University of Nigeria               Research Publications 

KAMALU, Beryl Perry

Aut

hor

PG/M.Sc/79/465

Title

Experimental Cyanide Toxicosis in Growing Male Dogs on Nutritionally Balanced Diets

Clinical Chemistry and Histopathology

Facu

lty

Veterinary

Dep

artm

ent

Veterinary Pathology and Microbiology

Dat

e

1989

Sign

atur

e

EXPERIMZNTAL CYANIDE T O X f COSIS IN GROWING MALE DOGS ON NUTRITIONALLY

BALANCED DIETS - CLINICAL CHEMISTRY AND HISTOPATHOLOGX

KAMALU , BERYL PEiiHY B.S. (Tuskegee) 1972 D.V.M. (~uskegee) 1974

k Dissertat ion presented for the Degree of Master of Science in the

Department of Veterinary Pathology and Microbiology,

University o f N i ~ e r ia , Nsukka

Supervisors: Professor M. Uzoukwu, B.V.M.S. (~lasgow) , MoSm ( N - S - U - ) , PhoD ( M ~ s ~ U . ) , MmR*CoVmS.

D r . 0. Obidoa, B . S c . , Ph.D (1badCm)

DEDICATION

Dedicated t o my l a t e mother #

BERYL ALBZBTHA VIRGIN.^ FERRY

t h i s .jr sng o ther University.

ii.

I wish t c express my sincerc thanks t o my s u p e r v i s o r s ,

P r o f e s s o r M. Uzoukwu, Department of Veterinary Fathology and

l i icrobiolol;;y, Faculty of Ve te r ina ry Medicine, and Dr . 01 Obiem,

Department o f Biocheni is t ry , F a c u l t y of Bicl logical Sciences.

I wish t o thsnk my family f o r p r o v i d i n ~ t h e reagents and

m a t e r i a l s for this p r o j e c t .

I wish t o thank Mr. F. Mwokike, N r . X. Cnuoha, Mr. E.O. Chinbo,

M r . C.C. Umeagudosu and Mr, 0. Ovaga f o r t e c h n i c a l a s s i s t a n c e .

BERYL PERRY KAWiLLU 1989

TABLE OF CCNTENTS

CE2T I F IC!:T I O N a

ACKNO'dLEDGLMENT

TjiBLE OF CONTENTS . L I S T OF TABLES a a

L I S T OF FIGURES a o

L I S T OF APPENDIX 0 .

ABSTRACT o o

INTRODUCTION o

LITERATURE REVIEW . O

Mf;PERII.LS X N D MZTHODS

RESULTS - .

D I S C U S S ION o o

CONCLUSION . REFERERCES e . APPLTDIX . a

LIST OF TABLES

Page

Compc:,sition o f d i e t s on d r y m a t t e r ca.lcu.lnt i o n s

Prcxi rn~. t e a n a l y s i s cf wet $ i e t s f e d t o a n i m a l s

E f f e c t :?f cyzni(1e c o n t a i n i n g d i c t s on a p ~ s a r e n t d r y incitter d i g e s t i b i l i t y

E f f e c t o f cyan ide c o n t a i n i n 3 d i e t e on we igh t of the t h y r o i d g l and a f t e r 14 wceks

2 f f ~ c t o f c y n n i e e c o n t a i n i n g d i e t s on plasma cnzymcs

E f f cc t of cyaniCc c o n t a i n i n g d i e t s on h m o ~ l o b i n c o n c e n t r a t i o n o f b l o o d a n d ~ a c k e c l cell volume nftcr 14 weeks

Effcct af cyanide c o n t a i n i n g c l i e t s on plasma e l e c t r o l y t e s

LIST OF FIGURES

F i g u r e Page

I Mean weekly p e r c e n t a g e change i n we igh t d u r i n g t h e e x p e r i m e n t a l p e r i o d 39

2 L i n e a r r e g r e s s i o n of p e r c e n t a g e change i n weight a g a i n s t t ime 40

3 V a r i a t i o n of mean (;tl S.E.M.) t o t a l sernm t r i ~ 6 d o - t h y r o n i n e (T ) c o n c e n t r a t i o n w i t h t ime during t h e e x p e r i m e a t a l p e r i o d

4 V a r i a t i o n of mean (2 SS,.blM.) plasma t h i o c y a n a t e c o n c e n t r a t i o n w i t h t ime d u r i n g t h e e x 3 e r i m e n t a l p e r i o d

5 L i n e a r c o r r e l a t i o n between plasma t h i o c y a n a t e c o n c e n t r a t i o n a n d t o t a l serum t r i i o d o t h y r o n i n e c onc en t rat i o n

6 V a r i a t i o n cbf mean (;t S.E,M.) u r i n e t h i o c y n n a t e c o n c e n t r n t i o n w i t h t i m e d u r i n g t h e e x p e r i m e n t a l p e r i o d

7 L i n e a r c o r r e l a t i o n between p lasma t h i o c y a n a t e c o n c e n t r a t i o n and u r i n e t h i o c y z n a t e c o n c e n t r a t i o n

8 V ~ . r i n t i o n of mean ( + s ,E,M.) u r i n e p r o t e i n c o n c e n t r n t i o n with Time d u r i n g t h e e x p e r i m e n t a l p e r i o d

9 Mean (2 s.E.M,) r a t e of c l o s u r e of e p i p h y s e a l p l a t e s d u r i n g e x p e r i m e n t a l p e r i o d

I 0 T h y r o i d g l a n d of a n i m a l on c o n t r o l d i e t

11 T h y r c i d g l a n d o f a n i m a l on c o n t r o l d i e t . Cubo ida l e p i t h e l ium l i n i n g f o l l i c l e f i l l e d with c o l l o i d

12 T h y r o i d g lnnd of a n i m a l nn c o n t r o l d i e t . Columnar e p i t h e l i u m l i n i n g f o l l i c l e c o n t a i n i n g s c a n t y c o l l o i d

13 T h y r o i d gland of an ima l on c o n t r c l d i e t . Desquamation of e p i t h e l i u m into lumen of empty f o l l i c l c

' v i .

F i g u r e

14 T h y r c i d g l a n l of an ima l on g a r i d i e t

15 T h y r o i d g l a n d of an ima l on g a r i d i e t . Low c u b o i d a l ep i the l iuml l i n i n g f o l l i c l e f i l l e d w i t h c o l l o i d

1 6 T h y r o i d g l a n d of zn ima l on r i c e + c y a n i d e d i e t

17 T h y r o i d gland of an imal on r i c e + c y a n i d e d i e t . Th ickened f o l l i c u l a r walls compcsed of c u b o i d a l c e l l s

1 8 Thyroid gland of an ima l nn r i c e + c y a n i d e d i e t . Desquarnntion of e p i t h e l i u m i n t 2 lumen of empty f o l l i c l e

19 E p i p h y s e a l growth ; ) l a t e of nnirnal on r i c e + c y a n i d e d i e t

20 L i v e r o f animal on g n r i d i e t . P e r i p o r t a l v a c u o l z t i o n

2 1 P a n c r e a s of an ima l on g n r i d i e t . F r a g m e n t a t i o n o f n c i n n r t i s s u e

22 P a n c r e a s of an ima l on g n r i d i e t Hemorrhage

2 3 P x n c r e a s @f anim2.l on g z r i d i e t . F i b r o s i s of i s l e t of Lnngerhans

24 F a n c r e n s c.f nriirncl :?n r i c e + cyanicie d i e t . F i b r o s i s i n ncinar t i s s u e

25 P m c r e n s of a n i m a l on r i c e + c y a n i d e d i e t . F i b r o s i s of islet of Langerhans

26 K i d n e y csf an ima l on g a r i d i e t . Cytoi;lasm from e p i t h e l i z l c e l l s p u s h i n g ou t a c r o s s t h e lumen of r r o x i m a l t u b u l e s

27 Kidney of an ima l on r i c e + c y a n i d e d i e t , Czsts i n lumen of p rox ima l t u b u l e s

vii.

L I S T OF APPENDIX

Appendix

1

Page

E f f e c t o f cy:nide c o n t a i n i n g ~ ~ i c t s on weight change ( l u r i n g 14 week expc r i rnen tz l p e r i o d 116

E f f e c t of cynn ide c o n t a i n i n g d i c t s on plasma T c o n c e n t r a t i o n

3 117

E f f e c t o f cynnicle c o n t a i n i n g d i e t s on plasma t h i o c y a n a t e c o n c c n t r n t i o n 1 18

E f f e c t of cyanicle c o n t a i n i n g C i e t s on u r i n e t h i o c y z n a t e c o n c e n t r a t i o n 119

E f f e c t cf c y a n i d e c o n t a i n i n g d i e t s on u r i n e y r o t e i n c c n c e n t r a t i o n 1 20

E f f e c t o f c y s n i d e c o n t , - . i n l n ~ d i e t s on ep iphysen l c l o s u r e 121

v i i i .

ABSTRACT

The e f f e c t s of p ro longed c y a n i d e c o n t a i n i n g d i e t s i n

growing male dogs were s t u d i e d o Three g roups of dogs were p l a c e d

on 3 d i f f e r e n t i s o c z l o r i c d i e t s c o n t a i n i n g 13% c r u d e p r o t e i n on

a dry m a t t e r basis. Group 1 r e c e i v e d t h e c o n t r o l d i e t ( I ) which

c o n s i s t e d of r i c e as t h e c a r b o h y d r a t e s o u r c e , group 2 w a s f e d gari

as the c a r b o h y d r a t e s o u r c e ( T I ) and group 3 r i c e t o which cyanide was

added t o t he l e v e l of t ha t found i n gari was the carbohydrate s c u r c e

(111). The fo l . iowing parameters were neasured: - a p p a r e n t d i g e s t i -

b i l i t y of t h e diets, weekly weight g a i n , r a t e of closure of

u n c a l c i f i e d ep ip l iy sea l growth p l a t e , t o t a l serum t r i i o d o t h y r o n i n e

c oncent r a t i o n , plasma t h i o c y a n a t e concentrat i o n , u r i n e t h i o c y a n a t e

c o n c e n t r a t i o n and s e l e c t e d p lasma enzyme a c t i v i t i e s , h e m a t o l o g i c a l

parameters and plasma e l e c t r o l y t e s . The dogs were e u t h a n i z e d and

necrops ied at t h e end of e x y e r i m e n t a l week 14. The t h y r o i d gland

was removed and weighed. The t h y r o i d g l a n d , t h e o l e c r a n o n

p roce s s , l i v e r , p a n c r e a s and kidney were p r o c e s s e d f o r h i s t o -

p a t h o l o g i c a l examina t ion .

Diet I ( r i c e o n l y ) produced no d e t e c t a b l e plasma

t h i o c y a n a t e whereas b o t h d i e t s I1 ( p r i d i e t ) and 111 ( t h e r i c e +

c y a n i d e d i e t ) p roduced plasm t h i o c y a n a t e w i t h d i e t I1 producing

s i g n i f i c a n t l y l e s s plasma t h i o c y a n a t e t h a n d i e t I11 ( r i c e +

cyanide d i e t ) . The h i g h e r l e v e l i n t h e latter p r o b a b l y i n d i c a t e s

that a greater q u a n t i t y of f r e e c y z n i d e was a v a i l a b l e f o r conversion

t o t h i o c y a n a t e , There was no s i g n i f i c a n t d i f f e r e n c e i n t h e e f f e c t

ax.

of d i e t s I , 11 and 111 on t h e f o l l o w i n g p a r a m e t e r s : d i g e s t i b i l i t y ,

b lood hemoglobin concent r a t i o n and packed c e l l volume. D i e t s I

and I1 d i d n o t d i f f e r s i g n i f i c a n t l y i n t h e i r e f f e c t on growth and

t h e t h y r o i d g l a n d b u t d i e t 111 s i g n i f i c a n t l y a f f e c t e d growth and

t h e t h y r o i d g land . D i e t I h a d no e f f e c t on t h e p a n c r e a s , b u t d i e t

I1 produced e f f e c t s on t h e p a n c r e a s t h a t were s i m i l a r t o d i e t 111-

S i m i l a r l y , d i e t I had no e f f e c t on t h e k i d n e y , however b o t h t h e gar5

d i e t I1 and r i c e + c y a n i d e d i e t I11 h a d ha rmfu l e f f e c t s on t h e

k idney . The re were no s i g n i f i c a n t d i f f e r e n c e s be tween d i e t s I and

111 on the l i v e r , plasma po ta s s ium and plasma c a l c i u m l e v e l s , b u t

p lasma p o t a s s i u m and p lasma c a l c i u m were s i g n i f i c a n t l y l o w e r i n t h e

animals on d i e t 11.

Die t 111 was more t o x i c t h a n d i e t I1 even though t h e l e v e l

of cyanide was the s m e , Growth, t h e t h y r o i d g l a n d , t h e p a n c r e a s

and t h e k i d n e y were a f f e c t e d by t h e r i c e + c y a n i d e d i e t . The

l i v e r , t h e p a n c r e a s and t h e k idney were a f f e c t e d by d i e t 11.

Growth and t h e t h y r o i d g l and were n o t a f f e c t e d by d i e t 11. The

d e g r e e of t o x i c o s i s t h e r e f o r e d i d n o t depend on t h e l e v e l of

c y a n i d e i n t h e d i e t p e r s e , b u t appeared to be r e l a t e d t o t h e

level of f r e e c y a n i d e r e l e a s e d by metabol ism, Bound c y a c i d e

a p p e a r e d t o e x e r t l i t t l e i n f l u e n c e . The v a c u o l a t i o n i n t h e

l i v e r a n d k idney and t h e hypacalcemia of t h e a n i m a l s on d i e t I1

b u t n o t i n t h e a n i m a l s f e d d i e t I o r 1x1 a p p e a r e d t o be s e c o n d a r y

t o t h e hypokalemia. The hypokalemia i n t he a n i m a l s f e d d i e t I1

canno t be e x p l a i n e d on t h e b a s i s of c y a n i d e t o x i c o s i s . Previous

workers have s u ~ ~ e s t ed t h a t l inarnar in , the cyanogenic glut os iGe

of g a r i e x e r t s a t o x i c e f f e c t on scclium-pctassium a d e n o s i n e

t r i p h o s ~ h n t a s e altering t h ~ s o c J i u n - y t a s s i u m t r a n s p o r t sys t em

i n such a way t h z t t i s s u e s Sccomc d e r l c t e d o f potass ium.

INTRODUCTION

The most w ide ly c u l t i v a t e d c a s s a v a p l a n t is known b o t a n i c a l l y

a s Manihot e s c u l e n t a C r a n t z and b e l o n g s t o t h e f a m i l y Euphorbiaceae .

The impor t ance o f t h e r o o t of t h e c a s s a v a p l a n t w a s f i r s t r e c o g n i z e d

i n t r o 9 i c a l America. Sometime a f t e r 1504, c a s s a v a was t a k e n by

t h e P o r t u g e s e t o t h e Cape Verde I s l a n d s and from t h e r e t o A f r i c a

a n d A s i a oran an, 1976). Four hundred y e a r s l a t e r i t s impor t ance h a s

n o t d imin i shed . Cassava i s a n i m p o r t a n t s t a p l e f o o d f o r a b o u t 800

million people i n eve ry p a r t o f t h e t r o p i c s . With few e x c e p t i o n s ,

t h e human f o o d market h a s been and c o n t i n u e s t o be t h e major

o u t l e t f o r c a s s a v a , Cassava i s u s u a l l y consumed i n e i t h e r t h e

f r e s h o r p r o c e s s e d form n e a r growing a r e a s ( ~ h i l l i p s , 1982)-

I n b o t h A f r i c a and L a t i n America c a s s a v a i s t h e second o r t h i r d

moet i m p o r t a n t c z l o r i e s o u r c e o Consumption is l o w e s t i n S o u t h e a s t

A s i a ( ~ h i l l i p s , 1982) -

N a t i o n a l stat i s t i c s of t e n mask t h e r e g i o n a l dependence on

c a s s a v a as z s t a p l e , T h i s w a s t r a g i c a l l y h i g h l i g h t e d by t h e

R i g e r i a n C i v i l War of 1967-69, I n N i g e r i a c a s s a v a i s more i m p o r t a n t

i n t h e s o u t h e r n p a r t of t h e c o u n t r y t h a n i n t h e n o r t h . I n a

s u r v e y o f s o u t h e r n N i g e r i a , N i c o l (1952) found t h a t 25-56% of t h e

d i e t a r y c a l o r i e s came from c a s s a v a as opposed t o t h e n a t i o n a l

f i g u r e of 14% ( N e s t e l , 1 9 7 3 ) ~

Because of i t s h a r d i n e s s a n d p r o d u c t i v e c a p a c i t y , c a s s a v a

p r o d u c t i o n t e n d s t o be r e l e g a t e d t o l e s s f a v o u r a b l e s o i l s . It

h a s r e l a t i v e l y good r e s i s t a n c e t o many d i s e a s e s , p e s t s a n d d rough t

2.

( ~ h i l l i ~ s , 1982). From t h e p o i n t of view of food ene rgy o u t p u t

v e r s u s l a b o u r i n p u t , c a s s a v a a p p e a r s t o be v e r y e f f i c i e n t (Chandra

e t a1 19741, - -- The f a c t t h z t c a s s a v a p r o d u c t i o n h a s k e p t p a c e w i t h p o p u l a t i o n

growth i s r e m a r k a b l e , e s p e c i a l l y when i t i s c o n s i d e r e d t h a t t h i s

e x p a n s i o n h a s been a c h i e v e d on small fa rms employing l a b o u r - i z t e n s i v e methods. It s h o u l d b e n o t e d t h a t t h i s is i n c o n t r a s t

t o d e c l i n i n g p e r c a p i t a c e r e a l p r o d u c t i o n i n t h e t r o p i c s ( ~ a d l e r

a n d Yang, 1976).

Because c a s s a v a c o n t a i n s cyanogen ic g l u c o s i d e s i t i s u s u a l l y n o t

e a t e n raw. The cyanogen ic g l u c o s i d e s a r e h y d r o l y z e d t o produce

hydrocyan ic a c i d ( H C M ) , T r a d i t i o n a l l y c z s s a v a i s p r o c e s s e d i n t o

v a r i o u s d r i e d p r o d u c t s , P r o c e s s i n g , l i k e p r o d u c t i o n is c a r r i e d ou t

on a small s c a l e , u s i n g s imp le t e c h n i q u e s such as s o a k i n g ,

f e r m e n t a t i o n and f r y i n g . P r o c e s s i n g is u n d e r t a k e n t o remove t h e

cyanogen ic g l u c o s i d e a n d t o c o n v e r t t h e c a s s a v a t o a s t o r a b l e form.

Accord ing t o Oke (19821, c a s s a v a f l o u r h a s been u s e d a s a

s u b s t i t u t e f o r b a k i n g f l o u r s i n c e t h e 1950 ' s . Bread baked i n B r a z i l a n d

Piadagascar c o n t a i n s abou t 10-13% c a s s a v a f l o u r by l a w . I n some

p a r t s of t h e t r o p i c s , c a s s a v a r o o t s a r e f e r m e n t e d t o make b e e r ,

and a l s o i n t h e p r o d u c t i o n of a l c o h o l , I n B r a z i l , t h e a l c o h o l

p roduced is b lended at a r a t i o of 20:80 w i t h g a s o l i n e f o r f u e l ,

I n Nor th America and Europe , c a s s a v a i s made i n t o g e l and u s e d

as a t h i c k e n e r i n conven ience foods. I t is a l s o made i n t o commercial

s t a r c h , It i e a good s u b s t r a t e f o r p r o d u c i n g s i n g l e - c e l l p r o t e i n s .

B e s i d c s i t s i m p o r t a n t r o l e i n human n u t r i t i o n , c a s s a v a also

3 0

h a s been u s e d as a f e e d s t u f f f o r l i v e s t o c k , and e s p e c i a l l y a s a n

ene rgy s o u r c e f o r swine i n t h e P h i l l i p i n e s , ~ f r i c a and many a r e a s

of L a t i n America. The whole c a s s a v a r o o t , t h a t i s , p u l p and p e e l ,

i s normal ly u s e d i n p r a c t i c a l an ima l f e e d i n g i n t h e s e a r e a s

(Waner a n d Gornez, 1973).

S i n c e t h e mid-1960's p r o c e s s e d c a s s a v a was p l a c e d among t h e

z o s t i m p o r t z n t c e r e a l s u b s t i t u t e s i n an imal f e e d w i t h i n t h e Yuropean

Community, I n 1981, 6.5 m i l l i o n t o n s of p r o c e s s e d c a s s a v a was impor t ed

into Europe. Germany, t h e N e t h e r l a n d s , a n d ~ e l ~ i u m / ~ u x e m b u r g a c c o u n t

f o r 90% of t h c Community i m p o r t s , S e v e n t y f i v e p e r c e n t of t h e

c a s s a v a impor t ed ia u s e d i n p i g r a t i ons , p o u l t r y and cattle ratians

s h a r e t h e r e s t e q u a l l y , Cassava c o m p r i s e s up t o 30-4Pk o f p i g a n d

p o u l t r y r a t i o n s , f o r d i a r y c a t t l e , t h e maximun c a s s a v a c o n t e n t is

20%, b u t o n l y 10% is norma l ly u s e d (Renshaw, 1982).

P r o l o n g e d i n g e s t i o n of c a s s a v a h a s been i m p l i c a t e d i n a

number of n u t r i t i o n a l i n s u f f i c i e n c e s a n d m e t a b o l i c d i s e a s e s i n b o t h

m m and l i v e s t o c k (Osuntokun and Nmekosso , 1969; Ekpechi e t a l e ,

1966, Geevarghese , 1968; Naner and Gonez, ,1973; Vogt and P e n n e r ,

1963, Delange, 1974). These s t u d i e s s u g g e s t t h a t t h c s e problems

a r e r e l a t e d e i t h e r t o n u t r i t i o n s 1 d e f i c i e n c i e s i n d i e t s c o n t a i n i n g

cassava as t h e c a r b o h y d r a t e s o u r c e or t o t h e consumpt ion of

p r o c e s s e d cassava c o n t a i n i n g e x t r e m e l y h i g h l e v e l s of r e s i d u a l

cyanogen ic gluc os ide s ,

Most dogs l i v e i n homos wi th man and a r e f e d from l e f t o v e r

food o r t a b l e s c r a g s . T h e r e f o r e t h e d i e t a r y p a t t e r n of t h e owner

may i n f l u e n c e t h e d i e t a r y p a t t e r n of t h e dog. I n f a c t , dogs have

been known t o s u f f e r from t h e same d i s o r d e r s as t h e i r owners , f o r

example g o i t r e ( M i l l e r , 1964). Some of t h e p a t h o l o g i e s o b s e r v e d

i n dogs i n v e t e r i n a r y c l i n i c s s u g g e s t p o s s i b l e i m p l i c n t i o n of

c y a n i d e t o x i c i t y . However, t h i s h a s n o t been conf i rmed o r

i n v e s t i g a t e d ,

The p u r p o s e o f t h i s s t u d y was t o de t e rmine t h e e f f e c t s of

n u t r i t i o n a l l y ba l anced d i e t s c o n t a i n i n g c y a n i d e t o t h e l e v e l of t h a t

found i n g a r i , on IGigcrinn mongrel dogs w i t h r e s p e c t t o g rowth ,

t h e t h y r o i d gland, t h e l i v e r , t h e k i d n e y , t h e p a n c r e a s and t h e

b lood .

