Effects of antifilarial drugs on the activities of the oxidative enzymes of mitochondria and microsomes of rat liver and kidneys

Vol. 106, No. 3, 1982 June 15, 1982

EFFECTS THE

BIOCHEMICAL

OF L4NTIFILARIAL

OXIDATIVE

ENZYMES OF RAT

Tamal

K.

Division and

of

Industrial Received

AND

DRUGS

Ray 1 , Sarita

ON T:JE ACTIVITIES

OF

AND MICROSOMES

AND ICIDNEYS

Agarwal

Biochemistry,

28,

RESEARCH COMMUNICATIONS Pages 888-894

OF MITOCHONDRIA LIVER

and

Central

Toxicology April

BIOPHYSICAL

Research

Deepak

Drug

K.

Agarwal2

Research

Institute

Lucknow,

India

Center,

226001

1'982

Centperazine or diethylcarbamazine, administered at various dose levels to rats inhibited the activity of succinate dehydrogenase significantly in 4 hrs in liver. Centperazine also inhibited the activity of cytochrome-c oxidase but stimulated the activity of benzo (a) pyrene hydroxylase in liver. IR kidneys, activities of succinate dehydrogenase, cytochrome-c oxidase and aniline hydroxylase were significantly inhibited by centperazine only, however, the activity of benzo(a)pyrene hydroxylase was inhibited by both the drugs. These drugs had no effect on the activity of aminopyrene N-demethylase and cytochrome P-45U contents of liver and kidneys. INTRODUCTION Hetrazan choice

against

perazine

filariasis

has

for

quite

recently

Institute,

satisfactory

some

a drug

time

five

(21.

Hetrazan

times

India,

has

value

more

has

pharmacological centperazine

by

Lucknow,

effects has

against

(1).

little

0006-291X/82/110888-07$01.00/0 Inc. reserved.

888

Drug

found

to

Res-

be

of

with than

studied

metabolism

background

Central

activity

hetrazan

regarding

its

(3,4),whereas except

1. Present address: Department of Pathology, Fort Collins, Colorado 80523. 2. Present address: MST Laboratories, Center ersity of Tennessee, Memphis, TN 38163

@ 1982 b? ,4mdernic Pre.s, o.f reproduclion in any form

Cent-

filariasis

extensively and

the been

pharmacologic

been

of

[4.4.0]

formulated

therapeutic

nearly

Copwghi All r;gh&

been

(3-ethyl-8methyl-1,3,8-triazabicyclo

decan-Z-One), earch

(diethylcarbamazine)

for

Colorado of Health

a few State

in-

University,

Sciences,

Univ-

BIOCHEMICAL

Vol. 106, No. 3, 1982 house and

investigations

from

the

been

body

of

to

enzymes

affect in

sister

that

of

and

enzymes

of of

-MATERIALS

(7). it

order

efficacy

was

and

of

rapidly

in

the

drug

form

has

of

been

its

recently

metabolizing

considered of

former

is

of the

the

interest

two

microsomal

elucidate the

eliminated

centperazine

effect

to

is

and

some

activity (5,6).

Since

mitochondrial in

higher

liver

comparative

renal rats

(3,4), of

hetrazan

the

hepatic

animals

activity pig

safety

hetrazan

products

RESEARCH COMMUNICATIONS

pharmacologic

drug

experimental

guinea

investigate

nism

found

the

analogue

its

versus

degradation

shown

to

quality

has

oxidative

BIOPHYSICAL

regarding

therapeutic It

AND

drugs

on

oxidative

the

possible

over

the

mechalatter

one.

AND METHODS -~

Male Charles-Foster rats (loo-150 gm) from Central Drug Research Institute colony were fasted overnight before administration of a single oral dose of 10, 25 or 50 mg/kg centperazine or hetrazan base. Control rats received equal volume of normal saline only. All the rats were then placed on pellet diet and water and -ad libitum, were sacrificed after 4 hrs of dosage by cervical dislocation. Liver and kidneys were quickly removed and homogenized in ice-cold O.ZSM sucrose. Mitochondria, 9,OOq x g supernatant and microsomes were prepared according to the method of Schneider (8). The activities of succinate dehydrogenase (9) and cytochrome-c oxidase (10) were determined in hepatic and renal mitochondria. The 9,000 x g supernatant was used to assay the activities of aminopyrene N-demethylase and aniline hydroxylase (ll), and benzopyrene hydroxylase (12). Cytochrome P-450 content was estimated in the microsomes and the total protein was determined by method of (13), Lowry c a. (14) using bovine serum albumin as standard. Statistical significance of the results was evaluated by student's 't' test as described by Fisher (15). RESULTS Administration produce of

any

Kidneys levels

sign

animals

during absolute

weights,

as well (data

of

not

of

centperazine

overt the

or

as protein shown).

