A study of vitamin B12 requirements for laying pullets and their progeny

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A STUB! OF VITAMHI B i* E^QUUiEMMTS FOE LAYING PULLETS AM) THEIR PROGENY

% John Lawrence M illig a n

T h e s is su b m itted t o th e F a c u lty o f th e Graduate S ch ool o f t h e U n iv e r s it y o f Maryland i n p a r t ia l f u l f i l l m e n t o f th e req u irem en ts f o r th e d eg ree o f D octor o f P h ilo so p h y I960

UMI Number: DP70493

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A c m o m jm m m s I w ish t o e x p r e s s agr a p p r e c i a t i o n t o D r, G. F . Combs f o r h i s v a l u a b l e a s s i s t a n c e and t i m e l y a d v ic e i n t h e s e t t i n g up o f t h i s s tu d y , I a® in d e b te d t o D r. M. A. J u l l f o r h i s co m p eten t c o u n s e l.

I w ish t o

th a n k Mr. C h a r le s D enton o f t h e B ureau o f Animal I n d u s t r y , U, S . D, A ,, B e l t s v i l l e , M ary lan d , f o r c o n d u c tin g t h e m ic r o b io lo g i c a l a n a ly s e s o f t h e e g g sam ples f o r v ita m in

I f e e l in d e b te d t o Mr, W arren S i l v e r f o r

a s s i s t a n c e i n ru n n in g t h e cup a s s a y s f o r v ita m in P o o le f o r o t h e r t e c h n i c a l h e l p .

an d t o Mr. C h a r le s

I am v e r y g r a t e f u l t o M iss J o s e p h in e

B l a i r , an u n d e r g ra d u a te s tu d e n t a t th e U n i v e r s i t y , f o r h e r g e n e ro u s a s s i s t a n c e i n c a r r y i n g o u t some o f t h e t e c h n i c a l f e a t u r e s o f t h e s tu d y . F i n a l l y , I am d e e p ly g r a t e f u l t o my w if e , P h y l l i s C. M illig a n , f o r th e en co u rag em en t and a s s i s t a n c e w hich sh e o f f e r e d th ro u g h o u t t h e c o u rs e o f th e re se a rc h . We a r e in d e b te d t o M erck and Company, I n c . , Bahway, Mew J e r s e y , f o r t h e e x p e r im e n ta l AFF su p p lem en t # 5 , v ita m in

8^

sodium c h l o r id e

t r i t u r a t e , an d o t h e r c r y s t a l l i n e v it a m in s ; S quibb and S o n s, Mew Y ork, f o r B nbram in; L e d e rle L a b o r a t o r i e s , P e a r l R iv e r , Mew Y ork, f o r s y n t h e t i c f o l i c a c i d ; A l l i e d M i l l s , I n c . , P o rts m o u th , V i r g i n i a , f o r so y b ean o i l ; A b b o tt L a b o r a t o r i e s , W orth C h ic a g o , I l l i n o i s , f o r H a liv e r o i l , and U* S . I n d u s t r i a l C h e m ic a ls , I n c . , f o r D L -m eth io n in e.

TABLE OF

contents

INTRODUCTION —------------------

Pm® i

R m m Of LITERATURE ■ ------

2

~~--------------- 12 EXPERIMENTAL PROCEDURE AMD R E S U L T S ---------------------A* S tu d ie s w ith L ay in g P u l l e t s - — — -----— ---------- — 12 B* S tu d ie s w ith C h ick s ---------------------87 DISCU3SI0K — --------------— ------------------------------------- ------------ -------— -----A* S tu d ie s w ith L ay in g P u l l e t s B* S tu d ie s on t h e P ro g en y — — ----- — —— —— — —---------

SB 68

06

BIBLIOGRAPHY --------------------- — ------------------------------------------------------------ 74 APPENDIX ------

82

LIST OF TABLES T a b le

Fag®

1

C o m p o sitio n o f P u l l e t B a s a l R a tio n s --------- —------------------

Z

B a s a l D i e t s and S u p p lem en ts Fed t h e P u l l e t G roups — ----

— 15 —

18

5

E x p e rim en t X*

4

E x p e rim e n t 1* E f f e c t o f D ie t Upon Egg P ro d u c tio n on H@n~Day B a s is — ------------------ ------— — — ----------- ------------- -------- —

19

5

E x p e rim en t I s

20

8

R a tio n s i&nployed i n Egg A ssay s w ith C h i c k s

7

R esponse o f C h ic k s i n Egg A ssay ( D ie t 5 ) —- — --------- ------------- 26

8

9

E f f e c t o f V itam in Bj# on P h y s ic a l C o n d itio n

F e r t i l i t y and H a te h a b ility

--------------- — — —

R esponse o f C h ic k s i n Egg A ssay ( D ie t 4 )

—

—

—-

25

27

V ita m in Bjjg A c t i v i t y P e r Egg a s I n f lu e n c e d by D ie t

29

E x p e rim e n t I t P e r c e n ta g e T ra n s m is sio n o f V itam in B i£ fro® P u l l e t D ie t t o Egg ------

51

11

E x p e rim e n t 2 % M o r t a l i t y and Egg P r o d u c tio n ——------- ------ ------

32

12

E x p erim en t 2 s

15

E x p e rim e n t Z i M a te rn a l D e p le tio n R a te a s Shown by P e rc e n ta g e H a t e h a b i l i t y o f F e r t i l e E g g s ----------------------------- 55

14

G ra d in g System f o r D egree o f F e a th e r in g — ■

15

L e v e ls o f S u p p le m e n ta l V ita m in B^ 2 Fed i n P rogeny S t u d i e s

16

E x p e rim en t I t I n i t i a l W eight o f C h ic k s a s I n f lu e n c e d By M a te rn a l D ie t ------ -------------------------- -— — ——

40

E x p e rim e n t I t P e r c e n ta g e L i v e a b i l i t y o f C h ick s a s In flu e n c e d By M a te rn a l D ie t ——-----•---- —

41

10

17

F e r t i l i t y and H a t e h a b i l i t y —

-------------- — —

—»■—

58 59

18

E x p e rim en t I t

19

E x p erim en t I t C hick W eight a t 6 Weeks Compared t o 4 Weeks E x p re s se d a s P e r c e n ta g e o f W eight o f £ C h ic k s i n Pen 4

47

E x p e rim en t I t R e l a t i v e E f f e c t i v e n e s s o f V itam in B1 2 F u rn is h e d t o C h ic k s By D ie t and v i a E g g -------------- — -------

48

20

D egree o f F e a th e r in g a s I n f lu e n c e d By D ie t —

55

45

T a b le

Page

£1

Experiment 1* R e la t iv e E f f e c t iv e n e s s o f V itam in B^g Fed in th e M aternal D ie t and i n th e Chick D ie t f o r Chick Growth — -----— -------------------— -------------------------------------------- 49

ZZ

Experim ent Zt

E f f e c t o f M aternal D ie t on I n i t i a l fcleight

£5

Experim ent Zt

P ercen ta g e o f C hicks A liv e a t 4 Weeks

£4

Experim ent Zt A Study o f th e M aternal D e p le tio n B ate a s Shown by F in a l Weight o f C hicks Fed Unsupplem snted B a sa l D ie t 5 — -------------------------------------------- ---------- — ---------

SI

55

LIST OF CHARTS Page Chart X

Standard Growth Curves f o r JSgg A ssays w ith 26

Chart 2

Experiment I s Vitam in Requirement o f C hicks f o r Growth a s A ffo o te d by M aternal D ieta ry 42

Chart 5

Experim ent 2* V itam in Requirement o f C hicks f o r Growth a s A ffe c te d by M aternal D ie ta r y SS

P im m

F ig u re 1

Experim ent I t E f f e c t o f T itam in Bxg In M aternal D ie t cm Chick Growth -

OTBDIHieTIOff The i s o l a t i o n a n d c r y s t a l l i s a t i o n o f v ita m in B^g b y B ic k e s , B r in k , SConiuasy, Wood and F o lk e r s {1948) and S m ith (1 9 4 8 a ) h a s opened a new f i e l d o f n u t r i t i o n a l re s e a r c h #

I t was soon, d e m o n s tra te d t h a t

t h i s v ita m in m s r e q u i r e d f o r t h e g ro w th o f c h ic k s #

I t m s a l s o found

t h a t t h e im provem ent i n g ro w th o b ta in e d by t h e a d d i t i o n o f a n im a l p ro ­ t e i n t o t h e c h ic k d i e t c o u ld

b e e x p la in e d i n l a r g e p a r t by it® c o n te n t

o f v ita m in B jg .

I t was s h o rn t h a t v ita m in B jg was r e q u ir e d

F u rth e rm o re

f o r h a te h a b ility . T h is s tu d y was u n d e r ta k e n t o d e te rm in e t h e v ita m in Bjg r e ­ q u ire m e n ts o f t h e p u l l e t an d

h e r p ro g e n y , an d t h e v ita m in Big c o n te n t

o f t h e egg a s a f f e c t e d b y p u l l e t d i e t .

a EJ5VISW o r LITERATURE H is to r y o f V itam in D u rin g W orld War I I , s i m p l i f i e d d i e t s d e v o id o f o r v e r y lo w i n a n im a l p r o t e i n s o u r c e s w ere f e d t o p o u l t r y from n e c e s s i t y .

P o o r g ro w th ,

p o o r v i a b i l i t y , and p o o r h a t e h a b i l i t y m arked t h e i r u s e , i n d i c a t i n g a de­ f i c i e n c y o f one o r more g ro w th f a c t o r s r e q u ir e d b y th e c h ic k and t h e p o u lt.

Hammond (1944 a an d b ) fo u n d d r ie d cow m anure and d r i e d rumen

c o n t e n t s s u p e r i o r t o a l f a l f a l e a f m eal f o r b o th th e c h ic k and th e p o u l t i n su ch a r a t i o n when t h e d i e t was a d e q u a te i n v ita m in A an d r i b o f l a v i n . F u r t h e r s t u d i e s on t h i s m a t e r i a l r e v e a le d t h e p re s e n c e o f an unknown g ro w th f a c t o r (R ubin and B ir d , 1946 a ) .

An e x c e l l e n t re v ie w o f t h e work

u p t o t h i s p o in t on t h e f a c t o r i n cow m anure was w r i t t e n by t h e e d i t o r s i n N u t r i t i o n R eview s, 1 9 4 6 .

In 1948 B ir d , M arsden an d K e llo g g r e p o r te d

a c r i t i c a l n e e d i n young t u r k e y s f o r t h e cow m anure f a c t o r . C o n c u rre n t r e s e a r c h i n o t h e r f i e l d s (S h o rb , 1947, C a ry , H artm an, B ryden and L i k e l y , 1 9 4 6 , e t c . ) beyond t h e scope o f t h i s p a p e r , r e v e a le d t h a t o t h e r a n im a ls and some m ic ro o rg a n ism s r e q u i r e d t h i s o r a s i m i l a r fa c to r.

C l i n i c a l work r e v e a le d t h e n eed f o r on© o r s e v e r a l unknown

f a c t o r s c o n ta in e d i n l i v e r f o r t h e r e l i e f o f A d d iso n ia n p e r n ic io u s anemia i n humans ( M inot and M urphy, 1 9 2 7 ).

W ith re m a rk a b le f o r e s i g h t C a s tle and

Ham (1 9 2 9 ) i n t e r p r e t e d r e s u l t s on s t u d i e s o f t h i s d is e a s e a s i n d i c a t i n g t h e e x i s t e n c e o f an i n t r i n s i c f a c t o r s e c r e t e d by c e r t a i n stom ach gland® , an d an e x t r i n s i c f a c t o r w hich h e d e s c r ib e d a s an e s s e n t i a l com plem entary f a c t o r fo u n d i n m e a t, e g g s , y e a s t , and l i v e r .

Hopes t h a t th e p r i n c i p a l

a n t i - p e r n i c i o u s anem ia f a c t o r c o n ta in e d i n t h e s e f e e d s tu f f ® had b een d is c o v e r e d w ith t h e I s o l a t i o n o f f o l i c a c id w ere q u ic k ly d ash ed by t h e

3 r e p o r ts o f B e t h e l l an d S t u r g i s (1 9 4 8 ) and B o ss, B o ld in g and P a e g e l ( 1 9 4 8 ). F i n a l l y , on A p ril. 1 6 , 1 9 4 8 , R ic k s s , B r in k , K oniuszy, Wood and F o lk e r s ( a ) r e p o r te d t h e i s o l a t i o n o f b i o l o g i c a l l y a c t i v e c r y s t a l l i n e m a t e r i a l from l i v e r c o n c e n t r a t e s c l i n i c a l l y a c t i v e i n t h e t r e a tm e n t o f p e r n i c i o u s a n e m ia .

S h o r t (1 9 4 8 ) c o n firm e d t h e a c t i v i t y o f t h i s m a t e r i a l ,

t e n t a t i v e l y named v ita m in

a s th e LLP f a c t o r f o r L a c to b a c illu s l a c t i s

D o m er, an d West (1 9 4 8 ) d e m o n s tra te d i t s a b i l i t y t o in d u c e f a v o r a b le b lo o d r e s p o n s e s i n p a t i e n t s s u f f e r in g from p e r n ic io u s a n e m ia .

On A p r i l

2 4 , 1 9 4 8 , Sm ith ( a ) in d e p e n d e n tly announced th e p u r i f i c a t i o n o f two am orphous fo rm s o f t h e a n t i - p e r n i c i o u s anem ia f a c t o r fro m l i v e r .

Sm ith

c i t e d u n p u b lis h e d c l i n i c a l s t u d i e s b y U ngley i n E ngland w hich demon­ s t r a t e d h i s m a t e r i a l t o b e e f f e c t i v e i n s ti m u l a t i n g b lo o d r e g e n e r a tio n an d t o be e f f e c t i v e a l s o i n t h e tr e a tm e n t o f t h r e e c a s e s o f s u b a c u te com­ b in e d d eg e n era tio n o f t h e s p i n a l c o rd a s s o c i a t e d w ith p e r n i c io u s an em ia. C h em ical and P h y s ic a l P r o p e r t i e s C r y s t a l l i n e v ita m in B^g i s a g a r n e t - r e d compound w hich c o n ta in s c o b a l t (S m ith , 1948 b , and R ic k e s , B rin k , K bnlussy, Wood an d F o l k e r s , 1948 b ) and PO, KH, and OH g ro u p s (R o b in so n , 1 9 4 8 ). t h e v ita m in y i e l d s S tS - d im o th y lb e n s iid d a s o le *

A cid h y d r o ly s i s o f

I t i s n o te w o rth y t h a t t h e

1 s 2 -d ia m ln o -4 85 -d ira e th y lb e n z e n e m o ie ty a p p e a r s i n t h e above d e g r a d a tio n p r o d u c t , i n v ita m in

i t s e l f , and i n r i b o f l a v i n , i n d i c a t i n g a p o s s i b l e

r e l a t i o n s h i p b etw een t h e two v ita m in s .

The v ita m in a p p e a r s t o b e a s i x -

g ro u p c o b a l t c o o r d i n a t io n com plex w ith a m o le c u la r w e ig h t o f 1 ,5 8 0 t o 1 ,7 5 0 .

A n a ly s is gave a c o m p o s itio n t y p i f i e d b y C q x -6 ^ 8 6 -9 2 $14 $15 P Co.

T h e re a r e s e v e r a l w eak ly b a s ic g r o u p s .

Upon a l k a l i f u s i o n , v ita m in Bx2

form ed p r o d u c ts which r e a c t w ith d h r l i e h *s r e a g e n t .

T h is and o t h e r

t e s t s a r e c o n s id e re d t y p i c a l o f c e r t a i n c y c l i c f i ve-memba red n i t r o g e n c o n ta in in g

com pounds

i n c lu d in g p y r r o le s *

The m e ltin g p o i n t was 500° C,

o r above.

The m ic r o b io lo g ic a l a c t i v i t y was n o t a f f e c t e d by a u io e la v in g

a t 1S1° C, f o r f i f t e e n m in u te s , b u t i t was i n a c t i v a t e d upon s ta n d in g a t room te m p e r a tu r e i n d i l u t e sodium h y d ro x id e o r h y d r o c h lo r ic a c i d {K ietees, B r in k , K o n iu s sy , Wood, and F o l k e r s , 1948 c , B rin k , W olf, K aczka, K ic k e s , K o n iu sz y , Wood, and F o l k e r s , 1 949, K aczka, W61f, and F o lk e r s , 1 9 4 9 , and B rin k and F o l k e r s , 1 9 4 9 ), When v ita m in Bjjg was c a t a l y t i c a l l y h y d ro g e n a te d , a p ro d u c t was o b ta in e d w hich was c a l l e d v ita m in

I t a p p e a re d t o have l e s s th a n

h a l f o f t h e b i o l o g i c a l a c t i v i t y o f v ita m in

f o r r a t s , c h ic k s , a n d

p e r n i c i o u s anem ia p a t i e n t s (K aczk a, W olf, and F o lk e r s , o£* c i t . ) . V itam in % 2 b

c r y s t a l l i z e d by P i e r c e , P ag e, S to k s ta d a n d

J u k e s (1 9 4 9 ) and showed s u b s t a n t i a l b i o l o g i c a l a c t i v i t y f o r c h ic k s and f o r L. le ic h m a n n ii 515.