LITERATURE REVIEW

Tox ic p r i n c i p l e i n c a s s a v a

Tissues of a l l c e s s a v a c u l t i v ~ r s s o f a r examined c o n t a i n

cyanogen ic g l u c o s i d e s , a l t h o u g h i n v a r y i n g c o n c e n t r ~ . t i o n s .

V a r i a t i o n s i n t h e cyanogen ic g l u c o s i d e c o n c e n t r a t i o n s i n t u b e r s as

w e l l as t h e morpho log ica l c h a r z c t e r i s t i c s of t h e p l a n t s , form t h e

b a s i s o f taxonomic d i f f e r e n t i a t i o n between b i t t e r ( h i g h HCN

p roduc ing ) and sweet ( low HCN p r o d u c i n g ) c u l t i v a r s (Roge r s , 1965) .

Nartey (1968) showed t h a t l i n a r n a r i n and l o t a u s t r a l i n ( m e t h y l l i n a m a r i n )

c o n s t i t u t e t h e cyanogen ic m a t e r i a l s of c z s s a v a , Xanihot e s c u l e n t a

C a n t z i n t h e p r o p o r t i o n s of 93% and 7% r e s p e c t i v e l y . -1

Chemis t ry of t o x i c p r i n c i p l e

L inamar in , 2-(B-D-glucopyranosyloxy)isobutyronitrile, h y d r o l y z e s

t o D-glucose , ac e t one a n d hydrogen c y a n i d e , w h i l e l o t a u s t r a l i n , 2-(B-D-glucopyranosyloxy)2-methylbutyronitrile, h y d r o l z e s t o

D-glucose, methyl e t h y l k e t o n e a n d hydrogen c y a n i d e ( B u t l e r , 1965,

N a r t e y , 1968; B i s s e t t - e t -= a1 7 1969). The a c t i o n of t h e two enzymes is

u s u a l l y i n v o l v e d i n t h e r e l e a s e of HCN, For l i n a m a r i n , t h e i n i t i a l

r e a c t i o n i n v o l v e s t h e h y d r o l y s i s by l i n a m a r a s e , a 3 - g l u c o s i d a s e ,

of t h e B-g lucos id i c bond between t h e s u g z r and t h e a g l u c o n e , i n

t h i s case , 2-hydrox~isobutyronitrile or a c e t o n e cyanohydr in , of

t h e g l u c o s i d e . Although cyanohydr ins ( / 3 - h y d r o x y n i t r i l e s ) p roduced

by t h e a c t i o n of pla.nt g l u c o s i d a s e s w i l l d i s s o c i s t e nonenzymnt i c a l l y , h y d r o x y n i t r i l e l y n s e s t h a t a r e p r e s e n t i n cyanogenic p l a n t s c a t a l y z e

t he d i s s o c i a t i o n of t h e s e compounds. I n t h e p r e s s n c e of h y d r o x y n i t r i l e

l y a s e , ace tone cyanohydrin d i s s o c i a t e s t o produce HCN and ace tone,

CN

+ H O C C

I CH

3

Linamarin D-glucose ace tone ( 2 - ( ~ - ~ - ~ l u c o ~ ~ r a n o a ~ l o x ~ ) ( ~ - ~ - ~ l u c o ~ y r a n o s e ) cyanohydrin - i s o b u t y r o n i t r i l e ) (2-hydroxyiso-

bu tyron i t r i l e )

hyd roxyn i t r i l e , % lyase

HCN + O=C-

I CH3

'43 ace tone

ace tone cyanohyd,rin (2-hydroxyisobutronitrile)

*

The r e a c t i o n i n v o l v i n g l o t a u s t r a l i n i s similar ( ~ o n n , 1969).

L i t t l e , i f any, f r e e HCIii would be expected t o 'accumulate i n the

p l a n t , s i n c e being v o l a t i l e i t would escape t o t h e atmosphere, HCN i s

a l a o a c t i v e l y metabol ized by plants, t he n i t r i l e group be ing connected

t o t h e amide group of asparag ine (Conn, 1969). Waceration of f r e s h p l a n t

t i s s u e i n i t i a t e s the enzymatic breakdown of t h e cyanogenic g lucos ide by

the p l m t anrymea to r o l s a s o HCN. It i s the release o f HCN that is

r e e p o n s i b l e for t h e toxicity r e s u l t i n g i n dea th of 'man and animals.

Theref ore , merely e a t i n g enough p l a n t material t h a t i s s u f f i c i e n t l y rich i n

cynnogenic ~ l u c o s i d e s and enzymes r e s u l t s i n p o i s o n i n g (Conn, 1973).

Cyanide and P o i s o n i n g

The po i sonous p r o p e r t i e s of c a s s a v a were f i r s t r e c o r d e d by

C l u s i u s i n 1605. Denths a f t e r heavy meals of c a s s a v a h a v e been

r e p o r t e d i n t h e N i g e r i a n P r e s s (Osuntakun, 1973) U n l e s s t h a

cyanogen ic g l u c o s i d e i s removed t h r o u g h p r o c e s s i n g , HCN i s

released when c a s s a v n is e 3 t en. C u l t i v a r s w i t h 1 ow cyanogen ic

l e v e l s are b c i l e d , f r i e d o r r o a s t e d , KCK h a s a b o i l i n g p o i n t of

26'~ and cook ing t e m p e r a t u r e s r e s u l t i n l o s s of HCN a s a g2s

(Piontgomery, 1980) . C u l t i v a r s w i t h h i g h cyanogenic l e v e l s a r e

p r o c e s s e d by s o a k i n g , f e r m e n t a t i o n and f r y i n g s o as t o g r e a t l y

r e d u c e t h e l e v e l of HCN b e f o r e cnnsumption (Oke, 1982). Osuntokun

(1968) s t z t e s t h a t HCK i n f r e s h c a s s a v a r o o t a t a l e v e l of 38 mg/lOO

g is r e d u c e d t o a b o u t 1 .? mg HCN/100 g i n 2 c a s s a v a p r e p a r a t i o n

known as g a r i .

Heat i n a c t i v o t e s t h e 3 - g l u c o s i d a s e , l i n a m a r a s e , bu t does n o t

a f f e c t l inamerii l . However, h c a t enhanccs t h e r e l e a s c of HCN from

a c e t o n e cyanohydr in . Any r e s i d u a l cyanogenic g i u c o s i d e mny be

h y d r o l y z e d i n t h a i n t e s t i n a l t r a c t of b o t h mnn and an ima l by t h e

m i c r o b i a l f l o r a ( r i i n k l e r , 1958) . The HCN r e l e a s e d i s r a p i d l y

a b s o r b e d from t h e g a s t r o i n t e s t i n , ? l t r a c t ( ~ o n n , 1973)

B a r r e t t e t al, ( 1977) were u n a b l e t o f i n d i n t a c t l i n a m n r i n i n -- t h e b lood of r a t s f o l l o w i n g a n o r a l d o s e of l i n n r n a r i n b u t found

a s i g n i f i c a n t amount i n t h e u r i n e , showing t h c t n p o r t i o n of t h e

o r a l l y admin i s t e r c d l i n a m a r i n up.s abso rbed i n t a c t . Barr c-t t

(1976) r e p o r t e d t h a t r n t s f e d 240 mg of l i n a m a r i n o v e r a 3-day

p e r i o d e x c r e t e d i n c r e a s e d u r i n a r y t h i o c y a n a t e which w2.s n o t

2 f f e c t e d s i g n i f i c s n t l g by t h e p r e s e n c e of me th ion ine i n t h e d i e t ,

T h i s is i n d i r e c t e v i d e n c e t h a t i n g e s t e d l i n a u l a r i n i n t h e absence

of a c t i v e l i n a m n r a s e i n t h e food , is p a r t i a l l y m e t a b o l i z e d t o

y i e l d c y z n i d e ( ~ h i l b r i c k -- e t al. , 1977) . Adewusi (1901) r e p o r t e d

t h a t most of t h e g l u c o s i d e i s e x c r e t e d unchanged i n t h e u r i n e .

Waduagwu a n d Umoh (1986) r e p o r t e d t h a t t h e l i v e r c o n t a i n s

enzymes c a p a b l e of h y d r o l y z i n g l i n a m a r i n , They a d m i n i s t e r e d a

s i n g l e dose of i i n a m a r i n i n t r n p e r i t ~ n e ~ d l y t o r n t s f o l l o w i n g

c a n n u l a t i o n of t h a b i l e d u c t and were a b l e t o r e c o v e r from t h e b i l e

12% of t h e a d m i n i s t e r e d dose as unchcnged l i n a m a r i n and 30-32% of

t h e dose i n t h e form of a cyanohydr in and a n i t r i l e w i t h i n 1 2 h o u r s

of a d m i n i s t r a t i o n . These m e t a b o l i t e s a r e cap2b le of d i s s o c i a t i n g

t o r a l e a s e c y a n i d e ion . bio l i n a m a r i n o r i t s m e t a b o l i t e s were

r e c o v e r e d frcm t h e b i l e a f t e r 1 2 h o u r s of a d m i n i s t r a t i o n . No

l i n a m a r i n o r i t s m e t a b o l i t e s were d e t e c t e d i n t h e f a e c e s of

u n c a n n u l a t e d t e s t rats from which b i l e was n o t removed,

The minimal l e t h a l dosc of KCT< t a k e n o r a l l y by humsns h a s been

e s t i m a t e d t o be between 0.5 2nd 3.5 mg/kg body we igh t (Chen

e t a1 1934; G y e t t l e r and Ba ine , 1938; Hals t rom and N o l l e r , - -- 9

1945). The l e t h a l dose of the a l k a l i n e c y a n i d e s i s a p p r o x i m a t e l y

t w i c e t h a t o f HCb? (Xontgomery, 1980). The lethal o r a l dose of

p u r i f i e d l i n n r n s r i n i n r a t s is 500 rcg/kg body v ~ e i g h t .

9.

I n plant t i s s u e , t h e l e t h a l o r a l dose of c y a n i d e i n t h e form

of cyanogen ic g l u c o s i d e s v a r i e s a c c o r d i n g t o t h e c o n d i t i o n

o b t a i n i n g i n t h e p l m t . The h i g h e s t c o n c e n t r a t i o n s of cyanogen ic

g l u c o s i d e s a r e s e e n i n young a c t i v e l y growing p l a n t s * The l e t h a l

dose a l s o v 2 r i e s a c c o r d i n g t o f a c t o r s o b t a i n i n g i n t h e a n i m a l a t

t h e t i m e of e a t i n g t h e p l a n t . For example, r u m i n a n t s a r e more

s u s c e p t i b l e t o p o i s o n i n g by cyanogenic p l a n t s than a r e h o r s e s a n d

p i g s , s i n c e t h e enzymes concerned w i t h t h e r e l e a s e of HCN a r e

d e s t r o y e d by g a s t r i c h y d r o c h l o r i c a c i d i n t h e l a t t e r . Sheep a r e

l e s s susceptible t h a n c a t t l e ( J o n e s , l952), Van d e r Walt (1944)

r e p o r t e d t h a t p o i s o n i n g i n r u m i n a n t s depends upon t h c q u a n t i t y of

p l a n t i n g e s t e d , t h e p r e v i o u s d i e t o f t h e a n i m a l , t h e pH of t h e

s tomach contents, t h e p e r c e n t a g e of t o t a l HCN p r e s e n t i n t h e f r ee

s t a t e i n t h e p l a n t and t h c t o t a l HCN c o n t e n t o f t h e p l a n t .

When p l a n t m a t c r i a l is r e s p o n s i b l z f o r d e a t h , g a s t r o -

e n t c r i t is may be observed . HCN p o i s o n i n g m a n i f e s t s a s c o n g e s t e d

b l o o d v e s s e l s , u n c l o t t e d and o f t e n b r i g h t red. b l o o d , c o n g e s t i o n

and Mmor rhagc of t h e l u n g s , and r e d d e n i n g and c o n g e s t i o n of t h e

mucous membranes of t h e stomzch. I n a c u t e p o i s o n i n g , dea th u s u a l l y

o c c u r s w i t h i n a few s e c o n d s wi thou t l e s i o n s i n thc organs. T h e r e

may be c o n v u l s i o n s , p a r a l y s i s , s t u p o r , and c e s s a t i o n of r e s p i r a t i o n

b e f o r e t h a t of t h e heartbeats ( C l a r k e and C l a r k e , 1975).

A f t e r i n g e s t i o n of cyanogenic p l a n t s , whe the r symptoms

occu r immed ia t e ly o r a r e d e l a y e d depends on t h e amount of t h e

cyanogen ic g l u c o s i d c i n g e s t e d , t h c r a t e of liberation o f c y a n i d e

from i t , t h e r a t e of a b s o r p t i o n of c y a n i d e from t h e gut and upon t h e

10.

r a t e o f d e t o x i f i c a t i o n , A l e t h a l dose may g ive r i s e t o d e l a y e d

symptoms o r t o n c symptoms wha t soeve r , if i t s a b s o r p t i o n is s u f f i -

c i d n t l y p r c l o n g e d by s low h y d r o l y s i s of t h e g l u c o s i d e . Death

w i t h o u t symptoms mzy o c c u r sometime a f t e r cyanogenic g l u c o s i d e s

have been consumed, o t h e r w i s e t h e r e may b e e x c i t e m e n t , p r o f u s e

s a l i v a t i o n ? c o n v u l s i o n s of v a r y i n g d e g r e e and d u r a t i o n ,

o p i s t h o t onus , j e r k y movements of t h e e y e b a l l a n d r e s p i r a t o r y

d i s t r e s s w i t h d e a t h o c c u r r i n g 15 t o 60 m i n u t e s a f t e r t h e o n s e t o f

symptoms ( C l a r k e ,and C l a r k e , 1975) .,

The s e v e r i t y of t h e r e a c t i o n t o KCN and l i n a m a r i n i n rats is

s t r o n g l y i n f l u e n c e d by t h e manner i n which t h e y a r e a d m i n i s t e r e d ,

h i x e d w i t h d i e t and f e d o v e r a p e r i o d of h o u r s , t h e y a r e t o l e r a t e d

i n l a r g e m o u n t s , whereas d e a t h r e s u l t s f rcm a s i n g l e d o s e ego when

50 rng of l i n a r n a r i n / l 0 0 g body weight was g i v c n t o one group of

rats d e a t h o c c u r r e d i n 4 h o u r s and when 0.6 mg KCN/100 g body

weight was g i v e n t o a n o t h e r g roup of r a t s , d e a t h o c c u r r e d i n

30 minutes . T ~ G symptoms produced by b o t h l i n a m a r i n and KCN were

a t a x i a , e p n e a , d o c i l i t y and p a r e s i s , XCN produced a s e v e r e

c y a n o s i s ( H i l l , 1877) .

The H C N , r e g a r d l e s s of i t s s o u r c e , a c t s by p r e v e n t i n g t h e

i n t r a c c l l u l a r o x i d h t i v e p r o c e s s , a l t h o u g h t h c b lood d ~ e s n o t l a c k

oxygen, The a c t i c n of cytochrome o x i d a s e w i t h i n t h e c e l l is

i n h i b i t e d by t h e c y n n i d e i o n and d e o t h is t h e r e s u l t o f g e n e r a l i z e d

c y t o t oxic anoxia.

E f f e c t s o f c y a n i d e on c a r d i o r e s p i r a t o r y system

Cyanide i s known t o a f f e c t h e a r t f u n c t i o n i n a c h a r a c t e r i s t i c

manner (Woolman and D r i p p s , 1970), When H i l l (1977) gave l e t h a l

dosee of KCN o r l i n a m a r i n t o r a t s , t h e r e were many s i m i l a r i t i e s

when t h e t r a c i n g s of t h e e l e c t r o c a r d i o g r a m s were compared.

S i m i l a r i t i e s i n c l u d e d c a r d i a c a r r h y t h m i a s , changes i n t h e

electrical s t i m u l a t i o n o f t h e h e a r t and a d e c r e a s e d h e a r t r a t e .

Compared wi th c o n t r o l v a l u e s , whole b lood l a c t i c a c i d was e l e v a t e d

i n ra ts dosed w i t h l i n z m a r i n o r KCM, whereas p y r u v i c a c i d was

e s s e n t i a l l y unchanged* The i n c r e a s e d l a c t a t e / p y r u v a t e r a t i o i s

i n d i c a t i v e of s e v e r e c a r d i o r e s p i r a t o r y f a i i u r e , T h i s phenomenon

h z s been z t t r i b u t e d t o an i n c r e a s e d a c t i v i t y of l a c t i c a c i d

dehydrogenase a c t i n g i n a r e v e r s e c a p a c i t y (Brode r a n d Wei l , 7964).

Decreased h e a r t t i s s u e c y t ochrome o x i d a s e a c t i v i t y was a l s o observed.

T h i s is a w e l l r e c o g n i z e d r e s u l t of c y a n i d e p o i s o n i n g and h a s

been c i t c d as a c a u s e of t h e a r r h y t h m i a s s e e n a f t c r c y a n i d e

a d m i n i s t r a t i o n ( ~ a l l a n t y n e e t a l . , 1972; Schube r t a n d B r i l l , 79681,

The p a r t i a l p r e s s u r e s f o r C 0 2 znd O2 i n t h e b l o o d and t h e b i c a r b o n a t e

i o n l e v e l were s i g n i f i c a n t l y a l t e r e d i n most c a s e s by l i n a m a r i n

d o s i n g , i n d i c a t i v e of m e t a b o l i c a c i d o s i s ,and a n a l t e r a t i o n i n t h e

b u f f e r c a p a c i t y of t h e b l o o d o Even a t a low s u b l e t h a l d o s e ,

( H i l l , 1977) l i n n m a r i n hcs similar e f f e c t s on t h e l a c t a t e / p g r u v a t e

r a t i o a n d c y t o c h r o n e ox idase .

When K i l l (1977) gave a s u . b l e t h a l b u t h i g h d o s e of l i n a r m a r i n

t o rats he found sodium-potassium a d e n o s i n e t r i p h o a p h a t a s e

a c t i v i t y c o n s i d e r a b l y r educed i n h e a r t t i s s u e b u t h e f e l t t h a t

t h i s e f f e c t wcs due t o 3 mechanism independex t of BCN s i n c c t h e same

e f f e c t c n sod ium-po tns s iun a d e n o s i n e t r i p h o s p h a t a s e i s produced by

t h e g l u c o s i d e d i g i t a l i s .

M c t a b l i c Pathways of s u b l e t h a l levels of HCN - The most w i d e l y s t u d i e d m e t a b o l i c pathway of s u b l e t h a l

l e v e l s o f c y a n i d e is i ts combina t ion w i t h t h i o s u l p h n t e t o form

t h i o c y a n a t e a d s u l p h i t e - The r e a c t i c n is c a t a l y z e d by t h e enzyme

- rhod-znese ( t h i o s u l p h a t e s l l - l phur t rnns f e r a s e LEC. 2.8.1.12)

L m g , I933a , b ; Cosby and Sumner, 1945). Rhodanese is a mi to-

c h o n d r i a l enzyme, and i s widesp read i n l i v i n g t i s s u e s , r e a c h i n g i ts

h i g h e s t c o n c e n t r a t i c n s i n t h e l i v e r , k i d n e y , t h y r o i d , a d r e n a l and

p a n c r e a s ( R o s e n t h ( ~ 1 , 1948). Thc r e s u l t i n g t h i o c y a n z t e i s e x c r e t e d

i n u r i n e and also i n t h e s a l i v a , from which i t nay e i t h e r be

r e a b s o r b e d o r me tabo l i zed i n t h e d i g e s t i v e t r a c t . T h i o c y a n a t e is

s l o w l y o x i d i z e d t o s u l p h a t e , b u t t h i s dces n o t a p p e a r t o be a n

i m p o r t a n t f a c t o r i n human t i s s u e s (Clemcdson et a l . , 1 9 6 0 ) ~

Knox (1972) showed t h a t t h e l i v e r i s t h e most a c t i v e t i s s u e

f o l l o w e d by t h e k i d n e y which had o n l y a b o u t 50% of t h e a c t i v i t y of

t h e l i v e r - Eimwich and Saunde r s ( $ 9 4 8 ) foune t h a t t h e r e was

s p e c i e s v a r i 2 , t i c n i n t h c l e v e l s of rhodnnese i n t h e l i v e r s o f m a m m a l s ,

and c o n c l u d e d t h a t m e t a b o l i c r e a c t i o n s may b e e x p e c t e d t o t a k e p l a c e

at d i f f e r e n t r a t e s . They found t h e l e v e l s o f rhodanese f o r t h e

l i v e r s i n t h e f o l l o w i n g s p e c i e s : -

Dog: 0 ~ 7 8 - 1 - 4 6 mgig

Rhesus Monkey: 70.08-15-16 r n ~ / ~

R a b b i t : 7.98-18.92 mg/g

R a t : 14.24-28.38 rng/g

Fiukcr j i a n d Smi th (1943) found t h a t n e a r l y a l l t h e c y a n i d e

i n g e s t e d by r a b b i t s was r e c o v e r e d i n t h e form ~f t h i o c y a n a t e i n u r i n e

i n 24 t o 48 h c u r s , wheraas i n dogs , l e s s t h a n 25% w a s r e c o v e r e d

i n seven days.

I n a second pathway c y s t i n c g i v e s r i s e t o I - c y s t e i n e a n d

t h e n by t r a n s a m i n a t i o n t o 3-rnercaptopyruvate , 3 -mercap topyruva te

r e c c t s w i t h c y a n i d e i n t h e p r e s e n c e of 3- rnercsp topyruvate

s u l p h u r - t r s n s f e r a s e (EC 2.8,1.2) t o form t h i o c g a n z t e and pyruvate

( F i e d l e r and Ilocd, 1956)- C y s t i n e may r e a c t more d i r e c t l y w i t h

c y a n i d e t o p roduce 2-amino-4 t h i o z o l i n e c o r b o x y l i c acid, a n i n e r t

c.3rnpound t h a t i s e x c r e t e d i n t h e u r i n e (wood and C o l l e y , 1956).

C y s t i n c is a s u l p h u r amino a c i d and a c t s as a s u l p h u r donor.

I n a t h i r d rne tnbol ic pathwoy, v a r i o u s forms of v i t a m i n

B q 2 a r e i n v o l v e d * Bydroxocabalamin r e a c t s w i t h c y a n i d e t o form

cyanocobalarnin (Kaczka &, , 1950). Mushet -- e t al. ( 1952) shotted

t h a t hydroxocob21arnin p r o t e c t e d mice agz - in s t c y a n i d e p o i s o n i n g .

