidant and immunostimulant activity [22].

Polysaccharides from leafy vegetables: chemical, nutritional and medicinal properties

575

3.4 Pectin Pectin represents a complex group of cell wall polysaccharides in plants that plays role in cell wall growth, intercellular communication and cellular defense [23]. It is synthesized in Golgi apparatus and gets transported to the middle lamella or space between two adjacent cells in a tissue [24]. Pectin content and its structure and chemical composition varies in different plants. Structurally they are hetero-polymers with galacturonic acid as the major component. The composition of these polysaccharides are often so complex that establishing their structure accurately is a daunting task. Nevertheless, pectic polysaccharides are classified as homogalacturonan, rhamnogalacturonan I, rhamnogalacturonan II, and xylogalacturonan (Fig. 25.3C) [25]. Though pectin is mostly extracted from fruits (e.g., apple, apricot, oranges, etc.) but it is also found in leafy vegetables especially in Brassica species. 3.4.1 Homogalacturonan Homogalacturonan (HG) is a straight chain polymer of a-1,4-linked D-galacturonic acid with a mean chain length of 100 units. It represents almost 65% of the total pectic substances. The galacturonan chain may contain monomers with esterified (C-6) carboxylic group and/or hydroxyl group (at O-2/O-3 position). The degree and type of esterification varies in different plant species [26]. The negatively charged unmethylated HG may form complex with Ca2þ. If proper atmosphere is provided to produce the network a stable gel forms through such complexation of unmethyl-esterified GalA residues. This process is called gelation of pectin [27]. 3.4.2 Rhamnogalacturonan These pectic polysaccharides are composed of disaccharide subunits of galacturonic acid and rhamnose. Rhamnogalacturonans-I and II (RG I and RG II)) are two different types of this heteropolymers. RG I is the major one which represents 20%e35% of pectin. In RG I, the galacturonic acid residue is connected with one neutral glucosyl residue and a-L rhamnose residues are connected with different polymeric side chains like a-(1,5)-L arabinans, b-(1,4)-D galactans, arabinogalactans-I and II and galacto-arabinans [26]. RG-I remains connected covalently with side chain of other pectic components such as HG and RG-II [27]. RG II is a minor pectin component with 0.5%e8% present in dicotyledons and >0.1% in monocotyledons. Usually in plant cell wall, rhamnogalacturonans-II exists as dimers interlinked via apiosyl residues [28]. The pectic polymers (HG, RG-I, and RG-II) give strength as well as flexibility to the cell wall through covalent linkages [13]. There are many leaves like Arabidopsis thaliana, Panax ginseng, Salvia officinalis, Impatiens

576 Chapter 25 parviflora, Artemisia princeps, Mesembryanthenum crystallinum, Gingko Biloba that contains rhamnogalacturonans and have high antioxidant activity. 3.4.3 Xylogalacturonan As the name suggests this pectic component contains a primary chain of galacturonic acid with xylose residues as side chain. Xylogalacturonan is mostly present in reproductive tissues along with other tissues including leaves [24]. In the “hairy” portions of pectin, xylogalacturonan is a side chain of rhamnogalacturonan I [29]. The ratio of b-xylose and galacturonic acid may vary significantly, depending on the origin of the xylogalacturonans [30].

3.5 Arabinogalactan Arabinogalactan comes under a class of heteropolysaccharides or more precisely proteoglycans (Fig. 25.3D). This molecule is part of hemicellulosic polysaccharide in the cell walls of plants but also capable of cross-linking with membrane proteins possibly to play a role in the intercellular signaling, healing of damaged tissues and many other cellular events [31]. They are categorized into two types; Arabinogalactan I (AG I) and Arabinogalactan II (AG II) based on the arrangement of galactose and arabinose units. AG I consist of galactose residues with combination of b 1-3, b 1-4, and b1-6 linkages. In AG II the galactose backbone (b1-3 linked) contains terminal arabinose or rhamnose residues. The arabinogalactan-protein (AGP) complex contains more than 90 % polysaccharide part with some Hyp/Pro, Ala, and Ser/Thr rich peptides. In the context of leafy vegetables, it was found that cabbage leaves mainly contain water soluble proteoglycan [32]. One of the characteristic feature of AGP in leaves of brassicaceous (cruciferous) plants, Arabidopsis, radish, rape leaves, and common plantain is the presence of L-fucose sugar in the peptidoglycan [33].

3.6 Functional attributes of the polysaccharides from leaves Various functional properties have been identified for the leaves originated polymeric compounds ranging from dietary fiber to immunostimulatory and antioxidant activities. The literature available on functional activities of these polymers are mostly in vitro studies. Also the identified structure of a polysaccharide is very much dependent on recovery methods so removal of any structural ambiguity is essential before initiating any functional studies. A discussion of the health benefits in connection to these polymers is attempted to encourage individual polysaccharide level studies for their functional properties. 3.6.1 Dietary fiber Dietary fibers are polymers derived from mostly plant sources which do not undergo hydrolysis by human enzymes. The main dietary fiber components of leafy vegetables are

Polysaccharides from leafy vegetables: chemical, nutritional and medicinal properties

577

cellulose, inulin, hemicellulosic polysaccharides and pectin substances. These fibers can be of two types water-soluble and water-insoluble fiber. Water soluble fibers have potential hypoglycemic properties because they can downregulate post-meal blood glucose level. Such hypoglycemic properties have been confirmed experimentally for certain watersoluble fibers like pectin. In addition to that, there are studies suggesting possibility of colon cancer can be reduced by consumption of pectin [34]. A dietary fiber may be active in their natural form with or without the other cellular components. Sometimes the polysaccharides may require chemical or enzymatic modification to become active. Generally, the cellulose and hemicellulose component can expedite gut movement without any treatment. On the other hand, inulin is incapable of accelerating gut movement in native condition and needs to be converted to short chain fatty acids by gut bacteria. Pectins, oligosaccharides, and b-glucans cause slower movement of food in the gut and increase viscosity of the feces. Net result of such slow bowel movement is the reduction of blood cholesterol level and reduction in sugar absorption from the food into the blood [35]. As a component of dietary fiber, arabinoxylans exists in many leafy vegetables such as spinach, parsley, cabbage, lettuce, etc. [36]. A section of the dietary fiber is called Prebiotics. They are various group of polysaccharides or more precisely oligosaccharides with following properties; they do not get hydrolyzed to monomers by acid or human enzymes in the gastrointestinal tract but get fermented by intestinal microflora and act as a preferred substrate for at least one of the beneficial bacteria species capable of colonizing in the colon. As a result these polysaccharides restore healthy composition of colonic microflora and exert health benefit to the host body holistically [37]. Xylans, fructans, galactans are well known prebiotics. However, many other polysaccharides from plant tissues are capable of showing prebiotic effects. For example, human enzymes are incapable of solubilizing arabinoxylans as it contains arabinose and xylose linkages. It enters the colon in intact form and acts as a source of food for saccharolytic bacteria along with Bacteroides, Bifidobacterium, Clostridium, Lactobacillus, and Eubacterium to maintain a balanced gut condition [38]. Prebiotic polysaccharides have been identified in leafy vegetables such as Lactuca sativa, Apium graveolens, Spinacia oleracea, Corchorus capsularis, etc. (refer Table 25.1). Since prebiotics support growth of a very narrow group of gut microflora unlike dietary fibers, establishing their health benefit is more challenging and requires an interdisciplinary approach. To establish the prebiotic roles, advance in situ microbial growth monitoring methods like fluorescence-probeebased techniques can be used and the obtained information can be further enhanced using simulated gut models. 3.6.2 Immunostimulant Immunostimulants are diverse group of chemicals that can augment biological defense system of an animal against various pathogenic bacteria or virus. These substances can be

Table 25.1: Polysaccharides and their functional properties from some selected leafy vegetables. Polysaccharide Leafy vegetables

Dietary Fiber

Prebiotic

Immuno-stimulant

Antioxidant

References

Pectin, homogalacturonan, hemicellulose, xyloglucan, cellulose, inulin Pectin, homogalacturonan, galacturonan, xyloglucan, rhamnogalacturonan, cellulose, arabinogalactan Xylan, xyloglucan, pectin, cellulose Pectin, hemicellulose, cellulose, xylan

Pectin, hemicellulose, homogalacturonan, inulin

Pectin, hemicellulose, homogalacturonan, inulin, xyloglucan

Inulin

Wagstaff et al. [39]; Hoffman et al. [40]; Williams et al. [41]

Pectin, homogalacturonan, arabinogalactan

Pectin, homogalacturonan, arabinogalactan, xyloglucan

Rhamnogalacturonan

Petrova et al. [42]; Chen et al. [43]; Zujovic et al. [44]

Xylan, hemicellulose, pectin Pectin, hemicellulose, xylan

Pectin, hemicellulose, xyloglucan Pectin, hemicellulose

Xylan

Miller [45]

Trigonella foenumgraecum

Galactomannans, pectin, hemicellulose

Galactomannans, pectin, hemicellulose

Pectin, hemicellulose, galactomannan

Sulfated polysaccharides, xylan, rhamnogalacturonan e

Brassica oleracea var. capitate

Arabinoxylan, pectin, xyloglucan, hemicellulose, arabinogalactan Pectin, cellulose, starch, hemicellulose, xylan Pectin, hemicellulose, xyloglucan Pectin, hemicellulose, homogalacturonan, xylogalacturonan, xyloglucan, glucuronoarabinoxylan, cellulose

Arabinoxylan, pectin, hemicellulose, arabinogalactan

Pectin, hemicellulose, xyloglucan, arabinogalactan

Rhamnogalacturonan

Eskander [46]; Sarkar et al. [47]; Katayama et al. [48] Majeed et al. [49]; Srichamroen et al. [50]; Aboughe-Angone et al. [51] Steven et al. [52]; Westereng et al. [53]

Pectin, hemicellulose, xylan Pectin, hemicellulose

Pectin, hemicellulose

Rhamnogalacturonan, xylan e

Lactuca sativa

Apium graveolens

Spinacia oleracea Corchorus capsularis

Morus alba Tamarindus indica Arabidopsis thaliana

Pectin, hemicellulose, homogalacturonan, xylogalacturonan, glucuronoarabinoxylan

Pectin, hemicellulose, xyloglucan Pectin, hemicellulose, homogalacturonan, xylogalacturonan, glucuronoarabinoxylan

Rhamnogalacturonan

Katayama et al. [48]; Anand et al. [12] Aboughe-Angone et al. [51] Yapo et al. [54]; Harholt et al. [9]; Zablackis et al. [55]

Panax ginseng

Arabinogalactans, pectins

Arabinogalactans, pectins

Arabinogalactans, pectins

Rhamnogalacturonan

Gingko biloba

Cellulose, hemicellulose, pectin, arabinogalactan, xylan, galactan Cellulose, hemicellulose, pectin, homogalacturonan Pectin, cellulose, arabinogalactan Pectin, hemicellulose, galactan, xylan, xyloglucan Pectin, cellulose, arabinogalactan, hemicellulose, galacturonan, galactan Pectin, cellulose, homogalacturonan arabinogalactan, hemicellulose Arabinogalactan, galactan Arabinoxylan, xylan, hemicellulose, galactomannan, galactoglucomannan Pectin, hemicellulose, galactan, glucuronoxylan, arabinogalactan Pectin, arabinogalactan, xyloglucans and heteroxylans Pectin, arabinogalactan Arabinogalactan, pectin, galacturonan, galactan

Xylan, hemicellulose, pectin, galactan

Hemicellulose, pectin, galactan

Rhamnogalacturonan, xylan

Xiu-zhen et al. [56]; Aboughe-Angone et al. [51] Kraus et al. [57]; Jin et al. [58]

Hemicellulose, pectin

Hemicellulose, pectin

Rhamnogalacturonan

Chen et al. [59]

Pectin, arabinogalactan

Pectin, arabinogalactan

Rhamnogalacturonan

Vagi, [60]

Pectin, hemicellulose, galactan, xylan, xyloglucan Pectin, hemicellulose arabinogalactan, galactan

Pectin, hemicellulose, galactan, xyloglucan

Xylan

Arribas et al. [61]

Pectin, hemicellulose, arabinogalactan, galactan

Glucomannan

Adom et al. [62]; Lukova et al. [63]

Pectin, arabinogalactan, homogalacturonan, hemicellulose, Arabinogalactan, galactan Arabinoxylan, xylan, hemicellulose, galactomannan, galactoglucomannan Hemicellulose, galactan, glucuronoxylan, arabinogalactan Pectin, arabinogalactan, xyloglucans and heteroxylans Pectin, arabinogalactan Arabinogalactan, pectin, galacturonan, galactan

Pectin, homogalacturonan, arabinogalactan, hemicellulose, Arabinogalactan, galactan Arabinoxylan, xylan, hemicellulose, galactomannan, galactoglucomannan Hemicellulose, galactan, glucuronoxylan, arabinogalactan Pectin, arabinogalactan, xyloglucans and heteroxylans Pectin, arabinogalactan Arabinogalactan, pectin, galacturonan, galactan

Rhamnogalacturonan

Msakni et al. [64]

Rhamnogalacturonan

Yamadat et al. [65]

Xylan

Gaillard et al. [66]

Rhamnogalacturonan, xylan

´dkova´ et al. [67] Hroma

Heteroxylans

Oliveira et al. [68]; Puri et al. [69]

e Rhamnogalacturonan

Raja et al. [70] Capek et al. [71]

Epimedium sp.

Thymus vulgaris Phaseolus vulgaris

Plantago major

Mesembryanthenum crystallinum

Artemisia princeps Trifolium pratense

Impatiens parviflora

Stevia rebaudiana

Moringa oleifera Salvia officinalis

580 Chapter 25 derived from natural sources or can be chemically synthesized. Often the immunostimulants may work specifically as antigens which can induce production of some specific antibody (e.g., vaccines). The other class of immunostimulants work differently without any antigenic property of their own. Instead they promote immune response of other antigens nonspecifically. There are several types of stimulants identified as complex hemicellulosic polysaccharides (e.g., arabinoxylan, arabinogalactan, etc.) in leafy vegetables [51]. Often a mixture of polysaccharide in crude extract showing immunostimulant activity cannot be explained by a single mechanism. For example, hot infusion of Maytenus ilicifolia could prevent gastric ulcer in rat models. The infusion contained heteropolysaccharide with residues of primarily arabinose and galactose along with small amount of galacturonic acid, 4-O-methylglucuronic acid, rhamnose, and glucose [72]. The mechanism behind nonspecific immune response of these polysaccharides requires further investigation of their phagocytosis, antibody synthesis, properdin and complement systems, interferon production, and lymphocytes synthesis properties. Particularly immunestimulant polysaccharides will be an important area because many of the existing antibiotics are no longer effective due to growing resistance of the pathogens plus their allergenicity, immune suppressive properties, and inefficiency toward viruses. 3.6.3 Antioxidant Essentially substances with reducing property and ability to stop or decelerate the oxidation of cellular components by either direct scavenging of free radicals or by recycling of other free radical scavengers are called antioxidants [73]. Leaves usually contain a huge pool of antioxidants in the form of pyrocatechol or a pyrogallol group (Fig. 25.4A), that is, ortho-disubstituted phenols. However, ortho-disubstituted phenolic compounds are often found to be toxic [74]. Since majority of the synthetic antioxidants have some limitations that offers scope to alternative natural antioxidants of plant origin often from leaves. Currently, to obtain naturally occurring antioxidants leaf polysaccharides are being explored. The structural properties like chemical composition, molar mass, glycosidic bonds, and conformation of the carbons controls the antioxidant activity of a polysaccharide [75]. Arabinoxylans exhibit antioxidant properties because of their linkages with ferulic acid (Fig. 25.4B). The polysaccharide extraction condition in leaves can be optimized using statistical methods in order to maximize their antioxidant activity [76].

4. Polysaccharide from different leafy vegetables Structure-function of the polysaccharides discussed in Section 3 is distributed abundantly in different leaves. In this section some of the leaf sources are enlisted along with the polysaccharides and their possible beneficial effect on health (Table 25.1).

Polysaccharides from leafy vegetables: chemical, nutritional and medicinal properties (A)

581

OH

OH

HO

OH O pyrocatechol

OH OCH3

OH

3,4-dihydro-2H-1-benzopyran-6-ol

2-methoxyphenol

hydroquinone

(B)

Lignin C

O

O CH2 O

O

O O

O

O

Arabinoxylan residue

O

O

O

O

O

O

O

O 2HC

2HC

O

O O

C

O

C

OH

Phenol OCH3 group

OH

CH3O

OH

OCH3

CH3O OH

C

C

O Arabinose residue

O 2HC

O

O

O

O O

O O

O

O

CH2

O O

O

O

O

Figure 25.4 (A) Basic chemical structure of phenolic antioxidants and (B) generic structure of arabinoxylans with antioxidant activity.

582 Chapter 25

5. Effect of processing condition on polysaccharide structure and functional relationship Polysaccharide properties from leafy vegetables or their derived products (cooked or chopped vegetables) is dependent on the method of processing as well as extraction. Extraction of polysaccharide is a sequential process which often starts from drying in case of dry leaf extraction procedures. Alternatively, direct fresh leaves can be used for extraction of different types of polysaccharide components. Drying is one of the primary steps in the extraction of polysaccharide from leaves, which not only reduces moisture content of the material but offers scope for plethora of reactions (e.g., hydrolysis, enzyme inactivation, cross-linking, etc.). Unfortunately not many in-depth studies on effect of processing conditions on leaf-polysaccharide qualities are there. Polysaccharide quality in economically important Aloe vera leaves has been reviewed recently. Starting from microbial quality during handling of leaves to filleting or crushing of the leaves, heating of the juice, and drying, every step altered the polysaccharide composition of the leaves [77]. It was also reported that different pulp fraction of the leaves show different polysaccharide composition. Like cell wall, disintegrated cell organelles and mesophyll cell fluid are respectively rich in polygalactose, galacturonic acid, and mannans in A. vera. [78] reported acemannan chains in A. vera leaves suffered significant changes when dried at higher temperature (70 C). The mean molecular weight of the polysaccharide increased from 45 to 75 kDa at higher temperature. Drying can be done via different mechanisms; solar drying, convective drying, vacuum drying, freeze drying, microwave drying, refractive window, etc. Each of these methods has its own merits and demerits but freeze drying is best in terms of disturbance of the polysaccharide molecules. Polysaccharides are separated from leafy vegetables with selected solvents (water/hot water/alcohol) or their mixture. Sometimes ultrasound, microwave, and enzymatic degradation is used to increase yield and/or selectivity of the polysaccharides. Similarly, processing conditions before consumption of leafy vegetables have major effect on structure of the polysaccharide molecules which alters their reactivity with the amylases and therefore bioavailability of the polysaccharide. The effect of processing condition is difficult to estimate because it varies dramatically depending on the polymer type and its exact structure. Also polysaccharide modifies its environment in solution during processing. Soluble polysaccharide increases the viscosity of a solution, subject to their structure and chain length, concentration, and presence of other species in the solvent [79]. Nonstarch polysaccharides (NSPs) with charged functional groups like pectic polysaccharides are more soluble in polar solvents including water. pH of the solvent determines the charge on the NSPs. The charge distribution of the polysaccharides may have multiple influence, namely, reduction of their solubility, gelation, or degradation of the polymeric chain in extreme case. Since food particle size and compactness hinders the

Polysaccharides from leafy vegetables: chemical, nutritional and medicinal properties

583

accessibility of the hydrolyzing enzymes, disruption of the cell structure by grinding and milling increases starch bioavailability. Moisture, cooking temperature, time, and pressure control the starch gelatinization and thus, the transformation of starch from resistant starch (RS) to slowly digestible starch and then to rapidly digestible starch occurs during cooking. RS starch is part of the wider group of dietary fibers. Physically inaccessible starch fraction is found in starchy foods that are not fractionated and refined. Ungelatinized starch is mainly structured as B-type crystal that are tightly packed and relatively dehydrated inside the granules. This dense structure hinders approach of amylases. Since, it is well known that thermal degradation promotes solubilization of starch, moist heat cooking methods like boiling, steaming, frying, microwaving, and autoclaving affect the content of resistant starch. In addition to that deacetylation, loss of side chains of nonstarch polysaccharides and their cross-linking with proteins (enzymatic and nonenzymatic browning) have also been established.