F ric k ® , L an iu s, D o-R ose, L ab id u a and F r o s t

( 1 9 SO) com pared t h e p o te n c ie s o f v ita m in Bjjg and v ita m in B ^ b *>or r a t s a n d f o r humans s u f f e r i n g from A d d iso n ian p e r n ic i o u s anem ia and found th em e q u a l l y e f f e c t i v e .

They c o n c lu d e d t h a t t h e Z compounds wore

a p p a r e n t l y c h e m ic a lly a lm o st id e n t ic a l*

S to k s ta d , J u k e s , Brockman,

P i e r c e and B r o q u is t (1 9 5 0 ) fo u n d e q u a l a c t i v i t y f o r t h e Z compounds f o r t h e c h ic k an d L. l e lc h m a n n il. (1 9 5 0 ) found v ita m in

Sm ith, C u lb e r ts o n , W alker and L e e s,

v ita m in

and d e s o x y r ib o s id e s o f c y t o s i n e ,

g u a n in e , h yp oxan th in a, a d e n in e and thym ine on p a r t i t i o n ch ro m ato g rap h y o f t h e v ita m in Bqg g ro u p o f f a c to r ® .

They s t a t e d t h a t pur® vita m in Big

sam p les gave 2 , som etim es 5 , zon es o f grow th due to a r t i f a c t s p ro d u ced p re su m a b ly b y o x id a tio n on t h e p a p e r .

5 S m ith (1948 b , 1 949) r e p o r te d t h a t t h e c r y s t a l l i n e m a t e r i a l w as v e ry s o lu b l e i n w a te r , s o lu b le i n n e a r l y a n h y d ro u s a l c o h o l , and in g l a c i a l a c e t i c a c i d , b u t i n s o l u b l e i n e t h e r , c h lo ro fo rm , o r non—p o la r s o lv e n ts . R e la tio n s h ip w ith O ther Growth F a c to r s C r y s t a l l i n e v ita m in By> h a s been shown t o r e p la c e a l l o r n e a r l y a l l o f t h e a c i d p r e c i p i t a t e f a c t o r { th e cow m anure f a c t o r ) ( L i l l i e , D enton and B ir d , 1 948) an d t h e a n im a l p r o te in , f a c t o r f o r growth o f c h i c k s , ( O t t , l i c k e a and Wood, 1 9 4 8 ).

I t re p la c e s n u tr ie n t X f o r rat

g ro w th and n o rm a l r e p r o d u c tio n (H artm an, Dryden and C a ry , 1949 a ) .

It

r e p l a c e s a n t i- p e m ic lo u s anemia l i v e r e x t r a c t s f o r p ig grow th (Jo h n s o n a n d H etm n n , 1949) and i s e s s e n t i a l f o r grow th o f a l e e f e d th y r o p r o ts in (H u ff, 1949) a s a ws t r e s s * f a c t o r .

C h ild re n s u f f e r in g from m a ln u tr itio n

an d grow th f a i l u r e w ere f e d d a i l y o r a l d o s e s o f v ita m in

w ith r e s u l t ­

a n t heightened, a l e r t n e s s and v ig o r , and a resum ption o f n orm al g ro w th , i n d i c a t i n g t h a t i t i s a g ro w th f a c t o r f o r humans ( t f e t s e l , F a rg o , Smith a n d H ellk e o n , 1 9 4 9 ).

A lth o u g h no c o n f irm a tio n h a s b een p u b lis h e d con­

c e r n in g t h e i d e n t i t y o f v ita m in

w ith "aoopherln1*, th e grow th f a c t o r

f o r r a t s d e s c r ib e d b y S u c k e r and Zueker, 1948 ( a an d b ) , th e s o u rc e s and e f f e c t o f i t in d ic a te a v e ry c lo s e r e la tio n s h ip .

I t I s p ro b a b le t h a t

v ita m in Bps i s C a s t l e ’ s p o s t u l a t e d e x t r i n s i c f a c t o r m en tio n ed p r e v io u s ly an d t h a t a b s o r p t i o n o f i t i s a id e d by t h e p r e s e n c e o f t h e i n t r i n s i c f a c t o r o f g a s t r i c j u i c e (B e rk , C a s t l e , W elch, M eln le, A nker and E p s te i n , 1 9 4 8 , and T em berg and E a k in , 1 9 4 9 ).

A lth o u g h v ita m in Bu> a cco u n ts f o r

a t l e a s t a m a jo r p o r t i o n o f t h e a c t i v i t y o f t h e a n iim l p r o t e i n f a c t o r , many groups o f w o rk e rs h av e p r e s e n te d e v id e n c e on t h e m u l t i p l e n a tu r e

6 o f th e l a t t e r .

C rav en s (1 9 5 0 ) h a s g iv e n an e x c e l l e n t re v ie w o f t h e w ork

on. t h i s q u e s ti o n . R e s u lts w ith m ic ro o rg a n ism s i n d i c a t e a c o m p lic a te d i n t e r r e l a ­ t i o n s h i p b etw een d e so x y rlb o si.d e© , re d u c in g a g e n ts and v ita m in B y*.

A

summary o f much o f t h e work i n t h i s f i e l d , w hich i s beyond th e sco p e c£ t h e p r e s e n t p a p e r , i s in c lu d e d i n a re v ie w by Hood3 , 1 9 4 9 .

S tu d ie s on

t h e d e s o x y r ib o s id e s h av e i n d i c a t e d s l i g h t i f an y a c t i v i t y on t h e i r p a r t a s s u b s t i t u t e s f o r v ita m in

f o r h ig h e r a n im a ls ( E e i s n e r and H a s t,

1 9 1 9 , an d L ew is, R e g i s t e r , Thompson an d S lv eh jem , 1 9 4 9 ). Some o f t h e v ita m in s seem t o b e a r a more d i r e c t r e l a t i o n s h i p w ith t h e l a t t e r v ita m in .

H igh l e v e l s o f c a lc iu m p a n to th e n a te and

p y rid o x in © h y d r o c h lo r id e f e d t© chick® ( B ird and R u b in , 1946) and h ig h l e v e l s o f r i b o f l a v i n f e d t o r a t s i n th e p re s e n c e o f s u lf& s u x ititn e ( H artm an, D ryden and C a ry , 1949 b ) seemed t o p a r t i a l l y o f f s e t a d e f i c i e n c y o f v ita m in

The r e l a t i o n s h i p ©f v ita m in Bi£ t o c h o lin e and m e th io n in e

w i l l b e d is c u s s e d i n t h e s e c t io n on th e g e n e r a l f u n c t io n s o f th e v ita m in . V itam in

d e s c r ib e d b y Novak and Kauge ( 1 9 4 8 ), i s a f a t -

so lu b le - s u b s ta n c e c l e a r l y d i f f e r e n t from v ita m in Byg.

V itam in B l4 ,

d e s c r i b e d fey M o rris and M ajn& rioh (1 9 4 9 ), w h ile a p p a r e n tly c l o s e l y r e l a t e d t o b o th f o l i c a c i d and v ita m in e n t i r e l y s e p a ra te e n t i t y .

i n f u n c t io n , i s a ls o a p p a r e n tly an

I t i s e x tre m e ly a c t i v e i n c e l l p r o l i f e r a t i o n

I n b o n e m arrow c u l t u r e s and h a s b e e n e f f e c t i v e i n c u r i n g e x p e r im e n ta lly in d u c e d an em ia o f r a t s .

The a u th o r s b e l i e v e t h a t i t , n o t v ita m in B jg ,

i s C a s t l e 's p o s t u l a t e d e x t r i n s i c f a c t o r . I t i s p o s s i b l e t h a t a n t i b i o t i c s may s p a re t h e l e v e l o f vita m in ®12 w hich m ust be s u p p lie d i n th e f e e d f o r th e c h ic k , by c r e a t i n g f a v o r ­ a b l e c o n d i t i o n s f o r a h ig h rat® o f i n t e s t i n a l s y n t h e s i s (S m ith arid

7 R o b in so n , 1945, and M cG innis, S te p h e n so n , L e v a d ie , C a r v e r , G a r i b a l d i , I j i c h i , S n a i l and L ew is, 1 9 4 9 ), b u t i t i s im p ro b a b le t h a t th e a c t u a l p h y s i o l o g i c a l re q u ire m e n t l o r t h e c h ic k i s r e d u c e d . F u n c tio n s - G e n e ra l V itam in B jg f u n c t i o n s i n t h e e f f i c i e n t u t i l i s a t i o n o f p r o t e i n a s shown by t h e i n t e n s i f i c a t i o n o f th e v ita m in d e f i c i e n c y when u n u s u a lly h ig h l e v e l s o f v e g e ta b le p r o t e i n s a r e f e d t o c h ic k s o r r a t s ( Rubin and B i r d , 1947 a , an d H artm an, Drydon an d C ary , 1949 a ) . The l e v e l o f n u c l e o p r o t s i n i n l i v e r b a s o p ia ilia o f dogs i s c o o r d in a te d w ith t h e l e v e l o f v ita m in B ^g.

C o n se q u e n tly i t i s b e lie v e d

a d e f i c i e n c y o f t h e l a t t e r r e t a r d s p r o t e i n s y n t h e s i s {S t e r n , T a y lo r , and R u s s e l l , 1 9 4 3 ).

T h is may be t h e f u n c t io n o f t h e v ita m in i n p r e v e n tin g

p e r n i c i o u s a n e m ia .

V itam in Hjjg may b e n e c e s s a r y t o c o n v e r t t h e th y m in e

(p ro d u c e d a s a r e s u l t o f c a t a l y t i c a c t i o n by f o l i c a c i d ) i n t o th y m id in e w h ich i s n eed e d f o r t h e m a n u fa c tu re o f n u c l e i c a c i d f o r new c e l l s , such a s r e d b lo o d c e l l s an d r e t i c u l o c y t e s (W rig h t, S kegge, and H u ff, .1348). The r e p o r t o f P o p p e r, Kochr-tfeeer and Ss&nt© (1 3 4 9 ) a l s o s u g g e s te d a p o s s i b l e r o l e o f v ita m in B ig i n t h e s y n th e s i s o f r ib o n u c l e ic a c i d i n t h e l i v e r o f th e r e t .

Hep o r t s i n d i c a t e an i n t e r p l a y b etw een c h o l i n e , d e t a i n s ,

m e th io n in e and v ita m in

i n w hich t h e v ita m in can re d u c e th e r e q u ire m e n t

f o r t h e o t h e r f a c t o r s f o r t h e c h ic k ( G l l l i s and M o r r is , 1343, B c h a e fs r, S alm on, and S t r e n g t h , 1 9 4 9 ). A c co rd in g t o J&asrson (1 9 4 9 ) and N ic k e l, D i e t r i c h , C ravena and b lv e h je m (1 9 4 9 ) v ita m in B ig can c o u n te r a c t a l l t h y r o t e x i c i t y , th u s r e ­ p l a c i n g l i v e r an d l i v e r e x t r a c t s f o r t h a t p u rp o s e .

T e s t a n im a ls u sed

w e re , r e s p e c t i v e l y , r a t s and c h ic k s f o r th e two s t u d i e s .

8 F u n c tio n s - In P o u ltr y k fh its o n , T i t u s and B ir d (1 9 4 6 ) rep orted t h a t t h e f e e d in g o f t h e cow m anure f a c t o r , i n a d i e t h ig h i n soybean m eal and c o n ta in in g no a n im a l p r o t e i n , s i g n i f i c a n t l y im proved h a t e h a b i l i t y .

T h e re was no i n ­

f lu e n c e on hen body w e ig h t, ©gg p ro d u ctio n , o r egg s is © .

Rubin and B ir d

(1946 a ) r e p o r t e d t h e cow m anure f a c t o r n e c e s s a r y f o r c h ic k g ro w th .

In

194? ( b ) , t h e y r e p o r te d t h a t t h e f a c t o r was t r a n s m i t t e d th ro u g h t h e ©gg t o th e c h ic k .

They showed t h a t t h e c o n te n t o f t h e f a c t o r i n t h e ©gg®

w as d i r e c t l y ' r e l a t e d t o t h e l e v e l o f t h e f a c t o r i n t h e h e n ’s d i e t .

B ird

(1 9 4 6 ) showed t h a t c h ic k s p ro d u ced on d i e t s d e v o id o f t h e cow manure f a c t o r ( v ita m in

) s u f f e r e d more th a n tw e n ty p e r c e n t g r e a t e r m o r t a l i t y

d u r in g t h e f i r s t week o f l i f e th a n u n d e p le te d c h ic k s d i d . and S lv e h je a (1 9 4 9 ), p r e s e n te d e v id e n c e t h a t v ita m in i n f l u e n c e on h em oglobin r e g e n e r a tio n i n c h ic k e n s .

N ic h o l, H a rp e r doe® have some

Chick© f e d on a f o l i c

a c i d - d e f i c i e n t d i e t and i n j e c t e d w ith p h en y lh y d raain © h y d r o c h lo r id e w ere t r e a t e d w ith f o l i c a c i d i n j e c t i o n s a l o n e , w hich r e s t o r e d hem oglobin v a lu e s t o n o rm a l.

V ita m in

i n j e c t i o n o f v ita m in tio n .

i n j e c t i o n s a lo n e gave no b e n e f i t , b u t a com bined and f o l i c a c i d p ro d u ced t h e m ost r a p id r e g e n e ra ­

The e f f e c t c o u ld e a s i l y be an i n d i r e c t a c t i o n through im proved

u t i l i s a t i o n o f f o lic a c id . L i l l i e , Denton an d B ird (1948). r e p o r te d b e t t e r f e a t h e r i n g o f c h ic k e n s when t h e y w ere s u p p lie d w ith v ita m in S j ^ .

L i l l i e , K & rsdan,

G ro sch k e and B ir d (1 9 4 9 ) fo u n d t h a t th e a p p a r e n t re q u ire m e n t o f th e g ro w in g c h ic k f o r v ita m in 3^2 d ro p p ed d u rin g th e l a t e r stage® o f g ro w th . Robbie© , l i c h o l , C ra v e n s, E lv eh jem and H a lp in (1 9 4 9 ) c i t e d work w hich i n ­ d i c a t e d t h a t even i f t h e m a te r n a l d i e t c o n ta in e d su p p o se d ly a d e q u a te

l e w i s o f v ita m in B-ys, t h e p ro g en y s t i l l r e q u ir e d s u p p le m e n ta l v ita m in B]j> i n t h e i r d i e t t o 4 w eek s. H ic h o l, D i e t r i c h , C rav en s and K lv e h jm (1 9 4 9 ) p r e s e n te d d a ta i n d i c a t i n g t h a t 15 ralcro g ram s o f c r y s t a l l i n e v ita m in

p e r k ilo g ra m

o f d i e t a p p e a re d t o be t h e iainlrau&k e f f e c t i v e d o se when h y p e r th y r o id c h ic k s w ere u s e d , a lth o u g h t h e e x a c t l e v e l w as n o t d e te r m in e d ,

J to k s t a d ,

d u k e s , F i e r c e , Page .and F r a n k li n (1 9 4 9 ) fo u n d t h a t a minimum o f 15 m icro * gram s o f c r y s t a l l i n e v ita m in By# p e r k ilo g ra m o f d i e t ( when f e d i n d i e t s c o n t a i n i n g s e v e n ty p e r c e n t o f soybean m eal t o d e p le te d $©w H am pshire c h ic k s ) m s a s a c t i v e in p ro m o tin g grow th a s f i s h s o lu b l e s and l i v e r ex­ tra c ts . O tt (1 9 4 9 ) h a s r e p o r te d t h a t d e p le te d c h ic k s u n d e r h i s con­ d i t i o n s r e q u ir e d a t l e a s t 27 laicro g ram s o f v ita m in B j# p e r k ilo g ra m o f r a t i o n f o r maximum g ro w th . I f t h e r e i s a n y n eed f o r v ita m in 3«y? f o r egg p r o d u c tio n , t h e re q u ire m e n t m is t b e low an d a t p r e s e n t i s unknown.

The q u a n t i t a t i v e r e ­

q u ire m e n ts o f t h e hen f o r t h e v ita m in f o r n o rm al h a t e h a b i l i t y , f o r v i a b l e o f f s p r i n g and f o r good g ro w th o f p ro g en y f e d v a r io u s l e v e l s o f v ita m in Bxa have n o t b een c l e a r l y d e f in e d . r e l a t i o n s h i p t o t h e v ita m in

F u rth e rm o re t h e r e i s a g e n e tic & l

re q u ire m e n t o f t h e c h ic k e n , w hich p e r m its

some b i r d s t o e x i s t s a t i s f a c t o r i l y on l e v e l s q u i t e d e f i c i e n t f o r o t h e r s , ( B i r d , R ubin arid G ro sc h k e , 1 9 1 7 ). A ssay M ethods b o th m i c r o b i o l o g i c a l and m a c r o b io lo g ic a l te c h n iq u e s have been em ployed f o r v ita m in B12 a s s a y s .

The f i r s t m ic r o b io lo g ic a l m ethod, t h e

LID a s s a y o f Shorb (1947 b ) p ro v ed t o be d i f f i c u l t t o u s e b e c a u s e o f th e

10 te n d e n c y o f t h e s p e c i e s t o d i s a s s o c i a t e .

I n t h e LL'D a s s a y m ethod d i s ­

c u s s e d by G ree n e, B rook, and McCormack i n 1 9 4 9 , th e oxygen t e n s i o n i s c o n t r o l l e d , s in c e i t a f f e c t s t h e a s s a y . S e v e r a l a s s a y m ethods em ploy L . le ic h m a n n il (H offm ann, S to k s ta d , H u tc h in g s , O om bush and J u k e s , 1 9 4 9 , P e e l e r , Y acow its an d M o rr is , 1 9 4 9 , and o t h e r s ) .