Ur ine s a m p l e s collected d u r i n g t h e s t u d y showed that w i t h i n 2.5

h c u r s 9.6% of t h e c y a n i d e a d m i n i s t e r e d c o u l d be a c c o u n t e d f o r as

v i t a m i n B I Z ancl 3.55 as t h i o c y a n a t e . Smith (1961) found t h a t

i n j e c t i o n s o f s u b l e t h a l d o s e s of c y a n i d e g i v e n t o rats czuse a

s i g n i f i c a n t d e p l e t i o n ?f t h e l i v e r s t o r e of v i te rn in B 12'

Vi tamin

B A 2 is s t o r e d i n t h e l i v c r as hydroxocobalamin. I t was a l s o

f ouns t h x t a d i e t a r y deficiency of v i t a m i n B l e a d s t o i n c r e a s e d 1 2

t h i o c y a n a t e e x c r e t i o n .

Some of t h e c y a n i d e t h a t ccmbines w i t h hgdroxocobclamin t o form

cyanccobnlamin i s l o s t t o s u p p l y t h e ? -carbon f ragment f o r t h e

s y n t h e s i s of i rnao r t an t compounds s u c h as c h o l i n c a n d o t h e r l a b i l e

methyl g roups and f o r t h e c a n v e r s i o n c f hcmocys t e ine t o n e t h i m i n e

( ~ r a t z e r , 1353; S t e h ~ l e t al., 79-53; S m i t h , 1954). Some of t h e

c y a n i d e i s l o s t as ca rbon d i o x i d e i n t h e b r e a t h , Eoxcr and R i c k a r d s

(1952) d e m o n s t r a t e d t h a t l a b e l l e d 1 4 ~ ~ g i v e n t o dogs c o u l d q u i c k l y b e

r c c o v e r e d as e x h a l e d c r r b o n d i o x i d e , and i n t h e u r e i d c c a r b o n of

a l l a n t o i n as w e l l as i n B an2 t h i o c y a n a t e o 12

V e g e t x i a n s who s r e s h o r t o f v i t a m i n B d e t o x i f y t h e i r c 3 a n i d e 12

t o t h i o c y ~ x n a t t ar id s o e x c r e t e e x c e s s t h i o c y a n a t e i n t h e i r u r i n e ,

Only d i e t s c o n t a i n i n g s i g n i f i c a n t zmount s of an ima l p r o t e i n s p r o v i d e

a n adequa te s u p p l y of v i t a m i n 9 12-

Umoh -- e t n l . (1985) r e p o r t e d t h a t t h i a m i n a i d s t h e metabol i sm

of l i n a m a r i n i n t h e ra t , T h e i r r e s u l t s showe6. t h a t d e f i c i e n c y o f

t h i a m i n causes a c c u m u l a t i o n o f unrnc tabol ized l i n a m a r i n and f r e e

cyan ide . The f r e e c y a n i d e was c o n v e r t e d t o t h i n c y a n a t e r e s u l t i n g

i n s i e ; n i f i c a n t l y h i g h e r c c n c e n t r a t i o n s of t h i o c f e ~ a t e

r e l a t i v e t o t h e c o n t r o l s ,

Umoh e t al. (1986) d e m o n s t r a t e d t h a t i n p r o t e i n d e f i c i e n c y i n -- ra ts b o t h t h e r a t e nf metabol i sm o f f r e e c y a n i d e t o t h i o c y a n a t e

and a l s o t h e e v e n t u a l c l e a r a n c e of t h i o c y a n a t e were p ro longed ,

A f t e r 2 s i n g l e c r a l dose of l i n a m a r i n , t h e c o n t r o l rats e x c r e t e d

most of t h e i r t h i o c y a n a t e i n t h e f i r s t 24 h o u r s w h i l e p r o t e i n

d e f i c i e n t rats e x c r e t e d t h e i r s i n 48 h o u r s of d r u g a d m i n i s t r a t i o n .

15-

Vesey and Ni l son ( 1978) r e p o r t e d t h a t i n a n e s t h e t i z e d dogs g i v e n

approx imate ly 1 mg KCN/kg body weight i n t r z v e n o u s l y , 7.5-99% of t h e

KCN r e c o v e r e d from t h e b lood was i n s a l i n e washed r e d c e l l s , t h e

p r o p o r t i o n p r e s c n t i n c r e a s i n g w i t h t ime a f t e r t h e KCN dose.

G o l d s t e i n and R e i d e r s (1951) r e p o r t e d HCK in t h e b lood cf dogs

po i soned w i t h MaSCN. It h a s been shown t h a t HCN d e r i v e d endogenously

from SCN- i s v i r t u e l l y u n i o n i z e d at p h y s i o l o g i c a l pH ( 1 z a t t e t a l , ,

1962; Vesey and b ' i l son , 1978) l i p i d s o l u b l e and r a p i d l y e n t e r s and

accumula te s i n r e d c e l l s . Here it appears t o have no t o x i c a c t i o n

and the r e d c c l l s a c t as a "cyanide s i n k " (Vesey e_t &*, 1976).

S i ~ ~ ; n i f i c a p c e of. Thiocya.na-+,e

S t o a ( 1957) s t a t e s t h a t a l t h o u g h cyanogenic g l u c o s i d e s may t o

some e x t e n t be hand led i n o t h e r ways, a c l e a r r i s e i n serum t h i o c y a n a t e

a f t e r t h e i n g e s t i o n of o rgan ic cynnogenic compounds is good ev idence

t h a t some d e g r a d a t i o n t o HCN must have t a k e n p l a c e .

The e s t i m a t i o n of t h i o c y a n a t e e i t h e r i n b l o o d , saliva o r i n

u r i n e is n a t a s imple i n i ' e s of t h e amount o f c y a n i d c i n g e s t e d f o r

a number of r easons . F i r s t l y , t h e r e i s t h e q u e s t i o n of o t h e r me tabo l i c

pathways. Second ly , t h e r e is a t h i o c y a n a t e m e t a b o l i c poo l i n t h e body,

which w i l l f l u c t u s t a wide ly w i t h t h e v a r i a t i o n s i n t h e d i e t a r y i n t a k e

o f preformed t h i o c g a n a t e o For example, S t oa ( 1957) dcmonst r a t ed

t h a t t h i o c y a n a t e i s p r e s e n t i n c o n s i d e r a b l e bu t v a r i a b l e amcunts i n

g reen v e g e t a b l e s , mi lk and beer. He a . l so found t h a t t h e u r i n a r y

e x c r e t i o n of t h i o c y a n a t e v a r i e d d i r e c t l y w i t h u r i n e volume and w i t h

c h l o r i d e e x c r e t i o n . Plasma l e v e l s of t h i o c y a n a t c a l s o v a r i e d w i t h

t he degree of h y d r a t i o n of t h e ~ u b j e c t . It may t h e r e f o r e be

d i f f i c u l t t o d e t e c t , o r a s s e s s t h e impor tance of f l u c t u a t i o n s i n

t h e t h i o c y a n a t e p o o l f o l l o w i n g t h e i n g e s t i o n of cyanogen ic

g l u c o s i d e i n f oodo

T h i c c y a n a t e is a w e l l r e c o m i z c d g o i t r o g e n . Th iocyana t e

f u n c t i o n s ss a g o i t r o g e n by i n h i b i t i n g i o d i d e t r a p p i n g . I o d i n J z a t i o n

of t y r o s i n c t o form t h y r o i d hormone clepends on t r a p p i n g b l o o d i o d i d e .

Pul lman -- e t al. ('1954) showed t h a t r e n d a x c r c t i o n of t h i o c y a n a t e i n

t h e normal m d c is s l aw due t o e f f i c i e n t r e a b s o r p t i o n of t h c t h i o -

c y a n a t e and t h a t t h i o c y a n a t e a c c u m u l a t e s when r e p e a t e d d o s e s are

g iven , Garv in ( 1939) showed t h a t t h e accumulation o f t h i o c y a n a t e

h a s t o x i c consequences .

Accord ing t o P'iontgomcry (1980) t h e r e is s t r o n g e v i d e n c e t h a t t h e

i n g e s t i o n of c y a n ~ g e n i c g l u c o s i d e s g i v e s r i s e t o chronic n e u r o l o g i c a l

d i s ea se i n man, w i t h a c c ompanyinf; r a i s e d plnsma a n d u r i n a r y t h i o c y a n a t e

l e v e l s as i n t r o p i c a l a t a x i c n e u r o p a t h y in N i g e r i a and Tanzan ia ,

F u r t h e r mcdicz.1 i n t e r e s t h a s been developed by t h e o b s e r v a t i o n t h a t

it is a powerful c a t a l y s t of n i t r o s a t i o n ( B ~ y l a n d a n d !sfalker, 1974)-

The n i t r a t e c o n t e n t of g a s t r i c j u i c e may be h i g h i n h y p o c h l o r h y d r i c

s tomachs ( 8 u d C e l l e t al., 1977). I n t h e p r e s e n c e o f i n c r e a s e d

t h i o c y a n a t e i n t h c s a l i v s and hence i n t h e g a s t r i c juice, t h e

p r o d u c t i o n of c a r c i n o g e n i c n i t r o s a m i n e is f a c i l i t a t e d , T h i s may be

relevant t o g a s t r i c c a n c e r and i t s r e l a t i o n s h i p t o d i e t , smoking

and h y p o c h l o r h y d r i a .

Toxic r e a c t i o n s i n t h e human r e c e i v i n g o r a l l y a d m i n i s t e r e d

sodium o r po ta s s ium t h i o c g n n a t e as a n a n t i h y p e r t e n s i v c d r u g i n c l u d e

depressed t h y r o i d f u n c t i o n , h e p a t i c n e c r o s i s , n e p h r o s i s , i r r i t a b i l i t y ,

motor aph:?s ia , h a l l u c i n a t i o n , d e l i r i u m , muscle c ramps , p r e c o r d i a l p:,in,

p a l p i t at i o n s , wezkness , anemia, f a c i a l edema, s k i n e r u p t i o n s .

D i a g n o s i s is e s t a b l i s h e d cn t h c b a s i s of b lood t h i o c y a n a t e l e v e l s

( ~ocdrnan and Gi l lman , 1970).

Hormonal f u n c t i o n of t he t h y r o i d g l a n d

PiacDonal:! (1980) s t c t e s t h z t t h o main f u n c t i o n of t h e thyroid

g l a n d is t o a c c u m u l a t e i o d i n c and c a u s e i t s n t t s c h m e n t t o t y r o s i n e ,

ail amino aci l? , i n o r d e r t o form t h c t h y r o i d hormone. The t h y r o i d

hormone r e g u l z t c s growth , d i f f e r e n t i z t i o n ,and o x i d a t i v e metabol i sm

(MscDonald, 1980). The t h y r o i d hormone i s known t o e x i s t i n two forms,

namely 3,4,3',5'-tctraiodothpnine ( t h y r o x i n o r T ) and 3,5 ,3'4ri iodo- 4 t h y r o n i n e (T ) (HacDonald, 1980). 3

Ingbar acd Braverman (4975) +educed t h a t T i s t h e a c t i v e form 3

of t h y r o i d hormoncl. The deductions were made on non-ruminants .

Ba r ry e t 31. ( 19831, work in^ on s h e e p showed t h a t T had a g r e a t e r 3

b i o l o g i c a l po tency t h a n T4 f o r r e g u l s t i r l g body growth and t h a t an ima l

p e r f ormancc- was n o t d e p r e s s e d u n t i l serum T c o n c e c t r a t i o n d e c l i n e d . 3

Choprn 2nd So1c)mon (1980) r e p o r t e 2 t h a t T b i n d s t o n u c l e a r 3

r e c e p t o r s w i t h grea ter a f f i n i t y t h a n T sac! i n mammals T is 3-5 4 3

t i m e s as p o t e n t as Tq i n t hcmogene f i i s . DeGroct and Rue (1980) r e p o r t e d

t h a t i n t h e n u c l c u s , T s t i m u l a t e s t h e s y n t h e s i s of messenger RNA, 3

p r ~ t e i n s y n t h e s i s a n d p h o s p h o r y l a t i o n , T h i s and o t h e r e v i d e n c e , h a s

lzcl t o t h e view t h a t mcst, b u t no t 311, t h e p h y s i o l o g i c a l e f f e c t s

of t h y r o i d hcrmones are a t t r i b u t a b l e t o T ( B e r n a l , 1980). 3

1 8 ~

The T h y r o i d g l a n d of t h e d o 6

Gieseckc (1985) d e m o n s t r a t e d t h a t t h e dog i s t h e s p e c i e s

most similcr t o man. M i l l e r -- e t a l , (1968) r e p o r t t h a t t h e t h y r o i d

g l a n d o f t h e dog is s u b j e c t t o a l l t h e p a t h o l o g i c a l c c n d i t i o n s known

t o man. It h a s been o b s e r v e d t h a t where g o i t r e was endemic i n man,

t h e same c o n d i t i o n was common i n 6ogs of t h e r e g i n n . Hypothyro id ism

may c c c u r w i t h o u t en l a rgemen t of t h e g l ~ n d i n s i m p l e g o i t r e , i n t h i s

c a s e t h e a c t i v e t i s s u e of t h e @rind is r e p l a c e d by i n a c t i v e t i s s u e .

I n c o l l o i d o r parenchymatous g o i t r e , which is t h e most common t y p e

i n d o g s , t h e gland a t t e m p t s t o compehsate f o r i o d i n e d e f i c i e n c y by

en la rgemen t , T h i s t y p e ~f g o i t r e i s found i n dogs f e d on t h y r o i d -

i n h i b i t i n g s u b s t a n c e s o The d c g r e e of a l t e r a t i o n of body s t r u c t u r e and

f u n c t i o n Zepends on t h e age a t which t h y r o i d a c t i v i t y becomes

d e f i c i e n t . C r e t i n i s m , which d e v e l o p s i n t h e t h y r o i d deficient young

is a more scvere c o n d i t i o n t h a n myxederna. P h y s i c d growth and m e n t a l

development a r c b o t h markedly r e t a r d e d i n c r i . t i n i s m p

E f f e c t s of c y a n i d e c o n t z i n i n ~ f o o d s on t h e t h y r o i d gland

Delange -- e t al. (1982) r e p o r t e d t h a t ~ l t h o u g h Cassava is

consumed on z l a r g e s c a l e w i t h i n t h e t r c ~ i c s , g o i t r a and c r e t i n i s m

a r c n o t found i n all p o p u l a t i o n s whose s t a p l e food i s cassava.

They f e l t t h a t t h e r e a p p e a r s t o be t h r e e p o s s i b l e e x p l a n a t i o n s f o r

t h e occurrence of g c i t r e and c r e t i n i s m , These a r c (1) method of

p r o c e s s i n g c a s s a v a and t h e m o u n t of r e s i d u a l cyanogen ic g l u c o s i d e

i n t h e p r o d u c t f o r consumpt ion , ( 2 ) low iocl ins c o n t e n t i n d i e t o r w a t e r ,

( 3 ) p r o t e i n c a l o r i e m a l n u t r i t i o n .

Nwokolo end Ekpech i (1966) found a high i n c i d e n c e cf v i s i b l e

g o i t r e i n t h e Nsukka D i v i s i c n of E a s t e r n N i g e r i a . Thc d i e t i n t h e

v i l l n g e s w i t h a h i g h i n c i d e n c e of g o i t r e was f o u n d t o c o n s i s t ma in ly

of u n f e r n e n t ed c a s s a v a o Ekpechi ( 1967) i n v a s t i g n t c d t h e e f f e c t s of

un fe rmen ted , d r y powdered c a s s a v a on t h y r o i d f u n c t i o n i n t h e rat .

A f t e r f e e d i n g t h e ? i e t f o r 7 days , he obse rved a s i g n i f i c a n t

i n c r e a s e i n t hy ro i ? . weight, markedly i n c r e a s e d p lasma-pro te in-bound

r a d i o a c t i v e i a d i n e , s e v e r e d e p l e t i o n o f t h e g l a n d ' s p r e c u r s o r and

hormone io r l i nc s t o r e s , an i m p a i r e d 13'1 t r a n s f e r from 3-rnonoiodatyrosine

(MIT) t o 3 , 5 - d i i o d o t y r o s i n e (DIT) i .e. a h i g h N I T t o DIT r a t i o , h igh

131 t r a n s f c r o f I t o i o d o t h y r o n i n e s , a f e a t u r e a l s o seen i n i o d i n e

d e f i c i e n c y , b u t n o t e l i m i n z t c d when t h s czssava d i e t was g i v e n w i t h

a d e q u a t e i o d i n e s u p p l e m e n t s o

B l a k e l y an$. Coop (1949) a d m i n i s t e r e 2 p o t a s s i u m c y a n i d e t o

sheep by rumen f i s t u l ~ and fcuncl a marked r i s e of t h i o c y a n a t e i n the

serum and u r i n e o Care (1954) r e p o r t e d t h a t l i n s e c d mca l , which

c o n t a i n s l i n a m a r i n f e d t o p regnan t ewcs, r e s u l t s i n t h e b i r t h of

g o i t r o u s l m b s . C o u r r i e r and Colonge ( -1960) r e p o r t e d l e s i o n s i n

thc t h y r o i d s of rcts f e d l i n s e e d meal. F l u x et al. (1960, 1963)

r e p o r t e d that lambs from ewcs grazed on p a s t u r e s c o n t a i n i n g cyanogzn ic

c l o v e r , m e . i n somc c a s e s t h e ewes, e x \ i b i t e d e n l a r g e d t h y r o i d s o

Flux -- e t al. (1960, 1963) found t h a t t h e p r o d u c t i o n of f r a n k

t h y r o i d en largement seemed t o depend on t h e l e v e l o f i o d i n e i n t a k e

r e l a t i v e t o t h e HCN i n t a k e . Frank t h y r o i d en la rgemen t d i d n o t

appear i f iocl inc i n t a k e w a s c o m p a r a t i v e l y high. Mo d e l e t e r i o u s

e f f e c t on weight g a i n s o r lambing per formance was o b s e r v e d i n any

sheep i n t h e s e e x p e r i m e n t s when i o d i n e i n t a k e was c o m p a r a t i v e l y

20 . high. Sihombing o t al. (1971) found t h c t admin i s t e r i ng t h iocyana t e

t o p i g s decreased growth an(? pro6uced en l a rged t hy ro id s . Lnnger

(7956) found a s i m i l a r e f f e c t with rats.

Delange m d Ermnns (1971) working i n Zaire, suggested t h a t t h e

i n g e s t i o n cf cassava reduces t h e t h y r o i d iod ine i n t a k e ,md i n c r e a s e s

r e n a l i c d i n c excre t ion . f'h f u r t h a r work i n Za i r e , Delnnge & 2.

(1982) showcd t h a t t h e prevalence 01 g c i t r e was i n v e r s e l y r e l a t e d t o

t h e p r o g r e s s i v e decrease of the u r i n a r y I/SCN r z t i o . They also

r epo r t ed t h a t i r i cne area( namely Kivu, t h e prevalence of g o i t r e

was seven t imes greater t h m t h e f i g u r e of 1.8% r e p o r t e d i n 1972

from t h e same vi l l :~ges . Tha cqcgrce of iod ine de f i c i ency was unchanged,

The i n c r e a s e i n p reva lence coPresponded with a marked i n c r e a s e i n t h e

ccnsumption of cassava i n kivu 2ue t o a d r a m t i c food s h o r t a g e

occu r r i ng within t h e area. The serum l e v e l s of su lphu r amino a c i d s +

p ~ ~ r t i c u l ~ r l y n e t h i o n h e ih a d u l t males were not lower than t h o s e found

i n t h e c o n t r o l s , but t h e r e Mew low l e v e l s of b r m c h e d amino n c i ? s ,

va l i f i e , l e u c i n e and i sn leud ine . T h i s p a t t z r n of scrum aminc a c i d s

is c h n r n c t e i - i s t i c uf s e v e r e p r o t e i n c a l o r i e m a l n u t r i t i o n and is

c o n s i s t e n t w i t h t h e low serum l e v e l s of albumin found i n t he se

p a t i e n t s ,

E f f e c t s of c y ~ n i d e c o n t a i n i n g foods on t h e ne rvous system

Endemic c r e t i n i s m i s def ined as 7n a s s o c i 2 t i o n of mental

de f i c i ency w i th a neu ro log i ca l syndrome c c n s i s t i n g of d e f i c i e n t

h c a r i n g an2 speech and d i s o r d e r s of stance L ~ n d g a i t o r hypo-

thyro id i sm and stunted growth. Pharoah e t al. (1980) working in -- Z a h e , have suggested thz t i od ine c?cf i c i c n c y i s respons ib le f o r

endemic c r e t i n i s m nnd that supplementat ion of d i e t a r y i od ine is

successful i n its preven t ion. I od ine def ic iency during pregnancy

would be r e spons ib l e f o r neuro log icn l damage. Benmiloud e t al. (1982)

r e p o r t e d t h a t t h e inc idence of c r e t i n i sm decreased without iodine

prophylax is when t h e economic c o n d i t i o n s improved i n affected areas

and sugges ted i od ine de f i c i ency was compounded by o t h e r factors,

some o f which were n u t r i t i o n a l .

Delange e t 21, ( 1982) r epo r t ed t h a t HCN-in?uced hypothyroidism

i n pregnant and nu r s ing rats was accompanied by a s t r i k i n g decrease

i n the p r o t e i n , RNA and c h o l e s t e r o l con t en t s of t h e cerebellum of t h e

pups at t h e end of t h e l a c t a t i o n per iod , i n d i c a t i n g a s lowing of

cellular growth. The BCN overload was accompanied by a h i g h l y

s i g n i f i c a n t i n c r e a s e i n serum th iocyana te concentra t ion. The s t u d i e s

confirmed t h a t severe hypothyroidism during t h e neona t a l pe r iod i n

t h e rat prcduces a l t e r a t i o n s i n t h e ma tma t inn of t h e central nervous

system. The s t u d i e s also demonstrated t h a t HCN docs not affect t h e

process of maturat ion of t h e c e n t r a l nervous system i n young rats

d i r e c t l y but acts i n d i r e c t l y by inducing hypothyroidism a f t e r

being conver ted t o th iocyannte .

Osuntown (1973) studied a disease known as tropical a t a x i c

neuropathy anong Nigerians. The e s s e n t i a l n e ~ r o l o g i c ~ d components of

t h e d i s e a s e are rnyelopathy , b i l a t e r a l o p t i c a t rophy , b i l a t e r ~ l perceptive

deafness a n d polyneuropathy. Go i t r e wns also found i n i n d i v i d u a l s

suffering from t r o p i c a l a t a x i c neuropathy, and t h e prevalence of

goitre among t h i s group was g r e a t e r than amcng o t h e r groups. The

22.

d i sea se exis ts mainly i n areas where a cassava prepa ra t i on , known

as purupuru, w i t h a h igh cyanick con ten t is commonly eaten. Plasma

th iocyana t e and u r i n a r y th iocyana te levels were found t o be about

fou r t imes as greet as levels found i n normal i nd iv idua l s , Levels

of a l l parameters fell t o normal a f t e r 6 weeks of h o s p i t a l i z a t i o n

and on h o s p i t a l d i e t .