6. Negative physiological impact of plant polysaccharides: food and drug interaction Many food components are known for their interactions with drug molecules and in turn affect clinical efficacy of the drug. However, these effects may be additive, synergistic, or antagonistic depending on the chemistry of the drug and the food after digestion. So nature and extent of the interaction is related to the bioavailability of the food component(s). One well-known case for leafy vegetables is interaction of coenzyme Q-10 with P-glycoprotein, the efflux transporter of the intestine. The coenzyme is present in many leafy vegetables and considered as a health supplement [80]. Another drug compound, warfarin is used for prevention of blood clots and their passage in the human body. However, its effectiveness is largely dependent on correct dosage to avoid unsafe levels of anticoagulation. Vitamin K interferes negatively with warfarin, hence leafy vegetables rich in vitamin K show erratic anticoagulation behavior during warfarin treatment. Especially presence of other vitamins A, E, or C may alter the steps in anticoagulation process [81]. Such interactions are very much dependent on leaf matrix and the particle size in which the leaves are consumed. Polysaccharides of leafy vegetables therefore have an important role in controlling these drug-food interactions. However very little is known about the precise mechanisms of leafy vegetable polysaccharides in the context of drug-food interactions. Generally, it has been observed that fiber-rich food products are capable of reducing the efficacy of drugs like simvastatin, ezetimibe, pravastatin, and fluvastatin. In presence of pectin, acetaminophen absorbs at a slower rate through intestinal mucosa. So in presence of polysaccharides, delay in bioavailability of drug molecules can be anticipated even if there is no reduction in their absolute bioavailability.

584 Chapter 25

7. Conclusions Recently polysaccharides and oligosaccharides from leafy vegetables are attracting researchers globally because of its beneficial properties on human health. Cell wallederived polysaccharides and few other poly-/oligosaccharides from leafy vegetables show promising prebiotic, immunostimulatory, and antioxidant effect. Although, there is a considerable amount of literature on application of plant polysaccharides in food and nonfood areas, polysaccharides from leafy vegetables are still lesser known in terms of their application. Basic information about the structure and function of such polysaccharides is still evolving. Also the broad definition of leafy vegetables encompasses over a thousand plant species which needs to be explored for functional polysaccharides. It is evident in the present review that most of the functional polysaccharides in leaves are complex heteropolysaccharides like pectin, glucuronoarabinoxylan, galactomannan, etc. The bioactivity of a polysaccharide is related to its monomers, molecular weight, chemical components, extent of branching, and even the type of glycosidic bonds. The knowledge about these bioactivities is important not only for their beneficial effect but also to predict their undesired effect like drug-food interaction. Another finding of the review is that the structural information of the polysaccharides is very much dependent on their extraction process and processing of the leaves. Indeed, these unique polysaccharides are promising but their large number and the inherent complexity of the molecules make their application challenging.

References [1] Xu Z. Introductory chapter: polysaccharides and their solubility. InTech; 2017. p. 4e5. [2] Carpita CN, Gibeaut MD. Structural models of primary cell walls in flowering plants: consistency of molecular structure with the physical properties of the walls during growth. Plant J 1993;3(1):1e30. [3] McCann MC, Roberts K. Architecture of the primary cell wall. In: Lloyd CW, editor. The cytoskeletal basis of plant growth and form. London: Academic Press; 1991. p. 109e29. [4] Eisenach C, Francisco R, Martinoia E. Plant Vacuoles. Curr Biol 2015;25(4):R136e7. [5] Pfister B, Zeeman SC. Formation of starch in plant cells. Cell Mol Life Sci 2016;73:2781e807. [6] Islam RM, Paul KD, Shaha KR. Nutritional Importance of some leafy vegetables available in Bnagladesh. Pakistan J Biol Sci 2004;7(8):1380e4. [7] Nwafor CI, Shale K, Achilonu CM. Chemical composition and nutritive benefits of chicory (Cichorium intybus) as an ideal complementary and/or alternative livestock feed supplement. Sci World J 2017:1e11. Article ID 7343928. [8] Ebringerova A, Heinze T. Xylan and xylan derivatives e biopolymers with valuable properties, 1; Naturally occurring xylans structures, isolation procedures and properties. Macromol Rapid Commun 2000;21:542e56. [9] Harholt J, Suttangkakul A, Scheller VH. Biosynthesis of pectin. Plant Physiol 2010;153:384e95. [10] Coimbra S, Pereira GL. Arabinogalactan proteins in Arabidopsis thaliana pollen development. Transgenic Plants Adv Limit 2012;16:329e52.

Polysaccharides from leafy vegetables: chemical, nutritional and medicinal properties

585

[11] Kays JS, Nottingham FS. Biology and chemistry of jerusalem artichoke: Helianthus tuberosus L. Taylor and Francis Group; 2008. p. 1e423. [12] Anand PAA, Vennison JS, Sankar GS, Prabhu GID, Vasan TP, Raghuraman T, Geoffrey JC, Vendan ES. Isolation and characterization of bacteria from the gut of Bombyx mori that degrade cellulose, xylan, pectin and starch and their impact on digestion. J Insect Sci 2009;10:1e20. [13] Ochoa-Villarreal M, Aispuro-Herna´ndez E, Vargas-Arispuro I, Martı´nez-Te´llez AM. Plant cell wall polymers: function, structure and biological activity of their derivatives. Intech; 2012. p. 63e86. [14] Rivie`re A, Moens F, Selak M, Maes D, Weckx S, Vuysta DL. The ability of bifidobacteria to degrade arabinoxylan oligosaccharide constituents and derived oligosaccharides is strain dependent. Appl Environ Microbiol 2014;80(1):204e17. [15] Das D, Maiti S, Maiti TK, Islam SS. A new arabinoxylan from green leaves of Litsea glutinosa (Lauraeae): structural and biological studies. Carbohydr Polym 2013;92(2):1243e8. [16] Darvill EJ, McNeil M, Darvill GA, Albersheim P. Structure of plant cell walls: XI. Glucuronoarabinoxylan, a second hemicellulose in the primary cell walls of suspension-cultured sycamore cells. Plant Physiol 1980;66(6):1135e9. [17] Hayashi T, Kaida R. Functions of xyloglucan in plant cells. Mol Plant 2011;4(1):17e24. [18] Fry CS. The structure and functions of xyloglucan. J Exp Bot 1989;40(1):1e11. [19] Zabotina AO. Xyloglucan and its biosynthesis. Front Plant Sci 2012;3:1e5. [20] Alonso-Sande M, Teijeiro-Osorio D, Remun˜a´n-Lo´pez C, Alonso JM. Glucomannan, a promising polysaccharide for biopharmaceutical purposes. Eur J Pharm Biopharm 2009;72:453e62. [21] Salinas P, Salinas C, Contreras AR, Zun˜iga EG, Dupree P, Cardemil L. Expression of a glucomannan mannosyltransferase gene (GMMT) from Aloe vera is induced by water deficit and abscisic acid. BioRxiv 2018:1e42. [22] Paulovicova´ E, Paulovicova´ L, Hrubisko M, Krylov BV, Argunov AD, Nifantiev EN. Immunological activity of synthetically preparedimmunodominant galactomannosides structurally mimicking Aspergillus galactomannan. Front Immunol 2017;8:1e14. [23] Jarvis MC. Structure and properties of pectin gels in plant cell-walls. Plant Cell Environ 1984;7:153e64. [24] Jensen KJ, Sørensen OS, Harholt J, Geshi N, Sakuragi Y, Møller I, Zandleven J, Bernal JA, Jensen BN, Sørensen C, Pauly M, Beldman G, Willats TGW, Scheller VH. Identification of a xylogalacturonan xylosyltransferase involved in pectin biosynthesis in Arabidopsis. Plant Cell 2008;20:1289e302. [25] Hamman HJ. Chitosan based polyelectrolyte complexes as potential carrier materials in drug delivery systems. Mar Drugs 2010;8:1305e22. [26] Wolf S, Mouille G, Pelloux J. Homogalacturonan methyl-esterification and plant development. Mol Plant 2009;2(5):851e60. [27] Darvill GA, Albersheim P, Mcneil M, Lau MJ, York SW, Stevenson TT, Thomas J, Doares S, Gollin JD, Chelf P, Davis K. Structure and function of plant cell wall polysaccharides. J Cell Sci 1985;2:203e17. [28] Mohnen D. Pectin structure and biosynthesis. Curr Opin Plant Biol 2008;11:266e77. [29] Zandleven J, Sørensen OS, Harholt J, Beldmana G, Schols AH, Scheller VH, Voragen JA. Xylogalacturonan exists in cell walls from various tissues of Arabidopsis thaliana. Phytochemistry 2007;68(8):1219e26. [30] Caffall HK, Mohne D. The structure, function, and biosynthesis of plant cell wall pectic polysaccharides. Carbohydr Res 2009;344:1879e900. [31] Tan L, Eberhard S, Pattathil S, Warder C, Glushka J, Yuan C, Hao Z, Zhu X, Avci U, Miller SJ, Baldwin D, Pham C. An arabidopsis cell wall proteoglycan consists of pectin and arabinoxylan covalently linked to an arabinogalactan protein. Am Soc Plant Biol 2013:1e18. [32] Yasufuku H, Kido S, Azuma J, Okamura K. Change of water-soluble arabinogalactan-proteins of cabbage accompanying its growth. Biosci Biotechnol Biochem 1994;58(2):225e9. [33] Samuelsen BA, Paulsena SB, Weld KJ, Knutsen HS, Yamadap H. Characterization of a biologically active arabinogalactan from the leaves of Plantago major L. Carbohydr Polym 1998;35:145e53.

586 Chapter 25 [34] Englyst NH, Bingham AS, Runswick AS, Collinson E, Cummings HJ. Dietary fibre (non-starch polysaccharides) in fruit, vegetables and nuts. J Hum Nutr Diet 1988;1:247e86. [35] Liu J, Wang W, Yu H, Cao X, Wang Y, Liu M, Yan X, Liu H. Antioxidant activity of polysaccharides extracted from Lentinus edodes mycelia and their protective effects on ins-1 cells. Biosci J 2015;32(6):1679e88.  [36] Dodevska M, Sobaji c S, Djordjevic B. Fibre and polyphenols of selected fruits, nuts and green leafy vegetables used in Serbian diet. J Serb Chem Soc 2015;80(1):21e33. [37] Carlson LJ, Erickson MJ, Lloyd BB, Slavin LJ. Health effects and sources of prebiotic dietary fiber. Curr Dev Nutr 2018;2:1e8. [38] Sinha KA, Kumar V, Makkar SPH, Boeck DG, Becker K. Non-starch polysaccharides and their role in fish nutrition e a review. Food Chem 2011;127:1409e26. [39] Wagstaff C, Clarkson JJG, Zhang F, Rothwell DS, Fry CS, Taylor G, Dixon SM. Modification of cell wall properties in lettuce improves shelf life. J Exp Bot 2010;61(4):1239e48. [40] Hoffman M, Jia Z, Penea JM, Cash M, Harper A, Blackburn RA, II, Darvill A, York SW. Structural analysis of xyloglucans in the primary cell walls of plants in the subclass Asteridae. Carbohydr Res 2005;340:1826e40. [41] Williams AB, Grant JL, Gidley JM, Mikkelsen D. Gut fermentation of dietary fibres: physico-chemistry of plant cell walls and implications for health. Int J Mol Sci 2017;18:2203. [42] Petrova I, Petkova N, Kyobashieva K, Denev P, Simitchiev A, Todorova M, Dencheva N. Isolation of pectic polysaccharides from celery (Apium graveolens var. rapaceum D. C.) and their application in food emulsions. Turk J Agric Nat 2014. [43] Chen D, Harris JP, Sims MI, Zujovic Z, Melton DL. Polysaccharide compositions of collenchyma cell walls from celery (Apium graveolens L.) petioles. BMC Plant Biol 2017;17:104. [44] Zujovic Z, Chen D, Melton DL. Comparison of celery (Apium graveolens L.) collenchyma and parenchyma cell wall polysaccharides enabled by solid-state 13C NMR. Carbohydr Res 2016;420:51e7. [45] Miller GJ, Buchanan JC, Eastwood AM, Fry CS. The solubilisation and hydrolysis of spinach cell wall polysaccharides in gastric and pancreatic fluids. J Sci Food Agric 1995;68(3):389e94. [46] Eskander ZM. Extraction of polysaccharides from the leaves of jews-mallow Corchorus olitorius L. and their potential anticoagulant activity. Basrah J Vet Res 2018;17(1):277e90. [47] Sarkar B,P, Mazumdar KA, Pal BK. The nature of the hemicelluloses of jute fiber Part I. Text Res J 1952:529e34. [48] Katayama H, Takano R, Sugimura Y. Localization of mucilaginous polysaccharides in mulberry leaves. Protoplasma 2008;233:157e63. [49] Majeed M, Majeed S, Nagabhushanam K, Arumugam S, Natarajan S, Beede K, Ali F. Galactomannan from Trigonella foenum-graecum L. seed: prebiotic application and its fermentation by the probiotic Bacillus coagulans strain MTCC 5856. Food Sci Nutr 2018;6:666e73. [50] Srichamroen A, Field JC, Thomson RBA, Basu KT. The modifying effects of gonalactomannan from Canadian-grown fenugreek (Trigonella foenum-graecum L.) on the glycemic and lipidemic status in rats. J Clin Biochem Nutr 2008;43:167e74. [51] Aboughe-Angone S, Nguema-Ona E, Boudjeko T, Driouich A. Plant cell wall polysaccharides: immunomodulators of the immune system and source of natural fibers. Curr Top Phytochem 2011:1e17. [52] Stevens HJB, Robert R, Selvendran RR. Pectic polysaccharides of cabbage (Brassica oleracea). Phytochemistry 1984;23(1):107e15. [53] Westereng B, Yousif O, Michaelsen ET, Knutsen HS, Samuelsen BA. Pectin isolated from white cabbage e structure and complement-fixing activity. Mol Nutr Food Res 2006;50:746e55. [54] Yapo MB. Rhamnogalacturonan-I: a structurally puzzling and functionally versatile polysaccharide from plant cell walls and mucilages. Polym Rev 2011;51(4):391e413. [55] Zablackis E, Huang J, Mu¨llerz B, Darvill CA, Albersheim P. Characterization of the cell-wall polysaccharides of Arabidopsis thaliana leaves. Plant Physiol 1995;107(4):1129e38.

Polysaccharides from leafy vegetables: chemical, nutritional and medicinal properties

587

[56] Xiu-zhen N, Bing-qing W, Yuan Z, Ning-ning W, Xu Z, Shan-shan L, Gui-hua T, Yi-fa1 Z, Ji-ming Z. Total fractionation and analysis of polysaccharides from leaves of Panax ginseng. Chem Res Chin Univ 2010;26(2):230e4. [57] Kraus J. Water-soluble polysaccharides from Ginkgo biloba leaves. Phytochemistry 1991;30(9): 3017e20. [58] Jin Z, Yeoup B, Iiyama K, Watanabe S. Changes in chemical components of leaf litter of Ginkgo biloba during mulching. J Arboric 2002;28(4):171e7. [59] Chen R, Li S, Liu C, Yang S, Li X. Ultrasound complex enzymes assisted extraction and biochemical activities of polysaccharides from Epimedium leaves. Process Biochem 2012;47:2040e50. [60] Vagi ME. Supercritical fluid extraction of plants and the functional properties of the extracts. Department of Chemical Engineering; 2005. [61] Arribas A, Revilla G, Zarra I, Lorences PE. Changes in cell wall polysaccharides during the growth of Phaseolus vulgaris leaves. J Exp Bot 1991;42(242):1181e7. [62] Adom BM, Tahera M, Mutalabisina FM, Amria SM, Kudosa ABM, Sulaiman WAWM, Senguptaa P, Susanti D. Chemical constituents and medical benefits of Plantago major. Biomed Pharmacother 2017;96:348e60. [63] Lukova KP, Karcheva-Bahchevanska PD, Bivolarski PV, Mladenov DR, Iliev NI, Nikolova MM, Medica F. Enzymatic hydrolysis of water extractable polysaccharides from leaves of Plantago major L. Folia Med 2017;59(2):210e6. [64] Msakni HN, Hatem Majdou H, Roudesli S, Picton L, Cerf LD, Rihouey C, Morvan C. Composition, structure and solution properties of polysaccharides extracted from leaves of Mesembryanthenum crystallinum. Eur Polym J 2006;42:786e95. [65] Yamadat H, Otsukat Y, Omura S. Structural characterization of anti-complementary polysaccharides from the leaves of Artemisia princeps. Planta Med 1986:311e4. [66] Gaillard EDB, Bailey WR. The distribution of galactose and mannose in the cell-wall polysaccharides of red clover (Trifolium pratense) leaves and stems. Phytochemistry 1968;7:2037e44. [67] Hroma´dkova´ Z, Kosta´lova´ Z, Vrchotova´ N, Ebringerova´ A. Non-cellulosic polysaccharides from the leaves of small balsam (Impatiens parviflora DC.). Carbohydr Res 2014;389:147e53. [68] Braz de Oliveira JA, Cordeiro CML, Goncalvesa CAR, Ceole FL, Ueda-Nakamura T, Iacomini M. Structure and antiviral activity of arabinogalactan with (1/6)-b-d-galactan core from Stevia rebaudiana leaves. Carbohydr Polym 2013;94:179e84. [69] Puri M, Sharma D, Barrow JC, Tiwary KA. Optimisation of novel method for the extraction of steviosides from Stevia rebaudiana leaves. Food Chem 2016;132:1113e20. [70] Raja W, Bera K, Ray B. Polysaccharides from Moringa oleifera gum: structural elements, interaction with b-lactoglobulin and antioxidative activity. R Soc Chem Adv 2016;6:75699e706. [71] Capek P, Hribalova V. Water-soluble polysaccharides from Salvia officinalis L. 2004. [72] Cipriani RT, Mellinger GC, de Souza ML, Baggio HC, Freitas CS, Marques ACM, Gorin JAP, Sassaki LG, Marcello Iacomini M. A polysaccharide from a tea (infusion) of Maytenus ilicifolia leaves with anti-ulcer protective effects. J Nat Prod 2006;69(7):1018e21. [73] Maestri DM, Nepote V, Lamarque AL, Zygadlo JA. Natural products as antioxidants. Phytochemistry. Adv Res 2006:1e34. [74] Wang H, Liu MY, Qi MZ, Wang YS, Liu XS, Li X, Wang JH, Xia CX. An overview on natural polysaccharides with antioxidant properties. Curr Med Chem 2013;20:2899e913. [75] Wang J, Hu S, Nie S, Yu Q, Mingyong Xie M. Reviews on mechanisms of in vitro antioxidant activity of polysaccharide. Oxid Med Cell Longev 2016:1e13. [76] Samavati V, Manoochehrizade A. Dodonaea viscosa var. angustifolia leaf: new source of polysaccharide and its anti-oxidant activity. Carbohydr Polym 2013;98(1):199e207. [77] Ramachandra TC, Rao SP. Processing of aloe vera leaf gel: a review. Am J Agric Biol Sci 2008;3(2):502e10.

