CHAPTER fI

O r g a n - i c constituents parti-

a carbohydrates, lipids and

proteins are of -dtmost importance

in. animal tissues, which for a

major source of energy b e s i d e s

being an integral component, in

structural frame work of animal

cells.

Carbohydrates, pro te ins and

lipids are sources of energy.

LLpids ape useful as direct and

stored energy, besides having

- . . - . other fy2-c:;z-s -:.sz Zcr- . - =-F--+-.- Cz , 3 p . ~ d . . A-,. . ?if f ~ r m t classes Of " 2 v ~ k e ~ k - - A * ~ - - - . - fsr: 7.2 jor source of ezergy and rr,aiinrfir.w sf ,:-.sse reser-.-e5 is an ixportant feature of

- - . - . c e l . Tke c l z r L c acid cycle acts as a final

C O ~ ~ ~ , ~ T v,=-q..-=:- - P P -'-D py i A - - - : ,, ,.,,,, -. Z,. ,,., ..-uS -LS~: of carbohydrates, lipids . .

and srstelrs , i s a's~dt the catabolism of acetyl

- . > . . residxes -~cers=:ns i ydrazsn eqiiivalents leading to the

release of ensrzv7 - - thrsugh $-msphorylation. Carbohydrate metaboliss is disturbed by the zction of toxic conpounds and

the cozpensatory shift frox aerobic to anaerobic segments is

an i~evita5le conseqcence in tissaes for their survivability

(Estler and Hein, 1960; Sporn et al. , 1970 ; I s m and Way,

1974; Varilos et al-, 1976). Utilisation of reduced co-

enzyzes leads to .:-'I? spthes is through oxidative

phosphorylztion (Martin, 1983). Under he stress conditions

HMP path a and operation of glycogenesis and

. . . gluconsogensis i r o ~ c i n o aclcs lmpart a great importanze to

the czrbohydrate ~.etabolis~,, xhich include pesticidal stress

also. The carbohydrate rnet~bolisrn is altered upon pesticide

toxicity, reducing the physiological activities of an

organism (~enzi et al. , 1978 ; Koundinya and Ramamurthi ,

1978; 1979 and Martin,1983).

?rcteir,s ir,-;zl..:e ir. r,~;:xlay architecture, metz'colic

- . C*'-- - r e , . , , . e . . Martin et a l . (1983)

. . . . - re~,3rte5 t5.3: zyc;err,s %re r.;-crc-jised t c z~!inoacias in the . . . +/,G-, ,,> . I. ,..- .r.- sr. arc f:;rt?er :eta5o1ised by Incorporation into

przteir. sr cie.-,y.at:cr. z r 2x~Sation. Anin0 acids are the

presTzs2rs f ~ r t;c.e ss..r.-,:-..zcis 35 various cellular proteins

and gl:czneo~enesis, zlycoqe~ synthesis and keto acids.

. . gy~~31y:sis sf aletar\r * r o - ~ + ,.,L,,r, and endogenous proteins result

, . in the for~.~ti.~.; of 27,ir.o aciB pool in the body. Amino acids

maintzin a ho~.eostasis in the protein environment because

, , aii.ino acids are xrlcier co?.scan-, movement between tissue and

plas~~a (Grinde and Seglen, 1981; Yutaka, 1984). Proteins

car! be t&en 2s an index of stress conditicns (Bano et al.,

1982 ; Assem and Hanki, 1983). Reports of Subbalakshmi and

Murthy (1984) confirz.zd the alterations ir, protein

rnetabolis:. upon pesr,icidal treatzent. Protein degradation

may be considered in assessing protein metabolic perturba-

tions awing tsxic ins2lts (Goldberg and Dice, 1974). Free

mino acids serve as ?recursors of important bionolecules

such as hornones, purines, pyrimidines, porpherins and some

vitai.ins. ?4oreover, they serve as a source of energy. They

act as a currency for ;he operation of protein metabolism.

The physiological state of the cell is dependent upon its

free x i i n o acid pool (Abidi, 1986).