The a c t i v i t y of t h e enzyme rhodanese i n t h e l i v e r of t h e s e

patients was normal ( ~ s u n t o k u n and Aladetoyinbo, 1970). Sulphur-

con t a in ing amino a c i d s methionine, c y s t e i n e and c y s t i n e were absen t

i n the plasma of 60% of t h e incl iv iduals seen and t h e concen t r a t i on

was g r e a t l y reduced i n t h e others . The q u a l i t a t i v e and q u a n t i t a t i v e

changes i n t h e plasma amino z c i d s are unlike t hose found i n kwashiokor,

the c l a s s i c a l diseese of p ro t e in - ca lo r i e def ic iency. Cyanocobalnmin

occurred i n h igher concen t r a t i ons i n t h e plosma t h a n i n normal

in8ividuals. Methylmalonic a c i d excreticln was normal inc l icz t ing t h a t

these i n d i v i d u a l s were not d e f i c i e n t i n v i t amin B at t i s s u e o r 12

c e l l u l a r l e v e l , The d i s e a s e has a l s o been i d e n t i f i e d among

Tanzanians i n a r e = where cassava is an important food i t em.

Moore (1934) proposed t h z t amblyopia in West A f r i c a is a

man i f e s t a t i on o f chron ic cassava poisoning. This form of amblyopia

i s commonly an isolated l e s i o n o r may be a feature of a more generalized

neuro log icn l syn2rorne- The d i e t among these i n d i v i d u a l s t ends t o

be low i n cvlimal p r o t e i n and t h e r e f o r e vi tamin B 12-

Wokes (1958)

proposed t h a t t h e causa l f n c t o r of ambylopin was cyanide and the

fa i lure t o de tox i fy via hydroxocobalamin.

E f f e c t s o f cyanide c o n t a i n i n g foods on growth

(i) Body growth

Maner and Gonez (1973) i n d i c a t e d t h a t cassava is a go

23

0 d

sou rce of energy i n swine r a t i o n s i f used a t levels of l e s g than

30-40% i n well-balanced diets. A t h i ghe r l e v e l s growth Sepression

is observed. They r e p o r t e d t h a t t h i s depress ion i n p i g performance

can be overcome by t h e u t i l i z a t i o n c f h igh-qua l i ty p r c t e i n and the

6uppLemcntation of C i s t s w i th adequate methionine. C~lderon -- e t d.

(1972) f e d a b a s a l d i e t o f cassava meal and c a s e i n t o growing rats

t o which cyanide was added as potassium cyanide and compared the

e f f e c t s of methionine supplementation. A t a l l l e v e l s of added cyanide,

rats f e d the methionine-supplemented d i e t s e x h i b i t e d a significantly

faster rate of growth t h a n those f e d unsupplemented d i e t s .

Maner and Gomez (1973) concluded t h n t among t h e reasons for t h e

response t o methionine supplementat ion of d i e t s c o n t a i n i n g high l e v e l s

of c a s sava o r cyanide w a s t h n t cassava p r o t e i n c o n t a i n s a very small

quantity of su lphur -con ta in ing aminc a c i d s and when f e d w i t h casein o r

soybean meal, both d e f i c i e n t i n methionine , there is a methionine

de f i c i ency which r e s u l t s from t h e combinztion of t h e two i n g r e d i e n t s ,

The p r o t e i n q u a l i t y , t h e r e f o r e was improved by the a d d i t i o n of

c r y s t a l l i n e DL-rnethionine.

(ii) Bone ~ r o w t h

HCN has a l r eady been shown t o induce hypothyroidism and s low

c e l l u l a r growth. Sage -- e t a l e (1968) r e p o r t e d t h a t thyroidectorny

had l i t t l e e f fec t on bone growth i n young p i g s f e d adequate m o u n t s

of calcium ;ad phosphorus, and t h a t plasma alkaline phosphatase

l e v e l s were p r o p o r t i o n a l t o p o w t h r a t e , They found t h n t

thyroidectorny a l t e r e d d r z s t i c a l l y t h e morrhology o f t h e growth

p l a t e . D i f f c r c n t i a t i o n was c o n s i d e r a b l y r e t a r d e d , and t h e r e s t i n g

c a r t i l a g e zone made up ss much as h a l f t h e width, The pr imary

spong iosa w a s vary narrow o r even absen t . Thyroxin ac?dit ion

p a r t l y r e s t o r e d t h e morphology of t h e growth p l a t e , It was t h e i r

view t h c t t h e dec rease i n plasma a l k a l i n e phosphatase f o l l o w i n g

thyroidectomy w a s due t o l a c k of o s t e o b l a s t i c a c t i v i t y i n t h e

pr imary spongiosa f o l l o w i n g t h e abrup t c e s s a t i o n of a c t i v i t y i n

t h e growth p l a t e . Zobrisky (1969) r e p o r t e d t h a t t h y r o x i n i s n o t

only invo lved i n bone growth bu t a l s o i n bone matura t ion.

D e f i c i e n c i e s i n t h y r o x i n impa i r c h o n 2 r o i t i n su lpha te synthesis

and t h u s t h e s y n t h e s i s o f c o l l a g e n of bone.

E f f e c t of cyan ide c c n t a i n i n g foods on t h e kidnex

Tnntum (1913) d c s c r i b e d hydropic i m b i b i t i o n o r v a c u o l n t i o n

i n t h c g lomerular t u f t e p i t h e l i u m i n exper imenta l ly induced

h y p ~ t h y r o i ~ i s m i n r a b b i t s - S i m i l a r changes a r e found i n other

p o r t i o n s o f t he nephron- Roess -- e t al. (1956) r e p o r t e d the

occurrence of mi ld p r o t e i n u r i a i n c l i n i c a l myxedema. C ~ S S - O

(1964) an? DiScala (1967) d e s c r i b e d t h i c k e n i n g of

g lomeru la r basement membranes and t h e accumulat i o n of a variety

o f e o s i n o p h i l i c i n c l u s i o n s w i t h i n t h e g l o m e r u l m c e l l s i n myxedema

p a t i e n t s . The glornerular c s l l s were a l s o no ted t o be e n l a r g e d ,

edemst ous an(? vacuclatecl. Cassanc (1 964) sugges ted t h a t t h e

e o s i n o p h i l i c i n c l u s i o n s r e p r e s e n t e d mucopolysaccharidos and hence

were an e x p r e s s i o n of myxedema. I n both exper imenta l hypothyroidism

25

( F o r t e , 1966) an6 c l i n i c a l rnyxedema (Di S c a l a , 1967) t h e r e are a l s o

l e s i o n s i n t h e r e n d t u b u l a r c e l l s and t h i cken ing of t h e basement

membranes,

Green -- et al. (1 974) showed t h a t exper imenta l ly induced

hypothyroidism a l s c produced mild p r o t e i n u r i a . Using e l e c t r o n -

microscopy, Green e t cl. (1974) foun? t h a t a l l c e l l s t y p e s of the

r e n a l glomerulus of r n t s wi th e ~ p e r i m e n t ~ i l l y induced hypothyroidism

show a sequence of Golg i appa ra tu s and o r g a n e l l e h y p e r p l a s i a , v e s i c l e

format ion and p rog re s s ive accumulation of e l e c t ron - lucen t m a t e r i a l i n

vacuo les i n t h e cytoplasm u n t i l f i n a l l y , thc e l e c t ron - lucen t m a t e r i d

g r adua l l y f i l l c c ? the c e l l whi le recognizab le c e l l u l a r organelles

g r a 6 u a l l y disappeared. An i n c l u s i o n occa s iona l l y developed. Foot

p roce s se s were sometimes l o s t o P e r i p h e r a l processes o f t h e e ~ i t h e l i u m

became g r o s s l y edematous o r accumulated vacuoles.

Onancgbu anc! Emole (1978) found swe l l i ng ,mcl v acuo l a t i on i n

t h e kidney c e l l s of a group of rats f e d d i e t s con t a in ing gari.

Effects of cyan ide contain in^ food on the liver

AGarns .- e t - nl. (1958) fed gari t o rats and mice. The d i e t contained

1% p r o t e i n cn a dry mat t e r basis, S l i g h t p e r i p o r t a l f a t t y changes

were seen i n mice kept on t h e d i e t f o r 2 or 4 weeks. I n the

rat, moderat? t o severe accumulat ions of p e r i p o r t a l fat were

seen from 7 weeks onwards. I n t he mouse l i v e r , a marked l o s s of

cytoplasmic p r o t e i n and RNA was noteZ after 4 weeks

on t h e d i e t . I n the r a t , g r o t e i n and RNA were reduced

after t h e an imals had been on t h e d i e t f o r 7 weeks. A l o s s o f

26.

35-40 p e r c e n t n i t r o g e n o c c u r r e d i n t h e l i v e r s of r e t s f e d cassava

f o r 7, 9 an6 15 wceks.

The a d d i t i c n of me th ion ine and c y s t i n c t o t h e d i e t o f mice

which h a d been f e d c a s s a v a f o r 14 days f a i l e d t o r e v e r s e t h e

changes t h a t h a d occur red . Supplements of o t h c r amino a c i d s

p a r t i c u l a r l y d e f i c i e n t i n cassava , namely, t r y p t o p h a n , phenyla-

lamine, h i s t i d i n e and v a l i n e a l s o f a i l e d t o have a n e f f e c t . Only

t h a a d z i t i o n of a comple te p r o t e i n containing a l l t h e e s s e n t i a l

amino a c i s s i n p r o p e r p r o p o r t i o n was e f f e c t i v e i n r e v e r s i n g a n d

p r e v e n t i n g t h e h i s t o l o g i c a l changes induced b y cassava , Dr ied

skim m i l k was t h e comple te p r o t e i n used. The mice gained weight

and t h e r e was an i n c r e a s e i n cy top lasmic p r o t e i n i n t h e l i v e r .

Ononogbu an< Emole (1978) found s w e l l i n g and v a c u o l a t i o n i n t h e

l i v e r c e l l s of rats f e d a d i e t of g a r i supplemented w i t h groundnut

cake , fish ueal and maize f o r f o u r weeks.

When changes from t h e normal occur i n t h e l i v e r , s y n t h e s i s

of a lbumin, an6 a l l o t h e r p r o t e i n s t h a t a r c s y n t h e s i z e d i n t h e

l i v e r parenchymal c e l l s i s impairecl end p l x m a c o n c e n t r a t i o n s

reduced. Depending on t h e degree cf ch?.nge w i t h i n t h e c e l l ,

t he re are ch-tnges i n t h e p e r m e a b i l i t y of t h e c e l l membraneo

When t h e c e l l is s u f f i c i e n t l y damaged, enzymes l e a k o u t of the

cell and t h e i r l e v e l s r i s e above normal u n t i l t h e c e l l s a r e

comple te ly exhaus ted and a r e u n a b l e t o s y n t h e s i z e more enzymes,

t hen t h e plasma l e v e l s f a l l below normal (Baron, 1982).

MATERIALS AN? METHODS

1. E x p e r i m e n t a l Animals - Management

The a n i m a l s f o r t h e p r o j e c t compr ised a group of 18 male

N i g e r i a n mongrel dogs of a p p r o x i m a t e l y 6 weeks cf a g e a c q u i r e d

randomly from v i l l a g e marke ts . I n o r d e r t o r e d u c e t h e i n c i d e n c e

of d i s e a s e ,

t r e a t e d f o r

t r e a t m e n t .

Te tmosol ( R )

as soon as t h e animals were b rough t i n , t h e y were washed,

e c t o p a r a s i t e s m d e n d o p a r a s i t e s and g i v e n a n t i b i o t i c

They were washed i n w a r m w a t e r w i t h med ica t ed s o a p ,

manufac tu red by Cnemical 2nd A l l i e d P r o d u c t s L t d - ,

N i g e r i z , t o w e l d r i e d , t h e n d ipped i n a 0.1% s o l u t i o n of

0,0-diethyl-O-(3-chloro-4-methyl-7-coumarinyl~-ph0sphorithioate,

( R ) k s u n t o l 50 manufac tu red by Bayer , Germany, as t r e a t m e n t f o r

e c t o p a r a s i t e s . They were g i v e n p rE .z iquan te l , D r o n c i t ' R' manufactured

by Bayer , Germany, a,osage, 5 mg/kg body w e i g h t , o r a l l y , once , f o r

tapeworms, an? p y r a n t e l pcmoat e , combant r i n ( R ) manufac tu red by

P f i z e r P r o d u c t s L t d . , N i g e r i a , dosage , 15 mg/kg body w e i g h t ,

o r a l l y , once , f o r hookworms and roundworms. They were given

o x y t e t r a c y c l i n e H C 1 5%, ~ d a r n ~ c i n ' ~ ) manufac tu red by "Assiaff

Chemical L a ! ~ o r a t c r i e s , I s r a e l , Gosage, 10 mg/kg body w e i g h t ,

i n t r a m u s c u l . ? r l y , d a i l y , f o r 3 Gays as a n a n t i b i o t i c , They were

a l s o g i v e n i r u n dex t ran , oni id ex'^) manufac tu red by Teva Ltd . ,

Israel, dosage , 12 rng/kg, c n c e , i n t r a m u s c u l a r l y , and v i t a m i n BIZ

manufac tured by Ge6.eon R i c h t e r L t d . , H u n g ~ r y , dosage, 200 mcg,

once , i n t r a m u s c u l a r l y . The a n i m a l s u e r e numbered by t a g g i n g a n d

housed i n d i v i d u a l l y i n m e t a b o l i c cages. Trea tment f o r e c t o p a r a s i t es

28.

and e n d o p a r a s i t e s were r e p e a t e d be fo re t h e start of t h e expe r imen t .

No t r e a t m e n t f o r e n d o p a r a s i t e s w a s done d u r i n g t h e e x p e r i m e n t a l

p e r i o d .

Mongrel dogs a r e l e s s r e l i a b l e s o u r c e s f o r normal d a t a u n l e s s

c a r e f u l l y s e l e c t e d f o r f reedom from o c c u l t d i s e a s e and c o n d i t i o n e d

f a r s e v e r a l months before d a t a a r c c o l l e c t e d ( ~ o a v e and Boyle , 1965)-

F o r t h e pu rpose of b r i n g i n g t h e a n i m a l s i n t o p r o p e r n u t r i t i o n a l

p l ane , a l l a n i m a l s were f c d a n u t r i t i o n a l l y b a l a n c e d d i e t f o r growing

a n i m a l s w i t h r i c e as t h e c a r b o h y d r a t e s o u r c e f o r 4 months b e f o r e

t h e stert of t h e expe r imen t ( ~ a i n e s Dog Resea rch C e n t e r , 1968).

The age of e a c h c m i m a l was d e t e r m i n e d by t e e t h e r u p t i o n a t

t h e end o f t h e p r e - e x p e r i n e n t a l p e r i o d and each a n i m a l was weighed.

2. E x p e r i m e n t a l Diets

The d i e t s fed i n t h i s exper iment were wet t y p e conked food-

I n t h e c o n t r o l d i e t , e x p e r i m e n t a l d i e t I , r i c e w a s u sed as t h e

s o u r c e of c a r b o h y C r a t e , w h i l e i n e x p 2 r i m e n t a l d i e t X I , g a r i was

u s e 6 as t h e c a r b o h y d r a t e Eource and i n e x p e r i m e n t a l d i e t 111, r i c e

t o which c y m i d e was added at t h e same l e v e l as found i n cooked

g a r i , was used as t h e c a r b o h y d r a t e s o u r c e -

The c l i e t s as ccmpounded were s u b j e c t e d t o p rox ima te a n a l y s i s

a c c o r d i n g t o t h e A.0.h.C. (1975) procedure , The cooked gari d i e t

was a l s o a n a l y z e d f o r t h e cyan ide c o n t e n t . The d e t p - i l s o f t h e

d i e t s a r e shown i n T a b l e s 1 and 2.

The d i e t s were i s o c a l o r i c , c o n t a i n e d approx i rne t e ly 13% crude

p r o t e i n an3 s u p p l i e d n u t r i e n t s i n a6ec,uate mounts znd proper

p r o p o r t i o n s f o r normal growth. P e d i a t r i c v i t a m i n and mineral syrup,

(R) T.M. ViDalyn-M , w a s a d e e d t o t h e d i e t of each an ima l d a i l y .

Approximnte ly 72% of t h e d i e t e n e r g y o f t h e gar i d i e t , cn a d r y

m a t t e r b a s i s w a s s u p p l i e d b y h i g h p r o t e i n i n t h e form of

lean pork because g a r i i s a lmos t devo id c f p r o t e i n , whereas

a p p r o x i m a t e l y 8% of t h e d i e t ene rgy of t h e r i c e d i e t , on a d r y m a t t e r

b a s i s , was s u p p l i e d by l e a n pork. Pork wns t h e p r o t e i n of c h o i c e

because of i ts h i g h t h i m i n (Vi tamin B ) c a n t e n t an? i n a d d i t i o n 1

gork f a t h a s a high l i n o l e n i c a c i d c o n t e n t (Gn ines Dcg Research

C e n t e r , 1968). The d i e t s were b a l a n c e d f o r c a l c i u m 2nd phosphorus

u s i n g bone meal.

B E 1 COMPOSITION OF DIETS ON DRY MATTER CALCULATIONS

Ingredients o r C a l c u l a t i o n s F e e 6 s t u f f s D i e t I D i e t I1 Die t 111

Gari , 74 - 70. 0 - Lean Pork, % 13.06 21-0 13-06 Bane meal , 7053 7.5 7 0 53

Vi tamin and m i n e r a l s u p p l amenta , m l I ml/kg food I ml/kg f o o d I ml/kg f o o d

Cyanide (MaCII 8.3 m g / m l ) , - - 2 ml/kg food

d ( R ) S u p p l i e d p e r ml of v i t a m i n and m i n e r a l s y r u p , ViDalyn-M - Vitamin 6, 600 I U. , V i t amin D , 80 I U ; Thiamine Hydrocho lo r ide ( E l ) , 0.3 mg; Riboflavin-5-Sodium Phospha te , 0.32 mg; ~ s c o r b i c k c i d ( C ) , 10.0 mg; Nicot inamir le , 2 - 0 mg; P y r i d o x i n e H y d r o c h l o r i d e ( B g ) , 0.2 mg; Vi tamin BIZ! 0.6 mcg; P a n t o t h e n o l , 1.0 mg; F e r r o u s Glucona tc , 5.2 mg; P o t a s s i u m I o d i d e , 1 9 ~ 6 mg; Calcium L a c t a t e , 30 r q ; Calcium Hypophosphi te , 16.5 mg; Manganese Glucona te , 0.9 mg; Zinc Glucohept ona t e , 0.84 mg; Msgnesium Glucon~ . t e l 11.1 mg; C h o l i n e B i t e r t r a t e , 2.08 mg; I n o s i t c l , 1.0 mg,

TABLE 2: PROXIMATE ANALYSIS OF WET DIET FED TO ANIMALS

D i e t I Diet I1 Die t I11

Crude p r o t e i n , % 12, 9 130 13 12,9

E t h e r e x t r a c t , % 4.6 5.30 4.6

R i t r c g e n F r e e Extrc.ct , % - Crude F i b r e , % 0.15 0.55 0.15

Ash, % - - - Energy, kcal/gm 6.03 6.36 6.03

3 Exper imenta l Design

A t t h e en2 of t h e pre-exper imenta l p e r i o d , t h e dogs were

randomly d iv ided i n t o 3 groups compris ing 6 i n d i v i d u a l s . The

an imals u c r e l e f t i n t h e ~ e t a b o l i c cages i n t o which t h e y wcre

i n i t i a l l y plncad. Dogs i n group 1 were f e d t h e c o n t r o l d i e t ,

experiment21 c'.iet I , group 2 t h e exper imenta l d i e t I1 and. group

3 t h e exper imenta l d i e t 111,

4. Feeding t h e Sxper imenta l Animals

For t h e d u r a t i o n o f t h e exper iment , t h e a n i n a l s were f e d

500 grn of r?ict once a dayo The amount of f c c d was based on t h e

body we igh t s at t h e beginning of t h e exper imenta l per iod. One

e x c e p t i o n a l l y l a r g e animal was f e d 1 kg of d i e t throughout t h e

exper imenta l p e r i o d , based on i t s weight compzred t o t h e weight o f

the c t h e r animals*

5. Apparent D i g e s t i b i l i t y of Diets

Apparent d i g e s t i b i l i t i e s o f t o t s 1 dry mat te r an2 of crude

p r o t e i n were de t erminefi u s ing t h e quant it8.t i v e f e c a l c o l l e c t i o n

method by Crampton and Lloyd ( 1 9 5 9 ) . Afer an adjus tment p e r i o d

cf s i x days f r c o the beginning of t h e exper imenta l f e ed ing , fecal

c c l l e c t i o n s were made over pe r iod of s i x days. Feces were

analyzed f o r proxirn2te f r a c t i o n s . The c o e f f i c i e n t of apparent

d i g e s t i b i l i t y of t o t a l d ry ma t t e r and of crude p r o t e i n were

c a l c u l a t e d u s ing d a t ~ from food i n t a k e , f e c a l e x c r e t i o n 2nd

proximate anal ys is.

6, De t e rmina t i a i of Weekly Weight Gains

Weighing cf t h e animals was don2 on day 1 of t h e exper imenta l

pe r i od and every 7 days t h e r e a f t e r u n t i l t h e end of t h e exper imenta l

pe r i od and t h e weekly weight ga ins c a l c u l a t e d .

7. Determinat ion cf t o t a l serum t r i i o d o t h y r o n i n e ( f ) concen t r a t i on 3-

Tota l serum t r i i o d o t h y r o n i n e ( T ) concen t r a t i on of all t h e 3

animals was determined be fo r e they were p laced on t h e e x p e r i m e n t d

d i e t and then a t s p e c i f i e d i n t e r v a l s u n t i l t h e animals were

sacr i f icecl. T o t a l serum t r i i o d o t h y r o n i n e (T ) w a s determined by 3

t he enzyme-linkecj. immuncsorbent assay p r i n c i p l e (ELIsA) u s i n g a kit

s u p p l i e d by Soehr inger Mannheim.

8. D e t e r m i n a t i o n cf plasma t h i o c y a n a t e c o n c e n t r a t i o n

Plasma t h i o c y a n z t e c o n c e n t r a t i o n of t h e a n i m a l s was

de t e rmined a t s p e c i f i e d i n t e r v n l s d u r i n g t h e e x p e r i m e n t a l p e r i o d

u s i n g t h e method Bowler (1944) as rncdi f ied by C h i l a k a (7984).

9, Determinnt i o n of u r i n e t h i o c y a n a t e c o n c e n t r a t i o n

T h i o c y a n a t e c o n c e n t r a t i o n of &-hour u r i n e c o l l e c t i c n

samples of t h e a n i m ~ l s w a s de t e rmined a t s p e c i f i e d i n t e r v a l s d u r i n g

t h e e x p e r i m e n t a l p e r i o ? u s i n g t h e method of Bowler (1944) as

m o d i f i e d by C h i l a k a (1984) .