588 Chapter 25 [78] nia A, Garcı´a-Pascual P, Simal S, Carmen Rossello´ C. Effects of heat treatment and dehydration on bioactive polysaccharide acemannan and cell wall polymers from Aloe barbadensis Miller. Carbohydr Polym 2003;51(4):397e405. [79] Lovegrove A, Edwards HC, Noni DI, Patel H, El NS, Grassby T, Zielke C, Ulmius M, Nilsson L, Butterworth JP, Ellis RP, Shewry RP. Role of polysaccharides in food, digestion, and health. Crit Rev Food Sci Nutr 2017;57(2):237e53. [80] Bushra R, Aslam N, Khan YA. Food-drug interactions. Oman Med J 2011;26(2):77e83. [81] Manzi FS, Shannon M. Drug interactionsda review. Clin Pediatr Emerg Med 2005:93e102.

Further reading [1] Hocq L, Pelloux J, Lefebvre V. Connecting homogalacturonan-type pectin remodeling to acid growth. Trends Plant Sci 2017;22(1):20e9. [2] Inaba M, Maruyama T, Yoshimi Y, Kotake T, Matsuoka K, Koyama T, Tryfona T, Dupree P, Tsumuraya Y. L-Fucose-containing arabinogalactan-protein in radish leaves. Carbohydr Res 2015. [3] Mark HF, Bikales NM, Overberger CG, Menges G, Kroschwitz JI. Encyclopedia of polymer science and engineering. John Wiley & Sons; 1988. [4] Meghwal M, Goswami KT. A review on the functional properties, nutritional content, medicinal utilization and potential application of fenugreek. J Food Process Technol 2012;3(9):1e10. [5] Shahbazi S, Bolhassani A. Immunostimulants: types and functions. J Med Microbiol Infect Dis 2016;4(3e4):45e51.

C H A P T E R 26

Electrospun natural polysaccharide for biomedical application Bor Shin Chee, Michael Nugent Athlone Institute of Technology, Materials Research Institute, Athlone, Co. Westmeath, Ireland

Chapter Outline 1. Introduction 589 2. Nanotechnology 590 3. Introduction of electrospinning

591

3.1 The influence of parameters on nanofiber morphology

4. Natural polysaccharide

591

593

4.1 Polysaccharide of human origin 594 4.1.1 Hyaluronic acid 594 4.2 Polysaccharide of plant and seaweed origin 4.2.1 Guar gum 598 4.2.2 Carrageenan 599 4.3 Polysaccharide of animal origin 600 4.3.1 Chitin 601 4.3.2 Chitosan 603 4.4 Polysaccharide of microbe origin 604 4.4.1 Xanthan gum 604 4.4.2 Pullulan 607 4.4.3 Dextran 609

596

5. Conclusion 611 References 612

1. Introduction Owing to ongoing environmental concerns, there has been an increasing focus on using environmentally friendly biomaterials found in natural polymers. The preference of using natural polysaccharide is because of the high cell affinity, low immunogenicity, practicality, moldability, flexibility, lightness, durability, chemical and physicochemical stabilities,

Natural Polysaccharides in Drug Delivery and Biomedical Applications. https://doi.org/10.1016/B978-0-12-817055-7.00026-1 Copyright © 2019 Elsevier Inc. All rights reserved.

589

590 Chapter 26 biodegradable and bioadhesive properties, abundance in nature, and low cost [1]. Unfortunately, natural polysaccharides have relatively low mechanical strength. Blending natural polysaccharides with either natural or synthetic polymers can improve the physical properties and biological performances, as well as delivering desired features of the resulting polymer composites [2,3] This chapter has demonstrated many types of research carried out for the advancement of electrospun nanofibers using selected natural polysaccharides for their potential in biomedical applications. Owing to a wide range of natural polysaccharide, it is impossible to cover each natural polysaccharide comprehensively. Therefore, one to two substances were selected to discuss in each category (i.e., polysaccharide from the human, plant, seaweed, animal origins). It is expected that this chapter will serve as a good reference tool for nanomedicine researchers, especially in researching the natural polysaccharide for electrospinning. Moreover, this chapter also highlights the factors that influence the formation of uniform and smooth nanofibers. This can give a better understanding to the researchers for future research activities.

2. Nanotechnology “Nano” comes from the Greek Nanos, meaning extremely small. Nanotechnology is any science, engineering, and technology conducted on a nanoscale. The measurement of nanoscale material is stated in nanometer [4]. Nanotechnology has enabled the researchers to explore the unique, enhanced, or totally novel properties of a material in the nanodimension, in which these properties cannot be discovered from either macro- or microsystems. The term “nanomedicine” indicates the integration of the knowledge of nanotechnology into biomedical applications. Nanomedicine involves several different types of applications and nanodevices: nanoparticles, nanofibers, nanorobots, and nanoelectronics biosensors. In this chapter, the focus is on reviewing the production of natural polysaccharide nanofibers using the technology of electrospinning for biomedical applications. The electrospun nanofibers are potentially used in tissue engineering sector because of biocompatibility, high porosity, and surface to volume ratio. These nanofibers can form scaffolds such as the bone, cartilage, skin, blood vessel, and cardiovascular stent with the ability to enhance cells attachment, migration, and proliferation for tissue regeneration. In addition, electrospun nanofibers are also useful in drug delivery system. The high drug encapsulation efficacy of the nanofibers prevents drug degradation and controls the drug release rate into the human body either in a controlled way or fast-dissolving rate. Moreover, in the medical diagnostic and therapeutic field, the surface area of electrospun

Electrospun natural polysaccharide for biomedical application 591 nanofibers acts as an ultrasensitive biosensor to provide many highly specific binding sites for the detection of cancer cells [5,6]. Besides the nano-sized therapeutic agent, encapsulated polymer is able to circulate in the bloodstream and allow them to reach the target site [7].

3. Introduction of electrospinning Electrospinning allows the processing of nanofibers using different polymers such as natural polymers, synthetic polymers, and blends of synthetic and natural polymers according to the requirements of the specific applications [1]. A typical electrospinning apparatus requires a syringe, a syringe pump, a high-voltage supply, and a collector. The electrospinning involves applying a high voltage at the metallic needle of the syringe and the collector plate. In addition, the syringe is attached to a pump to control the flow rate of the feed solution. When the polymer solution is pumped out from the syringe, it forms a conical shape called “Taylor cone” and accelerated toward the collector plate, forming a nanofibers mesh [8,9]. Electrospinning is considered as an electro-fluid-dynamic process because it is a progress of using an electrical charged polymer solution to form nanofibers [10]. There are two different electrospinning systems (Table 26.1) which included the solvent electrospinning and the solvent-free electrospinning that can be used to fabricate natural polysaccharide. The difference between both is that the solvent electrospinning uses polymer solutions while the solvent-free electrospinning uses polymer melts. Occasionally, solvent-free polymer processing is desirable especially in industrial mass production owing to several problems such as the cost of solvents, the toxicity and inflammability, their storage, disposal, and solvent recovery Therefore, the solvent-free electrospinning is used to produce nanofibers without any solvent residue [11].

3.1 The influence of parameters on nanofiber morphology The polymer solution parameters, processing parameters, and ambient parameters are three main groups of factors that can affect the nanofiber morphology such as the diameter of the nanofiber and the shape (i.e., bead, nonbead, flat) of the nanofiber [8]. The polymer solution parameters include solution viscosity, conductivity, chemical structure, concentration, molecular weight, and volatility of solvent. However, the process parameters consist of the diameter of needle tip, solution flow rate, and feed rate, the distance between capillary tip and collection plate. The ambient parameters include the solution temperature, relative humidity, surrounding air/gas, and vacuum condition [8,12]. The parameters mentioned above are closely related to the electrospinning behavior.

592 Chapter 26 Table 26.1: The solvent and solvent-free electrospinning apparatus. Solvent electrospinning

Horizontal electrospinning Vertical electrospinning Solvent-free electrospinning

Melt electrospinning writing

For example, the formation of uniform electrospun polysaccharide nanofibers is dependent on the degree of chain entanglements, the viscosity of the solution, and the most important requirement which is the weak shear thinning property to encourage the breakdown of liquid jet when dragged by the electric field [13]. The spinning solution is necessary to have a critical concentration and an appropriate viscoelasticity to cause polysaccharide chain entanglements. However, the viscoelasticity above the threshold will prevent the motion induced by the electric field. For instance, in the study of developing xanthan-based biomaterials, the xanthan solution concentration above 2.5% w/v was too viscous to be

Electrospun natural polysaccharide for biomedical application 593 electrospun at this high concentration, whereas the solution concentration below 1% w/v had a low viscosity, and beaded nanofibers were formed. Hence the concentration between 1.5% and 2.5% w/v had a suitable degree of viscoelasticity for electrospinning [14].

4. Natural polysaccharide Four major classes of biopolymers including proteins (e.g., collagen, silk fibroin [SF]), polysaccharides (Table 26.2), deoxyribonucleic acids (DNAs) (e.g., calf thymus Na-DNA), and lipids (e.g., lecithin) are possible to fabricate into nanofibers using electrospinning [3]. Polysaccharides can be categorized into synthetic polysaccharides and natural polysaccharides; the chapter is focused on natural polysaccharides. The term “saccharide” is derived from the Greek word for sugar. There are four types of saccharides, which includes monosaccharide, disaccharide, oligosaccharide, and polysaccharide. A monosaccharide is a simple sugar with the smallest units of saccharide.

Table 26.2: List of natural polysaccharide. Natu ral polysaccharide

Polysaccharide from human origin

Polysaccharide from plant and seaweed origin

Polysaccharide from animal origiin

Poly saccharide from microbe origi n

• •

• • • • • • • • •

•

• • • • • • • •

• •

Heparin Chondroitin sulfate Keratan sulfate Hyaluronic acid

• • • • • •

Pectin Inulin Guar gum Agarose Alginate Dextrin Amylose Xylan Locust Bean Gum Ulvan Carrageenan Porphyrin Fucoidan Psyllium Cellulose an d its derivative s

•

Chitin, Chitosan and its derivatives Glycogen

X anthan gum D extran S chizophyllan P ullulan Xylinan Alginates Gellan Curdlan

594 Chapter 26 Glucose, galactose, and fructose are some of the examples of monosaccharide. D-glucose is the most abundant monosaccharide in animal cells. A disaccharide is made up of two monosaccharides. Maltose, lactose, and sucrose are some of the examples of disaccharide. An oligosaccharide consists of short chains of monosaccharides, typically less than 20 monosaccharides, linked together. The example of oligosaccharide is maltotriose. When the oligosaccharide eventually exceeds 20 monosaccharides, it is called a polysaccharide or glycan. Additionally, polysaccharides can be subdivided into two types: (1) homopolysaccharide, which only contains a single type of monosaccharides, such as starch, dextran, cellulose, and glycogen made up of glucose, and (2) heteropolysaccharide, which contains two or more different monosaccharides. To make things a little bit more interesting, a polysaccharide can also be branched or unbranched; this goes for both homopolysaccharide and heteropolysaccharide. For instance, starch has two forms; it can be branched (i.e., amylopectin) or unbranched (i.e., amylose). Moreover, when looking into their functionality, polysaccharides can also be subdivided into two types: (1) storage polysaccharides (e.g., starch and glycogen) and (2) structural polysaccharides (e.g., cellulose, arabinoxylans, chitin, pectin).

4.1 Polysaccharide of human origin The polysaccharide derived from human includes glycosaminoglycans (GAGs). GAGs are classified into four groups: (1) heparin/heparan sulfate, (2) chondroitin sulfate/dermatan sulfate, (3) keratan sulfate, and (4) hyaluronic acid (HA). Table 26.3 has listed some of the research studies carried out by the researchers and their potential biomedical applications of natural polysaccharide derived from humans. In particular, keratan sulfate has not been paid for many attentions because no papers were found regarding this specific polysaccharide in electrospinning. Hence, it has the potential to be electrospun in future. 4.1.1 Hyaluronic acid 4.1.1.1 Chemical structure and applications of hyaluronic acid HA is a natural polysaccharide found in human tissues such as the skin, synovial fluids of joints, and connective tissues. It is composed of repeated disaccharide units with the alternation of D-glucuronic acid and N-acetyl-D-glucosamine (Fig. 26.1). Each monosaccharide is connected by alternating b-(1 / 4) and b-(1 / 3) glycosidic bonds. Its chains consist up to 30,000 repeating units with a high molecular weight range (1000e10,000,000 Da). HA is one of the most hydrophilic molecules because this large sugar molecule is capable to hold 500 times its own weight of water. HA is also known as a natural moisturizer. Hence, it is responsible for tissue hydration and lubrication.

Electrospun natural polysaccharide for biomedical application 595 Table 26.3: The researches of natural polysaccharide from human origin and their potential applications. Natural polysaccharide from human origin

Additive polymer

Heparin

Gelatin PLLAa and PA-6b PLLACLd PVAf/gelatin

Chondroitin sulfate

Gelatin PVA Silk fibroin

Hyaluronic acid

PCLh PEOi Type I porcine collagen and gelatin

Solvent Distilled water and acetic acid HFPc TFEe and distilled water Distilled water and acetic acid TFEg and distilled water Distilled water Formic acid and distilled water Chloroform and formic acid Acetic acid NaOHj, DMFk, and acetic acid

Potential applications Vascular tissue engineering Achilles tendon regeneration Blood vessel tissue engineering Tissue engineering Skin tissue engineering Tissue engineering Scaffolding and drug release Skin tissue engineering scaffolds Tissue regeneration Skin tissue engineering

References [15] [16] [17] [18] [19] [20] [21] [22]

[23] [24]

a

Polylactic acid. Polyamide-6. c Hexafluoro isopropanol. d Poly(L-lactide-co-ε-caprolactone). e 2,2,2-Trifluoroethanol. f Poly(vinyl alcohol). g Trifluoroethanol. h Polycaprolactone. i Polyethylene Oxide. j Sodium hydroxide. k Dimethylformamide. b

In the field of skin care, it is a powerful moisturizer that offers smoothness and softening effects to the skin. Furthermore, it can reduce the appearance of wrinkles. 4.1.1.2 Research in electrospun hyaluronic acid nanofibers

Owing to the low mechanical strength of HA, HA is suggested to be either chemically modified or chemically cross-linked or blending with another polymer. Dogan et al. [21] have fabricated the uniform and bead-free coaxial nanofibers with SF as the shell, while HA and olive leaf extract (OLE) as the core. The blend of 15% (w/v) SF, 0.5% (w/v) HA, and 12% (w/v) OLE nanofibers has the diameter of 468.19
161.51 nm. When the concentration of OLE increased to 15% (w/v), the diameter of nanofibers increased to 543.24
196.09 nm. It is explained that a higher voltage supply is needed to fabricate the nanofibers with the addition of OLE into SF/HA solution, resulting in an increase in

596 Chapter 26 D-gluc uronic acid

β-(1 3)

6

glycosidic bond b 5

6

4 D-glucuronic accid

1 3

1

4

5

3

2

2

6 5

6 5 1

4 3

1

4

2

3

N-acetyl-D- glucosamine

2

n β-(1

4)

glyco sidic bond N N-acetyl-D-glu cosamine

Hyaluroonic acid

Figure 26.1 Chemical structure of hyaluronic acid.

average nanofibers diameter. The OLE is used because it has the antibacterial and antifungal properties that expose their potential to be used as novel products for scaffolding and drug release applications [21]. In addition, Wang et al. [22] have generated the coreeshell polycaprolactone (PCL)/HA/ epidermal growth factor (EGF) nanofibrous scaffolds for the use in wound healing using emulsion electrospinning. The PCL/HA/EGF nanofibers have the smaller diameter (149
4.5 nm and a pore size 0.17
0.03 mm2) when compared with the PCL nanofibers (272
38 nm and a pore size of 0.56
0.19 mm2) and PCL/HA nanofibers (184
6 nm and a pore size of 0.16
0.02 mm2) [22]. As the viscosity of HA is very high, it is usually blended with other polymers for electrospinning. Furthermore, there are many studies that used HA for electrospinning such as fabricating the PEO/HA coreeshell electrospun nanofibers for tissue engineering scaffolds [23] and collagen/HA nanofibers for chronic wound healing [24].

4.2 Polysaccharide of plant and seaweed origin The polysaccharide derived from plant includes cellulose, dextrin, and its derivatives, amylose, xylan, starch, dextrin, guar gum, locust bean gum, inulin, and pectin, whereas the polysaccharide derived from seaweed includes agarose and carrageenan extracted from red algae, alginate (alginic acid) found in the cell walls of brown algae, fucoidan extracted from brown algae, porphyrin extracted from red algae, and ulvan extracted from green algae. Table 26.4 has listed out some of the research activities performed by the researchers. However, locust bean gum, porphyrin, and psyllium are not illustrated in the table because there are no studies that demonstrate electrospinning of these three types of polysaccharide. Therefore, they have an extensive interesting area that can be developed using locust bean gum, porphyrin, and psyllium in the future.