. , Grz&?~ ~ : ? 1 3 c i n ~ ~e~=:z:aes zce knos:i? to specifically

, ,,-ec- .CZ arL5 =-- .- - - - - - - 7 . - ,,,, ,I,,, -.. z7.c Irz:S-rlcn r e g l 3 r . s :Lindvall

and Jessop, 1979) .d .:zc=.-.er ~ r s c r z a r t and vlzel 3,rcz~ic

- . . . . . - nr3,

RESULTS :

- 7 . . . The , r u G L is albir.0 rats "Lrea~ed

. . - . :.:it,L: c;cllor~ane are zreser.zec rr, Table - 9, c l g . 7 . In =he , . . . - - . . 3ye5~3t ~ ~ ~ ~ 3 ~ y ~ r ~ t ~ 1~-.-1215 are f o > ~ ~ s -,o ze dezlf:ed

in 0 -,reate2 al$ir.s rats. The depic:ion ;.:zs

, , , . . . . CO~S~SY~R-LI:: ~TG;~OSS~-:E 3:;;~. incrcssing dose. i . ' , : ~ , ? sir-zle

dose hepatic - 4 - - I - . L 1 a 3 ~ ~ P C Z D ~ S ~ C l 2 . 6 9 per cer,t; double Case

, , hepatic tlssze sho;t:e5 36.55 Fer cent deplec~sn. 7.0 l i i . . D

aultiple dose a&.r.ir:istered ones showed 55.37 per cent

depletion, :.:hen coppared to nsrzal ones. The depletion xas

statistically significant (Pt0.001) in all doses (Table - 9 )

Following chlorjane a~r~inlstration, the total proteins

are decreased and an elevation ru'as observed in free xino

acid levels and it was presented in Table - 10 G Fig. 7.

After ini-' Lial dose, the total protein was found to decrezse

in the hepatic tissue and the decrease in protein content

was progressive In dol~ble and ir.ultiple doses. The naxi7.u~

decrease ( 140.76) vc:,-as feud in nultiple dose ad:.inistered

ones (Table - 10). All the decreased observations Fiere

statistically significant ( P QJ.001) . The total free xino acids were elevated in all experimental liver tissues

(Table - 11 G Fig. 7) . Significant increase was noted at single dose and this increase was continued till nultiple

dose. ?r,5 C C ~ C ~ T . ~ C T . Z Z Z ~ lr. free ZY.I~O acid c~ntent xas

- - - ?5.3Sk, 2 . - z . . 5er S T . r singie, doilsle and xc.,;sle ,.--c -4 - - -hr--- ̂.-

.A,=, cu -.-.:= .,. =. u-V.- res2ec-,L-:eiji in r a ~ s .

-,-. P - - . - . Tables - + s I.: ar.z :LC. 5. Ckl~rdane treated zriirr,als

- levels. e 2 r elevaxlon wzs consistent and

- - - was progressive iz z-, zcses. These changes under chiordane

. . - intoxicatix are szaz=s=lcsily significant (P

- . - -. . 13 " ; r r ' r s C - . , , ...- ..-CI..I- XI^V SIC ~~1~.5or~ydrat~s in liver under - - "b: - . . , ,..,orszne ;r,z3ji.:catlcr. s s z e s t z s i3u?ctional irr,?ortmce In

s>~thesising 3 sfcring carbohydrates. After the

pesticidzl e ~ c s . ~ e , t?--0 dezrease i r i carbohydrate levels is

coy-stznt azc 3 r 3 9 r ~ - ,=ialle -- - - A Ll-~ ' ' - nultlple dose. The depletion

of carbohydrate l e - 1 ~ 1 ~ sucsest - - possible utilization of - czrbohydrazes ,o r.eet the energy demands during stress

coT L,-,,,..s. A 4 - j -.- - res~:c:5e exsoixe . a y also be co~sidered a s a . . s'ress. Sirice 1: ~lters the physiology and ethology (see

chapter - I) cf z?-e anrzl. ,Fie decreased carbohydrates may

. - -0 U- - - - - t i q k e d tc t3.e e-src:~ crisis induced by the pesziclae.