A l l o f t h e s e a s s a y s have b e e n c o m p lic a te d by t h e a b i l i t y

o f th y m id in e ( S h iv e , R a v e l, end H a rd in g , 1948) and o t h e r d e s o x y r ib o s id e s ( K i ta y , McNutt an d S n e l l , 1949) t o r e p la c e o r s p a r e v ita m in Big u n d e r a ssa y c o n d itio n s . F o s t e r , t a l l y and W oodruff (1 9 4 9 ) have o u t l i n e d a cup a s s a y f o r v ita m in

w hich a v o id s some o f th e d i f f i c u l t i e s o f s ta n d a r d t u r b i -

d lm e tric o r t i t r i m e t r i c p ro c e d u re s.

The d i f f u s e grow th re s p o n s e o f t h e

t e s t c u l t u r e t o d e s o x y r ib o n u c le ic a c i d o r t o I t s c o n s t i t u e n t n u c le o s id e s i s e lim in a te d . The u s e o f t h e p ro to z o a n ik tg le n a g r a c i l u s lias b een s u g g e s te d f o r an a s s a y s in c e i t a p p a r e n t l y g iv e s a q u a n t i t a t i v e re s p o n s e t o v ita m in B ig b u t i s i n a c t i v e t o th y m id in e (H u tn e r, P r o v a s o l i , B to k s ta d , H offm ann, B e l t , F r a n k lin an d J u k e s , 1 9 4 9 ). M & erob iologieal te c h n iq u e s in v o lv in g t h e r a t an d t h e c h ic k w ere a l r e a d y i n u s e f o r t h e v ita m in when i t was i s o l a t e d (H artm an, D ryden, an d L i k e l y , 1946, and B ir d , Rubin and G ro seh k e, 1 9 4 8 , r e s p e c t i v e l y ) .

L a te r

(1 9 4 9 ) B o s s h a rd t, P a u l, 0*D o h e rty , H u ff an d B a rn e s em ployed a mouse assay.

A ll o f t h e s e m ethods in v o lv e d a p r e l im in a r y m a te rn a l d e p l e t i o n

p e r io d p r i o r t o u s e o f th e p ro g e n y f o r a s s a y s .

The above mouse a s s a y ,

some r a t a s s a y s ( F r o s t , F id c k e , and S p r u th , 1 949, R e g i s t e r , Ruegauer, an d i& vehjem , 1949 ) and some c h ic k a s s a y te c h n iq u e s h av e u t i l i z e d i o d i n a t e d c a s e i n a s a *s t r e s s * m echanism t o i n c r e a s e th e s e n s i t i v i t y o f

11 t h e a n im a l t o a d e f i c i e n c y o f t h i s v i ta m in .

The c h ic k a s s a y m ethod

d e s c rib e d , by B ir d , B abin and G ro sch k e (1 9 4 8 ) u t i l i z e s a m a te r n a l de­ p l e t i o n p e r io d and a Z week p r e lim in a r y f e e d in g p e r io d o f a v e r y h ig h l e v e l o f soybean m eal t o t h e p ro g e n y , fo llo w e d by a 2 week t e s t p e r i o d . G rowth i s t h e c r i t e r i o n i n a l l o f t h e s e a s s a y s .

Ko s i n g l e a s s a y , m icro ­

b i o l o g i c a l o r m a c r o b io lo g ic a l, i s s a t i s f a c t o r y i n e v e r y r e s p e c t .

S o u rc e s

D rie d cow m an u re, f i s h m e a l, m eat m e a l, m eat s c r a p s , co n d en sed f i s h s o lu b l e s and o t h e r a n im a l p r o t e i n su p p lem en ts h a v e been used, a s v a l u a b l e s o u r c e s o f v ita m in % g»

I n d i c a t i o n s t h a t th e d i e t a r y h i s t o r y

o f n o n -ru m in a n ts may h av e c o n s id e r a b le e f f e c t on t h e v a lu e o f t h e i r f l e s h a s a s o u rc e o f v ita m in Bx£ h av e been o b ta in e d b y l e g i s t o r , Lewi®, Thompson an d llv o h je m ( 1 9 4 9 ) .

S te p h e n so n , M cG innis, Graham and C a rv e r

(1 9 4 8 ) p r e s e n te d e v id e n c e f u r t h e r s u b s t a n t i a t e d by Cunha, B u r n s id e , Suschm an, G la s s c o c k , P e a rso n an d S h e a ly (1 9 4 9 ) t h a t s o i l s tim u la te d g ro w th when added t o v ita m in Big d e f i c i e n t d i e t s . C e r e a l g r a i n s a n d v e g e ta b le p r o t e i n su p p le m e n ts a r e q u i t e d e f i c i e n t in v ita m in B ig ( W h itso n , Hammond, T i t u s and B ir d , 1 9 4 b , and M cG innis a n d C a r v e r , 1 9 4 7 ). V a rio u s e o u s a e rc ia l c ru d e c o n c e n tr a te s o f v ita m in B\Z p ro d u ced fro m m i c r o b i a l f e r m e n ta tio n have a l s o become a v a i l a b l e ( S to k s ta d , P ag e, P i e r c e , J u k e s , H e in le , B p s te in an d W elsh, 1948, A n sb a c k e r, H i l l , Tlem&n, Downing and C a ld w e ll, 1 9 4 9 , and K ic k e s , B rin k , K o n iu szy , Wood an d F o l k e r s , 1948 c ) .

SXPERIHKNTAL PHOCSDURE AND RESULTS A.

S tu d ie s w ith L aying P u l l e t s

1.

G e n e ra l p r o c e d u r e .

The s t u d i e s w ith la y in g p u l l e t s in v o lv

a p r e l im i n a r y p e r io d and two e x p e r im e n ta l p e r i o d s .

D uring t h e p r e lim in a r y

p e r i o d r e c o r d s o f h a t e h a b i l i t y w ere o b ta in e d w hich s e rv e d a© a b a s i s f o r f o r m a tio n o f e q u a liz e d g ro u p s f o r t h e l a t e r work* d u c tio n an d f e r t i l i t y w ere a l s o o b t a in e d .

R ecords on egg p ro ­

B a s a l d i e t 1 , shown i n t a b l e

1 , was u se d t o s y s t e m a t i c a l l y d e p l e t e t h e p u l l e t s o f an y s t o r e s o f v ita m in B ig , d u r in g t h i s p e r i o d .

D uration® o f p e r io d s w ere A ugust 2 7 , 1949 t o

November 8 , 1 9 4 9 , November 8 , 1949 t o A p r il 8 , 1980 a n d A p r il 8 , 1950 t o d u l y 2 7 , 1 9 5 0 , r e s p e c t i v e l y , f o r t h e p r e lim in a r y p e r i o d , e x p e rim e n t 1 and e x p e rim e n t 2 .

S ix ty New H am pshire p u l l e t s , 22 w eeks o f a g e , wore h o u sed

i n i n d i v i d u a l l a y i n g cage® a t t h e s t a r t o f t h e p r e lim in a r y p e r i o d . h a d b een r e a r e d on c o n v e n tio n a l s t a r t i n g and gro w in g r a t i o n s .

They

S e le c tio n ®

w ere made p r i m a r i l y on t h e b a s i s o f s e x u a l m a t u r i t y , a lth o u g h c o n s i d e r a t i o n was a l s o g iv e n t o body w e ig h t an d o t h e r q u a l i f i c a t i o n s • Mew Hampshire male® w ere h o u sed i n b a t t e r i e s . same d i e t a s t h e p u l l e t s .

At t h e same tim e

They h ad b een r e a r e d on t h e

P u l l e t s fo u n d t o be u n s a t i s f a c t o r y d u r in g t h e

p r e l im i n a r y p e r io d w ere r e p la c e d by o th er® h e ld i n r e s e r v e on f r e s h l i t t e r i n l a y i n g p e n s u n d e r o th e r w is e s i m i l a r c o n d i t i o n s .

A f t e r te r m in a t io n o f

t h e p r e l im i n a r y p e r io d no f u r t h e r re p la c e m e n ts w ere m ade.

The p u l l e t s

w ere a r t i f i c a l l y in sem in a ted w ith p o o le d semen tw ic e w eekly th ro u g h o u t t h e e n t i r e s tu d y . in c u b a t e d .

Eggs w ere c o l l e c t e d d a i l y and s to r e d a t 50° F . u n t i l

S e t t i n g s w ere made a t tw o-w eek i n t e r v a l s .

D u rin g t h e p r e ­

l i m i n a r y p e r io d and f i r s t 4 weeks o f e x p e rim e n t 1 , th e e g g s w ere c a n d le d on t h e s i x t e e n t h d ay o f i n c u b a t i o n .

T h e r e a f t e r t h e y w ere c a n d le d on t h e

13

TABL3 1 C o m p o sitio n o f P u l l e t B a sa l R ation©

B asal D ie t 1 *

B asal D ie t 2 %

4 4 .9 7 5 5 0 .0 0 0 1 8 .0 0 0 —■ — —5*000 5 .0 0 0 0 .5 0 0 0 .0 2 5 0 .5 0 0

4 6 .9 7 5 5 0 .0 0 0 7 .0 0 0 4 .0 0 0 4 . OCX) 1 .0 0 0 5 .0 0 0 5 .0 0 0 0.5QQ 0 .0 2 5 0 .5 0 0

1 0 0 .0 0 0

1 0 0 .0 0 0

S u p p le m e n ta l i n g r e d i e n t s

m g ./ l b .

ia g ./lb .

C alcium p a n to th e n a te R ib o f la v in M enadione C h o lin e c h l o r i d e

0 .7 5 1 .0 0 0 .2 0 1 5 0 .0 0

0 .7 5 1 .0 0 0 .2 0 1 5 0 .0 0

In g re d ie n ts

Ground y e llo w c o m G round w heat iS x p e lle r so y b ean s e a l D r ie d b re w e rs y e a s t Menhaden f i s h m eal U n e x tr a c te d l i v e r m eal G round lim e s to n e D e f lu o r in a te d rock p h o s p h a te I o d is e d s a l t MnS04.4 HgO A & D f e e d in g o i l (2 2 5 0 A,400 D) T o ta l

A n a ly s is C rude p r o t e i n b y a s s a y M e th io n in e C y s tin e C h o lin e

1 4 .9 0 $ 0 .2 6 $ 0 .2 5 $ 6 0 9 .0 0 m g ./ l b .

1 5 .1 0 $ 0 .5 3 $ 0 .2 6 $ 6 3 7 .5 0 m g ./ l b .

14 e i g h t h d ay t o avoid. e x c e s s iv e f r a g i l i t y o f t h e v i t e l l i n e membrane*

T h is

d i f f i c u l t y a r o s e i n a l l t h e eggs p ro d u ced on d i e t 1 d e s c r ib e d i n t a b l e 1 , r e g a r d l e s s o f s u p p le m e n ta tio n w ith v ita m in

O b s e rv a tio n s by O lsen a t

B e l t s v i l l e , M ary lan d , i n d i c a t e t h a t a te n d e n c y to w a rd s f r a g i l i t y o f th e membrane o c c u r s when th e e g g s ar® p ro d u ced on soybean m eal a s th e s o le p r o t e i n su pplem ent*

A ll a p p a r e n t ly i n f e r t i l e eg g s w ere b ro k en o u t t o

a c c u r a t e l y d a te m i n e f e r t i l i t y .

The few eg g s w hich w ere p ip p e d b u t w hich

f a i l e d t o h a tc h by t h e tw e n ty -s e c o n d d ay w ere in c lu d e d a s dead g e m s . The chick® w h ich h a tc h e d w ere u s e d i n experim ent® t o b e d e s c r ib e d l a t e r * O b s e rv a tio n s w ere made on t h e p h y s ic a l c o n d itio n o f t h e p u l l e t s d u r in g th e t h r e e p e r io d s and on t h e i r r e p r o d u c tiv e perfo rm an ce*

B ir d s

w ore w eighed f i v e ti m e s , a s i n d i c a t e d i n t a b l e 5 , d u r in g th e p r e lim in a r y p e r i o d and e x p e rim e n t 1 .

H em oglobin v a lu e s w ere o b ta in e d d u rin g e x p e r i ­

m ent 1 b y us© o f t h e m ethod d e s c r ib e d b y E velyn ( 1 9 5 6 ).

M o r t a l i t y , egg

p r o d u c t io n , f e r t i l i t y and h a t e h a b i l i t y d a ta w ere r e c o rd e d d u r in g a l l th r e e p e rio d s . At t h e end o f t h e p r e lim in a r y p e r io d th© p u lle t© w ere d iv id e d i n t o f i v e e x p e r im e n ta l g r o u p s . A, B, 0 , B and E.

G ro u p in g s w ere mads

p r i n c i p a l l y on t h e b a s i s o f e q u a lis e d h a t e h a b i l i t y .

The g ro u p s w ere l e f t

u n ch an g ed e x c e p t b y n a t u r a l c a u s e s f o r b o th e x p e rim e n ts* F eed and w a te r war® s u p p lie d a d l i b i t u m *

The c o m p o sitio n o f

t h e b a s a l d i e t s u s e d f o r t h e p u l l e t s t u d i e s i s l i s t e d in t a b l e 1 .

B oth

b a s a l d i e t s c o n ta in ad d ed c h o lin e and on t h e b a s i s o f t h e l i m i t e d d a ta a v a i l a b l e on re q u irem e n t® o f m a tu re s to c k , a r e b e lie v e d to b e a d e q u a te i n m e th io n in e and c h o l i n e .

B a s a l d i e t 1 was f e d w ith o u t s u p p le m e n ta tio n

t o a l l p u l l e t s d u r in g t h e p r e lim in a r y p e r i o d .

I t was an a ll- m a s h b r e e d in g

u r a t io n composed o f p r a c t i c a l f o o d s t u f f s and m s c o n s id e r e d t o be- ad equ ate i n a l l knew*, n u t r i e n t s , ex c e p t v ita m in B j£ .

The vita m in B£g c o n te n t o f

such a d i e t \ m j b e assum ed on th e b a s i s o f u n i v e r s a l e x p e rie n c e t o b e a t b e s t v ery lo w .

T h e re a r e no r e l i a b l e a s s a y s f o r t h e v ita m in Bjyg. © onteni

o f i t s in g re d ie n ts .

B a sa l d i e t 2 in c lu d e s n a tu r a l so u r ces o f v ita m in %g

and. u n id e n t if ie d n u t r i t i o n a l f a c t o r s . p o t e n c ie s o f l i v e r m eal and f i s h and B ird ( 1 9 4 9 ).

Average f ig u r e s f o r the v ita m in m eal were o b ta in e d by l i l l i a , B enton,

A ccording t o t h e ir r e s u lt s w ith c h ic k a s s a y s t h e s e -feed**

s t u f f s m y fu r n is h a t o t a l o f about IB ^icrogm i& s o f v ita m in gm m o f b a s a l d i e t 2 .

p er k i lo *

V a r i a b ili t y i n th e r e s u l t s o f in d iv id u a l a s s a y s by

t h e s e w orkers in d ic a t e t h a t th«s©e v a lu e s mm o n ly a p p ro x im a tio n s. The m X m wera f e d a n o x m l b reed in g r a tio n iu c iu u in g so u r c e s o f v ita m in Bjjg throu ghou t th e s tu d y .

During b oth ex p erim en ta l p e r io d s ,

b a s a l d i e t 1 was fe d t o groups A, B , € and b , w h ile b a s a l d i e t 2 was fed t o group &»

C on seq u en tly group & fu n c tio n e d a s th # p o s it iv e

c o n tr o l*

T&fel# £ p r e s e n t s th© v a r ia t io n s i n supplem ents f o r th e d i f f e r e n t g ro u p s of

p u l l e t s d u r in g th e t h r e e p e r io d s .

The so u rce o f sup p lem en tal v ita m in

f o r a l l g ro u p s e x c e p t & was iftibram in, m a n u fa c tu re d b y Bqulbb an d S o n s. The l a t t e r p ro d u c t m® a h ig h ly p u r i f i e d p r o te ln - f r # © s o l u t i o n o f v ita m in B^g d e r iv e d fro m S treg to so rees M rlseu s.

A cco rd in g t o dster& aination w ith

L a c t o b a c illu s le f c iu a a n n ii by t h e m a n u f a c tu re r , i t c o n ta in e d So a l c r o g r a s s o f v ita m in B^g p e r c u b ic c e n t l n s t e r and m s added t o b a s a l d i e t 1 a s a h ig h ly d i l u t e a q u eo u s s o l u t i o n .

D uring sacperlwsxvt 2 g ro u p J£ m s f e d

b a s a l d i e t 2 supplem ented w ith Kerck an d Co. ex p erim en ta l APF c o n c e n t rut© #S.