or

toxicity

period relative contents

of

or

hetrazan

result

experiment. fresh

remained

weights unaffected

of

did

not

in

mortality

The

hody liver

at

all

and drug

Vol. 106, No. 3, 1982 TABLE 1: dehydrogenase

The

BIOCHEMICAL effects

of

AND

Centperazine

and cytochrome-c Succinate (nmoles minimg

G-TOUP

BIOPHYSICAL

Control

and

oxidase

DEC on

in rat

RESEARCH COMMUNICATIONS

the

liver

Dehydrogenase K3Fe(CN16 reduced/

activities

of

succinate

mitochondria. Cytochrome-c Oxidase (nmoles ferrocytochrome-c oxidised/min/mg protein)

protein)

39.19

+

3.61

160.08

+

11.90

29.18

zt 1.93

150.35

+

6.23

25.0

18.35

t

2.44a

123.70

T!Z

3.48'

50.0

15.25

zt

2.30a

117.17

zk

3.06b

DEC (mgj 10.0

38.48

t

3.24

164.70

+

6.90

25.0

29.75

zt

2.03

159.32

t

7.44

50.0

26.39

II=

2.77=

141.03

zt

3.16

Centperazine 10.0

All 'p<

(mg)

values are 0.01;bp<0.02

Tables hetrazan

in

cantly

rat

activities liver

effects

of succinate

both

the

and at 50 mgikg

had no perceptible

effect

effects of Centperazine and cytochrome-c oxidase Succinate (nmoles

and in

protein)

and

and cytochrome-c

Centperazine enzymes in kidney

at

signifi-

25 and 50 mg/kg

mitochondria,

on the activities

DEC on the rat kidney

Dehydrogenase K3Fe(CN)6 reduced/

minimg

centperazine

dehydrogenase

of

mitochondria

Group

of

activities

liver

TABLE 2: The dehydrogenase

the

mitochondria.

the

hetrazan

observations.

and kidney

inhibited in

whereas

+ SE from four and 'p< 0.05.

1 and 2 summarize

on the

oxidase

dose

mean

of

activities mitochondria.

of

ztz 2.66

284.48

zt 23.12

52.58

+ 3.92

251.36

LIZ 10.69

25.0

46.38

* 2.10

240.40

+ 19.69

50.0

22.14

i

206.46

zk 4.54b

DEC (mg) 10.0

54.24

zk 2.67

318.83

zk 23.20

25.0

47.49

zt

3.88

298.92

zt

19.29

50.0

51.56

zt 3.51

313.58

zt

15.87

Centperazine 10.0

All ap<

values 0.001

(mg)

are mean * SE from and 'p
four

2.28a

observations.

8%

succinate

Cytochrome-c Oxidase (nmoles ferrocytochrome-c oxidised/min/mg protein)

53.27

Control

these

BIOCHEMICAL

Vol. 106, No. 3, 1982 TABLE

3:

The effects Cytochrome

of Centperazine P-450 content

and rat

in

AND

BIOPHYSICAL

RESEARCH COMMUNICATIONS

IIEC on the activities liver microsomes.

of

micrasomal

nxidative

enzvmes

k 0.09

0.45

*

0.03

0.98

+ O.OL

0.10

0.57

*

O.OIC

I .l&

+ Cl.07

1.28

* 0.10

0.6/t

+ O.Ojh

1.03

* lU.06

zt 0.99

L.18

*

1.05

* o.06s'

u.,J9

* t1.02

11.88

* 0.79

L.14

zt 0.07

0.51

+ 0.03

o.qz

+ 0.02

25.0

10.41

* 0.62

0.8Q

*

0.03

0.51

+ 0.05

0.88

zk 0.03

50.0

9.67

k 0.86

0.99

+ 0.03

0.52

+ 0.04

0.89

k O.U3

11.76

+ 0.89

1.00

12.28

zt 1.06

1.10

l

25.u

11.92

* 1.00

50.0

11.65

DEC CrngJ iO.0

Control Centperazine 10.0

All values ap
(mg)

are

mean + SE from four 0.02 and =p< 0.05.