2 d

Decrczse in carbshydrate content nay be justified by

-- LAO . ways. O'Brien (1967) reported that the pecticidal

stress is k ~ o w n to induce h-ypoxic or anoxic conditions in

the animal by inpairing the r e sp i r a to ry centres of tissues

involved in breathing. Ranke-Rybicks and Bozena (1975);

Bhagyalakshmi (1981) showed respiratory distress, a sign of

pesticide toxicity.

^ . . - -;::,~r ssr: cf cr,;~ges I n carbohydrate levels have

beer. ro ; s r : r k ;r, seV.-erzl s-,ress ccr.d;t:m.s in -;ariccs non-

- fi,.-="- ^-- G -a . . (Linda and Charles, 1983 ; Satyaprasad,

1983; Sambzsiva Rao 1984; Ra&-aiah e t al., 1987; Azhar Baig,

1988; Vi jay doseph, 1989; Saila ja, 1991).

. - . a rzzid Cc;-stio? in car5chydrate levels in hepatic tissue

durirq cklcr5zr.e ai~;r,:surazlar, (Table - 9 & Fig. 7). This

. . may be & ~ e cc the ~csi~ced anoxiz c r hypoxic conditions and

increased clscogezzlysis, ?;hicn is observed ;u?der pesticidal

intoxication (O'Brien, 1967).

, . The aresezt investigation reveals that ;he total

protein ccnzent a decreased nider chlordane stress

(Table - 10 G Fig. 7 ) . Chlordane administered rats showed

reduced prozeir. c c r ~ t e n t in the hepatic tissue. Proteins

. , . occcpy a ur.iqze pcsxlon in the metabolism of the cell and

the ap.a~.ic szrcctural orcanization of cell is dependent on

L Aey. ( H a r p e r , 1985). ??oteirs may be degraded into amino

acids xhich are fed into TCA cycle through transferase

system. The low protein levels may be due to the inhibition

of arr,ino acid incorporation into their, (Roger, 1980). A

decrease in total proteins suggests their metabolic

utilization. However, Sandhu and Malik (1988) reported

increase in protein content during oral administration of

- - - - + -..--- - ^ ..^^ - -OD+ -'np T ~ n ~ p ~ n - ~ q - - = +a , vru..u-- . - = - , . .s , ,,, , Li,, ?.~cjh energy

:-. =**is fi,.,--.-.?. - - - -,-;=-= - - --- - - --. -Lo ; -nr02qc - - n p - - n m - .Ad* -.. 4 - . . - 4 - .. -..- -u..- " - " , , ,.,, ,,,, ,.. crs.=elns

d

car- 7 - - 2 ; - - - - - r i ? " + - - G ,... , 3 = u , 4 ...- . . - Ls. L4 - . i ate and - - .

- C r=--c= s- -.- - F Q qra-an- -m- . - zl:;csgf?~ - ie;:ek:?:. cs - . --.+-u -.. y* - 3 - * . b A,.., ~ s ~ l ~ ~ ~ l o n . r'orrp-ohr?,-f" - p - P C n=- , -C== ' - r -no nr" -c7" -2uu-4i 4 L . 4 u w -..- - - - - ---- +.. .,. d u - . . A L ccz:enc x e r e

- .- . - :.sS 2-7 ,?.ZTE2SE IF. ?..E CCZ?, :'Tee &-123 2 ~ 1 2 3301, ~ : h l ~ h -.rnqpq-c tr;- nr m v r i - r r ~ - = = - r ~ x r n n n 21_*zY .--.-:-_ d=ii.. ..-- -..;YYIU ~ T S V ~ O ~ Y S Z S :r. t h e l l v e r - . . - ? , e a a l n ~ t o ~zc:~,xlz::s. CT r r e e z~,:no a c i d s .

S i z i l a r s o r z of zza..i.:zs i n p r a t e i n l e v e l s have Seen

repar-led by nu~.ber of ;$:zrkers (Verma et a l . , 1979; Radhaiah

e t a l . , 1987; Rmamurthy, 1988; Vijay Joseph, 1989 and

Venkataswamy, 1991).