10. D e t e r m i n a t i o n of p r c t e i n c o n c e n t r a t i o n i n u r i n e

P r o t e i n concent r a t i o n i n u r i n e was d e t e r m i n e d by t h e dip-and-

ren8. t e c h n i q u e u s i n g ~ a ~ i ~ n o s t ~ To ta l -Sc reen A T e s t s t r i p s at

s p e c i f i e d i n t e r v a l s .

11.

l e f t

D e t e r m i n a t i c n of t he r a t e of e p i p h y s e a l closure by roen tgenography

Roentgenographs of t h e a n t e r i o r and l a t e r a l a s p e c t s of t h e

f o r e l i m b of a n i m c l s on e a c h d i e t were t a k e n a t s p e c i f i e d

i n t e r v a l s and t h c r a t e o f c l o s u r e of t h e p rox ima l e p i p h y s i s of t h e

u l n a measured. The wicXhs of t h e u n c a l c i f i e d e p i p h y s e a l p l a t e s

were measured u s i n g 3 m i c r o s c o p i c g r a t i c u l e , The r e a d i n g of the

measurement was done u s i n g a s t e r e o m i c r o s c o p e . The a n i m a l s were

groupec! a c c o r d i n g t o age f a r t h i s pa rame te r ,

12. Determination c f plasma a l k a l i n e phosphatase (EC. 3.1.3.1) a c t i v i t y

Using hepa r in i zed plasma, t h e a c t i v i t i e s of a l k a l i n e

phosphatase (EC 7.1.3.1) were determined us ing an opt imized

s t a n d a r d method based on the rccommen?ations of t he Deutsche

G e s e l l s c h a f t f u r K l in i s che Chemie (anon, , 1970; Anon,, 1972).

Tes t p r i n c i p l s :

kP p-nitrophenylphosphate + H20 ->phosphate + p-ni t rophenol

The de te rmina t ions were c a r r i e d out be fo r e t h e animals were

p l ace6 on t h e exper imenta l d i e t s and then at s p e c i f i e d i n t e r v a l s

du r ing t h e exper imenta l pe r i o? u s ing k i t s s u p p l i e d by Boehringer

Mannheirn,

13. Det erminat i m of Plasma a c t i v i t y of L - x - g l u t amyl t ransf erase ( X C 2.3.2.2)

Using heparinizecl plasma, t h e a c t i v i t i e s of L-% -glutamyl-

t r a n s f e r a s e (EC 2.3-2.2) were determine? f o r a l l t h e exper imenta l

animals b e f c r e t he an imals were p laced on t h e exper imenta l d i e t and

a t s p e c i f i e d i n t e r v a l s d u r i n g t h e exper imenta l pe r i od u s ing k i t s

s u p p l i e d by Boehringer Mannheirn accord ing t o t h e method of P e r s i j n

Test p r i n c i ~ l e :

L-X -glutnmyl- 3-carboxy-4 n i t r o a n i l i d e + g lycy lg lyc ine ' 4T L-2. - glu tamylg lycy lg lyc ine + 5-amino-2-nitrobenzoate.

I Determinat ion of p,lasma a c t i v i t y of n l a n i n e a m i n o t r a n s f e r a s e (EC 2.6.1.2)

Using h e p a r i n i z e d plasma, t h e a c t i v i t i e s of a l a n i n e amino-

t r a n s f e r a s e (EC 2.6.1.2) were determined by a n opt irnized s t a n d a r d

method conforming t o t h e recommendations of thc Deutshe G e s e l l s c h a f t

f u r K l i n i s c h e Chemie (Anon., 1970; Anon., 1972).

T e s t p r i n c i p l e :

p y r u v a t e + MADH + H +. LDHb L - l a c t a t e + NAD+ 7

The d e t e r m i n a t i n n s were c a r r i e d out b e f o r e t h e an imals were

p laced on t h e expcr imen ta l d i 2 t an< t h e n a t s p e c i f i e d i n t e r v a l s

during t h e exper imen ta l pcr ioeL u s i n g k i t s s u p p l i e d by Boehr inger

Mannhe i r n . 15. Det e r n i n a t i o n o f plasma i s o c i t r a t e dehy&ogennse (NADP')

(EC 1.1~1.42)

Using heparinizec! ~ l a s r n a , t h e a c t i v i t i e s of i s o c i t r a t e

dehydrogenase (NADP+) (EC 1.1 . 1-42) were determine? f o r a l l t h e

an imnls b e f o r e t h e s tart cf t h e experiment a n d t h e n a t s p e c i f i e d

i n t e r v a l s d u r i n g t h e expcr imen ta l p e r i o d u s i n g k i t s s u p p l i e d by

Boehr inger Fannheim a c c o r d i n g t o t h e method of Wolfsen e t a l . (1951). - m-

T e s t p r i n c i p l e :

i ~ o c i t r a t e + NI,DF+ -0xog1utarnte + CO + NADPH + H' 2

16. Determination of ~ l n s m a a c t i v i t y of lipase (EC 3 . 1 - 1 a

Using hepar inize? . plasma, t h e a c t i v i t i e s of l i p a s e (Ec 3.1.1.3)

were determinec! f o r a l l the m i n d s be fo r e t h e start of tihe experiment

and then a t s p e c i f i e l i n t e r v a l s dur ing t h c experimental p e r i o d using

k i t s s u p p l i e d by aoehr inger Mannheim acco r s ing t o t h e method of

Neurnann and Ziegenhorn ( 1979).

Tes t p r i n c i p l e :

li ase t r i o l e i n + 2H20 -PI monoglyceride + o l e i c a c i d ( 2 moles)

17. Determinat ion of hemato log ica l parameters

The hemoglobin concen t r a t i on and t h e packed c e l l volume of each

animal were determined by the cyanmethemoglobin method and t h e micro-

hem2t o c r i t metho2 r e s p e c t i v e l y a t t h e end of t h e exper imenta l per iod.

18. De t e rn ina t i on of t h y r o i d weiah t s cf t h e animals --

When t h e animals were s a c r i f i c e d at t he end of t h e exper imenta l

p e r i o d , t h e t h y r a i d g l a n d o f each animal was removed a n i weighed and

expressed as grn/kg body weight of animal.

19. Determinat ion of plasma calcium and qagnesium c o n c e n t r a t i o n s

Plasm concen t r a t i ons o f calcium and magnesium i o n s a t t he end

o f t h e exper imenta l peri.03 were Zeterrnined using Pye Unicam SP 90 A

S e r i e s Atcrntc Absorption Spectrcyhotometer.

20. Det ermr-na t i o n of Plasma inorgan ic phosphorus c o n c e n t r a t i o n

Plasma concen t r a t i on of inorgan ic phosphorus at t h e end of t h e

exper imenta l periocq was determined c o l o r i m e t r i c a l l y using F i s h e r

Electrophotorneter 11,

RESULTS

APPARENT DRY MATTER DIGESTIBILITY

The r e s u l t s of t h e d i g e s t i b i l i t y a s p e c t s cf ths experiment a r e

summarized i n Table 3 , Apparent d r y m a t t e r d i g e s t i b i l i t y f e r t h e r i c e ,

g a r i , and r i c e + cyan ide groups were 90.865, 91.00 and 88.95 pe r c en t

r e s p e c t i v e l y . These were n c t s t a t i s t i c a l l y d i f f e r e n t ( P 7 0.05).

These results suggest t h a t n e i t h e r cyanide a t t h e lev21 f e d nor cyan ide

i n gar i a f f e c t e d d ry m a t t e r d i g e s t i b i l i t y o

TABLE 3 : EFFECT OF CYANIDS COIVTAINING DIETS CN APPARENT DRY MATTER DIGESTIBILITY

D i e t a r y T r e a t ~ e n t - - -

Rice Gar i R i c e + Cyanide

91-63 91.09 92-05 95.14 89.24 92.27 89.24

90.34 9 0 ~ 8 4 82.68 90.77 97-79 76-46 91.79

92-84 90.46 91-80 91.25 86.46 95,O 86.46

88.66 91.10 8 8 3 2 y x 4 1 87.90 92.32 87-90

knimal Mean 90.86 90.87 88.86 93*14 88-85 89.012 89.017

Group Nean 90. 865a 91 .Ooa 88. 95a

S-E-M. 0,005 2, 14 0.056

a Means with t h e same s u p e r s c r i p t i n t h e rold a r e not s t n t i s t i c a l l y d i f f e r e n t (P > 0.05).

GROWTH Ri.TE

The e f f e c t s of t h e t r e a t m e n t s rm growth r a t e a r e shok!n i n

i;>ppendix 1, ant7. , s r n ~ h i c n l l y i l l u s t r 2 t e d i n F i g . 7 . The weekly i n c r e a

in weigh t were s imilar i n bo th t h e c c - n t r o l r i c e GrouF a n d t h e g n r i

group. However, t h e growth of t h c r i c e + cynn ide group was

s i g n i f i c m t l y &?pressed as e a r l y 2,s week one ( P -= 0.05). T h i s growth

r e t a r d i n g e f f e c t c o n t i n u e d u n t i l t h e end c f t h e exper iment . Thus

by t h e end cf t h e 1 4 t h week t h e mean p e r c e n t a g e i n c r e a s e i n weight

was o n l y 5 3 0 2 2 f c r t h e r i c e + c y a n i d e Grcup compare?. 50 80.43 and

83.35 f o r t h e r i c e and g a r i g roups r e s p e c t i v e l y . It i s n o t e w o r t h y th:

t h e g a r i g roup t e n 2 e d t c be s u p e r i o r t o t h e r i c e group. R e g r e s s i o n

analysis ( F i g u r e 2) s h c w ~ d t h a t growth w s s l i n e a r f c r a l l t h e groups;

and t h e r e g r e s s i o n c o e f f i c i e n t s of 5.966, 5.95 and 3.938 f o r t h e r i c e ,

g a r i 2nd r i c e + c y a n i d e g r c u p s r e s p e c t i v e l y ccnf i r rn t h ? t t h e growth

r a t e of t h c r i c e + c y a n i d e grcluF was s i g n i f i c a n t l y d i f f e r e n t from

t h e r e s t ( P c 0.05). These r.esu1t.s show t h a t a l t h o u g h t h e a d d i t i o n

of c y a n i d e t o r i c e d e p r e s s e d growth , t h e c y a n i d e i n gari h a d no e f f e c l

on growth.

Time '(weeks)

THE THYROID GLAND AND SECRETION

The e f f e c t of cyanide containing foods on plasma T toncen t ra t t a l 3 during the courae o f tho experiment and the weightti cf the thyro id

gland at the end of the experiment are shown in Table 4 and Appendix

2 and Figure 3. The e f f e c t of the experimental t r ea tmen t s on T 3 levels was h i ~ h l y s i g n i f i c a n t (P < 0.01). Lge o r t ime effect and

d i e t x t ime i n t e r a c t i o n were not statistically s i g n i f i c a n t (P> 0.05).

The gari group had a significantly higher T level a t the beginning of 3 t h e experiment (P < 0.05). However, t h e d i f f e rences disappeared

by the t h i r d week. From t h e n Onr t h e plasma T l e v e l s f o r the rice and 3 gari groups tenfled to rise while T l e v e l s f o r t h e rice + cyanide poup 3 tended t o eec l ine . h t termination of the experiment by t h e 74th

week, the mem T l e v e l s were 81.67 ng/dl, 316.67 ng/dl and 36.67 3

ng/dl for t he r i c e , gari and r ice + cyanide groups respsctively,

The di f ferences between t h e g a r i and r i c e + cyanide groups was

stat&stloally d g n i f icant (P c 0.05)

The mean t hy ro id gland weights expressed as g/kg body weight

were 179, 173 rcnd 212 f o r the g a r i , rice and r i c e + cynnic?e groups

respec t ive ly . The difference between t h e gari and rice + cyanide

p 0 u p s was s i g n i f i c a n t (PL. 0.05). The ranking of the thyroid gland

weights agrees with the ranking of t he T l e v e l s although in reverse 3

order. It is ev ident , however, t h a t although gari con ta ins cyanide,

a gar i d i e t af fec t s the thyroid gland i n a way different from free

cyanide added t o food.

TABLE 4: EFFECT OF C Y A N I D X COIYTiiINING DIETS ON WEIGHT OF THE T H Y R O I D GLLRD j,FTER 14 WEEKS

T h y r o i d w e i ~ h t s ( g / k ~ body weight) D i c t a

Dietary Treatments Rice Gari Rice + Cyanide

Mean

a b ' Means with t h e snme superscript i n t h e rcw are n c t s t n t i s t i c a l l y d i f f e r e n t (P 3 0,051.

PLASMA THIOCYfdiATE CONCENT1U.TIONS

Appendix 3 an?: F ig . 4 shaw t h e e f f e c t o f d i e t a r y treatment on

p la sms t h i o c y a n a t e c o n c e n t r a t i o n s and how i t v a r i e d ?.uring t h e c o u r s e

of t h e e x p ~ r i m e n t ~ Plasma from a r i c e group a n i m a l was u s e d as t h e

t h i o c y a n a t e f r e e stnnclnrcl s o as t o c a n c e l out l e v e l s of e n d o p n o u s

t h i o c y a n a t e , Before t h e a d m i n i s t r a t i o n of t h e v a r i o u s t r ez t rnen t s ,

p lasma t h i o c y a n a t e was no t d e t e c t a b l e i n a11 t h e groups . By t h e end

of t h e f i r s t weck of t h e e x p e r i m e n t a l p e r i o d , t h e p lasma t h i o c y a n a t e

l e v e l s had r i s e n t o 0,0908 u mole/ml and 0,0802 u mole/ml i n t h e gari

a n 6 r i c e + c y ~ n i c e [ y o u p s r e s p e c t i v e l y , The t h i o c y a n a t e c o n c e n t r a t i o n

i n t h e r i c e group remained u n d e t e c t a b l e t h roughou t t h e p e r i o d of

14 weeks. Tha plasma t h i o c y a m t e l e v e l s i n t h e r i c e + c y a n i d e group

r o s e t o 0.1794 u mole/ml by t h e 1 4 t h week, w h i l c t h e l e v e l s i n t h e

g n r i g r o q d c c l i n e d t o 0,0486 u rnole/ml d u r i n g t h e same p e r i o d .

S t a t i s t i c a l l y t h e c l i f f e r c n c e s i n res;mnsc were h i g h l y s i g n i f i c a n t

( F 4 0-01) A.lthough t h e feeding o f g n r i ani? r i c e + c y a n i d e w i l l

c l e v a t e plasm.? t h i o c y a n a t e c o n c e n t r a t i o n s , t h e p a t t e r n of change

appears t o bc d i f f e r e n t i n t h e two groups ,

Because o f t h c known e f f e c t of t h e t h i o c y a n a t e i o n on t h e t h y r o i d

g l a n d , plasma t h i o c y a n a t e m . 1 T c o n c e n t r a t i o n s were s u b j e c t e d t o 3

c o r r e l a t i o n a n a l y s i s ( F i r - u r e 5)- Thc c o r r e l a t i o n was n e g a t i v e , which

rzeans t h a t the c o n c c n t r n t i o n of T d e c r e a s e s as t h e c o n c e n t r a t i o n of 3

t h i o c y a n n t e i n c r e a s e s . The c o r r e l a t i o n c o e f f i c i e n t s of -0,874 2nd

-0,494 f o r t h e g n r i and r i c e + c y a n i d e grGups r e s y e c t i v c l y a r e

r e l a t i v e l y h i g h :lthouc.h n o t s t a t i s t i c a l l y s i k , n i f i c a n t (P O.O5) ,

I

0.00 Y A - - & - e 0 2 4 6 8 10 12 14

Time (weeks)

47 0

URIhE THIOCYAfiJATE CONCLN!!&TIONS

A-pyendix 4 and F i s . 6 show t h e e f f e c t of c y a n i d e c o n t a i n i n g f o o d s

on u r i n e t h i o c y a n a t e c o n c c n t r r t i o n s . Thc v ~ r i m s d i e t s r y t r e a t m e n t s

s i g n i f i c a n t l y a f f e c t e d t h s u r i n e t h i o c y a n a t e l e v e l s ( P -L. 0.01).

The r i c e + cyzniZc Trolly hscl t h e h i g h e s t u r i n e t h i o c y s n n t e c o n c e n t r a -

t i o n f o l l o w e d by t h e p r i t-;roup, The u r i n e o f a n a n i m a l of t h e r i c e

group w a s u sed as t h e t h i o c y a n a t c f r e e s tandarc? s o as t o c a n c e l o u t

l e v e l s of any endogsnous t h i o c y a n a t e . As a r e s u l t t h e r e was no

d e t e c t a b l e t h i o c y a n a t c i n t h e r i c e group t h r o u g h o u t t he expe r imen t ,

Time e f f e c t and t i m e x d i e t i n t e r a c t i o n were a l s o h i g h l y s i g n i f i c a n t

( P 0.01). 3rom F i g u r e 6 i t is s e e n t h a t f o l l o w i n g a n i n i t i a l high

r i s e i n u r i n e t h i o c y a n a t e l e v e l s , t h s c o n c e n t r a t i o n s g r ~ d u a I l y

d e c l i n e d as t h e cxpcr iment proceeded d e s p i t e c o n t i n u e d i n g e s t i o n of

c y a n i d e c o n t a i n i n g food , T h i s e x p l a i n s t h e s i g n i f i c a n t time e f f e c t

and t imc x d i e t i n t e r a c t i o n .

S i n c c u r i n e is o b t a i n e d from plasma f i l t r a t c t h r o u c h t h e g l o m e r u l u s ,

t h e e x i s t e n c e of a p o s s i h l s r c l n t i o n s h i p between p lasma and u r i n e

t h i o c y a n a t c c o n c e n t r a t i o n s was examined. R c s u l t s of t h e c o r r e l a t i o n

a n a l y s i s hctwcon rlasma and u r i n e t h i o c y a n a t e l e v e l s a r e shown i n

F i g u r e 7. The c o r r e l a t i o n i s p o s i t i v e f o r b o t h t h e g n r i and r i c e

+ c y a n i f c p c u p s . T h i s m a n s t h a t t h e u r i n e t h i o c y a n a t e c o n c e n t r a t i o n s

i n c r e a s e d as t h e p lasma t h i o c y a n a t c i n c r e a s e d , The z o r r e l a t i o n

c o e f f i c i e n t s of +0.782 and ~ 1 , 8 0 6 f o r t h e g n r i a n d r i c e + c y a n i d e

c r o u p s r c s p ~ c t i v e l y , are r e l a t i v e l y h i g h a l t h o u g h n o t s t a t i s t i c a l l y

s i g n i f i c a n t ( P : 0.05). These r e s u l t s i n d i c a t e t h a t t h e i n g e s t i o n

of cyani?.e c o n t ~ i n i n g f o o d s w i l l r e s u l t i n e l e v z t e d u r i n e t h i o c y a n a t e

Figure 6. Variation of mean (2 s.E.M.) urine thiocyanate concent rat ion w i t h t in@ during t h e experimental period. (+animals f e d control ( r i c e ) diet; ----Q-- animals f e d gari diet;--* anirn~ls f e d rice + cyanide d i e t ) .

Time (weeks)

Figure 7. Linear' correlat f on between plasma thiocyanate concentrat ion and urine thiocyanat e concentration, (--animals f e d con t ro l ( r i c e ) diet; + animals f ed gari d i e t ; animals f e d rice + cyanide diet).

URINE PK OFTKIN CONCENTRAT I O N S ~ - - ~~

H c s u l t s of t h e a n d y s i s of u r i n e p r o t e i n c o n c e n t r a t i o n s are

shown i n kpi2endix 5 and Fit, 8. The c f f e c t s o f d i e t and t ime on u r i n e

rotei in levels were h i g h l y s i ~ n i f i c a n t ( P < 0.01) ., Die t x t i m e

i n t e r a c t i o n was :also h i g h l y s i g n i f i c a n t (F < 0,01), T h i s means t h a t

u r i n e p r o t e i n l e v c l s were de t e rmined by the d i c t a r y t r e a t m e n t as well

as the l e n c t h of t i m e on t h e c!ict. From F i i ~ u r e 8 it can be s e c n t h a t

t h e cyanide c o n t a i n i n c d i e t s , , yz r i an? r i c e + c y a n i d e , c a u s e d p r o t e i n

t o ai3pear i n u r i n e , d l i l e t h e c o n t r o l , r i c e group showed no u r i n a r y

p r o t e i n t h r o u c h o u t t h c expcrirnen t a l perioc?. F o r t h e g a r i g roup ,

u r i n e p r u t e i n c o n c e n t r a t i o n r o s e from z e r o t o 15 mg/a by t h e 7 t h

day, ;mcl. t h e n rose t o 76 m g / d l by t h e f i f t h week and seemed t o h a v e

p e r s i s t e f . at t h i s h i g h l e v o l u n t i l t h e end of t h e expe r imen t , Fo r

t h ? r i c e + c y a n i d e group t h e s i t u a t i o n was t h a t of a g r a d u a l r i s e

i n u r i n e p r o t e i n c a n c e n t r a t i o n s from z e r o t o 48-33 mg/dl by t h e 1 4 t h

week. Thc eari p o u p t h e r e f o r e showccl t h e h i g h e s t u r i n a r y p r o t e i n level

Figure 8. Variation of mean (2 S.E.M.) urine protein concentration with time during the experimental period. (+animals fed control (rioe) diet;

+animals fed gar% d i e t ; + animals fed rice + cyanide diet).

4 6 8 Time (weeks)

EPIFHYSEAL CLOSURE

R e s u l t s af t h e measurement of t h e wiPLth of u n c a l c i f i e d

e p i p h y s e a l plates a r e shown i n Appendix 6 and F i g , 9, E p i p h y s e a l

c l o s u r e was s i s n i f i c a n t l y inf luencecl by c y a n i d e c o n t a i n i n g foods

( P .( 0,01), Tim< e f f e c t wzs a l s o found t o be h i g h l y s i g n i f i c a n t

( F < 0.31), I n t e r a c t i o n between t r ca t rncn t and t i n t : was n o t

s i g n i f i c a n t (P 7 0.05), G r o u ~ ccmpar ison en t h c t r e a t r n c n t s showed

t h a t t h e e f f e c t s o f r i c e and gari d i e t s were similar a l t h o u g h

d i f f e r i n g from r i c e + c y a n i d e which s i g n i f i c n n t l y i n h i b i t e d

c ~ l i p h y s e a l c l o s u r e ( P L 0.05)- By t h e en$ of t h e f i r s t week, t h e

i n h i b i t o r y e f f e c t of r i c e + cyani(Je idas a p p a r e n t . The i n h i b i t o r y

e f f e c t c o n t i n u e 6 u n t i l t h e end o f t h e expe r imen t , The r e s u l t s

in i1 . ica te t h a t a l t h o u g h g a r i c o n t a i n s c y a n o i ; c n i ~ g l u c osicjes i t

e x e r t s no i n h i b i t o r y e f f e c t on ec . iphysea1 c l o s u r e when f e d i n a

b d a n c c d c7ie t .