Electrospun natural polysaccharide for biomedical application 597 Table 26.4: The researches of natural polysaccharide from plant origin and their potential applications. Natural polysaccharide from plant and seaweed origin

Additive polymer

Pectin

PVA

Distilled water

Inulin

PEO PVA

Distilled water Distilled water

Guar gum

PVA

Purified water and nonalkaline pH or alkaline pH Deionized water and distilled water Distilled water Distilled water and chloroform Acetic acid and TFAa

Agarose Alginate

PVA PLA

Dextrin

Chitosan

Amylose Xylan

e PVA

Ulvan

PCL or PEO

PVA or PEO Carrageenan

Fucoidan Cellulose and its derivatives a

PCL PHBd and PHBVe Chitosan and PVA PVA

Trifluoroacetic acid. Dimethyl sulfoxide. c Dichloromethane. d Polyhydroxybutyrate. e Polyhydroxybutyrate valerate. f 1,1,1,3,3,3-Hexafluoro-2-propanol. b

Solvent

DMSOb NaOH and distilled water DMF and DCMc or distilled water

Denoised water, acetic acid DCM HFIPf and CHCl3 Deionized water and acetic acid Deionized water

Potential applications Skin regeneration or drug carriers Tissue engineering The treatment of digestive disorders, antiseptic sprays, or bandages’ fillers for wound infections, and many different types of bacterial infections Biodegradable wound dressing

References [25] [26] [27]

[28]

Drug delivery

[29]

Biomaterials Tissue engineering

[30] [31]

Antibacterial biomaterials Biomedical products Cardiac tissue engineering Tissue engineering scaffolds, wound dressings, or drug delivery systems Drug delivery systems

[32]

Tissue engineering Bone tissue engineering Vascular tissue engineering Bone tissue engineering

[37] [38]

[33] [34] [35]

[36]

[39] [40]

598 Chapter 26 4.2.1 Guar gum 4.2.1.1 Chemical structure and applications of guar gum Guar gum is a polysaccharide belonging to the group of galactomannans, being extracted from the endosperms of seeds of Cyamopsis tetragonolobus (Leguminosae family) [41]. It consists of linear chains of b-(1e4)-D-mannose units (backbone) with side units of a-(1e6) linked galactose (Fig. 26.2). The mannose backbone of guar gum with galactose side groups are suitable for human colon enzyme degradation, which can be developed to encapsulate a variety of bioactive drug ingredients for targeted drug delivery material [29]. For example, guar gum can be used as the colon-specific drug delivery application because of its resistance to human digestion and absorption, as well as susceptibility to microbial degradation in the large intestine. Its drug release retarding behaviors in colon, guar gum is mainly used for the

6

α-(1-6) link ed galactose s ide chain

5 4

1 2 3 6 3

5 4

1

4

1

β-(1-4)- D-mannosn ackbone

2

5 3

2 6

n

Figure 26.2 Chemical structure of guar gum.

4-linked 3,6-anhydroα-D-galactopyranose

–

O3SO 4

6

OH

5

O

O 3

6

1 2

HO

O

5

O 4

1

O 3

2

O

HO

3-linked β-D-galactopyranose

Figure 26.3 Chemical structure of k-carrageenan.

n

Electrospun natural polysaccharide for biomedical application 599 delivery of drug for the treatment of diseases associated with the colon which reduces the side effects and also the dose needed to reduce the dosing frequency [41e43]. In addition, guar gum is not only widely used in biomedical applications but also employed as food additives in food industries. 4.2.1.2 Research activities in electrospun guar gum nanofibers

Lubambo et al. [28] have investigated the guar gum/PVA electrospun membranes for better encapsulation of paramagnetic iron oxide (Fe3O4) nanoparticles in alkaline and nonalkaline condition. It is believed that the dispersion of Fe3O4 on the membranes can act as the bactericidal agent for the potential used as a biodegradable wound dressing. In nonalkaline condition, the nanofibers diameter of guar gum/PVA nanofibers ranges from 200 to 250 nm, whereas the diameter of the nanofibers decreased to 190 nm in an alkaline condition. It is because of the deprotonation of hydroxyl groups and the chelation of Fe2þ and Fe3þ ions which bring the molecules in the nanofibers closer together and resulted in the decrease in nanofibers diameter [28]. Shi et al. [29] have successfully fabricated the electrospun nanofibers membrane using guar gum, PVA, and citric acid. These nanofibers were proved to be insoluble in water by cross-linking them at high temperature (140 C) for 2 h after electrospinning. Hence, they have the potential to be used for drug delivery application and tissue engineering. The average diameter of the nonecross-linked guar/ PVA/citric acid nanofibers is between 194
23 nm and there is a slight increase in diameter after cross-linked to 204
18 nm [29]. 4.2.2 Carrageenan 4.2.2.1 Chemical structure and applications of carrageenan Carrageenan is a family of linear sulfated polysaccharides that are obtained by alkaline extraction from some species of red seaweeds (e.g., Gigartinales, Rhodophyta) [44]. There are three major types of carrageenan, which include iota-carrageenan (i-carrageenan), kappa-carrageenan (k-carrageenan), and lambda-carrageenan (l-carrageenan). Carrageenan is formed by disaccharide repeating unit which consists of alternating 3-linked b-D-galactopyranose or 4-linked a-D-galactopyranose or 4-linked 3,6-anhydro-a-D-galactopyranose (Fig. 26.3). In the food industry, carrageenans are used as thickener, food stabilizer, and gelling agents. Furthermore, over time, carrageenan as a microbiocidal compound was formulated in the form of wound dressings for successful wound healing [45]. 4.2.2.2 Research activities in electrospun carrageenan nanofibers

Basilia et al. [37] have fabricated electrospun PCL/i-carrageenan nanofibrous scaffolds for in vitro screening using simulated body fluids and in vivo screening using mice for tissue regeneration. Both screenings showed increased bioactivity and no invoke adverse

600 Chapter 26 inflammation, which proven to accelerate tissue healing. The nanofibers were not uniform, and it is reported to be influenced by the concentration of carrageenan and the glass transition temperature of the blend solution. The hydrogen bonded complex and intermolecular coadhesion might be the reason for having unstable electrospinning [37]. Furthermore, Goono et al. [38] reported the research in blending k-carrageenan with polyhydroxybutyrate (PHB) and polyhydroxybutyrate valerate (PHBV) to fabricate electrospun microfibers as scaffold materials in bone tissue engineering applications. Because of the higher molar weight of PHB exhibiting quicker jet transformation, the PHBV/k-carrageenan solution fabricated rough surface nanofibers, whereas PHB/kcarrageenan formed smooth surface nanofibers. The diameter of both fibers was at the range of 1.9
0.6 mm and 1.6
0.5 mm. k-Carrageenan was selected as it induced the higher production of the antiinflammatory interleukin-10 (IL-10) when compared with lcarrageenan. The results showed higher NIH/3T3 cell density and also determined an enhancement of the biomineralization using human osteosarcoma SaOS-2 cells, as well as showed osteogenic differentiation potential in the blend nanofibers, especially on PHBV/kcarrageenan nanofibers [38].

4.3 Polysaccharide of animal origin Apart from natural polysaccharide found in human, plants, and seaweed, polysaccharides can be obtained from animals. It includes (1) the storage polysaccharide, glycogen, which is normally found in the liver or muscle of animals and (2) the structural polysaccharide, chitin and its derivatives (e.g., chitosan), which is found in the exoskeleton of arthropods and the cell wall of fungi. Table 26.5 illustrates the studies of natural polysaccharide Table 26.5: The researches of natural polysaccharide from animal origin and their potential applications. Natural polysaccharide from animal origin Chitin

Chitosan

Additive polymer e

Gelatin PVA e

Solvent 1-Ethyl-3methylimidazolium [C2C1Im]þ and 1,3diethylimidazolium [C2C2Im]þ HFIP

HFIP TFA and DCM Acetic acid and distilled water TFA and DCM

Potential applications

References

Wound care

[46]

Wound healing and regeneration of oral mucosa and skin Wound dressing Skin tissue engineering Tissue engineering

[47]

Bone tissue engineering

[50]

[13] [48] [49]

Electrospun natural polysaccharide for biomedical application 601 derived from animals by the researchers and their potential applications. Unfortunately, there were no studies about the fabrication of the glycogen nanofibers using electrospinning found in the literature; an only similar study was found to use glycogen from oyster to form nanofibers using freeze-drying method [51]. Hence, it is a potential field to investigate in the future. In this chapter, chitin and its derivatives derived from shell animals are mainly discussed. 4.3.1 Chitin 4.3.1.1 Chemical structure and applications of chitin Chitin is a structural monosaccharide mainly found in the shell of arthropods such as insects, lobster, prawn, shrimp, and crab. It is the second most abundant of nature biopolymers after cellulose. Chitin is composed of a linear copolymer chain of N-acetyl-Dglucosamine residues with peptidoglycan b-1e4 linkage (Fig. 26.4A). Its physiochemical properties include odorless, white, or creamy-white powder. Chitin is generally used in food, cosmetics, biomedical, and pharmaceutical products. However, chitin has received little industrial-based attention because of its poor solubility in water and common organic solvents which resulted in a complicated network formation consisting of both inter- and intramolecular hydrogen bonding. Therefore, the deacetylation product of chitin called chitosan has been given an extensive interest to use in biomedical applications [47]. Because of its distinctive biological property of wound healing effect, chitin and its derivatives are widely used for the medical applications, specifically for wound management and tissue engineering. 4.3.1.2 Research activities in electrospun chitin nanofibers

Noh et al. [47] reported that relatively high human keratinocytes and fibroblasts cells attachment, migration, and proliferation were observed on electrospun chitin nanofibers. Owing to chitin’s poor solubility, halogenated compounds, hexafluoroisopropanol, were used to dissolve chitin. Other than that, strong acids such as methanesulfonic acid or mixed solvents such as lithium chloride/dimethylacetamide can be used to dissolve chitin [46]. The result showed that the chitin nanofibers have an average diameter of 163 nm. Additionally, the nanofibers coated with collagen significantly stimulated the cellular response. The overall results have indicated that the chitin nanofibers could be potential candidates for wound healing and oral mucosa and skin regeneration [47]. Another research activity reported by Min et al. [13] showed that the average diameter of chitin nanofibers is 110 nm. The study reported that certain chain entanglement in the chitin solution is essential to produce uniform nanofibers. The concentration of chitin solution higher or lower than the threshold can induce the formation of beaded nanofibers. The author also successfully performed the deacetylation of chitin

602 Chapter 26 N-acetyl group

(A)

N-acetyl-D-glucosamine

O

CH2OH 6 5

NH O

O

3

4 OH

2

4 OH

1

1

2

O

5

3

6

NH

O

CH2OH

O n

Alkali (hot, 40%-50% concentrated NaOH) Chitin-deacetylase (enzymatic deacetylation)

(B)

β-1.4-D-glucosamine

6

Amino group

CH2OH

5

NH2 O

O

3

4 1 OH

4

2 1

OH

2 5

3 NH2

O

O

6 CH2OH n

Figure 26.4 Chemical structure of (A) chitin and (B) chitosan.

nanofibers to chitosan nanofibers by chemically treated with a 40% aqueous NaOH solution at 60 C and 100 C. At high temperature (100 C), 85% of deacetylation completed within 2 h, while it took 1 day to complete 85% acetylation at low temperature (60 C). Both chitin and chitosan nanofibers can be used as wound dressings [13].

Electrospun natural polysaccharide for biomedical application 603 4.3.2 Chitosan 4.3.2.1 Chemical structure and applications of chitosan Chitosan is a modified natural polysaccharide produced by partial deacetylation of chitin under a hot alkali condition with 40%e50% concentrated sodium hydroxide (NaOH) solution or via enzymatic conversion of N-acetyl-D-glucosamine residue of chitin into D-glucosamine residues of chitosan in the presence of chitin deacetylase (Fig. 26.4B). Chitosan is obtained by eliminating enough number of acetyl groups, CH3eCO, and finally getting a linear polysaccharide consisting of randomly distributed b-1,4-Dglucosamine (deacetylated units) and N-acetyl-D-glucosamine (acetylated units). Basically, the actual difference between chitin and chitosan is the degree of acetylation (the ratio of N-acetyl-D-glucosamine to D-glucosamine structural units). The amount of N-acetyl-D-glucosamine greater than 50% is represented as chitin, whereas D-glucosamine level greater than 50% is represented as chitosan [52]. Chitosan is extensively used for bone tissue engineering because of its structural similarity to the GAG in bone. In addition, its low toxicity and biodegradability with excellent biological activities such as good immunological, intrinsic antibacterial activity, and low immunogenicity have provided many future perspective in development of biomaterials for wound dressing, drug delivery systems, and cell culture [45,53]. In detail, the deacetylated chitin derivative, chitosan, is a cationic polysaccharide because it possesses positive ionic charges from its free amino group. So, this has given chitosan the opportunity to chemically bind strongly with negatively charged cell surface, making it useful to formulate bioadhesive dosage forms. Also, chitosan is able to bind strongly to anions on the bacterial cell wall and subsequently altered the mass transport across the cell wall, resulting in the suppression of biosynthesis and accelerating bacterial death [49,54]. Furthermore, it is hard to electrospin chitosan because the cationic charge increases the excessive surface tension of the solution, which resulted in a high electrical force demand to fabricate nanofibers. Therefore, another polymer is mixed with chitosan to overcome the problem. 4.3.2.2 Research activities in electrospun chitosan nanofibers

Agrawal and Pramanik [49] reported a study related to the investigation of chitosan/PVA nanofibers for tissue engineering. A series of chitosan/PVA blend ratios in 10:90, 20:80, 30:70, 35:65, and 40:60 were tested, and to conclude all the analysis of experiment, it was proved that the 35:65 ratio of chitosan/PVA nanofibrous scaffold had the finest nanofibers in the average diameter of 260 nm with superior physicochemical analysis and in vitro study (i.e., high efficient for hMSCs proliferation). By figuring out the suitable ratio between chitosan and PVA, it is possible to get the appropriate morphology of nanofibers [49] Another example of using chitosan for electrospinning is coming from the work of Venugopal et al. [50] for bone tissue regeneration. They have prepared the

604 Chapter 26 electrospun chitosan/hydroxyapatite nanofibrous scaffold with the average diameter of the chitosan/hydroxyapatite nanofibers around 510
198 nm. Besides, it is also proved to show that these nanofibers were able to promote cell response and proliferation of human fetal osteoblast, hFOB cells, which further enhance the bone-forming ability [50,55]. A similar study has been conducted by Zhang and colleagues [56] where they fabricated chitosan/hydroxyapatite nanofibrous scaffolds together with ultra-high molecular weight poly(ethylene oxide) as a fiber-forming facilitating additive. Uniform nanofibers with a diameter of 214
25 nm were determined. Also, a significant level of bone cell formation ability was demonstrated at Day 15 with an increased level of hFOB cells proliferation [55,56].

4.4 Polysaccharide of microbe origin Microbial polysaccharide is also called exopolysaccharide as the polysaccharide is either obtained from the cell wall or secreted from the cell to form a layer over the surface of the microorganism such as bacteria, fungi, and yeast. Because of the absence of virtually no known toxic agents and very cheap to harvest in large quantities, these extracellularly produced natural polysaccharides by microorganisms are in food, pharmaceutical, and biomedical industries [57]. Microbial polysaccharides can be produced from either fungi or bacterium, including schizophyllan from the fungi Schizophyllan commune, pullulan from the fungus Aureobasidium pullulans, alginates from the bacterium Azotobacter vinelandii, xanthan gum from the bacterium Xanthomonas campestris, dextran from the bacterium Leuconostoc mesenteroides, gellan from the bacterium Pseudomonas elodea, and curdlan from the bacterium Alcaligenes faecalis. Table 26.6 illustrates the research activities of the natural polysaccharide from the microbe origin mentioned above. In this chapter, xanthan gum, pullulan, and dextran as the representative examples of microbial polysaccharides are mainly discussed. 4.4.1 Xanthan gum 4.4.1.1 Chemical structure and applications of xanthan gum Xanthan gum is a hetero-, branched polysaccharide formed during the fermentation process of the bacterium X. campestris originally isolated from the Rutabaga plant [80]. It has a chemical structure of five sugar repeating units in the molar ratio of 2:2:1 with two D-glucose units, two D-mannose units, and one D-glucuronic acid unit (Fig. 26.5). The backbone consists of b-1,4-D-glucose repeating units with additional sidechains of trisaccharide at every other glucose units at C-3. The sidechain is built up of (1,4)-b-Dglucuronic acid unit sandwiched between a-1,2-D-mannose with an acetyl group attached to C-6 and b-D-mannose [2]. The terminal mannose residues are usually pyruvated, and the

Electrospun natural polysaccharide for biomedical application 605 Table 26.6: The researches of natural polysaccharide from microbe origin and their potential applications. Natural polysaccharide from microbe origin

Additive polymer

Schizophyllan

PVA

Pullulan

e

DMSO, formic acid, and deionized water Redistilled water Distilled water Deionized water

Gelatin WPIa

Distilled water Distilled water

APIb

Formic acid and Tween 80 Distilled water Deionized water

Alginates

Xanthan gum Dextran

Gellan

Curdlan

PVA Chitosan and PEO PEO PEO

Whey protein isolate. Amaranth protein isolate. c Polyethylene glycol. d Polyurethane. e Polyvinylpyrrolidone. b

Deionized water Distilled water

PEO or PEGc e Chitosan PUd PU

Formic acid Formic acid THF and DMF DMSO and THF

PVPe PVA PVA

Distilled water Distilled water Deionized water

PCL

Chloroform, methanol and NaOH

PVA

Distilled water and formic acid Deionized water

PEO a

Solvent

Deionized water

Potential applications

References

Wound healing

[58]

Drug delivery, Bandages Water-resistant biomaterials Adsorption, separation, biomedical and tissue engineering Tissue engineering scaffold Bioactive compounds encapsulation matrices Drug delivery

[59] [60] [61]

Anti-ultraviolet packaging Tissue engineering scaffolds

[65] [66]

Tissue engineering Wound healing, regenerative medicine, and drug delivery systems Regenerative medicine and drug delivery applications Drug delivery Drug delivery Wound dressing Postmenopausal wound dressing Drug delivery Drug delivery Skin tissue regeneration, drug delivery, and regenerative medicine applications Nucleus Pulposus regeneration at intervertebral disc Wound healing

[67] [68]

Wound dressing and drug delivery

[62] [63] [64]

[69] [14,70] [53] [71] [72] [73] [74] [75,76]

[77]

[78] [79]

606 Chapter 26 OH

OH 6 Repeating β –1,4D-glucose units

5 O O

4

OH 1 3 2 OH O 6

H3C

6 5 O

4

O

3

H 3C

4 1 OH OH O 3 2

O

OH 1 3 2

4

O

3

O 6

1 2 OH n

OH

H 3C

O

β -D- mannose linked to pyruvate

5 O

5 O 4

1 2

O

C

6

OH

5 O α –1, 2 -D-mannose 4 OH 1 with acetyl group OH 3 COO– 2 6 O O 5 β –1, 4-D-gluc 4 1 glucuronic acid OH 2 O 3 OH O 6 COO– 5 O

OH

OH

6

O

O

HO 6

5 O 4 OH 1 OH 3 COO– 2 6 O O 5 4 1 OH 2 O 3 OH

5 O 4 1 OH OH OH 3 2

β -D- mannose

Figure 26.5 Chemical structure of xanthan gum, which consists of five sugar repeating units.

nonterminal residue may have an acetyl group at C-6 [81]. Xanthan gum is a highly electronegative or “polyanionic” molecule because of the presence of anionic sidechains (i.e., pyruvate and acetate). Xanthan gum is an ideal agent as an emulsifier, stabilizer, and thickener for many types of water-based products ranging from food, pharmaceutical, cosmetic, cleaner, agricultural, textile, coating, ceramic, and oil fields. However, in the biomedical point of view, xanthan gum in combination with other hydrophilic or hydrophobic polymers can form a more stable gel system [2,80,81]. 4.4.1.2 Research activities in electrospun xanthan gum nanofibers

There is a new breakthrough in fabricating xanthan gum nanofibers using electrospinning. Shekarforoush and his research team [14] have fabricated the xanthan gum nanofibers without using any additive polymers. By using formic acid as the solvent to dissolve xanthan gum, it is possible to obtain the ultrafine nanofibers with the average diameters ranging from 128
36.7 to 240
80.7 nm at the concentration of 1.0%e2.5% w/w. Owing to the unstable molecular conformation of xanthan solution in aqueous solutions, formic acid is discovered to have the ability to stabilize the helical conformation of xanthan by neutralizing the pyruvic charges in xanthan solution with formate groups in formic acid. The xanthan gum nanofibers have the potential to be used as a carrier for the encapsulation of bioactive compounds in drug delivery applications [14,70].