enzymes

in

liver

dehvdrogenase

As

evident,

hydroxylase any

of

cytochrome

liver

in

on

these

the

P-450

hetrazan

(Table

however,

significantly and alone

activity P-450

the

dependent

of content.

activity

manner

N-demethylase content.

of

Hetrazan

aminopyrine

remained

unaffected

activity

of

inhibited

at

the

succinate

dose.

mg/kg

cytochrome

aminopyrene

activity

The

that

of

of

but

had

no

and

aniline

had

no

effect

tested.

content 4).

on

stimulated

P-450

parameters

50

drugs

and

a dose

cytochrome

kidneys,

centperazine

on

inhibition at

these

significantly

activities

manner,

of enzymes

hydroxylase the

signigicant mitochondria

effects

oxidative

or

In

dent

the

centperazine

on

except

in

3 shows

benzo(a)pyrene

on

kidneys

microsomal

effect

observations.

activity

Table hepatic

or

O.O(+

of

aniline

25

and

by

N-demethylase by

the

hydroxylase 50

centperazine

benzo(a)pyrene both

mg/kg

and or

hydroxylase drugs was

in

was,

a dose

inhibited

depenby

dose.

DISCUSSION Centperazine gastro-intestinal

and tract,

hetrazan

are

rapidly

bio-distributed

aild

891

absorbed

through

almost

completely

the

and

Vol. 106, No. 3, 1982 TABLE

&:

The

effects

Cytochrome

8lOCHEMlCAL

of

Centperazine

P-450

content

and rat

in

AND

BIOPHYSICAL

RESEARCH COMMUNICATIONS

DEC CIII the activities kidney microsomes.

Aminopyrine N-demethylase Cnmoles HCHo/ minlmg protein)

Aniline Hydroxylase (nmoles p-amino phenollminlmg

3.03

Yt 0.34

0.41

3.27

*

0.32

25.0

3.67

50.0

of

microsomal

oxidative

iznzyrn~s

Benzo(a~pyrene Hydroxylase (relative flu0rescencel m%nlmg protein)

Cytochrome P-450 (nmoleslmg protein)

* 0.01

0.049

k 0.003

0.24

zt 0.04

0.37

+ 0.01

0.019

* o.ooza

0.31

* 0.03

E!Z0.31

0.28

*

o.02b

0.017

+

0.30

*

0.02

3.10

l

0.31

0.27

+ o.02b

0.016

+ O.OOla

0.29

l

0.02

DEC (rng) 10.0

2.89

zt 0.36

0.43

*

0.04

0.031

+

o.oo2b

0.18

*

0.02

25.0

3.30

*

0.24

0.45

*

0.04

0.022

*

o.oola

0.22

*

0.01

50.0

2.85

+ 0.31

0.41

*

0.03

0.018

zt O.OOla

0.21

*

0.02

Control Centperazine 10.0

All 'p<

(mg)

values 0.001

are mean & SE from and 'p
eliminated

from

(16,17).

The

little

the

two

the drugs

longer

present

four

observations,

body

of

experimental

show

no

organ

retention

in

study

represents

drugs

has

been

being

the

o.oola

animals

specific

the

accumulation

except

a

The

period

in

(16,17).

kidneys time

reported

when to

4-hour

maximum

be

24 hours

within

concentration

present

in

of

liver

and

kidneys

(16). Liver, and

other

xenobiotics,

toxicities mes

of

of

such

agents and

in to

the

toxicity

ention

in

the

kidneys.

Inhibition

Such to

and

a variety

hepatic

years

kidneys

seems

of of

benzo(a)pyrene

target of

This

(19,2(l). of

and to

be

new

drugs

a direct are

(21,22,23).

892

metabolizing kidneys

of

importance which

effect known

centperazine

drugs

ultimate enzyhas

have

of

longer

the

ret-

not

of

drugs. acute

stimulation is

well

enzymes

these

with

been with

oxidative

However, by

the

in

mitochondrial

enzymes

hydroxylase

is

of

of

drug

capabilities

oxidative

xenobiotics

disposition

Presence

evaluation

microsomal

metabolic

a vulnerable

(18).

of

inhibition

of

conjugative

recent

regard

liver

site

becomes

oxidative

established

rat

main

exposure

of understood

the

and

Vol.