0 2 4 6 8 10 12 t ime (wed@

PLASMA ENZYMES :

The a f f e c t s of cyan iSe c o n t a i n i n g f c o d s on s e l e c t e d p lasma

enzymes a r e summarized i n T a b l e 5 ,

I s o c i t r i c dehy2rogenase ( I C D H ) a c t i v i t i e s were n o t a f f e c t e d

by t h e d i e t a r y t r e a t m e n t s ( P =- 0 . 0 5 ) ~ However, t i m e e f f e c t was

h i g h l y s i g n i f i c a n t (P( 0.01). I n a l l t h e g roups ICDH a c t i v i t i e s

were s i g n i f i c a n t l y l ower a t week 14 t h a n a t week 0. The s i g n i f i c a n t

t ime e f f e c t a p p e z r s t o be t h e normal e f f e c t of age as o b s e r v e d i n the

human ( ~ l l i s and Gc ldbe rg , 1971).

D i e t a r y t r e a t m e n t s had no s i g n i f i c a n t e f f e c t s on t h e p l a sma

a l a n i n e a r n i n o t r m s f e r a s e ( ALT) a c t i v i t i e s (P .> 0.05). However,

t i m e e f f e c t w a s h i g h l y s i ~ n i f i c a n t ( P < 0 ~ 0 1 ) . ALT a c t i v i t i e s

f o r a l l t h e w o u p s a t week 0 were a lmos t doubled by week 1 4 , b u t was

w i t h i n t h e normal range.

A l k a l i n e phoa l~ha . t a se a c t i v i t i e s were n o t a f f e c t e d by e i t h e r

d i e t a r y t r e a t m e n t o r t ime (P > 0.05). However t r e a t m e n t compar ison a t

week 1 4 showed s t a t i s t i c a l s i g n i f i c a n c e between t h e g a r i and +

c y a n i d e grculss ( P < 0.05). T h i s e f f e c t i s r e l a t e d t o t h e d i f f e r e n c e

i n o s t e o b l a s t i c a c t i v i t y i n growth and c a l c i f i c a t i o n of t h e bones

of t h e an ima l i n t h e two g r o u p ,

G a m m a g l u t n m y l t r a n s f e r a s e a c t i v i t y w a s n o t i n f l u e n c e d by

e i t h e r t h e e x p e r i m e n t a l t r e a t m e n t s o r t i m e ; and t h e r e was a l s o no

s i g n i f i c a n t t ime x d i e t i n t e r n c t i o n . S i m i l a r l y t h e r e w a s no s i g n i -

f i c a n t 6 i f f e r e n c e between any of t h e t r e a t m e n t means (P > 0.05).

Plasma l i p a s e a c t i v i t y was n o t s i g n i f i c a n t l y a f f e c t e d by

dietary t r e a t m e n t s and time ( P > 0.05), However, t h e r e was a s i g n i -

f i c a n t d i e t x time i n t e r a c t i o n ( ~ 4 0.05). S i g n i f i c a n t i n t e r a c t i o n

56 0

o c c u r r e d because of t h e d i f f e r e n c e s i n t h e d i r e c t i o n of r e s p o n s e w i t h

t ime. Thus i n t h e g a r i group t h e r e was e l e v a t i o n of l i p a s e a c t i v i t y

from 64.17 u / l at week 0 t o 175.30 u / l at week 14, On t h e o t h e r

hand l i p a s c a c t i v i t y was Sepressed i~ t h e r i c e and r i c e + c y a n i d e

croups from 84,0 u / l t o 6 6 0 ~ u / l and 81.33 u / l t o 72.55 u / l L,

r e s p e c t i v e l y . Consequent ly mean ccrnparisons a t week 14 ehow

st st i s t i c a l s i g n i f i c a n c e between gari r i c e g roups , and gari

and r i c e + cy3nic.k g roups ( ~ d . 0.05).

These r e s u l t s show t h a t a l t h o u g h cyanide c o n t a i n i n g f o o d s may

have no effects on plasma i s o c i t r i c dehydrogenase, a l a n i n e

aminot rans fe rase a n d gamma @ l u t a r n y l t r a n s f e r a s e , t h e y may s i g n i -

ficantly i n f l u e n c e alakaline ~ h o s ~ h a e a s e and l i p a s e a c t i v i t i e s

when f e d over 2 l o n g period of timc. The r e s u l t s a l s o show t h a t

r e s p o n s e s t o gari and r i c e + cynniGe d i e t s may g e n e r a l l y b e d i f f e r e n t .

HEMATOLOGICAL CHANGES -- The e f f e c t cf cyani(:e cont:linin;; cl.iets on t h e hemogloSin

c o n c e n t r a t i o n of bloc( ' ?mG p?.cked c e l l volume a f t e r a 14-week f e e d i n g

p e r i r ~ d is shown i n Tab le 6 , None of t h e p n r a m e t e r s were s i g n i f i c a n t l y

affectei! !;y any n f t h c d i e t a r y t r e , ? tmen t s ( p . 7 0.05)

TABLE 6: EFFECT OF C Y A N I D E C O N T I ~ ~ I N I N G DIETS ON HEMOGLOBIN GUNCEN'1R?-TION OF 3 ~ 3 0 ~ AND PLCKED C E L L V O L U I ? ~ AFTiR 14 WEEKS

Parameter Die ta ry Trea tmen t

R e p l i c a t e s

(g/100 ml) Gnr i 16.k 12.0 13-6 14-45 I 14.0 14,06~+0.5€ -

Packed c e l l R i c e 44.2 42.0 49.0 50.0 40.0 40.0 44.2b+~.8~ - volume (%)

G a r i b 53.0 40.0 44.0 46,66 46.0 44,O 45.61 +-1.76

R i c e + Cyanide 40.0 51.0 40.0 52.0 41.0 42.0 44.3jbt2.2i -

n , b Mesns w i t h t h e same s u p e r s c r i r : t i n t h e scJournns are n o t s t a t i s t i c a l l y ;:if f e r e n t (P ; 0 0 0 5 ) .

PLASMA ELECTROLYTLS

Tab le 7 shows t h t c f f e c t of c y a n i e c c o n t a i n i n g foo6s on p lzsma

e l e c t r o l y t e s a f t e r 2 14-week feedin:; p e s i u d .

The d i c t a r y t r e a t m e n t s had a h i g h l y s i ~ p i f i c a n t e f f e c t cn plasma

c a l c i u m j e v e l s . The means f o r t h e r i c e , g a r i a n d r i c e + z y a n i d e

Grsups w e v e 3-58 mmole/l; 3 -08 mrnole/l an< 3.58 rnmol(./l r e s p e c t i v e l y ,

The g a r i group had a s i g n i f i c a n t l y l o w e r c a l c i u m l e v e l ( P L 0-05).

The g a r i grcup had t h c h i g h e s t i>hosphorus c o n c e n t r z , t i m ;

however , t h e group < i f f e r e n c c s were n o t s t a t i s t i c d l y ~ i ~ ~ n i f i c a n t

( P ) 0.05). D i f f e r e n c e s i n rnagnesimn and sodium c o n c e r A r a t i o n s were

n o t s t a t isticnlly s i g n i f i c a n t (P 7 O,O5),

P l a s na po ta s s ium c o n c e n t r a t i o n s was h i g h e s t i n t h e c o n t r o l , r i c e ,

(7rou-p (7.133 mmole/l) and lowes t i n t h e g a r i group (5,936 mmole/l) . The d i f f e r lnce between t h c s c two groups was s t e t i s t i c a l l y s i g n i f i c a n t

(P 4 0.05) - The r c s u l t s i n d i c a t e t h a t o n l y c a l c i u m an6 potassiu.m c o n c e n t r a t i o r

were s i g n i f i c a n t l y i n f l u e n c e d by t he e x p e r i m e n t a l t r e a t n e n . t s , The

d i f fe rence : . wei-c a c t u s l l y s e e n i n the g x r i g roup.

W W W 0 0 0

t"0-4 '.Jl

W W W P O 0

w l o w l

W U W a m .

0 0 0 V1 Cou W W W

IIISTOPATHOLOGICAL F ' INDIEGS - ----

I n t h c grouF of a n i m a l s f c d t h e c c n t r o l d i c t , t h e l u n i n a of

most o f t h e f o l l i c l c s were large, some were r e l a t i v e l y s m a l l , o r l y

a f e w f o l l i c l e s c c n t a i n e d c o l l o i d , most were e m ~ t y a n d i n some

f o l l i c l e s t h z c o l l o i d s t a i n e ? more b ~ ~ s o p h i 1 i . c t h a n i n a t h e r s

(F ig . 10)- The e p i t h e l i u m of t h e f o l l i c l e s t h a t were f i l l e d w i t h

c o l l o i d was c u b o i d a l , b u t t h e e p i t h e l i u m o f t h o s e f o l l i c l e s t h a t

were e r n p t ; ~ , or cont .2ined a s m a l l nmctunt 12f c o l l o i d was colurnr?,ar

( F i g s - 11 .?nd 12), There was d e s q u m a t i o n of c o l u r n a r e p i t h e l i d

c c l l s i n t o t h e lurninn of t h 2 emyty f o l l i c l e s (FiE. 13) .

In l h c group of z n i m s l s f e d t h e g a r i c l i c t , a l l t h e . f o l l i c l e s

c o n t a i n e d Co l l . ? id ( F i g , 1 4 ) - The e p i t h e l i a l c e l l s l i n i n g t h e

f o l l i c l s s F i l l c d w i t h c o l l o i d was c u b ~ i d a l ( F i g . 1 5 ) . The

e p i t h e l i a l c e l l s of f o l l i c l e s w i t h s c n n t y amounts of c o l l o i d were

a l s o c ; u b o i d d b u t l z r g e r t h a n t h o s e of thi : f o l l i c l e s f i l l e d w i t h

c s l l o i d , ,There was no c!esquarnation of t h e e p i t h e l i a l c e l l s i n t o t h e

1-umina cf f o l l i c l e so

I2 t h ? g roup of animals f ed t h e r i c a + c y ~ n i c ? e d i e t ; , t h e

l a m i n a of t h e f c l l i c l c s v a r i e d g r ? ? t l y i n s i z e ( P i g . 16 ) , ,

C o l l o i d wzs present i n on ly v e r y f m f o l l i c l c s , The c n l l o i d i n

t he f o l l i c l e s was p l e s t a i n i n g nnrq t e n d e d t o be s l i g h t l y b - ~ s o p h i l i c .

The f o l l i c l e s had much t h i c k o r w a l l s t h a n t h e f o l l i c l e s o f any o t h e r

grcup anc! t h i s w a s 6.uc t o a n i n c r e a s e i n t h e number of l a y e r s o f c e l l s

which fornet! t h e w d l s , Thc c e l l s i n t h e t h i c k e n e d walls were c u b o i d a l

(Rige 17) E Thcrc was desqunrnaticm of e ~ i t h e l i a l c e l l s i n t o t h e

lumina of enl) ty f o l l i c l e s ( F i g o 78).

64 .. Figure 12. Thyro id g l a n d o f an ima l on c o n t r o l d i e t . Columnar

e p i t h e l i u m l i n i n g f o l l i c l e c o n t a i n i n g s c a n t y colloid.

x 128,

F igu re 13. Thyro id g l a n d o f a n i n a l on c o n t r o l d i e t .

Desquamat i o n of cpi t l ie l i -urn i n t o lumen of empty

f o l l i c l e .

Figure 10, T h y r o i d gland of animal on c o n t r o l d i e t .

x 3 2 ,

Figure 11. Thyro id g lznd of a n i m a l on c o n t r o l d i e t . Cubo ida l

e p i t h e l i u m l i n i n g f a l l i c l e f i l l e d w i t h c o l l o i d .

x 128.

65 0

Figure 14- T h y r o i d g l a n d o f animal on g a r i d i e t ,

x 32.

F i g u r e i50 T h y r o i d g l a n d o f animal on g a r i d i e t , Low c u b o i d a l

e p i t h e l i u m l i n i n g f o l l i c l e f i l l e d wi th c o l l o i d p

x 128,

6 6 .

F i g u r e 16. Thyroid g l a n d of an ima l on r i c e + c y a n i d e diet.

x 32.

F i g u r e 17- T h y r o i d ~ l 9 n r . J of animal on r i c e + cyanide d i e t .

Th i ckened f o l l i c u l a r walls composed of cuboidnl

c e l l s ,

x 1 2 8 ~

Figure 18. Thyro id g l a n d of animal on r i c e + cyan ide d i e t .

Desquamat ion of epithelium i n t o lumen of empty

f o l l i c l e ,

x 128.

Epiphyseal growth p l a t e - I n t h e c o n t r d animzls and i n t h e an ima l s f e d t h e g a r i diet,

no e2 iphysea l ~ r c w t h p l z t a c c u l d be d i s t i n g u i s h e d . The epiphyseal

growth plate had been r e p l a c e d by bone, In t h e a n i m a l s f e d t h e

rice + cyanic-?c d i e t , t h c e p i p h y s e a l [yrowth p l a t e w a s present a n d

c o n s i s t e d o f a zcne of r e s t i n e ; c a r t i l ~ g e , a zone cf m a t u r i n g

c a r t i l a g e znc? a zone of c a l c i f y i n g c a r t i l a g e (Fig . 19).

6 9

Figure 19. Spiphyseal growth p l a t e o f a n i m a l on rice + cyanide

d i e t

x 80.

L i v e r - Thc group of animals f e d t h e ~ z r i c l ie t showed v a c u o l x t i o n

of t h e h e p a t i c tills which wns p e r i p w t a l i n d i s t r i b u t i o n

(Fig. 2 0 ) .

71 0

Figure 20, L i v e r o f an ima l on g a r i d i e t . F e r i p o r t a l

v a c u o l a t i o n ,

x 128,

/ L a

P a n c r e ~ s

I n t h e grcup af n n i m a l s f e d t h e g a r i d i e t , t h e p a n c r e a s showed

n e c r o s i s , hemorrha,;.;:e, f i b r o s i s and a t r o p h y . The edges of t h e sections

appeared i r r e g u l a r . T h i s ccnd i t i . on wss t h e r e s u l t o f f i b r o s i s a n d

a t r c p h y o f t h e Tancreas c ~ u s i n g t h e organ t o become sh runken a n d

t o h2.w an i r r e g u l a r o u t l i n e . I n s u m a r e a s , t h e c o n t e n t s of t h e

l o b u l e s h,?d unc!cri;one f r a g m e n t a t i o n ( F i g . 2 1 ) . The a c i n a r c e l l s

h a d becornc Cicloclged 3rid i n seme c e l l s , t h e cy top la sm had become

e o s i n o p h i l i c , i n o t h e r s t h e c y t op l~ , sm w a s r e l a t i v e l y srnal.1 i n

q u a n t i t y ,z.n$. b a s o r h i l i c an?. n u c l e i p y k n o t i c . The re were a r e a s

i n which i s l a n r k of f r a g m e n t i n g a c i n a r t i s s u e were s u r r o u n d e d

by hcmcrrhage (F ig . 2 2 ) . Thcrc were a r e a s of ghost l o b u l e s which

c o n s i s t e d of c o n n e c t l v c t i s s u c s t r n m a s u r r o u n d i n g very scanty d-ebris .

Bands of f i b r o u s c o n n e c t i v e tissue were s e e n between t h e a c i n i

f crrning i n t r a l o b u l a r f i b r o s i s . There w s s a n i n c r e a s e d amount o f

f i b r o u s t i s s u e betwecn t h e l o b u l e s f orrninr i n t c r l a b u l a r f i b r o s i s .

T h o walls o f t h e p a n c r e a t i c 6 u c t s were t h i c k e n e d by f i b r o s i s .

Some i s l e t s of Lnngerhans had a few f i b r o b h s t s i n d i c a t i n g the

b o g i n n i n g tif f i b r o s i s , o t h c r s were c o m p l e t e l y f i b r o t i c w i t h

f i b r c ! b l a s t c a b s e n t (Fig. 23).

The p o u ~ of a n i m a l s f e d t h e r i c e + c y a n i d e d i e t showed

n c c s o s i s , f i b r o s i s and a t r o p h y o f t h z paancreas, A s s e e n i n t h e

nn ime l s foe! t he g . n i d i e t , t h e edges of t h e s e c t i o n s o f t h e o rgan

were i r r e c u l a r as a r e s u l t of f i b r o s i s and a t rophy . There was

some f r a g n z n t a t i o n of a c i n a r t i s s u e , bu t f r a g m e n t a t i o n was n o t a s

prcnouncecl 2s in the ,mimsls fed t h e g n r i d i e t . F i b r o s i s fo l lowed

t h e samc p a t t e r n ,IS t h a t of t h e anirnal s f o d t h e gari d i e t bu t was

much more j:.ronounced (Figs. 24 and 25).

74,

F i g u r e 21. Pancreas of animal on gari d i e t . Necrosis and

f r a g m e n t a t i o n o f a c i n a r t i s s u e .

x 128.

F i g u r e 22. Pancreas of animal on gari d i e t . Hemorrhage.

x 128.

75 0

Figure 23. Pancreas of animal on gari diet, F i b r o s i s of i s l e t

of Langerhans .

Figure ?4, Pancreas of animal on rice + c y a n i d e dieto

F i b r o s i s in a c i n a r tissue,

76 0

Figure 25. Pancreas of animal on r i c e + cyanide d i e t .

F i b r o s i s cf islet of Langerhanc,

x 728.

77 0

I C i clney

I n t h e animals f 2 d tho gsri c ? i c t , thy-. e p i t h e l i a l c e l l s of

t h c proximal tuhuli :s hnd r u p t u r e d r e s u l t i n c i n stranc7.s 0::

cyto$o.sn push ing out a c r o s s thc; lumen of t h e tubules f r o m b o t h

s i d e s (Fi;;, 26). Thc s e l l s i n t hc se an ima l s h a d Seccme so swol len

that t h e y burL&. Thcr.2 vdas s e v e r e 2esqunrnation o f t h c c u l l s of t h e

t u b u l a r e p i t h c l i u r n and c a s t s o I n the group of a n i m a l s f e d rice

t. cyan ide , desquam>ti.on of the c e l l s c j f t h e t u b u l a r ep i t l l c l i um

w n s as marked as i n t h e animals f c d t h c ga r i . ditto Casts were

presen t i n t h e l u m i n a of t h e t u b u l e s (Fig. 2 7 ) .

e p i t h e l i a l c c l l s push ing o u t across t h e lumen of

proxirnzl t u b u l e s .

x 128,

Figu re 27. Kidney of an ima l on r i c e + cyanide diet. Casts i n

lurnina of p- oxirnal t u b u l e s ,

. *

DISCUSSION

R e s u l t s from t h i s s t u d y s u p p o r t t h e f i n d i n g s of p r e v i o u s

worke r s t h a t cynnic:e c o n t a i n i n g d i e t s have some p h y s i o l o g i c a l ,

m e t a b o l i c and p a t h o l o ( ; i ~ a l e f f e c t s i n t h e dog as i n o t h e r a n i m a l s

and man (Ekpech i & &. , 1966; Geevarghese , 1968; Osuntokun, 1970;

Mancr and Gomez, 1973; Del-ange, 1974; M i l l e r -- e t E X . , 19643. These

e f f e c t s were s e e n even when t h e cyani6e c o n t a i n i n g f o o d s w e r e f e d

as ba lanced d i z t s . The e f f e c t s producec'; by ~ a r i d i d not apy,t?ar t o

r e q u i r e p r c - i n g e s t i o n h y c ? r o l y s i s o f l i n a r n a r i a t o g i v e HGN o r t h e

s i m u l t aneo-us p r e s e n c e of a c t i v e l i n a m a r a s e .

I n t h i s s t u d y t h e r e were some d i f f e r e n c e s i n s p e c i f i c

p a r a m e t e r s n f f ec t ecl by t h e t v o cyaniclc c ontc?.ining d i e t s + and

also i n t h e d i r e c t i o n and i n t e n s i t y of t h e effects produced.

Scme of t h z s e d i f f e r e n c e s i n e f f e c t may have been i n f l u e n c e d by

t h e f a c t t h a t t h e g a r i d i e t c o n t a i n u d a lmos t twice as much an ima l

p r o t e i n a s t h e r i c e d i e t i n o r d c r t o o b t a i n a 13 p e r c e n t p r o t e i n

d i e t . The d i e t was b a l a n c e d f o r c r u d e p r o t e i n not f o r a r ~ i n o acids .

The c r u d e ? r o t e i n i n g a r i is n e @ i g i b l e compared t o r i c e b e i n g

1.6 g/100 g i n g a r i an? 7.5 g/100 g i n r i c e (FAO, 1970). P r o t e i n s

2 e r i v c d f r o m g r a i n a r e deficient i n some amino a c i d s e s s e n t i a l t o

a n i m a l s , and i f 2 p a r t i c u l a r e s s e n t i a l amino a c i d i s low i n

c o n c 2 n t r a t i o c , t h e o t h c r s become unusable a s w e l l because cells

s y n t h k s i z e whole p r o t e i n s o r none a t 211 (Guy tcn , 1981). Adarns

e t al, (1958) r ~ p o r t e d t h a t ui l ly t h e a l - ld i t ion of a comple t e -- p r o t e i n c o n t a i n i n s a l l t h e e s s e n t i a l amino a c i d s i n p r o p e r

p r o p o r t i o n was e f f e c t i v e i n r e v e r s i n g and p r e v e n t i n s t h e

h i s t o l o g i c a l c h a n g e s i n d u c e d by g x i .

Another f a c t o r i n f l u e n c i n g d i f f e r e n c e s i n m e t a b o l i c e f f e c t

is t h e chemica l n s t u r e of l i n n m a r i n . L inamar in is a glucoside and

e x p r e s s e s some of t h c c h a r a c t e r i s t i c f ea tu re s of g l u c o s i c ~ e s .

Johnson and Hzyncnd (1965) i n d i c a t e d t h a t ~ n r i consisted of

20% smylnse 2nd 70% arnylopec t in . C o n c l u s i o n s of t h e 1977 Int e r n n t i n n a l

Development Research C e n t r e c o n f e r e n c e on "Cassava as Animal Feed"

were t h a t cassava s t a r c h is l e s s r e a d i l y d i g e s t e d by rnonogas t r i c s

beczuse i t is h i g h e r i n m y l c p e c t i n and lower i n amylose.