Cata presented i~ Ta3le - 11 showed inc reased l e v e l s . . of f r e e c i n o aczds 22r:x~ chlordane adninis:ratlon t o

a l b i n o r a z s . ?h? inc reese ir, f r e e x . i n o a c i d content i s

s t a t i s t i c a l l y s i 2 r i f i c z - x (? r o t e i r c o ~ t e n t and t h e e l e v a t i o n i n f r e e

mino a c i d levels sho;c?d t h a t t h e r e was degradat ion of

p ro te in . James e t a l . , 1980 showed t h a t amino zcids may be

- - usef';, :or ,:r..e s:r.",-i.s:s sf nor: ?rote:ns 2nd enzyres tc face ',he stress "" "q-- "-;a 7..2 - m

+. . .wv3-- ,, I,,,,,,,,. ,,, ,, ? e s t l c l d a l 2c;ion t l s s c e S~-a;e rcs~l- , : r .s :r. e l e v s ~ e d Is-:els of z.iilno acid

(Dubale and Shaln, 1979; Jayantha Rao e t a l . , 1983) hss been

resoy ted . + j f e e . a c i 5 : ~ n t e z c

, . - . , . l i -der i p L ~ i c 2 t e 3 -c--P-: ,..,~a,,,,, - - 3;C+b~ - m - - - - - s1 ori zc ~:\.T~TccT;.: t he ~f feet - . . - . . . of chlordane toxlci-,y. -, ap?ears tr,~; :nz f r e e a r ~ i n c acid

. . - pos l xay fe rn a p s s l c - e source of energy =o nee; che energy

~ e q u i r e x e n z s . Sne xore p o s s i b i l i t y of a ~ ~ i n a t i o n of ke to

a c i d s nay a l s o be cors ide red t o provide ;,ore energy by t he

e l eva t i on of amino ac id s . This cay l eads LO increased l i p l d

content i n experirxentel a n i ~ a l s .

r ' o l l o ~ i n g chlordsne a u ~ ~ i n i s t r a t i o n , t h e pyruvace

l e v e l s a r e deple ted a d l a cza t e l e v e l s are e l eva t ed . The

decrease or inc rease xas progress ive i n a l l exper inenta l

anirnals (Tzbles 13 8 1 4 Pig .8 . ) . The l e v e l s of pyruvate and

l a c t a t e were e s t i m t t i n hepa t ic t i s s u e s of chlordane t o

understand t he r a t e of ae rob ic and anaerobic g lyco lys i s

during t he chlordane t o x i c i t y .

From the p resen t s tudy i t i s v i s u a l i z e d t h a t t he r e is

a rap id deple t ion i n pyruvate l e v e l s i n h e p a t i c t i s s u e

dur ing chlordane t r e a t n e n t when conpared t o norinal ones.

Pyruvate i s t h e t e r r i n a l metabol i te of g lyco ly s i s under

P' aDrfi": r ,.hnAL *: ,,- ,. vv+, u d . l . A - - 2 **.a. - 3f p17~~: :ate jndicste the - - . . . . , , ez= z1 PzC- i 8f 3u- - - - - --0 -"--i‘r' -- T4s i ~ - r ~ s = p d ~ ~ r u v ~ t e

2,- i .- ,..* . . = U . - - J b .... i--- -I"- ---- levels alsc, izdicate i-,s relp as a precursor for other

, , . . . pr~j,;c-,s 12 -,ne - e t a ' ~ c l i s y like ccr.lbjersj=n tc iac-,ate =r to - n- f crr , : PC: 5 3 , 1 cis, zr; z;;~cer; des (Sathyaprasad,

generating - sufficient 2dAD fc,r the reaccion catalized by

glyceraldehyde 3-phosphate dehydrogenase.

Several workers have reported sixi l a r decrease in

p-jruvate levels in various animals exposed to various

pesti cides (Sivaprasada Rao ( 1980) ; Satyappasad ( 1983) ; Azar

Baig (1988); Ramamwthy (1988) and Vijay Joseph (1989).