A c c o rd in g t o th^i m an u factu rer, t h i s c o n c e n tr a te c o n ta in e d 27.B

m illig r a m s o f v ita m in B^£ p e r k ilo g ra m when m easu red w ith t h e LL0 a ssa y *

16 n e i t h e r one o f t h e v ita m in B ig c o n c e n tr a te s was f o r t i f i e d w ith an y a n tib io tic * TABUS Z B a s a l D ie ts and S upplem ents Fed t h e P u l l e t Groups

G ro u p

B asal D io t

A

1

B

1

C

1

D

1 2*

B

S u p p lem en ts p e r K ilogram P r e lim in a r y E x p e rim en t E xperim ent P e r io d 1 Z

4

— — *****

—

3 meg. o f

meg. Of

4 m eg. o f BlZ

a »C«. o f %2

—'—

16 meg. o f Bl2

--- *

mm+mmm

0 .1 $ APF C o n c e n tr a te #S

* T h is g ro u p f e d b a s a l d i e t 1 d u r in g p r e lim in a r y p e r io d . F o r t h e sa k e o f b r e v i t y th e p h r a s e »l e v e l o f s u p p le m e n ta l v ita m in Bf£ p e r k ilo g ra m o f fe e d " w i l l b® re p la c e d , i n m ost i n s t a n c e s b y wl©v®l o f v ita m in B ig f e d " , o r a s i m i l a r p h r a s e , th ro u g h o u t t h e r e ­ m a in d e r o f t h e p a p e r .

I t w i l l b e u n d e r s to o d t h a t t h e a c t u a l t o t a l l e v e l

o f v ita m in B ig f e d w ould a l s o in c lu d e t h e c o n te n t i n t h e b a s a l d i e t s . 2.

R e s u lt s o f p u l l e t e x p e rim e n t J*.

E x p erim en t 1 was u n d e r ­

ta k e n t o s tu d y t h e e f f e c t o f g ra d e d l e v e l s o f v ita m in B1 2 on th e p h y s ic a l c o n d i t i o n and on t h e r e p r o d u c tio n o f la y i n g p u l l e t s .

T a b le 3 p r e s e n t s

t h e d a t a o b ta in e d d u r in g t h e f i r s t e x p e rim e n t on t h e p h y s ic a l c o n d itio n o f th e p u lle ts .

TABLE 8 Experim ent 1:

Group

E f f e c t o f Vitam in

Average P u lle t Weight in Kilograms August Hovenber December February 24,1949 2 7 , 1949 5 , 1949 I S , 1950

on P h y sic a l C o n d itio n o f P u l le ts

Ghu Hemoglobin p e r 1QQ e c . o f Blood A p ril 8 , 1950

ftem nber S , 1949

A p ril 8 , 1950

i M ortality Hov. 5 , 1949 to A p ril 8 , 1950

A

2 ,5 2 8 (7 }*

2.542

2.698

2.752

2.728

8 .1 0

8 .8 5

16.7

B

2 .2 6 1 (8 )

2.458

2.719

2.887

2.728

8 .1 ?

8.45

0 .0

€

2 .2 1 2 (5 )

2 .4 4 0

2.685

2.665

2.577

8.59

8 .7 0

8 .5

D

2 .2 2 8 (9 )

2.456

2.694

2.798

2.605

8.19

8.84

0 .0

S

2 .1 4 7 (8 )

2 .4 5 0

2.624

2.967

2.792

8 .6 0

9 .0 0

0 .0

# dumber i n p a r e n th e s is i n d i c a t e s number o f s u rv iv in g b ir d s from p re lim in a ry p erio d which were used f o r th e ex p e rim e n ta l p e r io d s , Bemaining b ir d s in th e groups f o r th e l a t t e r 2 p e rio d s were re ­ placem ents f o r whom no w eights were a v a ila b le f o r th e d a te o f August 27, 1949, Each group co n ta in e d 12 bird® a t th e b eg in n in g o f experim ent I ,

IB Body W eight o f P u l l e t s D u rin g t h e e a r l y m onths

experim ent 1 a v e ra g e w e ig h ts in c r e a s e d g e n e r a l l y i n o f t h e t o s t , fo llo w e d by a s l i g h t d e c lin e toward th e

en d o f th e t e s t * The f i n a l w e ig h ts w ere s t i l l above t h e s t a r t in g I n e v e ry g r o u p .

w e ig h ts

There was no i n d i c a t i o n o f a n y e f f e c t by v ita m in Bjjj on

t h e a v e ra g e w e ig h t. Hemoglobin V alu es The a v e r a g e hem oglobin v a lu e s r o s e i n a l l 5 g ro u p s d u r in g t h e e x p e r im e n t.

G roup A, f e d t h e unsupplem ented b a s a l d ie t X, showed t h e

seco n d h i g h e s t i n c r e a s e , b u t d i f f e r e n c e s betw een g ro u p s w ere a ssa il an d a l l o f t h e hem o g lo b in a v e r a g e s w ere w i th in a nona& l ra n g e . M o rta lity M o r t a l i t y was h i g h e s t I n g ro u p A, f e d th e unsupj&estented b a s a l d ie t 1 .

M o r t a l i t y a l s o o c c u r r e d i n g ro u p G, f e d 8 m icrograsis o f v l t s n i n

The r e s u l t s d id n o t i n d i c a t e an y d e f i n i t e n e e d f o r su p p lem en tal v ita m in Bxg t o p r e w a it e x c e s s iv e .m o r ta lity i n n a tu r e s to c k . £gg P ro d u c tio n f a b l e 4 p r e s e n t s t h e a v e ra g e p e r c e n ta g e egg p r o d u c tio n o f e a c h g ro u p on a hen-d& y b a s i s .

The d i f f e r e n c e s i n i n i t i a l p rod u ction aao n g

t h e g r o u p s , d u r in g t h e p re lim in a ry p e r i o d , a r e a ls o shown i n t a b l e 4 t o perm it c a lc u la t io n o f t h e r e l a t i v e d e c l in e d u r in g th e f i r s t ex p erim en t. Sgg p r o d u c tio n d e c lin e d f o r a l l g ro u p s, b u t t h e d ro p le s s e n e d a s th e l e v e l o f v ita m in

fe d i n c r e a s e d .

The egg p r o d u c tio n o f th e g ro u p fe d

?m m 4 Experim ent I t

Hen-d&y T o ta l Group

£

E f f e c t o f D ie t upon Egg P ro d u ctio n on Hen~Day B asis

Average % Egg, Production p e r Um

T o ta l Experim ent 1

P re lim in a ry P erio d

Experim ent 1

492

726

7 0 .1

41.9

28.2

1836

486

672

8 5 .5

56.6

26.9

086

mz

589

827

5 0 .7

5 5 .0

2 1 .7

714

1836

884

717

55 .8

89.1

14.7

719

1850

454

822

6 5 .1

44.8

1 8 .5

P re lim in a ry P e rio d

£xperi:aent 1

70S

1731

68?

P re lim in a ry P erio d

D ecline i n Average % Egg P ro d u ctio n During Experim ent 1

H to

20 b a s a l d i e t Z was i n t e r m e d i a t e betw een th e a v e ra g e p r o d u c tio n s o f t h e p u l l e t s f e d 8 m icrogram s and 16 m icrogram s o f v ita m in

R e s u lts i n

e x p e rim e n t 1 i n d i c a t e t h a t t h e l e v e l o f egg p r o d u c tio n was a f f e c t e d by t h e l e v e l o f v ita m in

fed .

F o llo w in g a d e la y t o p e rm it t h e n u t r i t i o n a l s t a t u s o f t h e p u l l e t s t o com® t o e q u i l i b r i u m a f t e r t h e d i e t a r y c h a n g e s , d a ta on f e r ­ t i l i t y and h a t c h a b i l i t y w ere c o l l e c t e d .

The p e r io d , r e p r e s e n t i n g e x p e r i ­

m ent 1 , s t a r t e d D ecem ber 5 0 , 1949 and te r m in a te d M arsh 1 6 , I 9 6 0 . F e rtility As shown i n t a b l e 5 t h e r e was a g e n e r a l d e c l i n e i n f e r t i l i t y o f t h e e g g s e x c e p t i n t h e e a s e o f t h o s e p ro d u ced b y g ro u p 0 , w here i t i n ­ c r e a s e d som ew hat.

The l e v e l o f s u p p le m e n ta l v ita m in

f e d had no

m e a s u ra b le i n f l u e n c e on i t . H a t c h a b i l i t y o f F e r t i l e Eggs The d a t a cm h a t c h a b i l i t y d u r in g th e f i r s t e x p e rim e n t i s a l s o in c lu d e d i n t a b l e S .

H a t c h a b i l i t y was m a in ta in e d o r im proved i n e v e r y

c a s e e x c e p t i n g ro u p A, f e d t h e u n su p p lem en ted b a s a l d i e t 1 . showed a d e c id e d d ro p i n h a t c h a b i l i t y .

The r e s u l t s i n d i c a t e d t h a t a

l e v e l o f 4 m icro g ram s o f s u p p le m e n ta l v ita m in ta in h a tc h a b ility .

Group A

w ould a d e q u a te ly m ain­

£1

I

&S g iA

p

CM

C"

m

u>

O

u

s

s> £0

4

£

vO

0*

f u r n is h e d by th e c h ic k d i e t and b y t h e egg i s shown i n t a b l e 2 0 .

The c a l c u l a t i o n s a r e b a s e d on

t h e a d m itte d ly a r b i t r a r y a s su m p tio n t h a t a l l c h ic k s consumed 461 gram s o f f e e d d u r in g t h e f i r s t 4 w eeks o f l i f e .

Vitam in

f u r n is h e d by th© c h ic k

d i e t was a t b e s t o n ly 25 p e r c e n t a s e f f i c i e n t l y u sed a s v ita m in f u r n i s h e d by t h e egg and th© r e l a t i v e e f f i c i e n c y o f u t i l i z a t i o n o f t h a t fu r n is h e d i n th© d i e t dropped a s h ig h e r l e v e l s w ere s u p p li e d . Tab©! 21 p r e s e n t s th® r e l a t i v e q u a n t i t a t i v e e f f i c i e n c y o f s u p p ly in g t h e v ita m in th ro u g h t h e c h ic k d i e t .

f o r t h e c h ic k th ro u g h t h e m a te r n a l d i e t an d The d a t a show t h a t t h e v ita m in

requirem ent

o f t h e c h ic k f o r g ro w th , on a q u a n t i t a t i v e b a s i s , can be more e f f i c i e n t l y m e tto 4 w eeks o f age by s u p p le m e n ta tio n in th© m a te r n a l d i e t .

S u p p le ­

m e n ta tio n i n t h e c h ic k d i e t was a t b e s t o n ly 51 p e r c e n t a s e f f i c i e n t .

TAfiLK 19 Ik p erim en t I t

Chick Pen

Chick Weight a t 0 Weeks Compared to 4 Weeks Expressed. a s P ercen tag e o f Average Weight o f I C hicks in Pen 4*

B

A

B

C

E

4th Week

8 th fcfesk

4 th Week

6 th Week

4 th Week

6 th le e k

4 th Week

0 th Week

4 th Week

1

44 .5 (4)**

4 6 .8 (4 )

41.5 (2 )

58.9 (2 )

6 2 .6 (7 )

6 4 .8 (7 )

85.7 (8 )

81.9 (8 )

7 4 .5 (8 )

75.7 (8 )

2

5 8 .5 (i)

5 9 .8 (1 )

69.6 (4 )

6 8 .0 (4 )

9 0 .0 (7 )

94 .8 (7 )

87.7 (9)

85.6 (9 )

85 .1 (7 )

85.2 (7 )

5

S6.7 (2 )

59 .8 (2 )

6 6 .7 (5 )

64 .4 (S )

88.0 (7 )

8 9 .1 (7 )

9 1 .4 (8 )

94.9 (8 )

70 .8 (7 )

79 .1 (7 )

4

8 5 .5 (2 )

6 5 .4 (2 )

89.6 (4 )

95.9 (4 )

97.9 (7 )

90 .6 (? )

100.8

75

4 2 .8

S

93

4 4 .1

ta bu ;

Sxperiraent 2 s

So. S ta rte d

i S u rv iv in g

P ercentage o f Chicks A liv e a t 4 Uteks

B Chicks

A Chicks Chick Group

25

So. S ta r te d

% S u rv iv in g

C C hicks S o. S ta r te d

0 C hicks

% S u rv iv in g

Ho. S ta r te d

J§ C hicks

i Ho. S u rv iv in g S ta r te d

$ S u rv iv in g

1

15

9 2 .5

57

91.9

H

92 .9

21

90.S

28

82.1

2

15

76.9

52

9 5 .8

11

100.0

21

8L.0

28

78.6

5

8

7S .0

51

90 .5

10

100.0

17

9 4 .1

16

100.0

4

15

9 2 .5

29

86.2

IS

80.0

21

9 0 .5

25

9 2 .0

S2 p e r k ilo g ra m t o t h e m a te r n a l d i e t *

Removal o f t h e v ita m in Bx2 su p p lem en t

fro m m a te r n a l g ro u p s C an d D d id n o t im p a ir t h e v i a b i l i t y o f t h e p ro g e n y . T hese r e s u l t s i n d i c a t e d t h a t th e dams* s t o r e s o f v ita m in

w ere s u f ­

f i c i e n t t o p r o t e c t t h e c h ic k s from e x c e s s iv e m o r t a l i t y f o r t h e d u r a tio n o f th© e x p e rim e n t. F i n a l W eight C h a r t 5 sum m arizes th e a v e ra g e f i n a l w e ig h ts a t t a i n e d by t h e c h i c k s , e x p r e s s e d a s p e r c e n ta g e o f t h a t a t t a i n e d by th e p o s i t i v e c o n t r o l g ro u p .

T h a t g ro u p c o n s i s t e d o f t h e E c h ic k s f e d th e h i g h e s t l e v e l o f

s u p p le m e n ta l v ita m in

The a v e ra g e w e ig h t o f t h e p o s i t i v e c o n t r o l

c h ic k s was p r a c t i c a l l y i d e n t i c a l w ith t h e a v e ra g e w e ig h t o f t h e 1 c h ic k s f e d 27 micrograms o f sup p lem ental v ita m in

in e x p e rim e n t 1 .

p ro d u ced b y p u l l e t s f e d 4 m icrogram s o f v ita m in

B c h ic k s ,

grew a p p ro x im a te ly

a s w e l l a s t h e c o n t r o l g ro u p when f e d l b mlerograms o f v ita m in

Th©

a v e ra g e w e ig h t f o r th© S c h ic k s f e d t h i s l e v e l o f v ita m in was h ig h e r th a n f o r th e c o n tr o ls .

T h is r e s u l t i n d i c a t e d t h a t no a d v a n ta g e c o u ld b e

g a in e d by f e e d in g h i g h e r l e v e l s t o S c h i c k s . C o n s id e rin g t h e r e s u l t s on a l l c h ic k d i e t s , t h e a v e ra g e w e ig h ts o f b o th C and D c h i c k s , whose dams w ere p resu m ab ly b e in g d e p le te d o f t h e i r v ita m in

s t o r e s , w ere lo w e r th a n t h e a v e ra g e w e ig h t o f t h e D c h ic k s .

Th® f i n a l w e ig h t o f t h e A c h ic k s w as g e n e r a l l y lo w e r th a n th e f i n a l w e ig h t o f any o t h e r ty p e o f c h i c k .

Th© r e s u l t s s u f f e r e d fro m t h e v a r i a b i l i t y

t o be e x p e c te d w ith s m a ll number® o f i n d i v i d u a l s , b u t g e n e r a l l y i n d i c a t e d t h a t IS m icro g ram s o f s u p p le m e n ta l v ita m in Big ad d ed t o such a c h ic k d i e t would s a t i s f y t h e requirem ent o f c h ic k s p ro d u ced on no more th a n 4 m ic ro gram s o f v ita m in Bjj>.

Chart 7 .ix n o r i-e n t 2 :

V ita ia ir : 3 i c H o r u i re- i.en t o f C h i c k s f o r G row th a s A f f e c t e d b y M a t e r n a l D i e t a r y H i s t o r t,

105

1 X)

control**

95

of positive

90

Percent

of weight

.85

eo

75

70

0

12 Meg. o f 3},° P ;;

* C a p i t a l l e t t e r s ir. d i e t s fed du rin g ** W eigh ted -average o f orogeny o f g r o u p per kg.

18

15 of ch ick

ra tio n

c o l u m n s r e f e r t o m a t e r n a l g r o u p w.-.ich p r o d u c e d t h e c h i c k s . M aternal e x p e r i m e n t 2 a r c 11 " t o d Lr. t a b l e 2 . f i n a l w e i g h t s a t 1 w eek s, e x p r e s s e d a s p e r c e n t o f t h a t p rod u ced by C p u l l e t s , on b a s a l u i - t s , su p p .. .-merited w i t h 2 7 n icg . o f B l 2

54 D e p le tio n Eat® f a b l e 24 p r e s e n t s , by d i v i s i o n i n t o Z p e r io d s , t h e f i n a l a v e ra g e w e ig h t o f t h e c h ic k s f e d o n ly t h e u n su p p lem en ted b a s a l d i e t 5 . P e r io d 1 i n c l u d e s t h e f i r s t 4 ? d a y s o f e x p e rim e n t 2 .

The seco n d p e r i o d

in c l u d e s r e s u l t s o b ta in e d on t h e s u b se q u e n t 42 days*

As shown i n th e

t a b l e , num bers o f i n d i v i d u a l s r e p r e s e n t i n g m ost o f t h a g ro u p s w ere v e r y s m a ll.

I t i s n o t i c e a b l e , h o w ever, t h a t th e g ro w th o f A c h ic k s in c r e a s e d

im m e d ia te ly a f t e r s u p p le m e n ta tio n o f t h e m a te r n a l b a s a l d i e t 1 w ith S m icro g ram s o f v ita m in

The B c h ic k s p ro d u ced on t h e same m a te r n a l

d i e t u s e d f o r e x p e rim e n t 1 , m a in ta in e d t h e same g e n e r a l r a t e o f g row th th ro u g h o u t e x p e rim e n t 2 .