W i c d and P h i l l i p s (1978) r e p o r t e d t h a t t h e d i g e s t i ' 3 i l i . t i e s

of r i c e and c a s s a v c d i e t s fed t o young N i g e r i a n men were t h e same

a l t h o u g h t h e t c t a l cruc:e f i b r e c o x t e n t of t h e r i c e d i e t was lower

t h a n t h n t of t h e c a s s a v a d i e t . T h i s p r e s e n t stur 'y shows t h n t i n

dogs, t h e d i ~ e s t i b i l i t i e s of r i c e and c a s 5 a v e d i e t s were t h e sameo

Th i s f i n d i n g s u p y o r t s t h e work of Wicol and P h i l l i p s ( 1978)

a l t h o u ~ h i t is at v a r i a n c e w i t h t h e I n t e r n z t i o n a l Development

Resea rch C ~ n t r c r e p o r t , It is sug[ystec! t h z t d i g e s t i b i l i t y p l a y e d

no r c l e i n t h e d i f f e r e n c e s obscrved i n t h i s s t u d y ,

The r e s u l t s of t h i s s t u d y show t h a t t h e i n s e s t i o n of e i t h e r

i n o r g a n i c cyani r ' e o r cy ,mcgen ic c l u c o s i i k w i l l e l e v a t e p l a sm3 and

u r i n e t h i o c y a n a t e c o n c e n t r a t i c n s a116 t h n t t h e r e is z c l e a r d i f f e r e n c e

i n t h e p a t t e r n s shown by t h e t u o forms of cyanide . S t b a (1957)

s t a t e s t h z t a c l e c r r i s e i n serum t h i o c y a n e t e a f t e r t h e i n g e s t i o n

of organic cyanog,nic c ompounCs is gocd e v i l e n c e t h n t some

d e g r a d a t i o n t o HCPJ must have t a k e n p l a c e . Maduagwu and Umoh (1986)

r e p o r t e d t h a t t h e l i v e r c o n t a i n s enzymes czpable of h y d r o l y z i n g

l i n o r n a r i n t o a m e t a b o l i t e which call d i s ~ o c i , ~ t e t o r e l e a s e c y a n i d e

i o n . T h i s m e t n b o l i t e is a c e t o n e cy2nchydr in ( 2-hydroxyisobutyro-

n i t r i l e ) .

Urnoh e t al. (1985) r e p o r t e d t h a t when a s i n g l e dose of -- l i n a r n a r i n w2s f e d t o g r o u p s of rats of g raded t h i a m i n s t z t u s , t h e

l e a s t amount of t h i o c y a n a t e w a s e x c r e t e d i n t h e u r i n e of a n i m a l s

r e c e i v i n g t h e h i g h e s t amount of t h i a m i n e i n t h a ? i e t , b u t t h e amount

of unrne taba l ized l i n a r r n a r i n was similar t o t h e c o n t r o l , This group

a l s o e x c r e t e d s i g n i f i c a n t l y h i g h e r f r e e cynnic3,0 i n t h e u r i n e compared

t o t h e c o n t r v l i n t h e f i r s t 24 hours of i n g e s t i o n , Umoh -- e t a l .

(7986) a l s o r e p o r t e d t h a t i n y % r c t e i n deficient r a t s bo th t h e r a t e

of metabol i sm of c y a n i e e t o t h i o c y a n a t e and a lsc t h e e v e n t u n l

c l e a r a n c e of t h i ~ c y a n a t e l:Iere p r c l o n g e 2 when a s i n g l e dose of

l i n a m a r i n was g i v e n o r a l l y . The s i p i f i c a n t l y h i g h e r p lasma and

u r i n a r y t h i g c y a n z t e of r i c e -i- c y a n i d e a n i m a l s compared t o g a r i

animals coulrl e v i d e n t l y b e r e l a t e c t t o a t l e a s t t h r e e f a c t o r s ,

F i r s t l y , the q u n n t i t y cf t h i a m i n e a n ? animal p r o t e i n i n cach d i e t :

t h e g a r i d i ~ t had t w i c e as much po rk a s t h e r i c e rf ic t i n orcler t o

bring t h e two d i e t s t o t h e same l e v e l o f p r c t e i n , a n d , a l s o , p o r k

has a v s r y h i g h l c v c l cf t h i m i n ( ~ a i n e s , 1968). Scccnd ly

t h e amount o f l i n a m a r i n m e t a b o l i z e d t o f r s e cyan iZe e i t h e r d u r i n g

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

of f r e e cyani1:e t o t h i o c y a n a t e mzy h s v e been d i f f e r e n t - T h i r d l y ,

t h ~ a b i l i t y of t h e !:idney t o r e a b s ~ r b t h i o c y s n a t e r e s u l t i n g i n

.2ccurnulation cf t h i o c g a m t e i n t h e plssmn ( ~ u l l r n a n & e,, 1954):

t h e swel l ink ; zn? r u p t u r i n r of t h e t u b u l a r e p i t h e l i a l ce l l f3 of t h e

kidney i n t h e g ? r i ~ n i r a n l s may have i m p a i r e d t h e r e a b s o r p t i o n of

t h c t h i o c y n n a t e inn.

Hich l e v e l s of plasma t h i m y a n 2 t e e r e knor~n t o c a u s e dec reased

p r o d u c t i o n o f t h y r ~ i d hor~ficnes. MacDcnnld (1980) s t a t e s that t h e

main f u n c t t o n cf t h i t h y r o i d g l and i s t o accumula t e i o d i n e and

cause i t s 3.ttachment t o t y r o s i n e , a n aminc a c i d , i n c r d e r t o form

t h e t h y r o i d hormone and t h e t t h i o c y a n a t e f u n c t i o n s by i n h i b i t i n g

i o d i n d t r a p p i n g by t h e t h y r o i d g l and , Delenge -- e t a l a (1979) showed

a c t i v i t y of t h i o c y a n s t e d e r i v e d f rom l i n e m m i n . When enough

t h y r o i d hormone i s not yro~:uccd t o meet t h e needs o f t h e a n i m a l ,

t h a t h y r o i l g l a n d uniergocs a compensa tary h y p e r t r o ~ h y i n an e f f o r t

t o xxoduce i n c r e a s e d s u r f a c e area t o t r a p t h e i o d i n e needed

(MacDonald ,1980)~ I n t h i s s t u d y t h e t h y r c i d g l and became e n l a r g e d

The l e v e l cf t o t a l sepum 3,5 + 3 ' - t r i i o d o t h y r o n i n e (T j p roduced 3

by t h e r i c e + cyzni?.c a n i n a l s was s i p i f i c a n t l y l o w e r t h m Ghat

produced by t h e p r i ~ n r l t h e ccnt ro l . an ima l s . The mean weight of

t h e t h y r o i d g l a n d s of t h e r i c e + cyznir le a n i m z l s was s i c n i f i c a n t l y

h ighe r t h a n t he mean w e i g h t s of t h e t h y r o i d g l n n d s of t h e gari and

c o n t r o l an ima l s , This is because enlnrgemcnt of t h i t h y r o i d g l a n d s

of t h e r i c e + c y a n i d e a n i m a l s w a s clue c h i e f l y t o ?n i n c r e a s e i n

t h e number ,an? size cf t h e e -p i the l . ia1 c e l l s of t h e f o l l i c l e s , that

i s , t h e parenchyma of t h e gl2n?., c a u s i n g parenchymatous g o i t r e ,

C o l l o i d ~ o i t r c and parenchymztous r o i t r e a r e t h ~ most commr,n t y p e s

of g o i t r e i n doc;s ( N i l l e r et &, , 7964) Thc r i c e + cyani ise group

of a n i m n l s had ,z plasma t h i o c y a n a t e l e v e l t h a t was s i ~ ; n i f i c a n t l y

hither t h m the ~ l a s r n a t h i o c y a n 2 t e l c v e l o f e i t h e r thz r i c e o r g a r i

graup of a-lirnals. Stba ('7957) reyiortcd t h - t 1,lasrna l e v e l s ::.f

t h i o c y a n a t c v n r i a d wi th t h e 6egree of h y d r a t i o n of t h e s u b j e c t .

Turner a n d Bagnara (7971) s t a t e d t h a t i n h y p o t h y r c i d i s ~ , b lood

volume is c o n s i d e r a b l y r educed , T h i s may p a r t i a l l y ex l : , l a in t h e v e r y

h i g h v a l u e s of ~ l a s m a t h i ~ c y ~ n a t e i n t h e r i c e + c y a n i d e an ime l s .

T r i t a l serum 3,5,3 ' - t r i i o d a t h y r o n i n c (T3) is produced at two l e v e l s

i n t h e bod)- - a t t h ~ t h y r f j i d elanrl l c v e l , and a t t h e p e r i p h e r a l

l e v e l by c c n v o r s i o n o f j,4,31,5t-tetraiodothyronine ( t h g r o n i n c T ) 4

t o 3,5 , j ' - t r i i o d c t h y r o n i n o (T3) i n t h c k idney mi1 l i v e r . Over 90%

c f t h e t h y r o i d hormcnc i s r e l e a s e d from t h e t h y r o i d g13nd a s

3,4,3',5'-tetraiod~th~ronine nn? l e s s t h a n 10% as 3 , 5 , 3 ' - t r i i o d o t h y -

r G n i n o . Therefore, any f a c t o r i n h i . ' l \ i t i n r t h e general yroc iuc t ion of

t h y r o i d hormones w i l l r e s u l t i n clepresseil 1.roc'uc t i o n of 3 , : , 3 I -

Growth. is i n f l u e n c e ? : by 3 , 5 , 3 ' - t r i i o d o t h y r o n i n e (T ) and growth 3

is no t depressed u c t i l t o t a l serum T c o n c e n t r a t i a n s declines 3

T s t i m u l a t e s s y n t h e s i s of messenger R N A , ) ? r o t e i n syn th .2s i s and 3

phosph i>ry la t ion .

T h i s study snowuc" t h a t growth r a t e , exi)ressec! as we igh t g a i n ,

of t h e g a r i a n i m d s l.-i;~~s s u p e r i e r t o t h s t : f t h e con t r c l l cmimals.

The e f f e c t was no t s ta t i s t i c a l l y s i g n i f i c a n t an<? was c v i c t e n t l y due

t o t h e h i r h e r t o t a l scrurr. 3,s ,j ' - t r i i o d o t h y x o n i n e (T,) c o n c e n t r a t i o n J

ccuyle!l w i t h t h e amount of ?,nirml ~ r o t e i n i n thc gnri d i e t compared

t o t h e t o t a l se rum 3,5,3 ' - t r i i oC .o thy ron ine (Tj) c o n c e n t r a t i o n of

t h e c o n t r o ? a n i m a l s and t h ~ ; j > r o t s i n c f t h e r i c e d i e t . Growth r z t e

of t h e n n i r m l s on t h e r i c e + c g a n i d ~ d i e t was s i g n i f i c a n t l y depressed

t o t h e c x t c n t of 33.83% by t h e 1 4 t h week when compared v i t h t h e

c c n t r o l m i m d s . T h i s may have o c c u r r e d becnuse t h e e s s e n t i a l d i e t m y

:imino ncic3s meant for grcwth were b e i n g used t o c7et:lxify t h e f r c e

cyaniOe t o t h i o c y n n a t e (F ' iect ler and Wood, 1956; Wood an? Cooley ,

- 1 9 5 6 ) ~ Osuntckun and Aladc t ~ y i n b ? (7970) reyor tec l t h a t *among p a t i e n t s

w i t h v c r y h i g h p l ~ s m a and. u r i n a r y t h i o c y a n a t e , m e t h i o n i n e , c y s t e i n e

and c y s t i n c were a b s e n t i n plasma of 60% of t h e inc i iv ic?ua ls and t h e

c o n c c n t r a t i c n s g r e q t l y r educcd i n o t h ~ r s and t h a t t h e q u e l i t a t i v e

anr? quan t i t - ~ t i v e changes of plssrna aminc 2c i d s were u n l i k e t h o s e found

i n p o t e i n - c a l c r i e c"cf i c i e n c y . Lelange e t al. ( 1982) s h o w d t h a t -- pups of hypothyrcl id r a t s s h o w ? liy;othyroic!ism and s l c w i n ~ o f

cellular growth duc t o s i i ; , n i f i c a n t l y h i g h serum t h i o c y a n a t e

c o n c e n t r a t i o n . The f i n d i n c s of t h i s p r e s e n t s t u d y , a r e t h e r a f o r e

s u p p o r t e d by t h e f i n c i n ~ s of rrepious worke r s and c o r r e s p o n d w i t h -- . . . t h e finclin[;s cf Maner and Gonez (1973) t h a t t h e u t i l i z a t i o n of

h i c h q u a l i t y p r o t s i r , w i l l ovcrcamc t h e d e p r e s s i o n i n growth c a u s e d

by t h e u s e of h i g h l e v e l s of c a s s a v a o r c y a n i d e i n diets.

I n t h i s s t u d y t h e u n c : : l c i f i ~ ( ~ , e p i p h y s e a l growth p l a t e of t h ?

z n i n n l s on t h e g a r i d i e t b e c m e c a 3 . c i f i ~ d o r c l o s e d a t a s l i g h t l y

f a s t e r r a t e t h a n th,ct of t h e c c n t r o l , r i c e f c ? an ima l s . The e f f e c t

wns n o t s t r t t i s t i c a l l y s i r ; n i f i c n n t and was av ic l en t ly due t o p r o t e i n

and t o t a l serum 3 3 , 3 ' - t r l i odo thy rcen i .ne (T ) c o n c e n t r a t ion. 3 By t h s enct of t h c e x p e r i m e n t , c , d c i f i c a t i o n :f t h e e p i p h y s e a l growth

p l a t e of t h e r i c e + cyanic?e f e d znirnals had n o t occurrecl* The

e f f e c t was s t a t i s t i c a l l y s i g n i f i c a n t . The n o n - c a l c i f i c a t i . o n o f t h e

e p i p h y s e a l growth p l a t e was r e l - t e d t o t h e hij;h l e v e l of ~ ~ R E I ~ X

t h i o c g a n ~ t e which incuced hypo thy ro id i sm and low l e v e l s of t o t a l

se rum 3 ,'j , 3 ' - t r i i o c ' o t h y r o n i n e c o n c e n t r n t i o n wi th r e s u l t in@; s l o w

c e l l u l a r growthl decreased o s t e n b i a s t i c z c t i v i t y i n t h e p r imary

s ~ : o n g i o s a , i m p a i r e d c h o n d r o i t i n s u l p h a t e s y n t h e s i s and sycthesis

of c o l l a s e n of bone ( ~ n g e & &.., 1968; Z o b r i s k y , 1969).

Cyanide i n r e s t i o n has been shown t n a f f e c t t h e metabol i sm

af r i b o f l a v i n ( v i t a m i n B ) an" v i t a m i n BIZ' 2 S p e c t o r e t ale (1943) --

demons t r a t e$ t h a t e x p e r i m e n t a l r i b o f l a v i n d e f i c i e n c y i n clogs l e d t o

a m i c r a c y t i c hypnchromic anemia. I n v i t a m i n B 2 e f i c i e n c y 12

m z c r o c y t i c e r y t h r o c y t e s a r e l.,roducecl and th:. t o t a l number of

e r y t h r o c y t e s r e s u l t i n g is markedly reducec!, Osuntokun (1973)

rel :or ter! low :-lasrna l e v e l s of r i b o f l a v i n a n d l o v u r i n a r y exc re t i on

of r i b o f l a v i n i n r a t i c n t s w i t h t r c r i c a l a t a x i c neuroyathy .

Edozien (1965) and U.S. Repor t (1967) i n d i c a t e 2 t h a t riboflavin

d e f i c i e n c y is commrn i n most s c u t h e r n Y i g e r i a n s . Braekhan e t al.

( 1957) dernonstrntecl t h a t c h r o n i c a 3 m i n i s t r n t i o n of cynni.de t o a n i m a l s

d e p l e t e s l i v e r Bq2. Goodman and Gilman (1970) l is ts anemia as one

of t h e t o x i c e f f e c t s of t h i o c y a n a t e . Schalm et &. (1975) s t a t e d

t h a t hypothyrcic!ism is c f - t e n a s s x i a t e : : w i t h a bo r t7e r l i ne no rmocy t i c

normochronic anemia p r o b a b l y because t h y r o x i n e is one of t h e hormones

invc: lved i n t h e c o n t r o l ~ f e r y t h r o g e n e s i s , T h i s s t u d y i n d i c a t e d that

t h e r e was n o anernin i n t h e a n i m a l s f e d e i t h e r t h e gari o r rice +

cyanide d i e t s o The blood hemoglobin values and t h e packed c e l l

volumes dic: n o t v a r y s i g n i f i c a n t l y frm t h a t of t h e c o n t r o l . Th i s

s t u d y a l s o show t h n t c y a n i c e o r t h i u c y a n a t e a t t h e c o n c e n t r a t i o n s

i n t h e ~ l a ~ m a d i d n o t a p p e a r t o i n t e r f e r e w i t h h e m ~ g l c h i n c o n t e n t

o r have any t o x i c a c t i o n on t h e e r y t h r o c y t e . The r e a s o n s a r e not

obv ious b u t i t may be t h n t t h e d i e t s a p p a r e n t l y s u p ~ l i e d enough r i b o -

f l a v i n 2nd v i tnrn in BI2 t c : overcome t h e a c t i v i t y o f any s u b s t a n c e

r e s ~ o n s i h l e f o r d e p l e t i n g s t o r e s cr a c t i n g as a n a n t a g o n i s t .

Bowever, t h e appa ren t l y normal hemoglobin c o n t e n t and packed cell

volume of t h e a n i m a l s f e d t h e r i c e + c y a n i d e d i e t c o u l d be c o n s i d e r e d

t o be c!ue t o h e m o c m c e n t r a t i o n . I n h y p o t h y r o i d a n i m a l s , b l o o d

volume i s c o n s i d e r a b l y r educed ( T u r n e r an2 Bagnara , 1971) -

Malharbc e t al. (1977) r e p o r t e d t h a t a n e l e v a t i o n of [Tarnma

g lu tamyl t r n n s f e r a s e a c t i v i t y in serum is t h e most s e n s i t i v e and

s p e c i f i c i n d i c a t o r of l i v e r damage of a l l t e s t s up t o the p resen t .

" ( 0

C o n d i t i o n s i n v o l v i n ~ i n t r a - or e x t r a - h e p t i c c h o l e s t n s i s g ive t h e

h i g h e s t values, w h i l e , f o r examl~ le n e c r o s i s of t h e l i v e r C e l l s due

t o i n t c x i c a t i a n w i t h c s r b o n t e t r n c h l a r i d e o r a c u t e v i r a l h e p a t it i s

p roduces less e l e v a t i o n of gamm~ g lu t amyl t r n n s f e r a s e t h a n an

enzyme such as alanine t r m s a m i n a s e . Gamma g l u t a m y l t r a n s f e r a s e

is more u s e f u l t h a n a l k a l i n e p h o s ~ h n t n s e i n t h e d i f f e r e n t a l

d i a g n o s i s o f c l i n i c a l jaunLicc- because i t is more s p e c i f i c . Gamma

g lu t amyl t r a n s f e r a s e is unaf f e c t er! by preg:anancy, bone disease a n d

i m m a t u r i t y , b u t i s c o n s t a n t l y e l e v a t e 2 i n a l l c h r o n i c l i v e r

c o n C i t i o n s an2 may p e r s i s t f o r many weeks i n c i r r h o s i s . Ir, general,

e x c e ~ t i n e a r l y a c u t e h e p a t i t is, gamma g lu t amyl t r a n s f e r a s e r i s e s

h i g h e r and s t a y s e l e v a t e d l o n g e r t h z n o t h e r erizyme a c t i v i t i e s .

Acute and c h r o n i c p a n c r e n t i c d i sease a l s o nkfec t s serum gamma

glutamyl t r m s f e r e s e l e v e l s .

I n this s t u d y the a c t i v i t y of plasma gamma g l u t a m y l

t r a n s f e r a s e of t h e an imals f e d t h e g n i 6 i e t and t h o s e f e d t h e

r i c e + c y a n i d s d i e t were no t s i g n i f i c a n t l y d i f f e r e n t from the

a c t i v i t y of t h e a n i m a l s f e d t h c c o n t r c l c ' i e te There was no

s i g n i f i c a n t d i f f e r e n c e i n t h e p lasma a c t i v i t i e s o f i s o c i t r i c

dehydrogcnaso anc! a l n n i n ~ t r a n s f e rase between t h e t h r e e groups,

The s i g n i f i c m t d i f f e r e n c e shown i n t h e plasma a c t i v i t y of the a l k a l i n e

phospha tnsc af t h e r i c e + c y a n i d e group was r e l a t e ? . t o t h e r e d u c e d

o s t e o b l n s t i c a c t i v i t y r e s u l t i n g from r e t a r d e ( ' ~ r o w t h and c z l c i f i c a t i c n

of the bones and no t t o h e p a t i c f u n c t i o n (Sage e t ol., 1968).

88.

Osuntckun (1968, 1969) r epo r t ed t k t l i v e r c i r r h o s i s d i d n o t

c c c u r i n p a t i m t s w i t h t r o ~ i c a l a t a x i c neuropathy . Goodman and

Gilmnn (1970) r e r o r t e d h e r , a t i c n e c r o s i s as one of t h e r e ? c t i o n s t o

i n c r e a s e d t h i o c y a n n t e i n t h e blood. Adnms et &. (1958) r e p o r t e d

p e r i p o r t a l f a t t y changes i n t h e l i v o r of n i c e sn? ra t s f e d a gari

d i e t c o n t a i n i n g 1% p r o t e i n on 3 2ry m a t t e r Sasis, f o r 4 weeks.

Ononoghu and Ernole (1978) fount?: v a c u 0 1 3 t i ~ n i n t h e l i v e r of r a t s

f ed ga r i f o r 4 weeks. Ths h i s t o p ~ t h c l o g i c d f i n c ' i n i ; ~ i n t h i s stucly

were p e r i ~ o r t a l hyclropic v n c u c l a t i o n i n t h e a n i m a l s f e d t h e gari d i e t .

The h i s t o p a t h o l o g i c a l f i n d i n g s i n t h e l i v e r seen i n t h i s study are n o t

s u p p o r t e d by t he f i n 6 i n g s of Adarns &. (1958) but are i n agreement

w i t h t h e f i n d i n p of O n o n o ~ b u and Emole (1978).

Lipasc (EC 3.1.1,3, t r i a c y l glycel-01 a c y l h y d r o l a s e ) is

secre ted by t h e pancreas. Plasma a c t i v i t y of l i l j a s e i n t h e a n i h d s

fed gari wns s i g n i f i c a n t l y h i g h e r t h a n plasma a c t i v i t i e s of l i p a s e

i n t h e a n i m a l s f e d t h e r i c e + cyan ide d i e t and t h e c o n t r o l d i e t ,

Davies (1948; 1952) s u g g e s t e d t h n t i n humans on a c a s s a v a d i e t and

suffering from k w a s h i o r k o r , t h e p a n c r e z s i s a f f e c t e Z b e f o r e t h e

l i v e r . He found t h n t p a n c r e a t i c a t r o p h y o c c u r r e d b e f o r e t h e o n s e t

of f a t t y l i v e r .

Geevsrghese ( 1982) r e p o r t e d t h a t i n r e c e n t y e a r s , e v i d e n c e

h a s accumulatec! t o show cassava b a s e d G i e t s a r e r e l a t e d t o t h e

development of t r o p i c a l c a l c i f y i n g p a n c r e s t i t is and d i a b e t e ~ m e l l i t u s .

oy.