Increased levels of lactate (Table - 13 & Fig. 8) may

be due to pjrruvate rr,oblllzaclon into Kreb's cycle. The

corverslon of pyruvate inzo lactate leads to reduction in

pyruvate levels. Cyclic AMP activates the phosphorylase

syster. during pesticide stress (~allicharan and Gibson,

1972) and inhibit he pyruvate levels (Anastasi and

Bannister, 1980) thus increasing l a c t i c acid (Nikinmaa,

1982) .

. . - .-?.q=> F 0 - n q 1 7 0 x - - -- ..-- __-_-,A- ,,.,.,. ,l4.. sf ?jrl-z7$ate t o l a c t a t e car! be . - - . -.. - - h a bw.-- - - . . e l z c t a t e l e v e l s i n h e p a t i c . . - . . . . . . . . , ~ s ~ . ; e s ~:~:zz :--.-:yzs-f t z j . ~ ~ l t : ; . The r a t e of production

d

- - . . . GI - 2 ~ L i c 2 : : ~ 15 Z . O T . E ~ ~ P T ~ ~ 2s a2 index of pnys io log ica l . - strsss ir. :kc c::-z-lzz- - s;:sz?:. (Thoye, 1971; Green e t a l . ,

1983; Michae l i s et a l . , 1983; Buono e t a l . , 1984; Kozlowski

e t a l . , 1985) . :e-,r.ezse ir. 2:iri~va-ue l e v e l s i n d i c a t e s t h e

conversicr- cr' r - . ~ x v - l e - - l n t o l a c t a t e . The inc reased l e v e l s

of l a c t i c zcid s;lzgests t h e prevalence of anae rob ios i s i n

t h e t i s s u e 2nd z i s o i n 5 i c z t e s s u s c e p t a b i l i t y t o ae rob ic

. . c o n d i t i o n s . ~ z z z i c ac id a i g h t be due t o t h e decreased

r : o b i l i z a t i c n of pyruvate i n t o t h e TCA c y c l e o r t o t h e

s t i m u l a t i o n c f axaerobic g l y c a l y s i s (Weil-Malherbe, 1937) .

I n order: t o -x?derstand t h e l a c t a t e pyruvate i n t e r

conversior?, the L3H a c t i v i t y has Seen s t u d i e d ( s e e

Chapter - 111).

The preser.t s tudy i n d i c a t e s t h a t changes i n c e r t a i n

l i p i d f r a c t i o n s 3ccur mcier chiordafie s t r e s s . The per cen t

i n c r e a s e vias z o r e i n mul t ip le dose adminis tered ones (85.41

per c e n t ) . c- , e r t a i n l i p i d f r a c t i o n s l i k e l i p i d s ,

phosphol ip ids and c h o l e s t e r o l were inc reased during

chlordane i n t o x i c z t i o n . D U T ~ I ~ ch lordane exposure inc reased

l i p i d c o n t e n t is observed. This nay be due t o increilsed

l i p o g e n e s i s through metabolic r e g u l a t i o n and compensation by

c e r t a i n Z p i d f r a c t i o n s l i k e phospho l ip ids , c h o l e s t e r o l ,

e t z . . $3 S - ~ ~ ~ C G T T ~ zke ~ Z Z Z C ; 35 ~ e s t i c l d e and t o p ro t ec t t he

- - - . - :e,,;-~y :r.z.t~r:=-; 2y.c r~r,c-,:cr.s. s t h e l i v e r ~s a z a j o r 2

- o - - - m 7 T. "cF-,-c - - . . . 1, sezer-sl ly e*.? bits Fighes t cho les=ero l 7-< m Q q C - C h ; 7 n' d m A P + D p - ,,,, ,, L LV..Lr..,~. Maptin (1983) repor ted e leva ted - Ic-:e1s sf cr.s;esxers, s;.:lr.g t~ I Z S f m c z i o n a l c a l i b r e of

q---..a-- - - p?, . - , 1 6 meta5c:izing. Increased l e v e l s of s=erc:ds l s d e r pezc,:c:de s z r e s s i s repor ted by many

;.;orkers (Agarwal, 1 9 8 1 Giri ja, 1987; Vi jay Joseph, 1989).