Electrospun natural polysaccharide for biomedical application 607 In addition, the xanthan gum has been successfully electrospun with a cationic natural polysaccharide, chitosan, as drug delivery carrier by Shekarforoush et al. [53]. The hydrophobic bioactive compound, curcumin, is encapsulated in the xanthan gum/ chitosan (X-Ch) nanofibers to overcome its low solubility and instability in a body fluid and to study its drug release in different pH buffers (i.e., pH 2.2, 6.5, and 7.6). The diameter of X-Ch nanofibers (750
250 nm) increased approximately 160 nm with the addition of curcumin in xanthan solution (910
440 nm). Moreover, the study showed a good curcumin encapsulation efficiency of 69.4
4.1% in X-Ch nanofibers with no burst release effect in all three pH buffers. There is a low curcumin release of 20% in the acidic condition, pH 2.2 after 5 Days. However, the amount of curcumin released from the X-Ch nanofibers was increased as the pH value increased, and 45% and 50% of curcumin was released in pH 6.5 and pH 7.6, respectively. Hence, it is suggested that these X-Ch nanofibers encapsulated curcumin is suitable for use as a long-term pH-stimulated drug release carrier [53]. 4.4.2 Pullulan 4.4.2.1 Chemical structure and applications of pullulan Pullulan is extracted from the fungus-like yeast, A. pullulans [59]. As shown in Fig. 26.6, it is formed by repeating maltotriose units. A maltotriose unit consists of three glucose units (i.e., a-1,4-glucan and a-1,6-glucan) connected by a-1,4 glycosidic bonds. In addition, the consecutive maltotriose units are bonded to each other via an a-1,6 glycosidic bond [45,82]. The consistent alternation of a-1,4- and a-1,6 glycosidic linkages gives rise to two unique properties of structural flexibility and improved solubility, which have huge benefits to the

OH

O 6

6 CH2 5 Maltotriose 4

OH

6

5

O

3

OH

1

4

OH 3

2

OH

5

O 1 2

O

OH α-1,6-glucan

4

O OH 3

O OH α-1,4-glucan

α-1,4 glycosidic bond

1 2 O

OH α-1,4-glucan n

α-1,4 glycosidic bond

C H2

α-1,6 glycosidic bond

Figure 26.6 Chemical structure of pullulan, which consists of three sugar repeating units.

608 Chapter 26 pharmaceutical industries [83]. Pullulan is potentially used as pharmaceutical coatings for tablets, pills, and granules because of its high water solubility and low moisture resistance. It also attracts great interest for the uses as predosed formulations wrapping materials for soft and hard capsule because it is colorless, transparent, and biodegradable [84]. Moreover, the nonimmunogenic, nonmutagenic, and noncarcinogenic nature of pullulan has been discovered as vaccine adjuvant in which was conjugated to various toxoids and showed very promising immunogenic results. A nasal vaccination research study conducted by Cevher et al. is using pullulan incorporated with chitosan derivatives to form a nanocomposite for the encapsulation of a model antigen, bovine serum albumin. The result showed an efficient nanoparticle uptake by macrophage [85,86]. Owing to its nontoxic, edible, and resistance to mammalian amylases characteristics, pullulan is also widely used in food industry as a low-calorie ingredient in foods. In addition, pullulan has a relatively low viscosity as compared with other natural polysaccharides; it is suitable to be used as low-viscosity filler in beverages and sauces [87]. 4.4.2.2 Research activities in electrospun pullulan nanofibers

The electrospun pullulan nanofibers have been successfully fabricated by Sun et al. [59]. In the study, by varying the pullulan solution concentration, applied voltage, flow rate, and needleecollector plate distance to 22 %w/w solution concentration, 31 kV voltage, 0.5 mL/ h flow rate, and 20 cm needleecollector plate distance during electrospinning, the finest and more uniform diameter of nanofibers can be produced between 100 and 700 nm. Additionally, the diameter of pullulan nanofibers can be reduced by decreasing the concentration of pullulan solution and flow rate while increasing the applied voltage and needleecollector plate distance. It is proposed that these pullulan nanofibers are potential to be used to manufacture bandages or act as drug carriers [59]. In addition, Qian and King [62] have investigated the gelatin/pullulan electrospun nanofibers to serve as a tissue engineering scaffold, particularly to mimic the extracellular matrix. The result showed an average nanofibers diameter of 152 nm. It was also discovered that not only the concentration and viscosity of the spinning solution but also the weight ratio of gelatin and pullulan influence the morphology and diameter of the nanofibers [62]. Besides, Li et al. [82] have fabricated the electrospun pullulan nanofiber membrane that has the potential utilized as a water-resistant biomaterial. The prepared pullulan nanofibers were cross-linked by ethylene glycol diglycidyl ether and ethanol absolute (1:7 ratio) to enhance its stability in water with lower water absorption. It is crucial to manage the cross-linking time. As the cross-linking time increased, the porosity of the nanofibers decreased [60]. Moreover, several types of pullulan nanofibers were prepared using electrospinning technique which includes pullulan-whey protein electrospun nanofibers [63], and curcumin-loaded amaranth-pullulan electrospun nanofibers [64] have a huge potential for biomedical applications.

Electrospun natural polysaccharide for biomedical application 609 4.4.3 Dextran 4.4.3.1 Chemical structure and applications of dextran Dextran is a natural polysaccharide that sucrose is extracted from specific lactic acid bacteria such as L. mesenteroides and S. mutans. It has a complex branched glucan structure as seen in Fig. 26.7. Its main chain composed of glucose monomers linked via a-1,6 glycosidic bond with branches from a-(1e2), a-(1e3), or a-(1e4) linkages. The biocompatibility and biodegradability characteristics of dextran nanofibers are very important properties in biomedical applications for the use as drug carriers, plasma expander, and tissue scaffolds [88]. Furthermore, dextran is soluble in both water and organic solvents including tetrahydrofuran, dimethyl sulfoxide, and acetone. Owing to its unique solubility characteristic, dextran can be used to prepare different types of formulations by blending with hydrophilic bioactive agents (e.g., penicillin) or hydrophobic bioactive agents (e.g., curcumin) or hydrophilic biodegradable polymers (e.g., PVA) or hydrophobic biodegradable polymers (e.g., poly(lactic-co-glycolic acid)). O 6 CH2 O 5 4 OH 1 3 2 OH O OH CH 6 2

6 CH2OH 5

O

α-1,6 glycosidic bond

O 5 OH α-1,6 4 OH 1 3 2 glycosidic bond 3 2 OH O O OH OH OH CH2 6 O CH 6 2 5 1 4 OH 5 3 2 4 OH O 3 OH α-1,3 OH branching

4

1

α-1,6 glycosidic bond

O 1 2

α-1,6 glycosidic bond

O OH CH 6 2 5

O

4 OH 1 3 2 OH OH

O n

Figure 26.7 Chemical structure of dextran, which consists of six sugar repeating units.

610 Chapter 26 The hydrophilic polymers such as dextran are said to be soft and flexible in a swollen state with high affinity to cells, yet owing to its low mechanical strength, it needs a copolymer like hydrophobic polymers, which have the high mechanical strength to assist them to improve the overall properties of electrospun nanofibers [89]. In addition, electrospinning has an advantage over other types of technology in allowing the use of blend polymers for nanofibers production. 4.4.3.2 Research activities in electrospun dextran nanofibers

Dextran encourages neovascularization and re-epithelialization in chronic wounds. It is a suitable natural polysaccharide used as wound dressings. Unnithan et al. [71] have conducted a study about developing polyurethane (PU)/dextran nanofibers loaded with a fluoroquinolone antibiotic, ciprofloxacin HCl, for postmenopausal wound dressing. Ciprofloxacin HCl is chosen as the model drug because of its low minimal inhibitory concentration for both Gram-positive bacteria such as Staphylococcus aureus and Bacillus subtilis and Gram-negative bacteria such as Escherichia coli, Salmonella typhimurium, and Vibrio vulnificus, which normally cause wound infections. The result presented a good antibacterial activity with the zone of inhibition around 15e20 mm for both types of bacteria. The diameter of the PU nanofibers also dramatically decreased from 401e100 nm to 101e300 nm with the addition of dextran and ciprofloxacin HCl [71]. Again in 2015, Unnithan led another research team to continue investigating the electrospun PU/dextran nanofibers for wound dressing mainly to treat the postmenopausal wounds. Thus, an estrogen, estradiol with potent antiinflammatory and good blood clotting properties, was loaded in the PU/dextran solution for electrospinning to accelerate the wound healing process. It was further proven in the study that the re-epithelialization property of dextran with the antiinflammatory property of estradiol has shortened the healing time with animal testing. The result also showed the continuous uniform nanofibers with a diameter range from 500 to 600 nm when blending the PU, dextran, and estradiol together [72]. Apart from using dextran for wound dressing materials, it is also potentially used in the drug delivery system. A study about oral fast-dissolving drug delivery was conducted by Maslakci et al. [73] by loading two different drugs, ibuprofen and acetylsalicylic acid, separately into the electrospun polyvinylpyrrolidone (PVP)/dextran nanofiber mats. The diameter of the PVP/dextran and PVP/acetylsalicylic acid nanofibers was not stable with beaded morphology, which is mainly due to the viscosity of the solutions. However, by adding the drugs into the PVP/dextran solutions, the PVP/dextran/ibuprofen and PVP/ dextran/acetylsalicylic acid nanofibers showed the diameter range from 300 to 650 nm using scanning electron microscopy with a uniform diameter and ultrafine surface. Therefore, to optimize the morphology, the ratio of PVP, dextran, ibuprofen, and

Electrospun natural polysaccharide for biomedical application 611 acetylsalicylic acid is highly modulated [73]. In another study, Moydeen et al. [74] reported the fabrication of electrospun PVA/dextran nanofibers loaded with ciprofloxacin and ciprofloxacin HCl drugs separately for in vitro drug release. The coaxial electrospinning and emulsion electrospinning were employed, and the transmission electron microscopy result indicated that uniform coreeshell nanofibers were observed by using the emulsion electrospinning. Owing to the uniformity, it showed a slow-release kinetics compared with the blended PVA/dextran/ciprofloxacin using conventional coaxial electrospinning. Furthermore, by introducing the drug into the PVA/ dextran solution, the diameter of the nanofibers has decreased from 400e600 nm to 200e300 nm, which further proves the flawless load of ciprofloxacin into the core [74].

5. Conclusion As detailed in this chapter, electrospun natural polysaccharide nanofibers derived from human, animals, plants, seaweed, and microbes have promising biomedical applications in different fields such as drug delivery, wound dressing, and tissue engineering. Some of the natural polysaccharides such as curdlan have proven to be very difficult for electrospinning [79]. Thus, different types of polysaccharide or polymer were chosen as a cospinning system to exhibit the formation of continuous nanofibers. In addition, the diameter of the nanofibers depends on solution concentration, applied voltage, surface tension of solution, spinning flow rate, etc. These nanofibers can be customized by modulating the electrospinning parameters. It is also important to draw some attention to the production of the natural polysaccharide nanofibers. There are a limited number of biomedical products that have commercially implemented electrospun nanofibers. Thus, there will be even less or no natural polysaccharideebased electrospun nanofibers. The most relevant commercialized nanofiber product that used natural polysaccharide is the “HealthSmart Personalized Wound Care System” from the provider PolyRemedy, Inc in 2013. The company has announced an electrospun wound dressing with the addition of HA [90]. As majority of these nanofibers are still in the research progress, a lot of efforts have been made by the researchers to transform the nanofibers from laboratory to commercial production. Other than that, the development of scale-up electrospun nanofibers production method is also a very important research area for the growth in the biomedical field. One approach for employing the electrospinning technology to the natural polysaccharide is to utilize the model systems or formulations that have already been established in other fields such as the food, textiles, and cosmetics sectors and adapting it to the requirements of the biomedical applications.

612 Chapter 26

References [1] Zanin M, Cerize NP, Oliveira A. Production of nanofibers by electrospinning technology: overview and application in cosmetics. Nanocosmet Nanomed 2011:311e32. [2] Brunchi C, Bercea M, Morariu S, Avadanei M. Investigations on the interactions between xanthan gum and poly ( vinyl alcohol ) in solid state and aqueous solutions. Eur Polym J 2016;84:161e72. [3] Liang D, Hsiao BS, Chu B. Functional electrospun nanofibrous scaffolds for biomedical applications. Adv Drug Deliv Rev 2007;59(14):1392e412. [4] Awan IZ, Hussain SB, ul Haq A, Khan AQ. Wondrous nanotechnology. J Chem Soc Pak 2016;38(6):1026e55. [5] Sapountzi E, Braiek M, Chateaux JF, Jaffrezic-Renault N, Lagarde F. Recent advances in electrospun nanofiber interfaces for biosensing devices. Sensors 2017;17(8):1887. [6] Zhang N, et al. Electrospun TiO2 nanofiber-based cell capture assay for detecting circulating tumor cells from colorectal and gastric cancer patients. Adv Mater 2012;24(20):2756e60. [7] Park JH, Lee S, Kim JH, Park K, Kim K, Kwon IC. Polymeric nanomedicine for cancer therapy. Prog Polym Sci 2008;33(1):113e37. [8] Chee BS, De Lima GG, Devine D, Nugent MJD. Electrospun hydrogels composites for bone tissue engineering. In: Applications of nanocomposite materials in orthopedics. Elsevier Inc.; 2018. p. 39e70. [9] De Lima GG, Lyons S, Devine DM, Nugent MJD. Electrospinning of hydrogels for biomedical applications. In: Hydrogels. Springer Singapore; 2018. p. 219e58. [10] Husain O, Lau W, Edirisinghe M, Parhizkar M. Investigating the particle to fibre transition threshold during electrohydrodynamic atomization of a polymer solution. Mater Sci Eng C 2016;65:240e50. [11] He HW, et al. Solvent-free electrospinning of UV curable polymer microfibers. RSC Adv 2016;6(35):29423e7. [12] Subbiah T, Bhat GS, Tock RW, Parameswaran S, Ramkumar SS. Electrospinning of nanofibers. J Appl Polym Sci 2005;96(2):557e69. [13] Min BM, et al. Chitin and chitosan nanofibers: electrospinning of chitin and deacetylation of chitin nanofibers. Polymer 2004;45(21):7137e42. [14] Shekarforoush E, Faralli A, Ndoni S, Mendes AC, Chronakis IS. Electrospinning of xanthan polysaccharide. Macromol Mater Eng 2017;302(8):1e11. [15] Wang H, Feng Y, Fang Z, Xiao R, Yuan W, Khan M. Fabrication and characterization of electrospun gelatin-heparin nanofibers as vascular tissue engineering. Macromol Res 2013;21(8):860e9. [16] Ye YJ, Zhou YQ, Jing ZY, Liu YY, Yin DC. Electrospun heparin-loaded coreeshell nanofiber sutures for achilles tendon regeneration in vivo. Macromol Biosci 2018;18(7):1800041. [17] Su Y, Li X, Liu Y, Su Q, Qiang MLW, Mo X. Encapsulation and controlled release of heparin from electrospun poly(L-lactide-co-ε-caprolactone) nanofibers. J Biomater Sci Polym Ed 2011;22(1e3):165e77. [18] Sadeghi A, Pezeshki-Modaress M, Zandi M. Electrospun polyvinyl alcohol/gelatin/chondroitin sulfate nanofibrous scaffold: fabrication and in vitro evaluation. Int J Biol Macromol 2018;114:1248e56. [19] Pezeshki-Modaress M, Mirzadeh H, Zandi M. Gelatin-GAG electrospun nanofibrous scaffold for skin tissue engineering: fabrication and modeling of process parameters. Mater Sci Eng C 2015;48:704e12. [20] Guo J, Zhou H, Akram MY, Mu X, Nie J, Ma G. Characterization and application of chondroitin sulfate/ polyvinyl alcohol nanofibres prepared by electrospinning. Carbohydr Polym 2016;143:239e45. [21] Dogan G, Bas‚al G, Bayraktar O, Ozyildiz F, Uzel A, Erdo gan I. Bioactive Sheath/Core nanofibers containing olive leaf extract. Microsc Res Tech 2016;79(1):38e49. [22] Wang Z, et al. Evaluation of emulsion electrospun polycaprolactone/hyaluronan/epidermal growth factor nanofibrous scaffolds for wound healing. J Biomater Appl 2015;30(6):686e98. [23] Chen G, Guo J, Nie J, Ma G. Preparation, characterization, and application of PEO/HA core shell nanofibers based on electric field induced phase separation during electrospinning. Polymer 2016;83:12e9.