Growth o f D c h ic k s rem ain ed r e l a t i v e l y c o n s ta n t

d u r in g e x p e rim e n t 2 , i n d i c a t i n g t h a t t h e dams had s u f f i c i e n t s t o r e s o f v ita m in Big re m a in in g from e x p e rim e n t 1 t o su p p ly t h e c h ic k s w ith enough v ita m in Big t o m a in ta in th e same r a t e o f grow th f o r a t l e a s t 89 d ay s on t h e c h ic k b a s a l d i e t .

The a v e ra g e w e ig h t o f D c h ic k s feci s u p p le m e n ta l

v ita m in Big ( c h a r t Z) i n d i c a t e d how ever t h a t some d e p l e ti o n had o c c u r re d i n p u l l e t g ro u p D.

Growth o f i£ c h i c k s , p r e s e n te d in t a b l e 2 4 , in c r e a s e d

f o llo w in g s u p p le m e n ta tio n o f t h e i r m a te r n a l b a s a l d i e t w ith 2 7 .5 m ic ro ­ gram s o f a d d i t i o n a l v ita m in B ig .

I t i s e v id e n t from t h e s e d a t a t h a t

a d d i t i o n o f a s m a ll am ount o f s u p p le m e n ta l v ita m in Big t o t h e d i e t o f d e p le te d p u l l e t s p ro d u c e s an im m ediate grow th s ti m u l a t i o n in r e s u l t a n t p ro g e n y .

I t i s a l s o e v id e n t t h a t s u p p le m e n ta tio n o f p u l l e t b a s a l d i e t 1

w ith 8 m icro g ram s o f v ita m in r e s e r v e s u p p ly .

d id n o t p e rm it s to r a g e o f a s a t i s f a c t o r y

S upplem ent o f t h e p u l l e t b a s a l d i e t 1 w ith 16 microgram©

o f v ita m in Big d id p e rm it s to r a g e o f a c o n s id e r a b le r e s e r v e .

TABLi 24 B x p eriaen t 2 5

A I tu d y o f th e M atern al D ep letio n Bat© a s Shown by F in a l W eights o f C hicks Fed Unsupplaraented B asal D iet 5 .

A C hicks P e rio d R ep resen ted

B Chicks

C Chicks

D C hicks

1 Chicks

So. C hicks

Weight

S o. C hicks

Weight

(pu)

Mo. C hicks

Weight

(gm.)

I® . C hicks

Weight

(g»u)

(gat.)

Mo. Chicks

Weight (p u )

51

155.8

59

225.4

62

256.9

62

269.6

67

2 6 2 .1

A p r il 21 May 25

8

190.1

21

215.8

9

201.2

14

245.0

15

2 5 1 .5

May 26 J u ly 6

4

214.8

15

257.8

4

221.8

5

262.4

8

509.5

F in a l av erag e w eig h t f o r experim ent 1

er* m

m 5.

Summary o f r e s u l t s w ith p rogen y.

D uring b o th e x p e r im e n ts ,

i n i t i a l w e ig h t o f t h e c h ic k s was u n a f f e c te d by m a te r n a l d i e t .

D u rin g

e x p e rim e n t 1 i t was shown t h a t a b s e n c e o f s u p p le m e n ta l v ita m in B ig i n d i e t 1 f o r t h e dams p ro d u ced c h ic k s o f p o o r l i v e a b i l i t y .

t h e e x c e s s iv e

m o r t a l i t y c o u ld be p re v e n te d by th e a d d i t i o n o f betw een 3 and 27 m icro ­ gram s o f su p p lem en tal v ita m in

p e r k ilogram o f c h ic k d i e t .

D uring

e x p e rim e n t 2 t h e r e was a p p a ren tly enough v ita m in

t r a n s m i t t e d t o th©

c h ic k t o p r e v e n t e x c e s s iv e m o r ta lit y i n a l l g r o u p s .

R e s u lts o f e x p e r i ­

m ent 1 i n d i c a t e d t h a t s l i g h t l y more th a n 8 m icrogram s o f s u p p le m e n ta l v ita m in

p e r k ilo g ra m o f p u l l e t f e e d w ould p e rm it p ro g en y t o grow a t

an optimum o r n e a r l y optimum r a t e , i f th e c h ic k d i e t c o n ta in e d 3 a i c r o grams o f s u p p le m e n ta l v ita m in 3 ^ .

R e s u lts o f e x p e r i m n t 2 I n d i c a t e d

t h a t t h e c h ic k s p ro d u c e d on 4 m icro gram s o f s u p p le m e n ta l v ita m in Big w ould grow a t an e x c e lle n t r a te on 15 mlerogreme o f s u p p le m e n ta l v ita m in b X2*

The g ro w th re s p o n s e o f b o th G and D c h ic k s was d e p r e s s e d some­ w hat by rem oval o f t h e s u p p le m e n ta l v ita m in B ig from th e m a te r n a l d i e t . D iv is io n i n t o 2 p e r io d s o f f i n a l w eig h t d a t a , o b ta in e d f o r c h ic k s f e d th® u n su p p le m e n te d b a s a l d i e t 5 , showed t h a t € c h ic k s im m e d ia te ly d ro p p ed i n grow th r a t e ,

D c h ic k s m a in ta in e d t h e i r r a t e of grow th f a i r l y w e l l .

T hese r e s u l t s i n d i c a t e d t h a t 8 micrograms o f s u p p le m e n ta l v ita m in Big i n t h e p u l l e t d i e t would p e rm it l i t t l e i f a n y s to r a g e i n t h e dam*a b o d y . The a d d i t i o n o f 3 m icro g ram s o f v ita m in Big t o t h e m a te r n a l d i e t o f A c h ic k s d id n o t r e s t o r e th e g ro w th r a t e t o n orm al i n t h e 89 day p e r io d r e p r e s e n t e d , when t h e c h ic k s r e c e iv e d a s much a s 18 m icrogram s o f v ita m in B ig p e r k ilo g ra m , b u t i t d id p ro d u ce an im m ediate grow th s t i m u l a t i o n .

5? L im ite d d a t a on grow th t o 6 week© i n d i c a t e t h a t t h e same l e v e l s o f s u p p le m e n ta l v ita m in

r e q u ir e d f o r th© f i r s t 4 weeks a r e a d e q u a te

f o r th® f i r s t 6 w eeks o f c h ic k l i f e .

D egree o f f e a t h e r i n g was p o o r e s t

i n A c h i c k s f e d th® u n su p p lem en to d b a s a l d i e t .

The d e g re e o f f e a t h e r i n g

was e a s i l y i n c r e a s e d by a d d i t i o n o f v ita m in B™ t o e i t h e r t h e c h ic k o r t h e m a te r n a l d i e t . The gro w th r e s u l t s i n d i c a t e t h a t v ita m in Bjg f u r n i s h e d th ro u g h t h e m a te r n a l d i e t i s more e f f i c i e n t l y u se d th a n when i t i© s u p p lie d th ro u g h t h e c h ic k d i e t .

sa DISCUSSION A.

S tu d ie s w ith L aying P u l l e t s

The v ita m in

c o n te n t o f t h e b a s a l d i e t was a p p a r e n tly

a d e q u a te f o r m a in te n a n c e o f body w e ig h t.

T h is r e s u l t agree® w ith th©

r e p o r t by W h itso n , T itu® an d B ir d (1 9 4 6 ) t h a t body w e ig h t was n o t a f f e c t e d by f e e d in g a s i m i l a r r a t i o n t o la y e r s * T h ere was some p u b lis h e d e v id e n c e t h a t v ita m in Bxg i n f lu e n c e d hem o g lo b in v a lu e s i n c h ic k e n s *

Th© r e p o r t b y M ie h o l, H a rp e r and E lv ehjem

(1 9 4 9 ) had i n d i c a t e d t h a t v ita m in

im proved t h e r a t e o f hem oglobin

r e g e n e r a ti o n i n f o l i c a c i d - d e f i c i e n t c h i c k s , a lth o u g h o n ly i n th e p r e s e n c e o f a d e q u a te l e v e l s o f f o l i c a c i d .

However, i n t h e s tu d y r e ­

p o r te d h e r e i n , hem o g lo b in v a lu e s o f p u l l e t s w ere n o t re d u c e d by t h e low d i e t a r y l e v e l o f v ita m in r e s u l t s w ith c h i c k s .

M ushett and O tt (1 9 4 9 ) r e p o r te d s i m i l a r

T h ese r e s u l t s p ro v e t h a t i f v ita m in Bxg i s r e q u ir e d

f o r m a in te n a n c e o f h em o g lo b in , t h e q u a n t i t a t i v e re q u ire m e n t m ust be ex ­ tr e m e ly lo w . A lth o u g h m o r t a l i t y was h ig h e s t i n t h e g ro u p w hich was f e d no s u p p le m e n ta l v ita m in Bxg i n e x p e rim e n t 1 and f e d a v e ry low l e v e l o f v ita m in

i n e x p e rim e n t S , th® l i m i t e d num bers in v o lv e d make i t im­

p o s s i b l e t o d e te rm in e i f t h e e f f e c t was due t o a v ita m in Bjjg d e f i c i e n c y in th e p u lle ts .

T h e re h a v e b een no r e p o r t s o f m o r t a l i t y in m a tu re b i r d s

a s a r e s u l t o f su ch a d e f i c i e n c y .

I t i s d o u b tf u l t h a t body w e ig h t would

b e m a in ta in e d when t h e d e f i c i e n c y f o r m a in te n a n c e a lo n e w ere so g r e a t t h a t i t caused in c re a s e d m o r ta lity . W hile egg p r o d u c tio n seem ed to be b e n e f i t e d i n e x p e rim e n t 1 by s u p p le m e n ta l v ita m in Bxg i n t h e d i e t , th© r e s u l t s in e x p e rim e n t 2 w ere

59 in c o n c lu s iv e .

S k in n e r and Q u isen b eriy (1 9 5 0 ) r ep o rted t h a t a b s e n c e o f

a n APF su p p lem en t o r a n im a l p r o t e i n fro® a h ig h e f f i c i e n c y la y i n g mash r e s u l t e d i n lo w e re d p r o d u c tio n .

H i l l , HcCon&chie, G a r tle y and B ran io n

(1 9 5 0 ) s t a t e d t h a t b i r d s f e d a c e r e a l g ra in -e o y b e a n m eal ty p e r a t i o n u n su p p le m e n te d w ith v ita m in Big came i n t o p r o d u c tio n l a t e r th a n o t h e r s w hich w ere f u r n i s h e d t h e v ita m in , and p ro d u ced fe w e r e g g s •

I t is not

c l e a r w h e th e r th© b i r d s l a i d fe w e r eg g s th an th© o ther® on a t o t a l q u a n t i t a t i v e b a s i s due t o slo w n e ss i n a t t a i n i n g s e x u a l m a tu r ity o r t h e egg p r o d u c tio n was a c t u a l l y l e s s i n t e n s e a f t e r th e y came i n t o l a y . G lees® , Couch and Lyman (1 9 5 0 ) have r e c e n t l y re p o rte d , t h a t th® a d d i t i o n o f v ita m in Big t o a su c ro se-so y b ea n p r o te in d i e t im proved egg p r o d u c tio n o f hens.

When s t a r c h r e p la c e d t h e s u c ro s e i n t h e a b se n c e o f v ita m in Big#

eg g p r o d u c tio n an d h a tc h a b iX ity im p ro v ed , p o s s i b l y i n d i c a t i n g in c r e a s e d i n t e s t i n a l s y n t h e s i s o f t h e v ita m in .

On th© o t h e r h a n d , W hitson, T itu s

an d B ird ( 1 9 4 6 ), F o r r e s t , B ie ly an d March (1 9 SO) and o t h e r s , w o rk in g w ith a l l - v e g e t a b l e p r o t e i n p r a c t i c a l d i e t s d e f i c i e n t i n v ita m in Big have found no r e d u c tio n

i n egg p r o d u c tio n . In o t h e r work c a r r i e d o u t by th® w r i t e r ,

an e x c e l l e n t

r a t e o f p r o d u c tio n was found t o b e m a in ta in e d fro m S ep tem b er

t o t h e f o llo w in g Ju n e (when th©t e s t was te r m in a te d f o r u n r e la te d

reaso n s)

on a s i m i l a r

used.

ra tio n .

More th a n IBS Rhode I s l a n d Bed p u l l at® w ere

They w ere m a in ta in ed on l i t t e r , how ever, and t h e r e fo r e p ro b a b ly r e c e iv e d s l i g h t l y more v ita m in B ig .

A r s c o tt (1 9 5 0 ) r e p o r t e d no s i g n i f i c a n t

d i f f e r e n c e s i n t h e egg p rod u ction b etw een p m s o f p u l l e t s m a in ta in e d on w ire w ith and w ith o u t su p p lem en tal v ita m in B ig . was b a s a l d i e t 1 . v ita m in

Th© b a s a l r a t i o n employed

F i f t y Hew H am pshire p u l l e t s w ere u se d p e r p e n .

Perhaps

1® r e q u i r e d f o r ©gg p r o d u c tio n , b u t i n no l a r g e r q u a n t i t i e s

t h a n a r e u s u a l l y c o n ta in e d in c e r e a l g r a in -so y b sa n m eal r a t i o n s w ith o u t

60 s u p p le m e n ta tio n w ith v ita m in

At an y r a t e th e e v id e n c e p ro d u ced in

e x p e rim e n t 1 f a v o r in g th e h y p o th e s is t h a t eg g p r o d u c tio n can be bene­ f i t e d by su p p lem en ta tio n o f such a r a t i o n w ith v ita m in Bq£ i s by no m eans c o n c lu s iv e b e c a u s e o f th e l i m i t e d num ber o f in d iv id u a ls and th e s t a l l d i f f e r e n c e s i n p r o d u c tio n . T h e re was c o n s id e r a b le v a r i a b i l i t y i n f e r t i l i t y among t h e g ro u p s a t d i f f e r e n t t i m e s , b u t t h e r e was no e v id e n c e t h a t £ e r t i l i t y was in flu e n c e d by v ita m in B i£ .

D uring e x p e rim e n t 1 , 4 microgronis o f v ita m in

Big w ere fo u n d t o s a t i s f y t h e re q u ire m e n t f o r good h a t c h a b ilit y when a normal l e v s ! o f p r o te in w as f e d and no s t r e s s f a c t o r s w ere in tr o d u c e d a s i d e from t h e e l i m i n a t i o n o f coproph& gy.

R e c e n tly s e v e r a l s t u d i e s on

h a t c h a b i l i t y h av e been r e p o r t e d which h av e a d i r e c t b e a r in g on th e work b e in g d i s c u s s e d .

P e t e r s e n , W iese, L&mpman an d Qahlstrom (1 9 5 0 ) s tu d ie d

t h e e f f e c t o f v ita m in Byg on h a t c h a b i l i t y by i n j e c t i n g g ra d e d l e v e l s o f t h e v ita m in i n t o w h ite leg h o rn p u l l e t s k e p t on w ire and p r e v io u s ly f e d a co m -so y b ea n meal d e p le tio n d i e t o f u n sta te d p r o te in l e v e l f o r 4 merith s . T h e ir d a t a i n d i c a t e d t h a t a p p ro x im a te ly 1 .0 laicrograjiis o f v ita m in Big i n j e c t e d p s r hen p e r week w ould r e s t o r e h a t c h a b i l i t y to 8 9 .6 p e r c e n t w i t h i n 5 weeks a© com pared t o 6 S .0 p e r c e n t h a t c h a b i l i t y f o r t h e b i r d s I n j e c t e d w ith 0 .5 xaicrogr&as o f

p e r w eek.

I f one assu m es t h a t each

hen consumed ap p ro x im a tely 800 gram s o f f e e d p e r w eek, t h e h ig h e r i n ­ j e c t i o n r a t e f u r n is h e d t h e v ita m in t o th e b i r d a t a r a t e o f 1«&& saicrograms p e r k ilo g ra m o f f e e d .

Growth o f p ro g en y was h ig h e s t when t h e hen was i n ­

j e c t e d w ith th© h i g h e s t l e v e l o f v ita m in Big u se d , 40 m ierogram s p e r hen p e r w eek, o r ap p ro x im a tely 5 .0 micrograms p e r k ilo g ra m o f f e e d .

The

p ro g e n y we .re f e d a v itam in , B is d e f i c i e n t a l l - p l a n t p r o t e i n d i e t f o r 4 w eek s.

S t ok s t a d , J u k e s , F i e r c e , Page and F r a n k lin (1 9 4 9 ) fo u n d t h e o r a l

61 p o te n c y o f v ita m in

t o bo 60 p e r c e n t t h a t o f th® i n j e c t e d p o ten cy *

I n 1960 Mange r e p o r t e d work i n ag reem en t e s s e n t i a l l y w ith t h i s f i g u r e . On th e b a s i s o f t h e s e c o n v e r s io n f a c t o r s , th e l e v e l s o f v ita m in BI2 w hich P e te r s e n e t a l found s a t i s f i e d t h e re q u ire m e n t f o r h a t c h a b i l i t y an d t h e r e q u ire m e n t f o r g ro w th w ere 2 . 5 m icrogram s a n d 10 sderogr& m s, r e s p e c t i v e l y , p a r k ilo g ra m when o r a l l y a d m in is te r e d . C a rv e r and M cG innis { i9 6 0 ) r e p o r t e d t h a t h a t c h a b i l i t y m s g r e a t l y re d u c e d when h e n s i n b a t t e r i e s ’w ere f e d an u n su p p lem en ted c e r e a l g r a in - s o y b e a n m eal ty p e o f d i e t o f u n s p e c if ie d p r o t e i n l e v e l .

was ad d ed p e r k ilo g ra m o f d i e t ,

ra ie ro g rsjs o f c r y s t a l l i n e v i t a m i n • h a t c h a b i l i t y m ark e d ly im p ro v ed .