Increase ir, choles te ro l and phospholipid content was

repor ted by nc..ber of ~ i o r ~ e r s i n a v a r i e t y of p e s t l c i d a l

in tox ica ted a n i n a l s (Madhu, 1983; Ramamurthy, 1988 and

Radhaiah, 1988; Vi jay Joseph, 1989) .

Increased l e v e l s of l i p i d f r a c t i o n s suggest t h a t t h e r e

i s an z l t e r a ~ i o n I'n t h e physiolo5y of t he animal under

chlordane i n t o x i c a t l on. However, a more c l e a r pi c t u r e

regarding t he energy sysxern and o the r a spec t s of metabolism

can be ~ c d e r s t o o d by s rudying some r e l a t e d enzymes.

Table - 9 : Tr.e z z c s l czrbsr.;.2rztes cf z e p a t i c tissue of p,-.---,-

- - ,,..--,1 ar-.Z zr.,,-r2ar.e t rez-,eZ albino rats.

zx/$ wcz wr Percent change

Control 1 1 - 1 4

- 1.0995 -

S i n c l e dose

Double dose

Xultiple dose 18.36

+ 0.611 P

Table - 10 : :r.g rzzal ;rsze:r esnzezt ke?atic tissue of - -

- 7 - w C -... h r r - n . . ,,..--, 1 -..Y -.-A b-US-L.e t r e a t e e aiD:no rats.

- - . , ~j i*.-e= h-z Percent change

Single dose 181.28

- 6.89 - P

, . --- " h Table - 11 : :he t c t a ; i ree -...I.., eclc cc r t en t i n he;a:ic - 7 w - - - u w = c . . t 0" -- - n * - r ~ l b U 4 1 L & ,ZJ;C ~ ~ ~ : ~ ~ t j ~ ~ e t r e a t & r a t s ,

Sinsle dose

Double dose

Percent c h g e

Note : Each value represents the mean of s i x i n d i v i d u a l

observations. i- ind ica tes the s t andard d e v i a t i o n . - 'P ' denotes the l e v e l of s t a t i s t i c a l s ignif icance.

. - Table - 12 : L E ; . E ~ s ~f "I~.-T:V~C - - 2~16 1 ~ : 5epstic tissue of

- - ?".--.-,-. 2-- ?"- n,----a -ye-- ,,..--, 1 ,..; " ..---- =..- i--~-e;i elbino rats.

c - 2 5 , ' ~ -,*re t ;%;; Percent change

Single dose 5 . 2 9

Double dose 4 . 5 8

P lu l t i 2 l e Ccse .,-.a 7 a7

- 0.282 P

Table - 13 : A c z l v i t y levels 25 lzctic acid in hepatic

tissues of ccctrol a ~ d cklordane treated albino

rats.

;?,cj/g wet wt Percent change

Control

Single dose

Double dose

Note : Each value represents the mean of six individual

observations. - + indicates the standard deviation. 'P' denotes the level of statistical significance.

. - + .-- 2 Table - 14 : The -,=:a- c3r.ter.: in hepatic tissue of cofizrcl a-5 cklor5ane treated albino rats.

ZG/G w2z wc Percent change

Control - r, l i i . 3 3 i 3 . 3 4 -

Single dose 85.8

Double dose 105.8

i 7 .35 - P < 0 . 001

Multiple dose 130.4

+ 7.1 P

. - Table - 15 : Levels of ckcs_sxc~ipids in hepatic tissue of

coctrol and c?-loreane treated albino rats.

z.ng/g wet i+;; Percent change

Control 27.04

Single dose 2 4 . 1 2

+ 0.731 P

Table - 16 : Levels of chcleszersl in kepatlc kissae of conzrol and chlorSane zreazed albino rats.

Percent change

Ccntrol 7.616

Single dose 8.60

+ 0.117 P

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