Electrospun natural polysaccharide for biomedical application 613 [24] Lai HJ, et al. Tailor design of electrospun composite nanofibers with staged release of multiple angiogenic growth factors for chronic wound healing. Acta Biomater 2014;10(10):4156e66. [25] Patra N, Salerno M, Cernik M. Electrospun polyvinyl alcohol/pectin composite nanofibers. In: Electrospun Nanofibers. Elsevier Ltd., 2016. p. 599e608. [26] McCune D, et al. Electrospinning pectin-based nanofibers: a parametric and cross-linker study. Appl Nanosci 2018;8(1e2):33e40. [27] Walaa Mohamed Ali Wahbi EG, Wael Mamdouh Sayed Sayed Ahmed EG, Rania Siam EG. Inulin nanofibers. 2016. WO2016086225A1. [28] Lubambo AF, et al. Tuning Fe3O4 nanoparticle dispersion through pH in PVA/guar gum electrospun membranes. Carbohydr Polym 2015;134:775e83. [29] Shi J, Yang E. Post-electrospinning crosslinking of guar/polyvinyl alcohol membrane. Therm Sci 2016;20(1):1e5. [30] Sousa AMM, Souza HKS, Uknalis J, Liu SC, Gonc¸alves MP, Liu L. Electrospinning of agar/PVA aqueous solutions and its relation with rheological properties. Carbohydr Polym 2015;115:348e55. [31] Xu W, Shen R, Yan Y, Gao J. Preparation and characterization of electrospun alginate/PLA nanofibers as tissue engineering material by emulsion eletrospinning. J Mech Behav Biomed Mater 2017;65:428e38. [32] Burns NA, et al. Cyclodextrin facilitated electrospun chitosan nanofibers. RSC Adv 2015;5(10):7131e7. [33] Kong L, Ziegler GR. Fabrication of pure starch fibers by electrospinning. Food Hydrocolloids 2014;36:20e5. [34] Venugopal J, et al. Xylan polysaccharides fabricated into nanofibrous substrate for myocardial infarction. Mater Sci Eng C 2013;33(3):1325e31. [35] Kikionis S, Ioannou E, Toskas G, Roussis V. Electrospun biocomposite nanofibers of ulvan/PCL and ulvan/PEO. J Appl Polym Sci 2015;132(26):1e5. [36] Toskas G, Hund RD, Laourine E, Cherif C, Smyrniotopoulos V, Roussis V. Nanofibers based on polysaccharides from the green seaweed Ulva Rigida. Carbohydr Polym 2011;84(3):1093e102. [37] Basilia BA, Robles AP, Ledda KA, Dagbay KB, St M. In-vitro and in-vivo screenings of electrospun polycaprolactone-carrageenan nanofibrous scaffolds for tissue engineering. TechConnect Briefs 2008;2:306e9. [38] Goonoo N, et al. k-Carrageenan enhances the biomineralization and osteogenic differentiation of electrospun polyhydroxybutyrate and polyhydroxybutyrate valerate fibers. Biomacromolecules 2017;18(5):1563e73. [39] Zhang W, et al. Electrospinning of fucoidan/chitosan/poly(vinyl alcohol) scaffolds for vascular tissue engineering. Fibers Polym 2017;18(5):922e32. [40] Chahal S, Hussain FSJ, Yusoff MM. Characterization of modified cellulose (MC)/poly (vinyl alcohol) electrospun nanofibers for bone tissue engineering. Procedia Eng 2013;53:683e8. [41] Manjunath M, et al. Guar gum and its pharmaceutical and biomedical applications. Adv Sci Eng Med 2016;8(8):589e602. [42] Krishnaiah YSR, Satyanarayana S, Rama Prasad YV, Narasimha Rao S. Gamma scintigraphic studies on guar gum matrix tablets for colonic drug delivery in healthy human volunteers. J Control Release 1998;55(2e3):245e52. [43] Yang W, Zhang M, Li X, Jiang J, Sousa AMM, Zhao Q. Incorporation of tannic acid in food-grade guar gum fibrous mats by electrospinning technique. Polymers 2019;11(1):141. [44] Pereira L, Van De Velde F. Portuguese carrageenophytes: carrageenan composition and geographic distribution of eight species (Gigartinales, Rhodophyta). Carbohydr Polym 2011;84(1):614e23. [45] Dragostin OM, et al. Polymeric materials as nanofibers used in the treatment of burns. Rev. Med. Chir. Soc. Med. Nat. 2015;119(3):903e9. [46] Barber PS, Griggs CS, Bonner JR, Rogers RD. Electrospinning of chitin nanofibers directly from an ionic liquid extract of shrimp shells. Green Chem 2013;15(3):601e7. [47] Noh HK, et al. Electrospinning of chitin nanofibers: degradation behavior and cellular response to normal human keratinocytes and fibroblasts. Biomaterials 2006;27(21):3934e44.

614 Chapter 26 [48] Dhandayuthapani B, Krishnan UM, Sethuraman S. Fabrication and characterization of chitosan-gelatin blend nanofibers for skin tissue engineering. J Biomed Mater Res B Appl Biomater 2010;94(1):264e72. [49] Agrawal P, Pramanik K. Chitosan-poly(vinyl alcohol) nanofibers by free surface electrospinning for tissue engineering applications. Tissue Eng. Regen. Med. 2016;13(5):485e97. [50] Venugopal JR, et al. Osteoblasts mineralization with Composite nanofibrous substrate for Bone tissue regeneration. Cell Biol Int 2011;35:73e80. [51] Vetrik M, et al. Biopolymer-based degradable nanofibres from renewable resources produced by freezedrying. RSC Adv 2013;3(35):15282e9. [52] Shaari N, Kamarudin SK. Chitosan and alginate types of bio-membrane in fuel cell application: an overview. J Power Sources 2015;289:71e80. [53] Shekarforoush E, Ajalloueian F, Zeng G, Mendes AC, Chronakis IS. Electrospun xanthan gum-chitosan nanofibers as delivery carrier of hydrophobic bioactives. Mater Lett 2018;228:322e6. [54] Majekodunmi SO. Current development of extraction, characterization and evaluation of properties of chitosan and its use in medicine and pharmaceutical industry. Am J Polym Sci 2016;6(3):86e91. [55] Chiara G, et al. Nanostructured biomaterials for tissue engineered bone tissue reconstruction. Int J Mol Sci 2012;13(1):737e57. [56] Zhang Y, Venugopal JR, El-Turki A, Ramakrishna S, Su B, Lim CT. Electrospun biomimetic nanocomposite nanofibers of hydroxyapatite/chitosan for bone tissue engineering. Biomaterials 2008;29(32):4314e22. [57] Lei S, Feng Edmund T. Polysaccharides, microbial. Ref. Modul. Life Sci. 2017:482e94. [58] Safaee-Ardakani MR, et al. Electrospun Schizophyllan/polyvinyl alcohol blend nanofibrous scaffold as potential wound healing. Int J Biol Macromol 2019;127:27e38. [59] Sun XB, Jia D, Kang WM, Cheng BW, Bin Li Y. Research on electrospinning process of pullulan nanofibers. Appl Mech Mater 2012;268e270:198e201. [60] Li Y, Ma X, Ju J, Sun X, Deng N, Li Z. Preparation and characterization of crosslinked electrospun pullulan nanofiber membrane as a potential for biomaterials. J Text Inst 2017;109(6):750e6. [61] Li Z, Pan T, Wu Y, Kang W, Liu Y. Preparation and characterization of long-term stable pullulan nanofibers via in situ cross-linking electrospinning. Adsorpt Sci Technol 2018. 026361741881301. [62] Qian Y, King M. Green electrospinning of gelatin and pullulan nanofibers to mimic the extracellular matrix. In: Front. bioeng. biotechnol. conference abstract: 10th world biomaterials congress; 2016. [63] Drosou C, Krokida M, Biliaderis CG. Composite pullulan-whey protein nanofibers made by electrospinning: impact of process parameters on fiber morphology and physical properties. Food Hydrocolloids 2018;77:726e35. [64] Blanco-Padilla A, Lo´pez-Rubio A, Loarca-Pin˜a G, Go´mez-Mascaraque LG, Mendoza S. Characterization, release and antioxidant activity of curcumin-loaded amaranth-pullulan electrospun fibers. LWT - Food Sci Technol 2015;63(2):1137e44. [65] Qian Y, et al. Incorporation of rutin in electrospun pullulan/PVA nanofibers for novel UV-resistant properties. Materials 2016;9(7):504. [66] Jeong SI, Krebs MD, Bonino CA, Samorezov JE, Khan SA, Alsberg E. Electrospun chitosanealginate nanofibers with in situ polyelectrolyte complexation for use as tissue engineering scaffolds. Tissue Eng 2011;17(1e2):59e70. [67] Jeong SI, Krebs MD, Bonino CA, Khan SA, Alsberg E. Electrospun alginate nanofibers with controlled cell adhesion for tissue engineering. Macromol Biosci 2010;10(8):934e43. [68] Kyzioł A, Michna J, Moreno I, Gamez E, Irusta S. Preparation and characterization of electrospun alginate nanofibers loaded with ciprofloxacin hydrochloride. Eur Polym J 2017;96:350e60. [69] Bonino CA, et al. Electrospinning alginate-based nanofibers: from blends to crosslinked low molecular weight alginate-only systems. Carbohydr Polym 2011;85(1):111e9. [70] Shekarforoush E, Mendes ACL, Chronakis IS. The electrospinning of xanthan gum: from solution to nanofiber formation. In: Applied nanotechnology and nanoscience international conference; 2017. p. 235.

Electrospun natural polysaccharide for biomedical application 615 [71] Unnithan AR, et al. Wound-dressing materials with antibacterial activity from electrospun polyurethaneedextran nanofiber mats containing ciprofloxacin HCl. Carbohydr Polym 2012;90(4):1786e93. [72] Unnithan AR, et al. Electrospun polyurethane-dextran nanofiber mats loaded with Estradiol for postmenopausal wound dressing. Int J Biol Macromol 2015;77:1e8. [73] Maslakci NN, et al. Ibuprofen and acetylsalicylic acid loaded electrospun PVP-dextran nanofiber mats for biomedical applications. Polym Bull 2017;74(8):3283e99. [74] Moydeen AM, Ali Padusha MS, Aboelfetoh EF, Al-Deyab SS, El-Newehy MH. Fabrication of electrospun poly(vinyl alcohol)/dextran nanofibers via emulsion process as drug delivery system: kinetics and in vitro release study. Int J Biol Macromol 2018;116:1250e9. [75] Vashisth P, Nikhil K, Roy P, Pruthi PA, Singh RP, Pruthi V. A novel gellan-PVA nanofibrous scaffold for skin tissue regeneration: fabrication and characterization. Carbohydr Polym 2016;136:851e9. [76] Vashisth P, Pruthi PA, Singh RP, Pruthi V. Process optimization for fabrication of gellan based electrospun nanofibers. Carbohydr Polym 2014;109:16e21. [77] Thorvaldsson A, Silva-Correia J, Oliveira JM, Reis RL, Gatenholm P, Walkenstro¨m P. Development of nanofiber-reinforced hydrogel scaffolds for nucleus pulposus regeneration by a combination of electrospinning and spraying technique. J Appl Polym Sci 2012;128(2):1158e63. [78] Basha RY, Sampath Kumar TS, Doble M. Electrospun nanofibers of curdlan (b-1,3 glucan) blend as a potential skin scaffold material. Macromol Mater Eng 2017;302(4):1600417. [79] El-Naggar ME, Abdelgawad AM, Salas C, Rojas OJ. Curdlan in fibers as carriers of tetracycline hydrochloride: controlled release and antibacterial activity. Carbohydr Polym 2016;154:194e203. [80] Eman A, Saber WI, El-Sawah MM, Hauka FIA, Kassem MM. Optimization of fermentation conditions for xanthan production by Xanthomonas campestris NRRL B-1459. J. Union Arab Biol. 2000;9B:211e28. [81] Palaniraj A, Jayaraman V. Production, recovery and applications of xanthan gum by Xanthomonas campestris. J Food Eng 2011;106(1):1e12. [82] Li R, et al. Electrospinning pullulan fibers from salt solutions. Polymers 2017;9(1):32. [83] Leathers TD. Substrate regulation and specificity of amylases from Aureobasidium strain NRRL Y-12, 974. FEMS Microbiol Lett 1993;110(2):217e21. [84] Singh RS, Saini GK. Biosynthesis of pullulan and its applications in food and pharmaceutical industry. In: Satyanarayana T. AP, Johri B, editors. Microorganisms in sustainable agriculture and biotechnology. Dordrecht: Springer; 2012. p. 509e53. [85] Yamaguchi R, et al. Conjugation of sendai virus with pullulan and immunopotency of the conjugated virus. Microbiol Immunol 1985;29(2):163e8. [86] Cevher E, et al. Development of chitosanepullulan composite nanoparticles for nasal delivery of vaccines: optimisation and cellular studies delivery of vaccines: optimisation and cellular studies. J Microencapsul 2015;32(8):755e68. [87] Islam S, Hyun J, Kumar A. Effect of pullulan/poly (vinyl alcohol) blend system on the montmorillonite structure with property characterization of electrospun pullulan/poly (vinyl alcohol)/montmorillonite nanofibers. J Colloid Interface Sci 2012;368(1):273e81. [88] Cengiz-Callioglu F. Dextran nanofiber production by needleless electrospinning process. E-polymers 2014;14(1):5e13. [89] Jiang H, Fang D, Hsiao BS, Chu B, Chen W. Optimization and characterization of dextran membranes prepared by electrospinning. Biomacromolecules 2004;5(2):326e33. [90] West Corporation. PolyRemedy launches HealSmartTM hyaluronic acid dressing enhancement. 2013 [Online]. Available: http://www.marketwired.com/press-release/polyremedy-launches-healsmart-hyaluronicacid-dressing-enhancement-1778690.htm. [91] Basha RY, Sampath Kumar TS, Doble M. Electrospun nanofibers of curdlan (b-1,3 glucan) blend as a potential skin scaffold material. Macromol Mater Eng 2017;302(4):1600417.

Index Note: ‘Page numbers followed by “f” indicate figures, “t” indicates tables’.

A Aceclofenac dental paste, 275e276 Acetobacter xylinum, 476 Acid peeling process, 205 Acridine orange-propidium iodide (AO-PI) staining, 21 Adipic acid dihydrazide (ADH), 67 Agarose, 521e522 Albendazole, 226e227 Alcaligenes faecalis, 604 Algal polysaccharides, 4e5, 5t Alginate-chitosan combinations implantable drug delivery system, 354e355 nasal controlled drug delivery, 352e353 ocular controlled drug delivery efficiency, 351 four model drugs, 351e352 gatifloxacin, 350e351 nanoparticles, 350e351 sodium alginate, 350 5-fluorouracil (5-FU), 351 oral controlled drug delivery liquid oral sustained release preparations, 341e344 particulate carriers, 344e348 outcomes of, 340, 341t tissue engineering, 355e356 transdermal controlled drug delivery, 348e350 vaginal controlled drug delivery, 353e354 wound dressing, 355e356 Alginate-hyaluronic acid (ALG-HA), 316 Alginate-polyacrylamide (ALG-PAAm) hydrogels, 46

Alginates, 500f, 550. See also Sodium alginate additives, 499 blocks, 371, 372f bone defects, 499 chemical structure, 499 defined, 371 “egg box”, 372, 372f gels, 372 marine polysaccharides, 514e515 monomers, 371, 371f nanoparticles, 386f formation, 386 nanoaggregates, 385e386 nanosphere formation, waterin-oil emulsions, 386e387 propylene glycol alginate (PrGA), 371 tissue engineering, 536e537 Alginic acid, 499 Alkaline phosphatase (ALP), 135e136, 475e476, 480, 503 Alzheimer disease, 109 5-Aminosalicylic acid (5-ASA), 432e433 Amnion tissue membrane, 17 Amphiphilic polysaccharide derivatives (AMPDs), 433 Anacardium occidentale, 264 Animal polysaccharides, 3e4, 4t Antibacterial textiles, 19 preparation of, 47e48 Antimicrobial food additives, 48e49 Antioxidants, 580, 581f Aplidin, 514 Arabica gum, 289 Arabinogalactan, 576 Arabinose, 1e2, 204e205, 406

617

Arabinoxylan, 406, 573 Arithmetic equations, 128 Ascophyllan, 522 Ascophyllum nodosum, 60, 514e515 Aspergillus flavus Af09, 47 Aureobasidium pullulans, 604 Azotobacter vinelandii, 604

B Bacterial cellulose, 476 BADSCs. See Brown adipose derived stem cells (BADSCs) Biodegradable packaging biopolymers, 19 Bioimaging HMSN-SS-CDPEI@HA, 44e45 QPG-pGFP labeled complexes, 44, 44f quaternized analogs of pectic galactan (QPG), 43 sodium carboxymethyl cellulose (CMCel), 42e43 Biological macromolecules, 423 Bioresponsive polymers, 378e379 Biotech drug delivery nucleic acid delivery, 318e319 protein delivery, 317e318 Bisodol, 62 BMP-2 growth factor, 501 Bovine lactoferrin (bLF), 458 Bovine serum albumins (BSA), 37, 270e273, 345 Brain drug delivery, 132e133 Branched chain polysaccharides, 532

Index Brown adipose derived stem cells (BADSCs), 20e21 AO/PI staining fluorescence images, 21, 22f GATA-4 immunofluorescence staining, 21, 23f Brown algae, 514, 515f Buccal drug delivery, 131 tamarind gum (TG), 297e298 Buccal systems, 276e277 Bulk erosion, 375

C Calcium alginate, 49, 62e63 egg-box model for, 63, 64f Calcium phosphate (CaP), 475e476 Cancer, 443e444 colorectal, 453e454 lung, 450 prostate, 454e455 therapy, 444e445 treatment approaches, 443e444 Candida albicans, 353e354 Carboxyethyl chitosan (CEC), 67 Carboxymethylcellulose, 550 Carrageenan, 36, 505e507, 507f, 599e600 marine polysaccharides, 516 Cashew gum buccal systems, 276e277 chemistry, 265 composition, 265 dental paste, 275e276, 276f gels, 274e275, 274f microparticles, 271f bovine serum albumins (BSA), 270e273 chitosan/carboxymethyl, 270e273 scanning electron microscopy micrographs, 269e270, 272f swelling, 270e273 in vitro isoxsuprine HCl releasing, 269e270, 273f zinc sulfate, 269e270 nanoparticles, 273e274 pharmaceutical biopolymeric excipients, 264

properties, 266 source and extraction, 264e265 tablets, 267e269 Cassia obtusifolia seed mucilage, 32e33, 33t Celecoxib, 213 Cell-based therapy, 19e20 Cell encapsulation acridine orange-propidium iodide (AO-PI) staining, 21, 22f advantages, 19e20 brown adipose derived stem cells (BADSCs), 20e21 cell-based therapy, 19e20 drug localization, 19e20 fluorescence microscopy images, 24, 26f GATA-4 immunofluorescence staining, 21, 23f human adiposeederived stem cells (hASCs), 24e27 morphology and attachment, 28e29, 28f NT2 cells encapsulated within alginate beads, 21, 24t optical microscopy images, 24, 25f spatial distribution of, 27e28, 27f Young’s moduli, 20e21 Cellular immune response, 191e192 Cellulose, 503 tissue engineering, 539 Cellulose acetate (CA), 503 Cell walls, 568e569 Charged polysaccharides, 403 Chemotherapy (CT) drugs, 446te449t chitosan (CS), 445e450 doxorubicin (DOX), 445e450 hyaluronic acid (HA), 450 MDA-MB-231, 450 methoxy poly(ethylene glycol) (mPEG), 450 paclitaxel (PTX), 450e451 Chicory, 571e573 Chitin, 3e4 Chitosan (CS), 3e4, 18, 38, 315