BIOCHEMICAL

106, No. 3, 1962 at present.

There

of hetrazan

by rat

shown

to exhibit

has been liver

observation

that

homogenate

kidney

hetrazan

(24)

but

The more hetrazan bolism,

over

microsome,

did of

pronounced

increased probably

enzymatic

N-demethylation

and the

enzymatic

system

to aminopyrene

4) further

not

RESEARCH COMMUNICATIONS

on the

similar (Table

also

on enzymes,

(16) 9 which

a report

properties

The present

AND BIOPHYSICAL

mediate

supports the

has been

demethylase

(24).

the

report

earlier

demethylation

not

only

of

centperazine. effect

studied

bio-availability contribute

here,

of centperazine is

as compared

consistent

and higher to higher

with

its

therapeutic

efficacy

of

to

slower

meta-

efficacy centperazine

hetrazan.

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6. 7.

8. 9.

10.

Hewitt, R-I., Kushner, S., Stewart, H.W., White, E., Wallace, W.S., and Subba Row, Y., (1947) J. Lab. Clin. Med. s, 13141329. Saxena,R., Sharma, S., Iyer, R.N., and Anand, N., (1971) J. Med. Chem. g, 929-931. Bangham, D.K., (1955) Brit. J. Pharmacol. 2, 406-412. Chandrasekaran, B.,and Harinath, B.C., (1980) Indian J. Exp. Biol. g, 722-724. Saxena, R., Iyer, R.N., Anand, N., Chatterjee, R.K., and Sen, (1970) J. Pharm. Pharmacol. z, 306-307. A.B., Central Drug Research Institute. (1980) Annual Report, pp. 41-42. Chandrasekaran, B.,and Harinath, B.C., (1980) Indian J. Exp. Biol. 18, 1298-1300. Schneider, W.C., (1972) in Manometric and Biochemical Technique, (W.W. Umbreit, R.H. Burris and J.F. Stauffer, ed.), pp. 196-212, Burgess Publishing company, Minneapolis. Slater, E.C., and Banner, W.D., (1952) Biochem. J. z, 185-195. Cooperstein, S.J., and Lazarow, A., (1951) ,J. Biol. Chem. lS9, 665-670.

11.

Johnson, Biochem. 12. Dehnen, 53,

R.K., Mazel, P., Donahue, J.D., Pharmacol. 3, 955-966. W., Tomingas, R., and Ross, J.,

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T'., and Sate, T., (1964) ,J. Biol. Chem. x, 2370-2378. O.H., Rosebrough, N.J., Far-r, A.L., and Randall, R.,J., (1951) J. Biol. Chem, 193, 265-275. 15. Fisher, R.A., (1948) Statistical Methods For Research Workers, 10th Edition, (1liver and Boyd, Edinburgh. 16. Roy, T.K., Sharma, S., and Srivastava, V.M.L., (1981) Tndian LT. Med. Res. E, 565-571. li. Srivastava, V.M.L., and Roy, T.K., (1981) Indian J. Exp. Biol. lJ, IS.

785-i86.

Kemmer,

19. Litterst, (1975)

H., (1970) Amer. J. Med. 3, 617-629. C.L., Mimnaugh, E.C., Reagan, R.L., Life Sci. E, 813-818.

and Cram,

(1971)

T.E.,

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BIOCHEMICAL

AND BIOPHYSICAL

RESEARCH COMMUNICATIONS

2@. Jacobson, S.V., and Cinti, D.L., (1973) J. Pharm. Exp. Therap. E, 226-234. 21. Srivastava, S.P., Seth, P.K., and Agarwal, D.K., (1975) Environ. Physiol. Biochem. 2, 178-183. 22. Agarwal, D.K., Kaw, J.L., Srivastava, S.P., and Seth, P.K., (1978) Environ. Res. g, 333-341. 23. Agarwal, D.K., Agarwal, S., and Seth, P.K., (1982) Drug Metab. Dispos. g, 77-80. 24. Srivastava, V.M.L., and Khan, M.M., (1976) Indian J. Exp. Biol. g, 504-505.

894