Whan 1 .0

B a ta w ere i n s u f f i c i e n t t o i n d i c a t e a

q u a n t i t a t i v e re q u ire m e n t f o r h a t c h a b i l i t y .

In December, 1949, L i l l i e ,

O lso n , and B ir d , r e p o r te d t h a t h a t c h a b i l i t y o f e g g s fro m d e p l e te d h en s was r e s t o r e d by i n j e c t i n g 0 .6 a ic ro g ra m o f 8 ^ i n t o ea c h egg. r e s u l t i n g c h ic k s a l s o i n c r e a s e d m ark ed ly f o r a t l e a s t 6 w eek s. in v o l v i n g a d m i n i s t r a t i o n o f v ita m in

Growth of .In s tu d i e s

t o t h e dam t h e l e v e l o f p r o t e i n

f e d t h e hen m ust b e c o n s id e r e d a s an im p o rta n t f a c t o r .

Y&covdtz, F e e l e r ,

M i l l e r , C a r ls o n , H e u se r and M o rris (I9 6 0 ) r e p o r t e d t h a t when b i r d s w ere f e d a n a l l - v e g e t a b l e p r o t e i n ty p e o f d i e t c o n ta i n i n g no s u p p le m e n ta l v ita m in 3qg,

h e n s on a l e v e l o f 25 p e r c e n t p r o t e i n p ro d u ced eg g s o f 26

p e r c e n t h a t c h a b i l i t y a f t e r 12 w eeks o f d e p l e t i o n .

I f th.* p r o t e i n l e v e l

i n t h e hm. d i e t was 16 p e r c e n t, h o w ev er, h a t c h a b i l i t y w as 53 porear.it a f t e r 15 w e ek s.

They e s tim a te d t h a t t h e re q u ire m e n t f o r h a t c h a b i l i t y on

a 25 p e r c e n t p r o t e i n d i e t was 2 m icrogram s o f s u p p le m e n ta l v ita m in Bi2 p a r k ilo g ra m o f f e e d . W hile 4 saicrogr&HiS o f v ita m in Bqg p e r k ilo g ra m o f f e e d s u p p o rte d s a t i s f a c t o r y h a t c h a b i l i t y on t h e 15 p e r c e n t p r o t e i n d i e t 1 , th e r e s u l t s

62

w ith 5 m icro g ram s p e r k ilo g ra m i n e x p e rim e n t 2 w ere n o t c l e a r .

T ru e ,

t h e o v e r a l l h a t c h a b i l i t y o f t h a t g ro u p d id im prove v e ry s l i g h t l y a f t e r t h e v ita m in B is was ad d ed , b u t h a t c h a b i l i t y i n £ g ro u p d e c lin e d con­ s id e r a b l y an d o v e r a l l h a t c h a b i l i t y i n 6 g ro u p , from w hich a l l s u p p le ­ m e n ta l v ita m in Big h ad b een rem oved, i n c r e a s e d s l i g h t l y .

Group B, f e d

4 m icro g ram s o f v ita m in Big d e c lin e d c o n s id e r a b ly i n o v e r a l l h a t c h a b i l i t y . In s p i t e o f t h e f a c t t h a t h a t c h a b i l i t y in A g ro u p rem ain ed lo w e r th a n t h a t i n a n y o t h e r g r o u p , t h e s p r e a d betw een t h e p o s i t i v e c o n t r o l gro u p £ , g ro u p A and g ro u p B was so n a rro w a s t o i n d i c a t e t h a t a l l o f t h e f l u c t u a t i o n s m ig h t h av e been t h e r e s u l t o f o t h e r f a c t o r s .

I t i s q u ite p o s s ib le , th e re ­

fo r® , t h a t 5 m icrogram s o f v ita m in Big su p p lem en ted t o such a r a t i o n may m a in ta in good h a t c h a b i l i t y .

In d eed c o n c u r r e n t work c o n d u c te d a t t h i s

s t a t i o n by A r s c o tt ( I9 6 0 ) s t r o n g l y i n d i c a t e s t h a t on t h e same b a s a l d i e t , b i r d s m a in ta in e d u n d e r f l o c k c o n d iti o n s on w ire r e q u i r e s l i g h t l y more th a n 2 m icrogram s o f v ita m in B ig p e r k ilo g ra m o f d i e t f o r optimum h a tc h ­ a b ility .

T h is f i g u r e i s i n c l o s e ag reem en t w ith t h e a d j u s t e d f i g u r e o f

2 .S m icro g ram s c i t e d f o r P e te r s e n e t a l . The s tu d y on th© r a t e o f d e p le ti o n o f p u l l e t s f o llo w in g rem oval o f s u p p le m e n ta l v ita m in B i£ was h a n d ic a p p e d by l i m i t e d egg p r o d u c tio n . Th© r e s u l t s d id show a h im m ed iate d ro p i n h a t c h a b i l i t y when t h e 8 m ic ro ­ gram s o f s u p p le m e n ta l v ita m in Big w ere removed fro m t h e p u l l e t d i e t .

Th©

r a p i d d e c l i n e i s e v id e n c e t h a t th o s e p u l l e t s war© u n a b le t o s t o r e a r e ­ s e rv e o f v ita m in B ig s u f f i c i e n t t o c a r r y them o v e r even a te m p o ra ry s h o r ta g e o f t h e v ita m in .

I t in d ic a te ® t h a t 8 m icrogram s of s u p p le m e n ta l

v ita m in Bi£ i n such a d i e t i s in a d e q u a te i f s to r a g e i s d e s ir e d a s a s a fe ty f a c to r. The r e s u l t s o f th© a n a ly s e s o f th e eggs f o r v ita m in 012 c o n te n t d e m o n s tra te th® c u r r e n t d i f f i c u l t y in a s s a y in g an y c ru d e f e e d s t u f f s f o r

es t h i s v ita m in .

The r e s u l t s w ith t h e d i f f e r e n t e h ie k a s s a y s show good

c o rre la tio n .

The m i c r o b i o l o g i c a l a s s a y r e s u l t s a r e a l s o r e a s o n a b ly

c o n s is te n t.

But 9 0 .S p e r c e n t o f t h e v ita m in Bxg a c t i v i t y fo u n d i n B

e g g s an d 8 3 .5 p e r c e n t o f t h e v ita m in B ig a c t i v i t y fo u n d I n IS e g g s by c h ic k a s s a y w ere m i c r o b i o l o g i c a l l y u n a v a i l a b l e .

In 1947 Rubin an d B ird

r e p o r t e d t h e o c c u r re n c e o f t h e cow m anure f a c t o r in e g g s .

They a s s a y e d

t h e egg f o r p o te n c y w ith c h ic k s , fe e d in g w hat l a t e r p ro v e d t o bo a v ita m in B12 c o n c e n t r a t e a s a p o s i t i v e c o n t r o l .

S in c e t h a t tim e L i l l i e and B ird

(1 9 4 9 ) have d e te rm in e d t h e v ita m in Big c o n te n t o f t h a t c o n c e n t r a t e .

By-

a p p ly in g t h e now known v ita m in B1 2 p o te n c y one f i n d s t h a t Eubin and B ird fo u n d a p p r o x im a te ly 1*£S m icrogram s o f v ita m in Bi# a c t i v i t y p e r eg g when i t was p ro d u ce d on a v ita m in B i g - d e f i c i e n t r a t i o n s i m i l a r t o b a s a l d i e t 1 . They d e te rm in e d n o rm al eg g s t o c o n ta in ro u g h ly S . 89 m icrogram s o f v ita m in a c t i v i t y , a p ie c e .

S in c e t h e i r la y in g b i r d s w ere m a in ta in e d on l i t t e r

and had a c c e s s t o t h e i r d r o p p in g s , i t i s l o g i c a l t h a t t h e v ita m in B1 2 a c t i v i t y o f t h e i r e g g s s h o u ld be s l i g h t l y h ig h e r f o r b o th t h e n o rm al eg g s and f o r t h e d e f i c i e n t o n e s .

The c o r r e l a t i o n b etw een t h e r e s u l t s o f t h e i r

s tu d y and t h e o n es r e p o r te d h e r e i n i s re m a rk a b le .

The p r e lim in a r y v a lu e

f o r v ita m in Big c o n te n t o f e g g s produced by u n d e p le te d c h ic k e n s a© d e­ te rm in e d b y t h e r a t a s s a y , r e p o r te d by L e w is, R e g i s t e r , Thompson and B lvehjem (1 9 4 9 ) c o n f l i c t s w ith t h e c h ic k r e s u i t s .

T h e ir r e s u l t showed

good c o r r e l a t i o n w ith t h e m i c r o b io lo g ic a l r e s u l t s r e p o r te d h e r e i n .

They

fo u n d 2 b>2 m illim ic ro g ra m s o f v ita m in B1 2 p e r e g g , w h ereas th e l a t t e r r e ­ s u l t s i n d i c a t e d 262 m illim ic ro g ra m e o f v ita m in

p e r eg g .

R e s u lts o f t h e m ic r o b io lo g ic a l a s s a y s r e p o r te d i n t h i s t h e s i s v e r i f y t h e r e s u l t s r e p o r t e d by X a c o w its, F e e l e r , M i l l e r , C a r ls o n , Heuser an d M o rris (1 9 6 0 ) •

They fo u n d 158 m illim ic ro g ra m s o f v ita m in Bjjg a c t i v i t y

p e r "normal** e g g .

V a r i a t i o n s i n e x p e r im e n ta l p ro c e d u re r e c o n c i le d th e

s m a ll d i f f e r e n c i s s b etw een t h e m ic r o b io lo g ic a l r e s u l t s .

The q u e s tio n

oT w hich .r e s u l ts a r e n e a r e s t th e a b s o lu t e v a lu e , th o s e o b ta in e d by m icro ­ b i o l o g i c a l , r a t o r c h ic k a s s a y s , p la g u e s r e s e a r c h w o rk e rs to d a y , t y p e o f a s s a y p ro b a b ly m e a su re s th e a c t u a l v ita m in m a t e r i a l s more a c c u r a t e l y th a n any o t h e r a s s a y ,

j&ach

c o n te n t o f some

d to k s ta d , HoTfi&aim and

J u k e s (1 9 4 9 ) r e p o r te d d i s c r e p a n c i e s betw een m i c r o b io lo g i c a l and c h ic k a s s a y s f o r v ita m in

lie s u i t s o f r a t and m ic r o b io lo g ic a l a s s a y s

f r e q u e n t l y d i s a g r e e w id e ly .

C e r t a i n l y a s f a r a s c h ic k n u t r i t i o n i s con­

c e r n e d , v a lu e s d e te rm in e d w ith c h ic k a s s a y s a r e t h e m ost r e l i a b l e .

The

v a r i a b l e r e s u l t s b etw een s p e c ie s p ro b a b ly a r e c a u se d by ( 1 ) d i f f e r e n t i a l a b i l i t y o f t h a s p e c i e s t o u t i l i s e bound fo rm s , ( 2 ) d i f f e r e n t i a l s ti m u l a t o r y o r s p a r in g e f f e c t o f v a r io u s com pounds, o r ( 5 ) a c o m b in a tio n o f th e tw o . Among f a c t o r s o t h e r th a n v ita m in

w hich m ig h t c o n c e iv a b ly a f f e c t a s s a y

r e s u l t s w ith t h e c h ic k s i s t h e whey f a c t o r .

I t was c o n s id e r e d l i k e l y t h a t

t h e whey f a c t o r w ould be p r e s e n t i n t h e eg g s w hich were a s s a y e d .

In

a d d i t i o n i t was f e l t t h a t t h e whey f a c t o r m ig h t e x e r t a s tim u la to r y 1" e f f e c t on gro w th w hich c o u ld b e i n t e r p r e t e d a s b e in g cau sed by t h e p r e s e n c e o f v ita m in Bi2*

F o r t h a t re a s o n 5 p e r c e n t o f d r ie d whey m s added t o d i e t 5 .

I t m s n o t added t o d i e t 4 .

S in c e th e r e s u l t s o f t h e tw o ty p e s o f c h ic k

assay © showed v e r y c l o s e a g re e m e n t, t h e whey f a c t o r i s b e li e v e d t o have no e f f e c t on t h e r e s u l t s o f c h ic k a s s a y s f o r v ita m in Bjjj ^ben c h ic k s o f such d i e t a r y back g ro u n d a r e u s e d . h e s u i t s o f b o th m ic r o b io lo g i c a l and c h ic k a s s a y s d e m o n s tra te a d e f i n i t e c o r r e l a t i o n betw een th e l e v e l f e d th e hen and t h e l e v e l fo u n d in th e egg.

S tu d y o f t a b l e 8 r e v e a l s t h a t t h e c h ic k a s s a y s fo u n d in c r e a s e d

s t o r a g e i n th e egg a t a d im in is h in g rat® w ith h ig h e r l e v e l s o i v ita m in Bjjg

65

i n th© p u l l a t d i e t .

The e f f i c i e n c y o f t r a n s m is s io n o f i d t a s d n Big from

t h e p u l l e t r a t i o n t o t h e egg d e c r e a s e d w ith h ig h e r le v e l© o f su p p le m e n ta ­ t i o n a s shown i n t a b l e 1 0 .

I t i© probable t h a t t h e v ita m in Big co n ten t

o f g ro u p C e g g s a s d a te m in e d by th e c h ic k a s say i s to o lo w .

I f more

c h ic k a s s a y s had oeen c o n d u c te d , i t i s b e lie v e d t h a t a h ig h e r v ita m in B ig c o n te n t w ould have b e en d e te rm in e d f o r C e g g s .

The grow th o f th e

c h ic k s h a tc h in g from such e g g s i n d i c a t e t h a t t h e v ita m in Bjg c o n ten t i n them i s i n t e r m e d i a t e betw een th a l e v e l i n g ro u p 8 and g ro u p 8 eggs* B. The d a ta

S tu d ie s on th e Progeny

on l i v e a b i l i t y i n d i c a t e

t h a t a d e f i c ie n c y o f v ita m in

B ig i n t h e m a te r n a l d i e t l e a d s t o e x c e s s iv e m o r ta lit y among th© c h ic k s w hich can o n ly be c o u n te r a c ta d by th© us® o f h ig h l e v e l s o f v ita m in Mjz i n th® ©hick d i e t . m it.

II©

A r e c e n t r e p o r t by B ird (1950.) s u b s t a n t i a l ©3 t h i s r e ­

s u g g e s ts t h a t t h e m o r t a l i t y i s c a u se d by

th© p h y s ic a l i n a b i l i t y

o f t h e c h ic k t o i n g e s t enough -vitam in Bxg d u r in g th e f i r s t few days o f l i f e f o r s u rv iv a l.

I n tr a m u s c u la r in j e c t i o n o f th© v ita m in would p reven t

t h e e x c e s s iv e m o r t a l i t y . I n t e r e s t i n g ev id e n ce f o r t h e in flu e n c e o f m a te r n a l d i e t on t h e re q u ire m e n t o f t h e c h ic k f o r v ita m in 8^*> f o r n o rm al grow th h a s been ob­ ta in e d .

The f i n a l w e ig h t o f A c h i c k s , p ro d u ced on a m a te r n a l d i e t i n ­

c lu d in g no s u p p le m e n ta l v ita m in B^g and fe d a c h ic k d i e t c o n ta in in g Z? micrograms o f su p p lem en tal v itam in B jg , was 5 p e r c e n t l e s s th a n th® f i n a l w e ig h t o f c h ic k s p ro d u ced on 10 m icrogram s in t h e m aternal d i e t and fe d o n ly 3 m icro g ram s in t h e c h ic k d i e t ,

linen A p u l l e t s s u b s e q u e n tly

r e c e iv e d 3 micrograms o f v ita m in B ^ , growth o f th© p ro g en y was un­ s a t i s f a c t o r y on 18 m icrogram s o f v ita m in Big* th e h i g h e s t l e v e l u sed in

e x p e rim e n t 2 .

By c o n t r a s t B c h ic k s p ro d u ced b y dams f e d on 4 ssicro g raias

o f v ita m in Bxg th ro u g h o u t b o th e x p e rim e n ts , a p p a r e n tly r e q u ir e d o n ly lb m icro g ram s o f v ita m in Bxg i n t h e d i e t f o r good g ro w th .

The q u a n t i t a t i v e

re q u ire m e n t o f c h ic k s f o r v ita m in Bxg i s t h e r e f o r e seen t o be in f lu e n c e d tre m e n d o u s ly by t h e m a te r n a l d i e t .

A lthough th e f i g u r e s p re se n te d , in

t a b l e s 2 0 and 21 a r e b a se d on assum ed v a lu e s f o r p u l l e t and c h ic k f e e d co n su m p tio n , th e y c l e a r l y show t h a t v ita m in Bxg s u p p lie d th ro u g h t h e m a te r n a l d i e t i s much more e f f i c i e n t l y u s e d th a n i f s u p p lie d i n th e c h ic k d i e t .