618

chemical properties, 374 biomedical applications, 105t characteristics of, 104e105 chemical structure of, 102e103, 103f deacetylaction of, 373, 373f derivatives, 103, 105t functional groups, 102e103, 103f marine animals, 518, 519f micro- and nanoparticles, 387 coacervation/precipitation, 388 desolvation, 389 emulsion cross-linking/ emulsion solvent diffusion, 388 emulsion-droplet coalescence, 389 nanofibers, 389e390 polyelectrolyte complexation/ ionic gelation, 388 reverse micellar method, 389 sieving method, 388 spray drying, 388 modifications of, 105 chemical, 106e107 enzymatic, 107e108 radiation-induced, 108e109 pharmaceutically active compounds nucleic acids, 112e113 proteins and antigens, 110e111 small-molecule drugs, 109e110 tissue engineering, 537e539 Chitosan-g-poly(ethylene glycol) (CS-g-PEG), 318e319 Chitosan-ZnO nanocomposites, 482 Chondroitin sulfates, 3e4, 420 marine animals, 520 Ciprofloxacin, 346, 610e611 “Clickable” zwitterionic starch hydrogels, 20e21 Clostridium perfringens, 48e49 Clove oil (CO), 40e41 Coacervation, 385, 404

Index Cochlospermum kunth, 225 Coil-helix transitions, 148e149 Colloidosomes brilliant blue-loaded, 72 preparation, 72, 73f, 75f production yield, 74 Colon-specific drug delivery, 231f, 299e300 capsular body, 229 carriers, 228e229 microflora-triggered drug delivery systems (MCDDS), 229 Coloplas, 555 Contact lenses, 18 preparation of, 45e46 Controlled/living radical polymerization (C/LRP), 384e385 Controlled release microparticles/ beads, 291, 292f ConvaTec, 555 Core-shell nanocomposites, 473 Corn starch, 48e49 Cosmetics, 136e137 Critical aggregation concentration (CAC), 312e313 Cross-linked hydrogel, 61e62 CS. See Chitosan (CS) Curcumin (Cur), 36 Current good manufacturing practices (cGMPs), 145e146 Cyclodextrins (CDs), 352e353, 455 Cytochrome C (CC), 458

D Deferoxamine (DFO), 434 Delayed drug release, 152e153 Dental paste, 275e276, 276f Dermal patches, 134 Dermatan sulfate, 3e4, 520 Desolvation, 405 Dextran, 540 Diabetes mellitus (DM), 251e252 Dialysis, 382 Diels-Alder reaction, 70

Dietary fibers, 576e577, 578te579t Diffusion chamber, 17 Diltiazem-Indion 254 complex, 291e292 Dilute alkali-water extraction method, 6 Direct compression method, 209e211 DM. See Diabetes mellitus (DM) Doxorubicin (DOX), 310, 429, 473 Doxorubicin hydrochloride, 37 Doxycycline hyclate, 241 Drug delivery systems (DDSs), 102, 308, 311e316, 367 Drug-laden contact lenses, 18 Drug-loaded gellan beads, 155e156 Drug loading, 367 Drug release (DR), 368

E Ecteinascidin 743, 514 “Egg box”, 372, 372f Electropositivity, 150e151 Electrospinning, 389e390, 400 nanofiber morphology, 591e593 solvent and solvent-free electrospinning apparatus, 591, 592t Taylor cone, 591 Electrospray, 405 Electrospun nanofibers, 590e591 Electrospun natural polysaccharides animal origin chitin, 601e602, 602f chitosan, 603e604 researches of, 600e601, 600t electrospinning, 591e593 human origin hyaluronic acid, 594e596 researches of, 594, 595t microbe origin pullulan, 607e608, 607f researches of, 604, 605t xanthan gum, 604e607, 606f nanotechnology, 590e591 plant and seaweed origin

619

carrageenan, 599e600 guar gum, 598e599 researches of, 597t, 599 Embelia ribes, 135 Emblica officinalis, 129e130 Emulsification-evaporation method (EEM), 380, 380f Emulsification/solvent diffusion, 381 Emulsion polymerization microemulsion, 383e384 miniemulsion, 383 Emulsions lamivudine, 330e332 multiple, 330e332 primary, 329e330 Tween 80, 330e332 viscosity, 329e330 Encapsulation, 17 curcumin, 36 vitamin B2, 77 of water-soluble drugs, 74 Enhanced permeability and retention effect (EPR), 444e445 Enzymolysis method, 6e7 Erythema, 42 Escherichia coli, 353e354, 482 Exopolysaccharides, 5e6, 146 External gelation, 65 Extracellular matrix (ECM), 308e309 Extracellular polysaccharides, 5e6

F Faecalibacterium prausnitzii, 251 Fickian diffusion mechanism, 230 Fick’s Law, 561 Field-responsive polymers, 379 Flat systems, 549e550 applications, 550 “living” surface, 550e551 mass transfer processes, 550, 551f oral disintegration films (ODF), 551 Floating beads, 297

Index Fluid-handling capacity (FHC), 558 5-Fluorouracil (5-FU), 189e190, 351 Food safety, 48, 480e483 Fourier transform-infrared (FTIR) spectroscopy, 332e333 Fructans, 1e2, 571e573, 572f Fucoidans, 503e505, 506f marine polysaccharides, 516, 517t Fucus vesiculosus, 516

G Galactan, 1e2 Galactomannan, 126e127, 574 Galactose, 1e2, 204e205 Galactoxyloglucan, 287e288 Galectin-3, 253e254, 254f Gastroesophageal reflux disease (GERD), 93 Gastrointestinal tract (GIT) microbes, 187e188 Gastroretentive drug delivery, 232f drug loading efficiency, 230e232 Fickian diffusion mechanism, 230 hydroxyl propyl methyl cellulose (HPMC), 233e234 Korsmeyer-Peppas exponential model, 230 GATA-4 immunofluorescence staining, 21, 23f Gaviscon, 62 Gelatin, 289 Gelation acetylated and deacetylated, 148e149 acyl groups, 149 coil-helix transitions, 148e149 hydrogels, 148 hydrophilic ingredients, 151e152 ion type and concentration, 150e151 pH, 151 sol-gel transition, 148e149

sterculia gum (SG), 225e226 “true hydrogel”, 148e149 “weak gel”, 148e149 Gellan gum (GG), 173t delayed drug release, 152e153 dosage forms, 146 gelation acetylated and deacetylated, 148e149 acyl groups, 149 coil-helix transitions, 148e149 hydrogels, 148 hydrophilic ingredients, 151e152 ion type and concentration, 150e151 pH, 151 sol-gel transition, 148e149 “true hydrogel”, 148e149 “weak gel”, 148e149 nasal drug delivery, 163e167, 168te169t ophthalmic drug delivery, 156e163 periodontal drug delivery system, 171 peroral drug delivery, 153e154 liquid oral in situ gelling systems, 154e155 multiparticulate drug delivery systems, 155e156, 157te162t tablets and capsules, 154 properties, 148e152 routes of administration, 146 in situ vaginal gel formulations, 172 sources and structure, 146e147, 147f sustained drug release, 152e153 topical drug delivery, 170te171t cutaneous, 167e170 dermal, 167e170 transdermal, 167e170 tissue engineering, 540 Gels, 2 skin permeation, 274e275 in vitro lidocaine HCl permeation profile, 274e275, 274f

620

Gemcitabine hydrochloride (GEM), 30e32 Gene delivery, 33 curdlan-based nanoparticles, 34 glycosaminoglycans (GAGs), 34e36 RNA interference (RNAi), 34 siRNA, internalization of, 34, 35f siRNA-loaded 6AC-100 nanoparticles, 34, 34t in vitro silencing, of endogenous mRNA, 34, 35f General extraction methodologies benefits and drawbacks of, 7, 7t dilute alkali-water extraction method, 6 enzymolysis method, 6e7 hot water extraction method, 6 selection of, 5e6 Generally Recognized as Safe (GRAS), 68, 224e225 Gene therapy, 451 Genipin, 406 GG. See Gellan gum (GG) Glassy carbon electrode (GCE), 484e485 Glaucoma, 106 Glucan, 1e2 Glucomannan, 574 Gluconic acid, 266 Glucosamine, 499e501 Glucose, 1e2, 204e205 Glucuronic acid, 64e65, 265 Glucuronoarabinoxylan, 573e574 Glutaraldehyde (GA), 190e191, 212 Glycanes, 420e421 Glycerine, 275e276 Glycosaminoglycans (GAGs), 3e4, 34e36, 420, 517 Glycosyltransferase, 123e124 Glycyrrhiza glabra, 129e130 Gold nanoparticles (GNP), 30e32 Growth factors delivery importance, 498 polysaccharides for alginate, 499

Index carrageenan, 505e507, 507f cellulose, 503 chitin/chitosan, 499e501, 502f fucoidan, 503e505, 506f hyaluronic acid (HA), 501e503, 504f ulvan, 507 Guar gum (GG) buccal film, 195 chemical structure and applications, 598e599, 598f gelling network, 188 grafted systems, 192e193 hydrogel systems, 193e195 microparticles glutaraldehyde (GA), 190e191 mebeverine hydrochloride, 189e190, 190f scanning electron microscopic (SEM), 189e190 5-fluorouracil (5-FU), 189e190 X-ray diffraction (XRD), 189e190 nanoparticles, 191e192 properties, 188 research activities, 599 structure of, 187e188, 188f tablets for, 195e198, 197f Gum karaya (GK), 30e32 Gum odina dosage forms, 328 Odina wodier Roxb., 328e329 pharmaceutical excipient emulsions, 329e332 tablets, 332e334 properties, 328e329 source, 328e329, 328f Gums, 225

H HA. See Hyaluronic acid (HA) Halloysite clay, 48e49 Helicobacter pylori, 346 Helix aggregation, 149 Helmholtz free energy, 72 Hemicellulose, 569e571

Heparan sulfates, 3e4, 520e521 Heparin, 3e4, 420 Herbicide, 483e484 Heteroglucans, 496 Heteropolysaccharides, 3e4, 421, 422f, 496 High-performance liquid chromatography (HPLC), 195e196 Higuchi model, 561 HMSN-SS-CDPEI@HA, 44e45 Homogalacturonan (HG), 249e250, 575 Homopolysaccharides, 421, 422f, 496 Hot water extraction method, 6 Human adiposeederived stem cells (hASCs), 24e27 Human Embryonic Kidney (HEK293) cells, 475 Hyaluronans. See also Hyaluronic acid (HA) marine animals, 518e519 Hyaluronic acid (HA), 3e4, 407, 408f active targeting drug delivery, 310e311, 312t chemical structure, 308e309, 309f chemical structure and applications, 594e595, 596f cross-linking of, 501 derivatives, 309 drug delivery systems (DDSs), 311 assembly of, 314e316, 317t biotech drug delivery, 316e319 nanogels and hydrogels, 311e314 hydrogel implantation, 502e503, 504f research, 595e596 Hydrogel contact lenses, 45e46 Hydroxyl propyl methyl cellulose (HPMC), 233e234 Hypocholesterolemia effect, 252e253 Hypoglycemic effect, 251e252

621

I Immunostimulants, 577e580 Immunotherapy, 443e444 Implantable drug delivery system, 354e355 Implant preparation, 46e47 Injectable delivery artemisinin nanocapsules, 79 doxorubicin, 79e81 photosensitizer, 77e79 single-wall carbon nanotubes (SWCNTs), 79e81, 80f Intercellular cell-adhesion molecule-1 (ICAM-1), 474 Interfacial polymerization, 384 Internal gelation, 65 Interpenetrating polymer network (IPN), 269e270 Intracellular polysaccharides, 5e6 Intraocular lens (IOLs), 45 Inulin, 571e573 Ion-exchange coefficient, 64e65 Ionic gelation, 385 Ionotropic gelation, 63 external, 65 internal, 65 Ionotropic gelation method, 269e270

J Joint Expert Committee for Food Additives (JECFA/FAO), 225

K Karaya, 224e225 K-carrageenan, 208 Keratan sulfates, 420, 521 Korsmeyer model, 562 Korsmeyer-Peppas model, 212, 274e275, 292e293 Kuhn’s modeling, 287e288

L Laminaran, 522 Laminaria digitata, 514e515 Laminaria hyperborea, 514e515

Index Laminaria hyperborean, 60 Laminaria japonica, 514e515 Langmuir mechanism, 367 LBG. See Locust bean gum (LBG) Leafy vegetables, 571f, 580 arabinogalactan, 576 fructans, 571e573, 572f functional attributes of antioxidants, 580, 581f dietary fibers, 576e577 immunostimulants, 577e580 hemicellulose, 569e571 mannans galactomannan, 574 glucomannan, 574 negative physiological impact, 583 pectin homogalacturonan (HG), 575 rhamnogalacturonan, 575e576 xylogalacturonan, 576 polysaccharide localization, 568e569, 570f processing condition, 582e583 xylans, 572f arabinoxylan, 573 glucuronoarabinoxylan, 573e574 xyloglucan, 574 Leuconostoc mesenteroides, 604 Linear chain polysaccharides, 532 Liquid oral dosage form, 129e130 Liquid oral in situ gelling systems, 154e155 Liquid oral sustained release preparations effect of, 342e344, 343f gelling capacity and gelling strength, 342e344 hypoglycemic effect, 344 ion-induced cross-linking, 342 preparation of, 341e342 Listeria monocytogenes, 48e49 “Living” surface, 550e551 Locust bean gum (LBG) applications of, 210f

gel formulation, 213e214 microcapsules, 211e212 microparticles, 213 microspheres, 212 modified tablet, 209e211 nanoparticles, 213 polymeric beads, 214e215 polymeric films, 214 solubility enhancement, 215 tissue engineering, 215, 216te217t composition and properties, 206 impurities, 206 neutral galactomannan polymer, 204e205 processing of manufacturing aspect, 205 manufacturing principle, 205 synergistic interaction, 207 K-carrageenan, 208 xanthan gum (XG), 207e208 in vivo biodegradation, 208e209, 209f Lower critical solution temperature (LCST), 377e378 Lyophilization technique, 70e72, 83 Lysozyme, 49

M Macrocystis pyrifera, 60, 514e515 Magnesium aluminum silicate (MAS), 76 Magnetic nanoparticles (MNPs), 135e136 Magnetic responsive polyelectrolyte complex hydrogels (MPECHs), 42 Mannans, 1e2 galactomannan, 574 glucomannan, 574 Mannose, 1e2, 204e205 Mannuronic acid, 64e65 Marine algae, 4e5 Marine animals, 517 chitosans, 518, 519f chondroitin sulfates, 520 hyaluronans, 518e519

622

Marine polysaccharides, 402, 403f alginates, 514e515 antitumoral properties, 514 biological properties and structures, 514 brown algae, 514, 515f carrageenans, 516 fucoidans, 516, 517t glycosaminoglycanlike polysaccharides agarose, 521e522 ascophyllan, 522 dermatan sulfate, 520 heparan sulfates, 520e521 keratan sulfates, 521 laminaran, 522 ulvans, 516e517 Membrane technology, 550 Mesenchymal stem cells (MSCs), 314 Mesoporous titania nanoparticles (MTNs), 310 Methicillin-resistant Staphylococcus aureus (MRSA), 434 Methoxypolyethylene glycol (mPEG), 314e315 3M HealthCare, 555 Micro- and nanoparticle synthesis, 365e366 alginates blocks, 371, 372f defined, 371 “egg box”, 372, 372f gel, 372 monomers, 371, 371f nanoparticles, 385e387 propylene glycol alginate (PrGA), 371 applications alginate nanoparticles, 390e392 chitosan-alginate nanoparticles, 394 chitosan nanoparticles (CS NPs), 392e393 chitosan, 387e390 chemical properties, 374 deacetylaction of, 373, 373f

Index controlled release (CR), 366 formulation method, 366 loading and realease, 367e368 nanoencapsulation, 366 nanoparticle sources biodegradable NPs, 369 polymeric matrix, 369e370 polymers used, 369, 369t surface charge, 370 nanosphere vs. nanocapsule, 366, 366f polymeric nanoparticles, 367 preparation of, 379 ionic gelation/coacervation, 385 polymerization methods, 383e385 preformed polymers, 380e382 smart polymers, 377t bioresponsive polymers, 378e379 bulk erosion, 375 factors, 375, 375t field-responsive polymers, 379 pH-responsive polymers, 378 polymer matrix, 374e375 stimuli-sensitive, 375e376 stimulus responsible, 375e376, 376f surface degradation, 375 temperature-responsive polymers, 377e378 Microbial polysaccharides, 4, 5t, 146 Microemulsion polymerization, 383e384 Microencapsulation bioactive compounds, 77 polyphenolic extracts, 77 Microneedle system (MNs), 111 Microfludization, 126 Miniemulsion polymerization, 383 Modified citrus pectin (MCP), 253e254 “Moist healing” products, 82e83 Moringa olifera, 135 Mucilages, 225 Mucoadhesive microparticles/beads

scanning electron microscopy (SEM), 293e294, 294fe295f in vitro drug release, 293e294, 295fe296f Mucoadhesive nanoemulsions (mNNEs), 106 Mucoadhesivity, 66e67 Multiparticulate drug delivery systems, 155e156, 157te162t Multiwalled carbon nanotubes (MWCNT), 432e433

N NABDs. See Nucleic acidse based drugs (NABDs) N-acetyl glucosamine, 499e501 Nanofibrillated cellulose with alginate (NFC/A) bioink, 92e93 Nanodevices, 590e591 Nanoencapsulation, 400 Nanomedicine, 590e591 Nanophase forsterite, 479 Nanoprecipitation, 380e381 Nanostarch (NS), 38 Nanotechnology, 590e591 Nasal drug delivery, 81e82, 135 gellan gum (GG), 163e167, 168te169t tamarind gum (TG), 299 Natural polysaccharide-based composites, 420e421 biological macromolecules, 423 biopharmaceutical applications, 430e434, 431t classification of, 423e426, 426t drug delivery, 430e434 heteropolysaccharides, 421, 422fe423f homopolysaccharides, 421, 422fe423f hydrolysis, 426 improving composites, 433e434 methylation, 426 molecular formula and structure, 423, 424te425t nanocomposites, 427e428 and nanomaterials, 428e429

623

cardiovascular disease (CVD), 429e430 nanogels, 429 pharmaceutical and biomedical industries, 435e437, 435f preparation of, 427e428, 427f, 428t purification, 426 resources, 421, 422f self-assembly amphiphilic polysaccharide derivatives, 433 stimuli-responsive properties, 431e433 Nearinfrared (NIR) photothermal effect, 432e433 Neoteric polymers (NPs), 102 Nisin, 49 Nonwoven fabric, 553, 553f Nucleic acid delivery, 318e319 Nucleic acidsebased drugs (NABDs) chitosan (CS), 454 cyclodextrins (CDs), 455 hyaluronic acid (HA), 453 multidrug resistance impacts, 454 polysaccharide polymers, 451, 452t RNA-inducing silencing complex (RISC), 451 small interfering RNAs (siRNAs), 451 Nylon, 550