H i s t o p a t h c l o g i c a i l y , t h e r e was d i l ~ . t i o n of p n c r e a t i c d u c t s

c o n t a i n i n g p r o t e i n a c e o u a c a l c i f i e d m a t e r i a l , acinar a t r o ~ h y and

i s l e t c e l l d c s t r u c t ion . He r e ~ o r t e d t h a t e p i d e m i o l c g i c a l d a t a i n

s u p p o r t of Gassave as a oause of t r o p i c a l c a l c i f y i n f ; p a n c r e a t i t i s

show t h a t t h i s d i s e a s e i s most prevalent i n countries where c a s s a v a

i s e a t e n and t h c i n c i d e n c e <)f endemic g o i t r e r e l a t e 6 t o cassava

t o x i c i t y is a l s o comr.on. Exzmples are RTigcrin ( O s m t o k u n 1970);

t1~nnd.a (Shape r 1964) , I n d i a (Gecvarghese 1968) , I n d o n e s i a (Zuidema

' l g r g ) and B r a z i l (Dani and Noguei ra 1976). Davic?son e t al. (1969)

showed t h a t d i a b e t e s m e l l i t u s i s mcrc common i n p e o p l e e a t i n g

cassava-basc-d d i e t s . Davidson r e p o r t e d a 1% i n c i d e n c e of d i a b e t e s

m e l l i t u s ih i n h a b i t a n t s of Knlenc P i l l , Zambia , where cassava made

up 93% of t h e 2a i ly c a l o r i c i n t a k e whereas i n Malawi and Zimbabwe

population s u r v e y s have shown t h e i n c i Z e n c e of d i a b e t e s t o be o n l y

0-1-0013$+

The re i s e x p e r i m e n t a l e v i d e n c e i n s u p p o r t of cassava-based

d i e t s as a c a u s e of t r o p i c a l c a l c i f y i n g p a n c r e a t i t i s . Fushpa

(1980) found p a n c r e a t i c changes similar t o t h o s e seen i n t r o p i c a l

c a l c i f y i n g p a n c r e a t i t i s i n rats f e d on a d i e t of 22.8 g of cassava

c o n t a i n i n g 73 mg/g c y a n i d e d a i l y f o r -18 months.

The s t a t i s t i c a l l y s i g n i f i c a n t h i g h plasma i i p a s e a c t i v i t y i n

t h e gari group o f animals i n t h i s s t u k y when compared wi th the r i c e

and rice + cyan ide groups may i n d i c a t e t h a t t h e cyanogenic g l u c n s i d e

cf gari i n d u c e s p a n c r e a t i c i n j u r y even in a b a l a n c e d d i e t , c a u s i n g

90.

1arli;i: q u s l n t i t i e s of' l i p a s e t o l e a k o u t of t h e c e l l s o r a c i n i o r

d u c t s . This was conf irrned by t h o h i s t o p a t h o l o g i c p i c t u r e . The

F a n c r e a s of t h e dogs f e d t h e g a r i d i e t showed n e c r o s i s , €&ma,

hemorrhage , f i b r o g i s , a c i n a r a t r c ~ h y , d e s t r u c t i m of i s l e t cells.

It appears t h a t t h e e f f e c t on t h e p a n c r e a s was m i l d a n d s l o w l y

p r o p e s s i v c r e s u l t i n g i n a c t i v e d e s t r u c t i o n of p m c r e s t i c t i s s u e

and in t h e main tenance of a r e l n t i v e l y h i g h l i p a s e a c t i v i t y after

14 weeks. Plasma t r i a c y l g l g c e r o l lipase (EC 3 . I. 1.3) a c t i v i t y is

n s p e c i f i c p a r a m e t e r f o r t h e measurement of p c n c r e a t i c integrity,

S i m i l a r h i s t o p a t h o l o g i c l e s i o n s were obse rved i n t h e pancreas

of t h e a n i m a l s f e d r i c e + cyhnideo The finding of normal eerum

l i p a s e a c t i v i t y i n r i c e + cyanide mimals a f t e r 14 weeks on d i e t

i s n o t unusual . Stsombeck e t a l . (1984) found normal l i p a s e and -- gamma g l u t a m y l t r a n s f e r a s e a c t i v i t i e s i n dogs w i t h s e v e r e c h r o n i c

p a n c r e a t i c p a t h o l o ~ i c a l change&. It is p r o b a b l e t h a t t h e l i p s @

a c t i v i t y i n t h i s group had become e l e v a t e ? e a r l i e r in t h e

experiment and h a d r e t u r n e d t o a low l e v e l . T h i s p a t t e r n was seen

by Strornbcck I& &. (1984).

In b o t h t h e g a r i f e d a n d t h e r i c e + c y a n i d e f e d a n i r n d s t h e r e

was f i b r o s i s of the i s l e t of Lzngerhans. F i b r o s i s w a 6 more pronounced

i n t h e r i c e + c y a n i d e f e d an ima l s .

It appenrs t h a t t h e p a t h o l o g i c changes seen i n t h e p a n c r e a s i n

t h i s s t u d y were c a u s e d by t h e t h i o c y a n a t e i n t h e plasma a n d t h a t t h e

d i f f e r e n c e s seen between t h e t h r e e g roups were r e l a t e d t o the

plasma c o n c e n t r a t i o n s of t h l o c y a n a t e. T h i s f inGing is i n agreement

w i t h t h c work of HcMil lan and Gcevarghese (1979) who showed t h a t

t h e a e m i n i s t r a t i o n o f c y a n i d e (KCN) o r s l l y o r i n t r a p e r i t o n a a l l y t o

rats cnused a p e a k i n g of plasma g l u c o s e up t o 300 mg/dl w i t h i n

30 minutes. T h i s war assoc ;a ted w i t h temporary g l ~ c o s u r i a . The

f inc ' ings c f t h i s s t u d y a r e a l s o i n agreement with t h e work of

Fushpa (1980) an?. t h e work of Dav ie s (1948, 1952).

I n h e a l t h y a n i m a l s , z small amount of' p r o t e i n f i l t e r s i n t o

the g l o m e r u l a r f i l t r a t e e a c h day. T h i s rotei in is n o t e x c r e t e d i n

urine b u t r e tu rne i ! t o t h e body f l u i d s a f t e r i t h a s been a b s o r b e d

t h r o u g h t h e b r u s h b o r d e r o f t h e p rox ima l t u b u l a r e p i t h e l i u m by pino-

c y t o s i s (Guyton, 1981). The d e g e n e r a t i o n seen in t he proximal tubular

e p i t h e l i a l c e l l s i n t h e g s r i f e d a n i r m l s r e s u l t e d i n t o t n l loss

of f u n c t i o n cf t h e s e c e l l s , and t h e r r o t e i n of t h e glornerulnr f i l t ra te

was not absorbed bu t was observed t o be e x c r e t e d i n t h e t~r ine . The

great i n c r e a s e i n u r i n a r y p r o t e i n from week 1 t o week 5 seemed t o

i n d i c a t e t h n t t h e :!recess of d e g e n e r a t i o n became progressively more

severe w i t h t i m e u n t i l a l i m i t was r e a c h e d o Goodman and Gilman (1970)

r e p o r t e d t h n t t h i o c y a n a t e causes n e p h r o s i s . The l c s i o n s s e e n i n t h e

k i h e y of t h e r i c e + c y a n i c e a n i m a l s were d i f f e r e n t from t h o s e

seen i n t h o s n i m a l s f e z g a r i and t h e p r o t e i n e x c r e t e d was ~ i g n i f i c ~ a n t l y

much lcss and f o l l o w e d a d i f f e r e n t p a t t e r n from t h e a n i m a l s f e ?

gzri i n t i c x t i n g t h n t some f a c t o r i n a d d i t i o n t o t h e l e v e l of

t h i o c y a n a t c i n t h e b lood was r e s p o n s i b l e f o r t h e q u a n t i t i e s of

p r o t e i n i n the ur ine* Al though plasma t h i o c y a n a t e was s i g n i f i c a n t l y

h i g h e r i n t h e r i c e + c y a n i d e f ed an ima l s t h a n i n t h e ga r i f e d animals

( F i c,OI) t h a u r i n e p r o t e i n e x c r e t e d was s i p i f i c a n t l y l o w e r i r . t h e

r i c e + c y n n i d e f e d a n i m a l s t h a n i n t h e g a r i f e d a n i m a l s (P ~f 0.01).

I n t h e l i t e r s t u r e rcv iewed t h e r e was no i n f o r m a t i o n on p r o t e i n i n t h e

u r i n e i n r e l a . t i o n t o c a s s a v a d i e t s o r l i n a m a r i n .

Low p o t a s s i u m l e v e l s i n t h c c a r d i a c muscle h a v , p r e v i o u s l y been

assoc i a t e d w i t h l i n a m a r i n and c y a n i d e i n g e s t i on . P h - l b r i c k , H i l l and

Alexande r (-1977) r c p o r t e d t h a t t h e b l o o d t a k e n j u s t b e f o r e d e a t h

from rats closed o r a l l y w i t h a l e t h a l amount of l i n a m a r i n (500 mg/kg)

o r KCN ( 6 rng/kg) h a d i n c r e z s e d serum potz.ssium, *h;ch ~ y ~ g s t a t i s t i c a T 1 4 s i g n i f i c a n t o n l y i n t h e c y a n i d e dose2 group. Serum sccliun. c o n c e n t r g -

t i o n s remained u n a f f e c t e d , They a l s o r c ; ~ o r t e d t h a t c a r d i a c t i s s u e

showed a decrease i n p o t a s s i u m i o n c o n c e n t r a t i o n and an i n c r e a s e i n

sndium i o n c o n c e n t r a t i o n which w a s no t s t a t i s t i c a l l y s i g n i f i c a n t .

Decreased c a r d i a c p o t a s s i u m c o n c e n t r a t i o n s a n d h y p e r k d e m i a have been

r e p o r t e d i n cyanide-dosed c a t s by K a r z e l -- e t a l e (1974) . L inamar in

and c y a n i e e had p a r a l l e l e f f e c t s on t h e h e a r t . Large d o s e s of

d i g i t a l i s , a non-cyanogenic g l u c c s i d e , a r e known t o cause 2zrdiac

a r r h y t h m i a s (Von C a p e l l e r e t al., 1959) an2 a t r i a l f i b r i l 1 , i t i o n and

f l u t t e r (p i ck , 1957; Delman and S t e i n , 1964). However, thrr o n l y

enzymat ic sys t em i n t h e h e a r t known t o be a f f e c t e d by l i g i i a l i s is

t h e sodium-potassium a d e n o s i n e t r i p h o s p h a t a s e a s s o c i a t e d w i t h t h e

cat ion t r m s ~ o r t (Mat s u i and Schwar t z, 1 9 6 7 ) ~ It t h e r e f o r e a p p e a r s

93 0

t h a t t h e m e t a b o l i c e f f e c t s c a u s r d by t h e i n g e s t i o n of g a r i may

n o t bc e n t i r e l y due t o t h e r e l e a s e of cyan ide .

H i l l (1977) r e p o r t e d t h a t c a r d i a c sodium-potassium a d e n o s i n e

t r i p h o s p h a t a s c was a f f e c t e d by a 25 rng/100 g body weight dose of

l i n a m a r i n which was s u b l e t h a l , He a l s o r e p o r t e d t h a t i n general

a d e n o s i n e t r i p h o s p h a t nse a c t i v i t i e s were s i g n i f i c a n t l g r e d u c e d by

l i n a m a r i n i n e i c s t i n g t h a t l i n a m a r i n may e x e r t a t o x i c e f f e c t by a

mechanism ~ n d e p e n d e n t o f HCNo It seems t h e r e f o r e , t h , ? t r e p e a t e d

i n g e s t i o n of s u b l e t h a l d o s e s of g a r i c o u l d a f f e c t t h e ,sodium-

po ta s s ium ad-enosine t r i p h o s p h n t a s e th roughou t t h e body t h u s a l t e r i n g

t h e sodium-potassium t r m s p o r t sys tem i n such a way t h a t t h e t i s s u e s

become dep lc t c d of p o t a s s i u m r e s u l t i n g i n i t i a l l y i n a hgperka lern ic

c o n d i t i o n u n t i l a s i t u a t i o n is r eached when hypokalemia r e s u l t s and

c e l l s begir.. t o swell. T h i s is demons t r a t ed i n t h e l i v e r c e l l s

where v a c u o l e s are seen. The ~hcnomenon of s w e l l i n g o f k i 6 n e y

and l i v e r c z l l s i n tk? rat f e d a d i e t c o n t a i n i n g gnri hae been

r e p o r t e d by Ononogbu and Emole (1978). Smi th , J o n e s and Eunt (1972)

s t a t e d t h a t v a c u o l a r changes i n t h e ez . i t he l ium of t h e p rox ima l

t u b u l e s o f <he k i d n e y i s n c h a r a c t e r i s t i c of p o t a s s i u m def ic iency .

The animals f e d g a r i h n d n plasma po ta s s ium c o n c e n t r a t i o n that w a s

s i g n i f i c a n t i y l ower t h a n t h e plasma po ta s s ium c o n c i - n t r a t i o n o f t h e

c o n t r o l a n i m a l s ( P L Op05). The a n i m a l s f e d t h e r i c e + c y a n i d e d i e t

had a p l a s m po tns s ium c o n c e n t r a t i o n t h a t was l o w e r , b u t n o t

s i g n i f i c a n t l y s o , than t h e plasma p o t a s s i u m c o n c e n t r a t i o n of t h e

94.

a n i m a l s f e d t h e c o n t r o l list. T h i s f indin[; s u p p o r t s t h e work o f

F h i l b r i c k -- c t a1. (1977) who reportef i t h a t l i n z m a r i n and cyanide

had p a r a l l e l e f f e c t s i n decreasing t i s s u e po ta s s ium and that t h e

m e t a b o l i c e f f e c t from l i n s r s r i n i n g e s t i o n may n o t be c a u s e d e n t i r e l y

by t h e re lease of cysnj.de.

I n t he l i t e r a t u r e r ev i ewed no ment ion has been made of

hypocalcernin i n r e l a t i o n t o g a r i o r l i n a r n a r i n o ? i y ~ ~ o c a l c e m i n i s known

t u occur i n n e ~ h r o s i s 2us to l o s s of p r o t e i n 2nd i n p a n c r e a t i t i s

due t o t h e f o r m a t i o n of ca l c ium sonrrs ( T i e t z , 1 9 8 2 ) ~

The r e s u l t s of t h i s stuc?y i n ~ ? i c a t e t h a t ~ l a s m a c a l c i u m

c o n c e n t r a t i o n w e r e s i g n i f i c a n t l y lower i n t h e group o f a n i m a l s fed

t h e ga r i C i a t compzred t o t h e r i c e + c y a n i d e a n d c c n t r o l group o f

a n i m a l s ( P L 0 ~ 0 5 ) ~ Biochemica l examinat i o n showed t h a t a s i g i f i c a n t l y

h i c h l e v e l of p r o t e i n was l o s t i n t h e u r i n e of t h i s g roup of an ima l s .

k ~ p r o x i m a t z l y 41% of p l ~ s m n ca l c ium is bound t o p r o t e i n and any

decrease i n serum ? r o t e i n s f r e q u e n t l y r e s u l t s i n a d e c r e a s e i n t h e

t o t a l serum cal.ciurr l e v e l . S i n c e t h i s d e c r e a s e a f f e c t s main ly t h e

n o n c l i f f u s i b l e f r a c t i o n of c a l c i u m , t e t a n y i n r a r e l y a s s o c i a t e d w i t h

t h i s form of c a l c i u m l o s s (Guyton, 1981). The hypoca lcemia i n t h i s

group of an ima l s may t h e r e f o r e t o b e r e l a t e d t o p r o t e i n l o s s by

n e p h r o s i s o

95

CONCLUSION

T h i s s t u d y showed t h a t f o r t h e d u r s t i o n of t h c experimental p e r i o d

of 14 weeks t h 2 r i c c d i e t produced no ? e t e c t a b l e plasma t h i o c y a n n t e

whereas b o t h t h e p r i d i e t and t h e r i c e + c y a n i d e d i e t p roduced

?lama t h i n c y n n a t e b u t t h e g n r i c?ie t 1 ro?.uced s i ~ n i f l c a n t l y l e s s

p lasma t h i o c y n n n t e t h m t h c r i c e + c y a n i d e e i e t . The h i g h e r leva1

i n t h e l n t t e r yroba.bly i n c l i c a t e s t h a t a g r e a t e r q u a n t i t y of f r e e

c y a n i d e w::s a v a i l a b l e f o r ccmvers ion t o t h i o c y a n a t e . The re was

no s i g n i f i c a n t d i f f e r e n c e i n t h e e f f e c t cf t h e r i c e d i e t , t h e gari

d i e t and t h e r i c e + c y a n i d e d i e t on t h e f o l l o w i n g p a r a m e t e r s :

d i g e s t i b i ! i t y , b lood hemoglobin c o n c e n t r a t i o n and b l c o d packed

c e l l volulne. The r i c c 6 i e t and t h e g a r i d i e t d i d n o t d i f f e r

s i ~ i f i c a n t l g i n t h e i r e f f e c t cn growth and t h e t h y r o i d g l a n d b u t

t h e r i c e + c y a n i d e d i e t s i g n i f i c a n t l y a f f e c t e d grcwth and t h e t h y r o i d

g land . The r i c e d i e t h a d no e f f e c t on t h e p n n c r c a s , b u t t h e g a r i d i e t

produced e f f e c t s on t h c p a n c r e a s t h a t were s imilar t o t h e r i c e +

c y a n i d e d i e t . S i m i k r l y , t h e r i c e d i e t ha6 no e f f e c t on t h e kidney,

however b o t h t h e ear i d i e t and t h e r i c e + c y a n i d e d i e t had ha rmfu l

e f f e c t s on t h e kidney. The re were no s i g n i f i c a n t d i f f e r e n c e s

between t h ? r i c e d i e t and t h e r i c e + c y a n i d e d i e t on t h e l i v e r ,

~ l a s m a po tns s ium and plasma ca l c ium l e v e l s b u t p lasma p o t a s s i u m

2nd plasma c a l c i u m were s i g n i f i c a n t l y l ower i n t h e animals on t h e

gari d i e t .

The r i c e + c y a n i d e d i e t was more t o x i c t h a n t h e g a r i d i e t even

though t h e l e v e l of c y a n i d e wa.s t h e same. Growth, t h c t h y r o i d

g land, the p a n c r e a s and t h e k idney were s f f e c t e d by t h e r i c e +

cyanide d i e t , T h c l i v e r , t he pancreas and the k i d n e y were a f f e c t e d

by t h e g a r i d i e t . Growth and t h e t h y r o i d g l a n d were n c t a f f e c t e d

by t h e g a r i d i e t . The deg ree of t o x i c c s i s t h e r e f o r e d i d n o t depend

on t h e l e v e l of c y a n i d e i n t h e d i e t p e r s e , bu t a p p e a r e d t o be

r e l a t e d to t h a l e v e l of f r e e cyan ide releascd by metabol ism. Bound

c y a n i d e a p p e a r e d t o e x e r t l i t t l e i n f l u e n c e . Thc hypcca lcemia i n t h e

animals f e d t h e g a r i d i e t wes seconda ry t o t h e l o s s o f r r o t e i n i n t o

t h e u r i n e . The v a c u o l a t i o n i n t h e l i v e r , t h e l o s s of p r o t e i n i n t o

t h e u r i n e ,znd the hypokalemia i n t h e an ima l s f e d t h e gar i d i e t

canno t be e x p l a i n e d on t h e b a s i s of c y a n i d e t o x i c c s i s . P r e v i o u s

worke r s have suggested t h a t l i n a m a r i n , t h e cyanogen ic g l u c o s i d e of

g a r i e x e r t s n t o x i c e f f e c t cn sc2 iu rn -~o tns s iu rn a d e n o s i n e t r i p h o s -

phntase a l t e r i n g t h e sodium-potassium t r a n s p o r t s y s t e m i n s u c h a way

t h a t t i s s u c s become d e p l e t e d of po tass ium.

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a n d Geogrnyhicd Medicine 11: 70-74.

APFENDIX I: THE EFFECT OF CYANIDE CONTAINING EIETS ON WEIGHT CHANGE

DUitING 14 WEEK EXPERIMENTAL PEHIOD

Percentage (%) increase i n weight (Mean - + s.E.M.) F e r i o d (weeks) D i e t a r y Treatment

Rice Gar i Rice + Cyanide

a'b~eans with t he same superscript in t he row f o r a s p e c i f i c week are nc,t s t a t i s t i c a l l y d i f f e r e n t (F 0.05).

AFTENDLX 2 : EFFECT OF CYILNIDE CONT:LINIMG DIETS ON PLASMA T CUMCEPiTRfLTIONS 3

Plasma T c m c e n t rat ion Dietary

Week Treatment (nZ/dl Mean 2 S.E.M.

0 Rice

Gari

Rice + Cyanide

3 Rice

Gari

Rice + Cyanide

R i c e

Gar i

Rice + Cyanide

14 Rice

Gari

Rice + Cyanide

40 70 60 70

70 70 90 60

40 60 40 70

50 80 70 80

70 76 80 80

40 50 30 80

60 80 80 go

60 250 90 90

30 40 60 80

80 80 PO 90

100 loo 100 go

30 20 40 30

a'b~eens wi-lih t h e same superscript in the columns f o r o s p e c i f i c week ere not statistically d i f f e r e n t (F 0.05).

APFENDIX 3 : EFFECT CF CYANIDE CONTf.IN I N G DIETS ON FLhSMA TRIOCYANATE CONCENTRfiTTON

Die ta ry Plasma Thiocyanate Concentration (u rnole/ml) Treatment Mean + S.E.1 -

Repl i ca te s

I Rice

G a r i

Rice + Cyanide

3 Rice

Gar i

Rice + Cyanide

14 R i c e

G a r i

R i ce + Cy,znide

a,b c ' Menns w i t h the same superscript in the column f o r a s p e c i f i c week are n o t s t a t i s t i c a l l y d i f f e r e n t (P 0.05).

0 0 - d W O ~ A 0 u l N 0 ~ N 0 \ D O . N - 4 .

0 0 . . o m . o f . 0 3 L % F &

- d O M N O A 2 O ~ N O A W O W N * 2 0 3 . u r n . N W .

g o * 3 m . 0 . * O < - 0 \D C n W 0 -r -I= 0 \O cn u 0

APPENDIX 6: EFFECT OF CYtJJIDE C O N T A I N I N G DIETS ON EPIFHYSEAL CLOSURE

Width of u n c a l c i f i e d Dietary epighyseal j l a t e (u) ,

Week Treatment Moan SoEmFI. Rep l i ca t e s

Rice

Gzr i

Rice 4 cyanide

Rice

Gnri

Rice + cyanide

Rice

Gar i

N i c e + cyanide

Rice

Gar i

Rice + cyanicla

a'b~sans with t h e same superscript in t he crlumn f o r e s p e c i f i c week are not statistically s i g n i f i c a n t ( F 0.05).