O b v io u sly t h e l e v e l f u r n is h e d i n t h e c h ic k d i e t w ould i n ­

c r e a s e i n im p o rta n c e a s th® b ird ® became o ld e r# T h e re i s f u r t h e r e v id e n c e o f an a d m itte d ly in c o n c lu s iv e n a t u r e f o r a l i m i t i n g e f f e c t on c h ic k grow th o f low l e v e l s o f v ita m in Bjjg i n t h e m a te r n a l d i e t , r e g a r d l e s s o f l e v e l o f s u p p le m e n ta tio n i n t h e c h ic k d i e t . D u rin g experim ent 1 , t h e grow th o f B, C, an d 1 c h ic k s t unde a t o p l a t e a u a t a © lig h tly lo w e r l e v e l th a n t h e grow th o f B c h ic k s a s shown i n c h a r t Zm

D u rin g experim ent 2 t h e grow th o f A c h ic k s a l s o te n d e d t o p l a t e a u a t

& low l e v e l .

I t seem s u n l i k e l y t o th e w r i t e r t h a t such a l i m i t i n g a f f e c t

a c t u a l l y e x i s t s i r r e s p e c t i v e o f ch ick d i e t .

E s th e r d o e s i t seem p ro b a b le

t h a t h i g h e r l e v e l s o f v ita m in Bx£ th a n w ere f e d i n th® c h ic k d i e t w ere r e ­ q u i r e d f o r maximum g row th o f such p ro g en y t o 4 w eek s.

The d a ta o b ta in e d

by B ird (1 9 5 0 ) on e x c e s s iv e m o r t a l i t y i n d e p le te d c h ic k s s u p p o rt t h i s b e lie f.

As p r e v i o u s l y m e n tio n e d , he found t h a t e x c e s s iv e m o r t a l i t y

w as p r e v e n te d by i n j e c t i o n o f v ita m in Bxg.

The r e s u l t s w ith D and & c h ic k s

i n e x p e rim e n t 1 do p r e s e n t t h e p o s s i b i l i t y t h a t h i g h e r l e v e l s o f v ita m in B ig t h a n a r e u s u a l l y fo u n d i n b r e e d in g r a t i o n s may p ro d u ce c h ic k s t h a t v / i l l grow a t an i n i t i a l l y f a s t e r rat© on cu sto m a ry b r o i l e r r a t i o n s .

I t is

67

e v id e n t t h a t t h e l e v e l o f v ita m in B^g f e d i n th e b re e d in g pen i s a s im p o r ta n t t o t h e bro ilerra& n a s t h e l e v e l f e d i n t h e b r o i l e r house* The r e s u l t s o f t h e s e e x p e rim e n ts a l s o p ro v e t h a t sorae s u p p le m e n ta l v ita m in Big i s n eed ed i n c h ic k d i e t s composed p r i m a r i l y o f c e r e a l g r a i n s and so y b ean p r o t e i n .

When a good, b r e e d in g r a t i o n was

u s e d , t h e re q u ire m e n t o f & c h ic k s f o r supp lem en tal v ita m in Big was no l a s s th a n 5 m ic ro g ra m s, p o s s i b l y as- much a s 16 m icro g ram s, f o r optimum g ro w th .

S in c e t h e s e c h ic k s w ere r e a r e d in b a t t e r i e s th e y d id n e t have

t h e advantage- o f a c c e s s t o n u t r i t i o n a l f a c t o r s in dropping;# an d l i t t e r , w hich m ost b r o i l e r c h ic k s w ould h a v e .

( M ic r o b io lo g ic a l a n a ly s e s o f hen

f e c e s and o f d eep l i t t e r i t s e l f by H&lferook, S u tto n , and W in te r (I9 6 0 ) i n d i c a t e d t h a t th e y may a c t a s l i m i t e d s o u rc e s o f t h e v i t a m i n .)

N ever­

t h e l e s s such an a d v a n ta g e i s o f d u b io u s Im p o rta n c e d u r in g th e c r i t i c a l f i r s t 4 w eeks o f I l f © .

A t t h i s tim e th© a b i l i t y o f t h e c h ic k t o i n g e s t

l a r g e q u a n t i t i e s o f b u lk y m a t e r ia l i s p a r t ic u la r ly l i m i t e d , and th e am ount o f v ita m in

a v a i l a b l e i n t h e l i t t e r would be a t th e lo w e s t

p o i n t u n l e s s o ld l i t t e r w ere u s e d t o s t a r t new c h ic k e n s .

T h e re f o re , th e

a v a i l a b i l i t y o f v ita m in 8 ^ i n t h e m a te rn a l and c h ic k m ashes would p r o b a b ly be n e a r l y a s im p ortan t f o r c h ic k s r e a r e d u n d e r com m ercial con­ d itio n s .

E o b 'b lee, K ic h o l, C ra v e n s , S lv eh jem and H a lp in (1948) d e s c r ib e d

w ork w hich i n d i c a t e d t h a t oven i f th e m a te r n a l d i e t c o n ta in e d su p p osed ly a d e q u a te l e v e l s o f v ita m in B^g, th e pro g en y s t i l l r e q u ir e d supplem ental v ita m in 812 t o 4 w eeks o f a g e .

S te v e n s , i i i e l y and M arch (1 9 4 9 ) r e p o r te d

t h a t ev en on a co m m ercial c h ic k s t a r t e r c o n ta in in g m ixed g r a i n s , soybean m e a l, f i s h m eal an d d r i e d whey, th e fe e d e f f i c i e n c y was m r k e d ly im proved b y t h e a d d i t i o n o f t h e su p p lem en tal v ita m in s tu d y was u n a f f e c t e d .

a lth o u g h grow th in t h e i r

T h e ir b a s a l r a t i o n i n t h i s c a s e was c e r t a i n l y a s

h ig h i n vit&min 3x2 from n a tu r a l s o u rc e s a s c o u ld bo exp)a c te d .

Un­

f o r t u n a t e ly t h e e x a c t co m p o sitio n o f i t i s n o t g iv e n and th e c h o lin e an d m e th io n in e l e v e l s a r e n o t s t a t e d .

A d e f i c i e n c y o f th e s e two com­

po unds i s q u i t e p o s s i b l e i n such a r a t i o n .

A low l e v e l o f th e s e two

compounds i n t h e c h ic k d i e t h a s b e e n shown by B r ig g s , H i l l and G ile s (1 9 5 0 ) and o th e r s t o i n c r e a s e th e requirem ent f o r v ita m in iii£ *

it

shou ld b e p o in te d o u t a l s o t h a t th e g ro w th r e l a t i o n s h i p s e x p r e s s e d f o r t h e f i r s t 4 w eeks, p ro b a b ly a l s o h o ld t o 0 w eeks a s shown on t a b l e 1 5 . T h is d o es n o t mean t h a t t h e c h ic k re q u ire m e n t i s a s h ig h a t 3 weeks a s it

i s a t 4 w eek s, b u t i t d o e s mean- t h a t a l l grow th r e q u ire m e n ts a r e

s a t i s f i e d w ith th ss® l e v e l s .

L i l l i ® , M&rsden, Groschke and Bird. (1949)

fo u n d t h a t t h s a p p a r e n t re q u ire m e n t o f th e grow ing c h ic k e n f o r v ita m in 2 d ro p p ed d u r in g t h e l a t e r s ta g e s o f grow th* The d e g re e o f fe a th e r in g i s c l e a r l y .in flu e n c e d by v ita m in E i t h e r 4 microgr&ms o f v ita m in B^g i n t h e b r e e d in g r a t i o n o r 8 m icrogram s i n t h e c h ic k d i e t s u p p o rte d good fe a th e r in g * i s so low t h a t t h e e f f e c t o f v ita m in l i t t l e p r a c t i c a l im p o rta n c e . in g and v ita m in

T h e refo r© th e requirem ent

on d e g re e o f f e a t h e r i n g i s o f

A p o s s i b l e r e l a t i o n s h i p betw een f e a t h e r ­

i s e a s i l y d is c e r n e d .

F e a t h e r s c o n ta in a h ig h l e v e l

o f c y s t i n e , w hich may b e p ro d u ced from m e th io n in e .

The work o f B riggs*

H i l l and G ile s ( 1 9 5 0 ), G i l l i s and M o rris (1 9 4 9 ), S c h a e f e r , Salmon and S tr e n g t h (1 9 4 9 ) and o t h e r s h av e i n d i c a t e d t h a t t h e r e i s a m u tu ally s p a r i n g a c t i o n b etw een t h e v ita m in and m e th io n in e .

On© may assum e

t h a t p re s e n c e o f a h ig h l e v e l o f v ita m in Bjji would i n c r e a s e t h e am ount o f m e th io n in e a v a i l a b l e f o r s e c o n d a ry p h y s io l o g i c a l f u n c t i o n s , w h ile a v ita m in B\Z d e f i c i e n c y w ould c o r re s p o n d in g ly d e c r e a s e th e s u p p ly o f m e th io n in e a v a i l a b l e f o r such f u n c t io n s .

The r e s u l t s o f t h e d e p l e t i o n

ms tu d y show t h a t more th a n 8 m icrograras o f s u p p le m e n ta l v ita m in

is

r e q u i r e d i n t h e m a te r n a l d i e t t o p e rm it a d e q u a te s t o r a g e , a s a s a f e t y fa c to r. The f i g u r e s c i t e d f o r t h e v ita m in Bqg requirem ents have n o t c r e d i t e d t h e b a s a l d i e t s w ith c o n te n t o f an y v ita m in

The re a so n

i s t h a t t h e r e i s no r e l i a b l e v ita m in Bx£ a s s a y a v a i l a b l e f o r c e r e a l g r a i n s , so y b ean m eal and d r i e d b rew ers* y e a s t .

T hese f e e d s t u f f s a r e

b e lie v e d t o c o n ta in u n i d e n t i f i e d grow th f a c t o r s (Mange, 19BO) w hich may causa Microbiological assays to y i e l d inflated v a l u e s .

I f the r e s u l t s

o f a m ic r o b i o l o g i c a l a s s a y ( p u b lis h e d by P e e l e r , l a c o w i t s an d M o rr is , 1950) a r e u s e d , b a s a l d i e t 1 may b e c o n s id e re d t o c o n ta in a p p r o x im a te ly 8.L -10 m ic ro g r a a s o f v ita m in Bqg p e r k ilo g ra m . B a s a l d i e t 2 may b© e s tim a ts d t o c o n ta in 25 m icro.gram s o f v ita m in Big p e r k ilo g ra m .

A p p ro x im a te ly 15

m icro g ram s o f t h e t o t a l c o n te n t o f b a s a l d i e t Z h a s been d e t e m i n e d w ith c h ic k a s s a y s , and th e r e fo r © i s b e lie v e d d e f i n i t e l y t o b e p r e s e n t . e x i s t e n c e o f t h e o t h e r 8 mlcrograma I s q u e s ti o n a b l e .

The

I t i s f e l t th a t

t h e c a l c u l a t e d c o n te n t o f v ita m in 3 j£ i s much to o h ig h where v a lu e s ob­ t a i n e d b y m i c r o b io lo g ic a l a s s a y s o f t h e s e f e e d s t u f f s a r e em ployed.

For

t h a t r e a s o n th e y h av e n o t b een u s e d in i n t e r p r e t i n g th e s e r e s u l t s . The r e q u ire m e n ts d e te rm in e d f o r t h e p rog en y a r e b a s e d on re­ s u l t s o b ta in e d w ith d i e t 5 w hich c o n ta in e d , s l i g h t l y more p r o t e i n th a n i s n orm a lly f e d .

S in c e a h ig h l e v e l o f p r o t e i n i n t h e d i e t in c re a s e * "

t h e n e e d f o r v ita m in Big f o r g ro w th , t h e a c t u a l r e q u ire m e n ts o f th e p ro g e n y f o r v ita m in Bqo ®ay be s l i g h t l y lo w e r th a n th o s e d e te r m in e d . On t h e o t h e r hand n o rm al l e v e l s o f p a n to th e n ic a c i d ( a v ita m in w hich h a s b een shown by Y & cow ilz, M o rris and H e u se r, 1 950, t o s p a re v ita m in Bl2 f o r c h i c k s ) , m e th io n in e and c h o lin e war© f e d i n t h e p u l l e t d i e t s

70 an d i n c h ic k d i e t 5*

T h e r e f o r e , i t i s b e lie v e d t h a t an y undue e f f e c t

o f t h e s e compound© on t h e v ita m in

re q u ire m e n ts h a s been e l im in a te d .

I t sh o u ld b e s t a t e d t h a t p o s s i b l y t h e p u l l e t s W iich w ere s e ­ l e c t e d f o r e x p e r im e n ts ! an d 2 w ere r e l a t i v e l y r e f r a c t o r y t o a v itam in d e fic ie n c y .

B ir d , R u b in , an d G roschke (1947) h a v e shown t h a t t h e r e

i s a g e n e t i c r e l a t i o n s h i p t© th e v ita m in Bjjg re q u ire m e n t o f t h e c h ic k e n w hich p e r m its some b i r d s t o f u n c tio n s a t i s f a c t o r i l y on l e v e l s q u i t e d e f i c i e n t f o r o th e r s *

The t e s t b i r d s war® s e l e c t e d on t h e b a s i s of' an

a v e r a g e h a t c h a b i l i t y o f 8 0 .8 p e r c e n t .

Only 1 b i r d i n each p u l l e t g ro u p

had a r e c o r d o f l e s s th a n 68 p e r c e n t o v e r a l l h a t c h a b i l i t y , when e x p e r i­ ment 1 was s t a r t e d .

P e rh a p s i t w ould h av e b e e n w i s e r to p e r a i t b i r d s

w ith r e c o r d s o f lo w e r h a t c h a b i l i t y d u r in g t h e p r e lim in a r y p e r io d t o b e u se d i n t h e 2 e x p e rim e n ts*

The r e s u lt© o f e x p e rim e n t 2 s u f f e r fro m

v a r i a b i l i t y a s a r e s u l t o f l i m i t e d num bers, and p o s s i b l y from s e a s o n a l e ffe c ts .

71

summary

The v ita m in

re q u ire m e n ts o f Hew H am pshire p u l l e t s and t h e i r

p ro g en y w ere s tu d ie d , u s in g a c e r e a l g r a in - s o y b e a n m eal ty p e o f d i e t .

In

t h e p u l l e t s t u d i e s a v e ra g e w e ig h ts , hem oglobin v a lu e s and m o r t a l i t y b o re no m e a s u ra b le r e l a t i o n s h i p t o t h e l e v e l o f v ita m in

i n th e d i e t .

D u rin g e x p e rim e n t 1 , eg g p r o d u c tio n d e c lin e d in d i r e c t r e l a t i o n s h i p t o d e c r e a s in g l e v e l o f v ita m in Big s u p p le m e n ta tio n .

The r e s u l t s o f e x p e r i­

m ent 2 d e m o n stra te d no c l e a r r e l t i o n s h i p o f t h e d ie t a r y l e v e l o f v ita m in B^g t o egg p r o d u c tio n .

Reason® d is c u s s e d p r e v io u s ly le a d th e

w r i t e r t o c o n c lu d e t h a t an y e f f e c t o f v ita m in b 12 on egg p r o d u c tio n c a n n o t be m easu red w ith su ch t e s t r a tio n ® .

Mo r e l a t i o n s h i p was p ro v en

b etw een f e r t i l i t y and t h e l e v e l o f v ita m in B-^ f e d t o th e p u l l e t s . H a t c h a b i l i t y d e c lin e d s h a r p ly i n e x p e rim e n t 1 when no s u p p le m e n ta l v ita m in

m s fu r n ish e d i n p u l l e t d i e t 1 .

The r e s u l t s o f experim ent

1 showed t h a t t h e re q u ire m e n t f o r h a t c h a b i l i t y was a d e q u a te ly met by 4 micrograms o f s u p p le m e n ta l v ita m in B^g p e r k ilo g ra m u n d er th e con­ d i t i o n s o f th e t e s t .

In e x p e rim e n t 2 , f l u c t u a t i o n s i n h a t c h a b ilit y

o c c u r re d betw een g ro u p s in such a m anner t h a t an y r e l a t i o n s h i p o f h a t c h a b i l i t y t o a lo w e r l e v e l o f v ita m in

was o b s c u re d .

R ate o f

d e p l e t i o n s t u d i e s w ith h a t c h a b i l i t y i n d i c a t e d , how ever, t h a t even 0 micrograms o f s u p p le m e n ta l v ita m in

m ig h t n o t p e rm it enough s to r a g e

i n t h e m i l l e t s * b o d ie s f o r s a f e t y . The v ita m in

a c t i v i t y o f e g g s was m easured by b o th m ic ro ­

b i o l o g i c a l and c h ic k a s s a y s .

I t was shown, t o b e a r a d i r e c t r e l a t i o n s h i p

to th e le v e l fe d th e p a l l e t .

Mien p u l l e t s w ere f e d a p r a c t i c a l b re e d in g

m ash, t h e v ita m in Bqg a c t i v i t y o f t h e a v e r a g e egg was 1 .5 6 0 mic ro c rams by c h ic k a s s a y .

When th e y w ere f e d a c e r e a l g ra in -so y b e a n . p r o t e i n ty p e

72

o f d i e t unsuppl^raented 'w ith v ita m in Bx2 t h e c o n t e n t was U.604 m icrogram s o f v ita m in

a c t i v i t y by c h ic k a s s a y .

The m ic r o b i o lo g ic a l a s s a y r e ­

s u l t s showed o n ly 9 ,5 t o 16*7 p e r c e n t &s much v ita m in t h e e g g s a s d id t h e c h ic k a s s a y s .

a c t i v i t y in

The r e s u l t s o f t h e c h ic k a s s a y s i n d i ­

c a t e d t h a t betw een SO and 48 p e r c e n t o f th e v ita m in w as t r a n s m i t t e d t o t h e e g g .