O Ocular delivery, 81 tamarind gum (TG), 298e299 O-glycosidic linkages, 1e2 Olive leaf extract (OLE), 595e596 Ophthalmic drug delivery, 130, 156e163, 164te166t Oral delivery colloidosomes, 72, 73f cross-linked alginate hydrogel, 70, 71f Diels-Alder reaction, 70 drug release profile, 70

Index Oral delivery (Continued) grafting reaction mechanism, 77, 78f liquid oral sustained release preparations, 341e344 lyophilization, 70e72 magnesium aluminum silicate (MAS), 76 microcapsules, 69e70, 69f microencapsulation, 77 mucoadhesive films, 76 nanocarriers, 72 particulate carriers, 344e348 reversibly cross-linked SA hydrogel, 70 risperidone, 70e72 Oral disintegration films (ODF), 551 Organic nanocomposites biomedical engineering bioglass, 480 high-density polyethylene (HDPE), 478 nanophase forsterite, 479 polycaprolactone (PCL), 479 Young’s modulus, 478 diagnosis (600), 477e478 drug delivery alkaline phosphatase (ALP), 475e476 bacterial cellulose, 476 calcium phosphate (CaP), 475e476 chitosan, 473 core-shell, 473 doxorubicin, 473 endosomal internalization, 475 examples of, 473, 474t graphene oxide, 476 intercellular cell-adhesion molecule-1 (ICAM-1), 474 poly(lactic-co-glycolic acid) (PLGA), 474 silk fibroin (SF), 475e476 vascular endothelial growth factor (VEGF), 475e476 venlafaxine hydrochloride, 474 effectiveness and safety, 473

enrichment, 480e483 food safety, 480e483 inorganic ions, 472e473 matricial structure, 471e472, 472f packaging systems, for food products, 481, 481t phytopharmacy and ecology, 483e485 shelf-life extension, 480e483 toxicity, 485e487

P Packaging materials, 48e49 Papers barrier properties, 19 features, 19 preparation of, 47e48 Parkinson disease, 24 Particulate carriers bovine serum albumin (BSA), 345 capsule, 344e345 ciprofloxacin, 346 hemoglobin (Hb), 346e347 limitations, 344e345 microspheres, 344e345 simulated gastric fluid (SGF), 346e347 structure and size, 344e345 swelling behavior, 346 PDSs. See Polysaccharides-based delivery systems (PDSs) Pectin-honey hydrogel (PHH), 258 Pectins, 403 controlled and directed release matrix blinking and lacrimation, 257 pectin/arabinoxylan beads, 255e256, 256f pectin-honey hydrogel (PHH), 258 skin applications, 258 homogalacturonan (HG), 249e250, 575 hypocholesterolemia effect, 252e253 hypoglycemic effect, 251e252

624

metastasis and apoptosis, in cancer cells antineoplastic drugs, 254e255 galectin-3, 253e254 modified citrus pectin (MCP), 253e254 rhamnogalacturonan I (RG-I), 255 prebiotic effect, 250e251 properties of, 249e250 rhamnogalacturonan, 575e576 tissue engineering, 540 vegetable sources, 250 xylogalacturonan, 576 PEGylation, 106, 315e316 Peppas-Sahlin model, 562e563 Peptides and proteins (P-PDS) anticancer activity in vitro and in vivo, 457e458, 459t bovine lactoferrin (bLF), 458 cytochrome C (CC), 458 drawbacks, 457, 457t granzyme B (GrB), 458 mechanism of, 456, 457f rituxan, 456e457 Peroral drug delivery, 153e154 liquid oral in situ gelling systems, 154e155 multiparticulate drug delivery systems, 155e156, 157te162t tablets and capsules, 154 Phaeophyceae, 340 Pharmaceutical applications, 20 antibacterial textiles/papers, 47e48 antimicrobial food additives/ packaging materials, 48e49 bioimaging, 42e45 cell encapsulation acridine orange-propidium iodide (AO-PI) staining, 21, 22f advantages, 19e20 brown adipose derived stem cells (BADSCs), 20e21 cell-based therapy, 19e20 drug localization, 19e20 fluorescence microscopy images, 24, 26f

Index GATA-4 immunofluorescence staining, 21, 23f human adiposeederived stem cells (hASCs), 24e27 morphology and attachment, 28e29, 28f NT2 cells encapsulated within alginate beads, 21, 24t optical microscopy images, 24, 25f spatial distribution of, 27e28, 27f Young’s moduli, 20e21 contact lenses, 45e46 gene delivery, 33e36 implant preparation, 46e47 pharmaceutics/drug delivery A549 human lung cancer cells, 30e32, 32f Cassia obtusifolia seed mucilage, 32e33, 33t clonogenic assay, 30e32, 32f FITC tagged MR/CS nanoparticle absorption, 29e30, 30f gemcitabine hydrochloride (GEM), 30e32 gold nanoparticles (GNP), 30e32 gum karaya (GK), 30e32 mauran-chitosan (MR-CS), 29e30 Sypro-ruby tagged MR/CS nanoparticle absorption, 29e30, 31f protein binding, 36e37, 37f tissue engineering, 41e42, 43f wound healing, 38e41 Phosphate buffer saline (PBS), 190e191 Photolysis, 483e484 Photosensitizer, 77e79 pH-responsive polymers, 378 Pickering emulsion, 72 Plant polysaccharides, 2e3, 3t Plastic packaging materials, 19 Platelets, 498 Platelets rich plasma (PRP), 498 Poly(acrylic acid) (PAA), 432e433, 472e473

Polycaprolactone (PCL), 479 Polyethylene glycol (PEG), 315e316 Polyhexamethylene biguanide (PHMB), 555e557 Polyholosides, 420e421 Poly(3-hydroxybutyrate-co-3hydroxyvalerate) (PHA), 482e483 Poly(lactic acid) (PLA), 472e473 Poly(lactic-co-glycolic acid) (PLGA), 472e473 Polyosides, 420e421 Polysaccharide1459, 122 Polysaccharides, 153. See also Polysaccharides-based delivery systems (PDSs); Polysaccharides nanoparticles classification, 497 in drug delivery, 498 functional groups, 16e17 functionalization of, 534 gene delivery, 17 locust bean gum, 204e205 natural sources, 264 physicochemical properties, 16e17 sources of, 534, 534t structure and properties, 496e497, 497f Polysaccharides-based delivery systems (PDSs) biofunctionalization, 462, 462f cancer therapy, 444e445 for cancer treatment, 445 CT drugs, 445e451, 446te449t nucleic acidsebased drugs (NABDs), 451e456, 452t peptides and proteins (PPDS), 456e461 clinical trials of, 461, 461t Polysaccharides nanoparticles for biomedical applications, 405 arabinose, 406 arabinoxylan, 406 biopolymer-inspired gold nanoparticles synthesis, 407, 407f

625

genipin, 406 hyaluronic acid (HA), 407, 408f physical/chemical crosslinking, 406 of health products biomaterials, 402 functions, 401 marine polysaccharide, 402, 403f monosaccharides, 401 reserve compounds, 401 separation and purification, 401e402 oral delivery systems, 408 adverse effects, 409 drug loaded, in chitosan nanoparticles, 410, 410f examples of, 410, 411t size distribution, 409 for oral insulin delivery dextran sulfate/chitosan, 413 DMT1 and DMT2, 411e412 glucose-lowering drug (GLD), 410e411 low-molecular-weight chitosan (LMWC), 413, 413f pancreas, 412 sulfonylurea (SU), 410e411 preparation of charged polysaccharides, 403 complex coacervation, 404 desolvation, 405 electrospray, 405 emulsification, 404 factors, 402 ionic gelation, 403 pectin, 403, 404f self-assembly, 402 tripolyphosphate (TPP), 403 Poly(sebacic acid) (PSA), 472e473 Polyurethane, 550 Poly(vinyl pyrrolidone) (PVP), 38 Porcine reproductive and respiratory syndrome virus (PRRSV), 353 Porphyrin, 596

Index Porphyromonas gingivalis, 47 Prebiotic effect, 250e251 Prebiotics, 577 Propylene glycol, 349 Propylene glycol alginate (PGA), 36, 371 Protein binding, 36e37, 37f Protein delivery, 84e89, 86f, 88f hyaluronic acid (HA), 317e318 Proton Sponge (PS) effect, 475 PRP. See Platelets rich plasma (PRP) Pseudomonas aeroginosa, 61, 298 Pseudomonas elodea, 146e147, 604 Psyllium, 596 Pullulan, 540

Q Quantum dots (QDs), 18, 42e43 Quaternized analogs of pectic galactan (QPG), 43

R Radiation therapy (RT), 443e444 Rapid expansion of supercritical solution (RESS), 382 Reactive oxygen species (ROS) assay, 237, 238f Regenerated cellulose (RC), 503 Rhamnogalacturonan, 575e576 Risperidone, 70e72 Rituxan, 456e457 RNA interference (RNAi), 34 Roseburia intestinalis, 251

S SA. See Sodium alginate (SA) Salting out, 381e382 Sandalwood oil (SO), 40e41 Sargassum stenophyllum, 516 Scanning electron microscopy (SEM), 189e190, 227, 292e293, 502f Schizophyllan commune, 604 Self-assembled nanoparticles, 273e274 Self-assembly, 402 Serratiopeptidase, 131e132 SG. See Sterculia gum (SG) Silk fibroin (SF), 475e476

Simulated colonic fluid (SCF), 196e197 Simulated gastric fluid (SGF), 346e347 Single-wall carbon nanotubes (SWCNTs), 79e81, 80f Skin edema, 42 Skin permeation, 274e275 Small interference RNA (siRNA), 34, 34t, 451 “Smart” nanogels, 400 Smart polymers, 377t bioresponsive polymers, 378e379 bulk erosion, 375 factors, 375, 375t field-responsive polymers, 379 pH-responsive polymers, 378 polymer matrix, 374e375 stimuli-sensitive, 375e376 stimulus responsible, 375e376, 376f surface degradation, 375 temperature-responsive polymers, 377e378 Sodium alginate (SA) applications, 68e69 cell delivery and implants, 83e84, 85f gastroesophageal reflux disease (GERD), 93 injectable delivery, 77e81 nasal delivery, 81e82 ocular delivery, 81 oral delivery, 69e77 protein delivery, 84e89, 86f, 88f tackling obesity and weight management, 82 tissue engineering, 89e93, 91fe92f vaccine delivery, 89 wound dressings, 82e83 chemical structure, 60, 60f cross-linked hydrogel, 61e62 extraction of, 61 physicochemical properties biocompatibility, 67e68, 68f cross-linking and gel formation, 63e65, 64f molecular structure, 62

626

mucoadhesivity, 66e67 pH sensitivity, 65e66 solubility, 62e63 viscosity, 63 weight, 62 properties of, 61 Pseudomonas aeroginosa, 61 regulatory status, 68 Sodium caseinate (SC), 67e68 Soft contact lens (SCL), 45 Sol-gel transition, 148e149 Solid oral dosage form, 129 Solvent displacement procedure, 380e381 Soxhlet extractor, 5e6 Soybean protein isolate (SPI), 36 Spherical self-assembled nanoparticles, 273e274 Spheroids, 291 Sphingomonas elodea, 146e147 Sphingomonas paucimobilis, 146e147 Staphylococcus aureus, 482 Staphylococcus enterica, 482 Starch, 38 tissue engineering, 539e540 Sterculia gum (SG) applications, 226 chemical composition, 225 colon-specific drug delivery, 228e230 dissolution enhancement albendazole, 226e227 doxylamine succinate, 226e227 drug dissolution rate, 227 pure nimodipine, 227, 228f rizatriptan benzoate, 226e227 gastroretentive drug delivery, 230e234 gelation, 225e226 hydrogels, 224e225 IPN microparticles, 234e235, 234f modified SG-based hydrogels ciprofloxacin-loaded b-CDSG-cl-carbopol hydrogels, 243e244, 243f doxycycline hyclate, 241 drug release rate, 239 ornidazole, 242

Index polyvinyl alcohol (PVA), 239 polyvinylpyrrolidone (PVP), 237e239 ranitidine HCl, 241 scanning electron micrographs, 239, 240f swelling equilibrium protocol, 237 nanoparticles, 237, 238f source, 225 sustained release tablets, 235e236, 236f wound dressings, 224e225 Sterculia setigera, 225 Sterculia villosa, 225 Stimuli-responsive properties, 431e433 Streptococcus mutans, 47 Sulfated polysaccharides, 514 Sun protection factor (SPF), 136e137 Supercritical antisolvent (SAS), 382 Supercritical extraction, 400 Supercritical fluid (SCF) technology rapid expansion of supercritical solution (RESS), 382 supercritical antisolvent (SAS), 382 Super paramagnetic iron oxide nanoparticles (SPION), 432e433 Surface degradation, 375 Surgical Dressing Manufacturers Association (SDMA), 555 Sustained drug release, 152e153 Sustained release tablets, 235e236, 236f Synergy, 207 K-carrageenan, 208 xanthan gum (XG), 207e208

T Tablets diclofenac sodium tablets, 267e268 direct compression method, 267e268

disintegration time, 267 gum odina effectiveness, 332 formulation, 332 Fourier transform-infrared (FTIR) spectroscopy, 333e334 matrix-forming agent, 332e333 uses of, 333e334, 334t swelling behavior, 268e269 tamarind gum (TG), 289e291, 290t tensile strength and brittle fracture index values, 267 Tacrolimus (TCS), 310 Tamarind gum (TG) buccal drug delivery, 297e298 characteristics, 285e286 chemical composition, 287e288 colon-targeted drug delivery, 299e300 drug delivery applications, 288e289, 288f emulsions, 289 isolation, 286e287 molecular structure, 287f nasal drug delivery, 299 ocular drug delivery, 298e299 oral multiple-unit systems controlled release microparticles/beads, 291, 292f floating beads, 297 interpenetrating polymer network microparticles, 291e293, 293f mucoadhesive microparticles/ beads, 293e297 spheroids, 291 properties, 285e288 sources, 286e287 suspensions, 289 tablets, 289e291, 290t wound dressing, 300 wound healing, 300 Taylor cone, 591 Temperature-responsive polymers, 377e378

627

TERM. See Tissue engineering regenerative medicines (TERM) Terminalia belerica, 129e130 Terminalia chebula, 129e130 Tetrabutylammonium (TBA) salt, 62e63 TG. See Tamarind gum (TG) Therapeutic contact lens (TCL), 45 Thermal peeling process, 205, 206f “Thiol-ene” Michael addition reaction, 20e21 Timolol maleate (TML), 45 Tissue engineering, 41e42, 43f, 89e93, 91fe92f alginate-chitosan combinations, 355e356 alginates, 536e537 cellulose, 539 chitosan, 537e539 dextran, 540 gellan gum, 540 locust bean gum (LBG), 215, 216te217t marine polysaccharides. See Marine polysaccharides pectins, 540 pullulan, 540 starch, 539e540 strategy for, 535, 535f xanthan gum (XG), 135e136, 540 Tissue engineering regenerative medicines (TERM), 532, 532f Titanium, 18 Topical drug delivery, 131e132 gellan gum (GG), 170te171t cutaneous, 167e170 dermal, 167e170 transdermal, 167e170 Transdermal drug delivery, 134 Trimethylation, 106 Tripolyphosphate (TPP), 403 Tuberculosis, 191e192 Turbinella rapa, 129e130 Two-dimensional systems, 549e550

Index U Ulvans, 507 marine polysaccharides, 516e517 Undaria pinnatifida, 505 Upper critical solution temperature (UCST), 377e378 Uronic acids, 1e2, 204e205 U.S. Food and Drug Administration (USFDA), 68

V Vaccine delivery, 89 Vacuoles, 568e569 Vascular endothelial growth factor (VEGF), 63e64, 475e476 VEGF. See Vascular endothelial growth factor (VEGF) Venlafaxine hydrochloride, 474 Vibrio anguillarum, 111 Viscoelasticity, 591e593 Vitamin B12, 274

W Wound dressings, 82e83, 224e225 adhesivelike flat system, 552, 552f alginate-chitosan combinations, 355e356 characteristics, 554 commercial names and producers, 555, 556t drugs, 555e557, 556t flat systems applications, 550 “living” surface, 550e551 mass transfer processes, 550, 551f oral disintegration films (ODF), 551 frequent polysaccharides, 559e560, 560f mathematical models first order model, 562 Higuchi model, 561 kinetic models for drug release, 561, 562t

Korsmeyer model, 562 Peppas-Sahlin model, 562e563 methodologies for, 553, 553t nonwoven fabric, 553, 553f other polysaccharides, 560, 561f polyhexamethylene biguanide (PHMB), 555e557 properties, 557 fluid-handling capacity (FHC), 558 healing activity, 558 scientific papers, 558e559, 559t Surgical Dressing Manufacturers Association (SDMA), 555 tamarind gum (TG), 300 Wound healing collagen fiber, 39e40 cutaneous, 38 dextran-based bionanocomposite films, 40e41 dressing materials, 38 hair follicle cells, 39e40 healing efficacy, 39e40 histological analysis, 39e40, 40f starch, 38 tamarind gum (TG), 300 in vivo, 38e39, 39f xanthan gum (XG), 133e134

X Xanthan gum (XG), 36, 42, 195e196 applications of, 122e123, 124f advanced drug delivery, 132 brain drug delivery, 132e133 buccal drug delivery, 131 cosmetic uses, 136e137 dermal patches, 134 food, 137e138, 138te139t healthcare systems, 128, 128f liquid oral dosage form, 129e130 nasal drug delivery, 135 ophthalmic drug delivery, 130 solid oral dosage form, 129 tissue engineering, 135e136 topical drug delivery, 131e132 wound healing, 133e134

628

biochemistry, 123e124 biocompatibility, 122e123 chemical structure, 122e123, 123f factors carbon sources, 126 galactomannan, 126e127 high pressure, 126 pH, 125 polymer concentration, 126 salts, effect of, 126 temperature, 125 viscosity, 126e127 production, 127 kinetics, 128 media used, 127 properties, 125 structure of, 122 synergy, 207e208 tissue engineering, 540 water solubility, 122e123 X. arboricola, 122 X. axonopodis, 122 Xanthan membranes (XM), 135e136 Xanthomonas campestris, 123e124, 128, 139e140 Xanthomonas campestris NRRL B-1459, 122 XG. See Xanthan gum (XG) XG-chitosan combination (XG-CH), 135e136 X-ray diffraction (XRD), 189e190 Xylans, 1e2, 572f arabinoxylan, 573 glucuronoarabinoxylan, 573e574 xyloglucan, 574 Xylogalacturonan, 576 Xyloglucan, 574 Xylose, 1e2, 204e205

Y Young’s modulus, 478

Z Zinc oxide (ZnO), 482e483 Zinc sulfate, 269e270 Ziziphus mauritiana, 297