812

f e d t o th e p u l l e t s

The tr a n s m is s io n was more e f f i c i e n t a t low

l e v e l s o f s u p p le m e n ta tio n i n th e p u l l e t d i e t . I n i t i a l w e ig h t o f p ro g e n y was u n a f f e c te d b y t h e v a r i a t i o n s i n th e d ie t*

M o r t a l i t y was c o n s id e r a b ly h ig h e r I n p ro g en y o f t h e p u l l e t

g ro u p f e d t h e u n su p p le m en te d b a s a l d i e t 1 d u r in g t h e f i r s t experim ent* The e x c e s s iv e m o r t a l i t y among p ro g en y c o u ld be a v o id e d by t h e a d d i t i o n o f S microgram® o f v ita m in

to p u lle t b a sa l d ie t 1 o r th e a d d itio n

o f b etw een 9 and 27 m icrogram s o f v ita m in Bxg t o c h ic k b a s a l d i e t S . The a f f e c t on l i v e a b i l i t y o f lo w e r l e v e l s o f s u p p le m e n ta l v ita m in 3x2 i n t h e m a te rn a l d i e t i s unknown. When th e r e was no s u p p le m e n ta l v ita m in 3x2 in b o th th e m a te rn a l d i e t 1 and t h e c h ic k d i e t 5 , d e g re e o f f e a t h e r i n g was poor*

When 4 m ic ro -

gram s o f s u p p le m e n ta l v ita m in Bx2 w ere fe d i n th e m aternal d i e t 1 , o r $ m icrograsts w ere added t o t h e c h ic k d i e t S , d e g re e o f f e a t h e r i n g was s a t i s ­ fa c to ry .

The e f f e c t o f lo w e r l e v e l s o f supplem ental v ita m in B1 2 h a s n o t

b een s tu d ie d . G rowth o f p ro g en y was d i r e c t l y a f f e c t e d by b o th t h e m aternal and c h ic k d i e t a r y l e v e l o f v ita m in B ig .

F in a l w e ig h t o f t h e ch ick

pl& teaued a t a n e a r l y optimum p o i n t when a kilogram, o f t h e m a te rn a l d i e t c o n ta in e d

8

micrograms o f sup p lem en tal v ita m in Bx2 and a k ilogram , o f t h e

c h ic k d i e t c o n ta in e d 5 m icro g ram s o f s u p p le m e n ta l vitam in Bxg*

Thus©

l e v e l s o f v ita m in Bx2 a r e b e l i e v e d t o mark t h e lo w e r l i m i t s f o r b e s t

?$ g ro w th on su ch a r a t i o n .

Mien 16 m icrogram s o f s u p p le m e n ta l v ita m in

B^2 w ore f e d t o t h e p u l l e t s i n s t e a d o f 8 , t h e f i n a l w e ig h t o f th e p ro g en y p la te a u e d a t a s l i g h t l y h ig h e r l e v e l .

The p o i n t o f p l& te a u in g

in v o lv e d t h e same l e v e l o f s u p p le m e n ta l v ita m in i n th e c h ic k d i e t f o r b o th g ro u p s .

C o r r e l a t i o n o f t h e r e s u l t s o f th e egg a s s a y s by c h ic k s

and t h e grow th o f p ro g en y showed t h a t v ita m in

f u r n is h e d by th « egg

m s f a r more e f f e c t i v e l y u t i l i s e d th a n v ita m in B is f u r n is h e d i n th e c h ic k d i e t .

L e ss v ita m in B^g was r e q u ir e d , i p i a n t i t a t i v e l y , f o r good

g ro w th o f c h ic k s when m ost o f i t was f u r n is h e d v i a t h e m a te r n a l d i e t th a n i f f u r n i s h e d th ro u g h t h e c h ic k d i e t . t h e l e v e l o f v ita m in

T hese r e s u lt® i n d i c a t e t h a t

f e d t o t h e da® i s o f v i t a l im p o rta n c e f o r t h e

p r o d u c tio n o f r a p i d l y g ro w in g c h ic k s .

Growth I n o n e s e r i e s t o 8 weeks

i n d i c a t e d t h a t t h e l e v e l s determ ined f o r s a t i s f a c t o r y grow th t o 4 w eeks o f ag e may a l s o b e u se d f o r t h e f i r s t 8 w eeks o f l i f e . S ix te e n m ic ro grains o f s u p p le m e n ta l v ita m in p e r m it t e d th e p u l l e t s t o b u i l d up a r e s e r v e . m e n ta l v ita m in

p e r k ilo g ra m

E ig h t microgram© o f s u p p le ­

d id m a in ta in h a t c h a b i l i t y and d id m a in ta in grow th o f

p ro g e n y f e d S micrograms o f t h e v ita m in .

The im m ed iate d ro p i n h a tc h ­

a b i l i t y o f e g g s and d ro p in g ro w th f o llo w in g o m iss io n o f t h e v ita m in , h o w ev er, showed t h a t such a l e v e l d id n o t p e rm it b u ild - u p o f an a d e q u a te r e s e r v e in th© p u l l e t f o r e m e rg e n c ie s .

74

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A

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7?

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Com m ercial APF cone a n t r a t e s 4 6 8 -4 6 7 .

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V itam in

814

and c e l l

Novak, A. F . and 3 . M. H&uge, 1 9 4 8 . I s o l a t i o n o f t h e u n i d e n t i f i e d g row th f a c t o r ( v ita m in B is ) i n d i s t i l l e r s * d r i e d s o l u b l e s . J . B i o l . Chem. 174s 6 4 7 -6 5 1 . O lc e s e , 0 . , J . R. Couch and C. M. lym an, 1950. V itam in Big c o n c e n tr a te s i n t h e n u t r i t i o n o f t h e m a tu re d o m e stic f o w l. J . N u t r i t i o n 4 1 1 7 5 -8 7 . O ls e n , M. ¥ . , 1 9 4 9 .

P e r s o n a l co m iau n icatio n .

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19 4 8 T b T ”

?itai?iin "'% £ a c o b a lt co m p lex .

S c ie n c e i S i

15 4 .

1946 ^(c) • C o M p a ra tiv e ^ d a ta on v ita m in Bjg from l i v e r and from a new s o u r c e , Streptom ycea g r is e u e . S c ie n c e 106s 8 5 4 -6 5 5 .

H o b b le s , A. R ., C. A. M ic h o l, W. W. C ra v e n s, C. A. E lvehjem and J . J . H a lp in , 1 9 4 8 . The e f f e c t o f hen d i e t an d c h ic k d i e t on t h e need o f c h ic k s f o r a n u n id e n t if ie d grow th f a c t o r . P o u ltr y S c i. 27s 4 4 2 447. R o b in so n , R o b e r t, 1 9 4 8 . A dvances i n b io c h e m is tr y . P roblem s o f n u t r i t i o n and d i s e a s e . C hem ical T rad e J . 125s 6 8 5 -0 6 6 * R o ss, J . P . , H. B a ld in g , an d B . L. P a e g e l, 1948* Developm ent and p ro ­ g r e s s io n o f s u b a c u te com bined d e g e n e r a tio n o f s p i n a l c o rd i n p a t i e n t s w ith p e r n ic io u s anemia t r e a t e d w ith s y n t h e t i c p t e r o y l glut& m ic ( f o l i c ) a c i d . B lood 5s 6 8 . R u b in , Max, and H. R. B ir d , 1946 ( a ) . A c h ic k g ro w th f a c t o r i n cow m an u re. I . I t s n o n - id e n t it y w ith c h ic k g ro w th f a c t o r s p r e v i o u s ly d e s c rib e d .. J . .B io l. Chem. 165s S8 7 -592• R u b in , Max, and H. R. B ir d , 1946 ( b ) . A chick; grow th f a c t o r i n cow m anure. I I . The p r e p a r a tio n o f c o n c e n tr a te s a n d t h e p r o p e r t i e s o f t h e f a c t o r . J . B i o l . Chem. 1 6 5 1 595—4 0 0 . R u b in , Max, and H. R. B ir d , 1947 ( a ) . The c h ic k grow th f a c t o r o f cow isa n u re , a c o u n t e m e t a n t o f e f f e c t s o f e x c e s s iv e soybean m e a l. F e d . P ro c . 6s 4 1 9 . R u b in , M. and II. R. B ir d , 1947 ( b ) . A c h ic k grow th f a c t o r i n cow m anure. I I I . I t s o c c u r re n c e i n e g g s . P o u l t r y S c i . 26s 5 0 9 -5 1 0 . S c h a e f e r , A. E . , W. D. Salmon and D. E . S tr e n g th , 1 9 4 9 . R e la tio n o f v ita m in By? t o c h o lin e re q u ire m e n t o f t h e r a t and th e c h ic k . F e d . P ro c . 8s 595. S h iv a , W., J . M. H av el and W. M. H a rd in g , 1 9 4 8 . An in t e r r e la t i o n s h ip o f p u r in e s and v ita m in B ^ . J* B io l. Chem. 176s 9 9 1 -9 9 2 .

S h o rb , Mary S . , 1947 { a ) . U n i d e n t if ie d e s s e n t i a l grow th f a c t o r s f o r L a c t o b a c i l lu s l a c t i s fo und in r e f in e d l i v e r e x t r a c t s and i n c e r t a i n n a t u r a l m a t e r i a l s . J . B a e t. 55s 6 6 9 . _____________ , 1947 ( b ) . U n id en tifie d grow th f a c t o r s f o r L a c t o b a c i l lu s l a c t i s i n r e f i n e d l i v e r e x t r a c t s . J . B io l. Chen. 169s 4 6 5 -4 5 6 . » 1 9 4 8 . A c t i v i t y o f v ita m in Big f o r th e grow th o f L a c t o b a c i l lu s l a c t i s . S c ie n c e 10 7 : 5 9 7 -5 9 8 . S k in n e r, J . L. and J . H. Q u ls e n b e rry , 1950. Some a f f e c t s o f h ig h e f f i c i e n c y and i n c h ic k e n b r e e d e r r a t i o n s . P o u ltr y S c i. 2 9 ? 780. S m ith , S . L e s t e r , 1946 ( a ) . P u r if ic a t io n o f a n t i —p e r n i c io u s anaem ia f a c t o r s from l i v e r . M ature 161s 6 5 6 -6 5 9 . * 1946 ( b ) . P re se n c e o f c o b a l t i n th e a n t i —p e r n ic io u s an aem ia f a c t o r . M ature 1 6 2 : 1 4 1 -1 4 5 . _______________ , 1 9 4 9 . L e c tu r e . M ary lan d . D ec. 1 2 th .

U. S . P u b lic H e a lth D e p t., B e th e s d a ,

S m ith , S . L e s t e r , W. F . J . C u lb e r ts o n , A. M&lker and K. A. L e es, 1950. P a r t i t i o n c h ro m a to g ra p h y o f t h e v ita m in gro u p o f f a c t o r s . F e d . Proc* 9 s 2 5 0 . S m ith , D o ro th y G. and. B a rry J . R o b in so n , 1945. The i n f l u e n c e o f s tr e p to m y c in and s t r e p i o t h r i c i n on t h e i n t e s t i n a l f l o r a o f n ic e . J . M e t . 50s 6 1 5 -6 2 1 . S te p h e n s o n , S . L ., 1 . M cG innis, W. D. Graham and J . S . C a rv e r, 1946. E f f e c t o f s o i l on c h ic k g ro w th . P o u ltr y S c i . 2 7 : 8 2 7 -8 2 0 . S t e r n , J . R ., M. ¥ . T a y lo r and U". C. R u s s e ll , 1949, R e la tio n o f v ita m in Byg t o l i v e r b a s o p h i l i a . P ro c . S o c. &xp. B io l, and Med. 70? 5 5 1 552. S te v e n s , l o a n , Ja c o b R ie ly an d B. M arch, 1 9 4 9 . A n o te on th e wa n im a l p r o t e i n f a c t o r " and f e e d e f f i c i e n c y . P o u ltr y S c i . 2 0 : 9 2 1 -9 5 2 . S t o k s t a d , JS. L. R ., C. II. H offm ann, and T . H. J u k e s , 1 9 4 9 . C o m p arativ e m i c r o b i o l o g i c a l , a n im a l, and c l i n i c a l a s s a y s o f p r e p a r a t i o n s con­ t a i n i n g v ita m in B ig . Abs* 1 1 6 th m e e tin g , Amer. Chem. S o c ., p . 54A. S t o k s ta d , E . L . R ., T. H.J u k e s , John A. Brockm an, J . 7 . P i e r c e and H. P . B r o q u is t, 1 9 5 0 . R e la tio n o f v ita m in Bfgb t o v ita m in tejg and b i o l o g i c a l a c t i v i t i e s o f t h e s e com pounds. F e d . P ro c . 9s 2 5 5 . S t o k s t a d , £ , L . R ., T . H. J u k e s , J . P i e r c e , A. C. P ag e, J r . , and A. L. F r a n k l i n , 1 9 49, The m u l t i p l e n a tu r e o f t h e a n im a l p r o t e i n f a c t o r . J . B i o l . Chem. 180? 6 4 7 -6 5 4 .

S to k s ta d , S . L. R*, A. P ag e , J r . , J . P i a r c s , T. H. J u k e s , R. W. H e in le , M, E p s te in and A. 1). W elsh, 1948. A c t i v i t y o f m ic r o b ia l anim al p r o t e i n f a c t o r c o n c e n t r a te s i n p e r n ic i o u s a n e m ia . J . L ab. C lin . Med. 35s 880—864• To m b e r g , J . L. and R. IS. E a k in , 1949. S r y th e in and a p o e r y th e in an d t h e i r r e l a t i o n t o t h e a n t i - p e r n i c i o u s anem ia p r i n c i p l e . J . Amer. Chem. S o c. 7 1 ; 3 8 58. W est, R ., 1948. A c t i v i t y o f S c ie n c e 1 07; 398.

v ita m in

i n A d d is o n ia n

p e r n ic io u s an e m ia .

W e tz e l, N oraan C ., W arren C. F a rg o , I s a b e l II. Sm ith and J o s e p h in e H e llk s o n , 1 9 4 9 . Growth f a i l u r e i n s c h o o l c h i l d r e n a s a s s o c i a t e d w ith v ita m in Big d e f i c i e n c y - re s p o n s e t o o r a l t h e r a p y . S c ie n c e 1 1 0 ; 6 5 1 -6 5 3 . W h itso n , D ., J . C. Hammond, H. W. T it u s and H. R. B ir d , 1945. The u s e o f soybean o i l m eal i n t h e d i e t o f grow ing c h ic k s . I I . E f f e c t o f d i f f e r e n t g r a i n s . P o u ltr y S c i . 2 4 ; 408—4 1 6 . W h itso n , D ., H. T i t u s , and II. B ir d , 1 946. The e f f e c t o f f e e d in g cow m anure on egg p r o d u c tio n and h a t c h a b i l i t y . P o u ltr y S c i . 2 5 ; 1 4 5 147. Woods, R u th , 1 9 4 9 . R ecen t advances i n th® know ledge o f a n ti- a n e m ia f a c t o r s . B o rd e n 1s Review o f N u t r i t i o n a l R e s e a rc h . X( 9 ) i 1 - 1 2 . W rig h t, L . D ., H. R. Skeggs and J . ¥ . H u ff, 1 948. The a b i l i t y o f th y m id in e t o r e p la c e v ita m in Byg a s a grow th f a c t o r f o r c e r t a i n L a c t o b a c i l l i . J . B i o l . Chem. 17 5 ; 4 7 5 -4 7 6 . Y & cow iiz, H ., L. C. N o r r is an d G. F . H a u se r, 1 950. E vidence o f i n t e r ­ r e l a t i o n s h i p s b etw een v ita m in and r i b o f l a v i n , p y rid o x in ® an d p a n to th e n ic a c i d , P o u ltr y S c i . 2 9 ; 78 7 . Y a c o w itz , H a ro ld , H. T. P e e l e r , R. F . K i l l e r , G. W. C a r ls o n , G. F . H e u se r and L. G. N o r r i s , 1950* S tu d ie s o f t h e e f f e c t s o f 3^2 on h a tc h a b ility . In fo rm a l P o u ltr y N u t r i t i o n C o n fe re n c e , A t l a n t i c C i t y , A p r i l 1 7 th . E u c k e r, L o is M. and T h eo d o re F . t u c k e r , 1948 ( a ) . E o o p h e rin ; & n u t r i t i o n a l f a c t o r f o r r a t s a s s o c i a t e d w ith a n im a l p r o t e i n s o u r c e s . A rch . B iochem . 1 6 ; 1 1 5 -1 2 9 . E u c k e r, T . P . , an d L . K. E u c k e r, 1946 ( b ) . B io s y n th e s is o f s o o p h e r in . A bs. 1 1 4 th m e e tin g Amer. Chem. S o c ., pp 52C - 55C.

82 ArrM4DlA TABU'S I C o m p o sitio n o f S a l t Mix and V itam in Mix f o r S a s a l .Diet 4 D ie ta r y I n g r e d i e n t CaCO$ K2 HPO4 v Ca*( PO4 X0 la C l NsgHFC* MgSC;4*7 % Q Fe{C 6 % 0 7 ) o . 8 KI H nS04.4 HgO ZnClg H5 BO4 C0 SO4 . 7 HgO GUSO4 % po4

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