COMPARATIVE EFFECTS OF POLYCHLORINATED BIPHENYLS AND ORGANOCHLORINE
PESTICIDES IN INDUCTION OF HEPATIC MICROSOMAL ENZYMES 1
Joseph C. Street, Francis M. Urry,
D. Jesse Wagseaff and Adrian 0. Blau
Department of Animal Science
Utah State University
Logan, Utah 84321

Chlorinated biphenyls (PCB), triphenyls (PCT) and other related
compounds hav<* important Industrial uses and have been produced in large
vivlumoA for many years.
ment.

Evidently some amounts reach the general environ­

Pesticide residue chemists became Interested In such materials

following a report from Sweden of their presence in wildlife tissues.^
This has been shown to interfere with the routine residue analyses of
many organochlorine pesticides since complex patterns are produced in
2 i
EC-GLC. *

Moreover, routine cleanup procedures fail to eliminate such

Interferences unless special precautions are taken.

These materials un­

doubtedly contributed to some of Che "apparent" DDT found in biological
specimens predating usage of DDT in agriculture or public health and
constitute part of the present background of llpoidal organic chlorine
in the biota.
Polychlorinated biphenyls produce acute toxic effects in mammals
similar to those of chlorinated naphthalenes and other chlorinated aro­
matics.

The special vulnerability of the liver to such agents is well

documented.^

Many llpld-soluble chemicals induce various enzymes of the

hepatic endoplasmic reticulum (microsomal enzymes) including the drugmetabolizing hydroxylative system.

Several organochlorine pesticides are

potent Inducers of such enzymes, with the result that treated animals
metabolize drugs, insecticides and other foreign compounds more rapidly.5
1 Presented at the 158th meeting, American Chemical Society, Pesticide
Chemistry Dlviolou, September 8, 1969.

MCNS

063107

)
In some situations such stimulation Is beneficial as, for example, in
causing a more rapid detoxication of dleldrin and a lover residue stor­
age in the body.

The induction could likewise be detrimental because

certain toxifying reactions, such as activation of organothlophosphate
insecticides, are also stimulated.

Enhanced steroid degradation also

occurs which raises possibilities of disturbing certain physiological
processes due to altered endocrine relationships.

The latter has been

suggested es the baslB for the thin egg shell phenomenon end poor repro­
duction in certain birds chronically exposed to DDT, dleldrin and some
other orgenorhlovine Insecticides.**

Some polychlorinated biphenyl matcr-

hnvo already been reported to Induce microsomal enzymes.^

The pre­

sent investigation was undertaken to compare the induction potencies of
several types of polychlorinated biphenyls and triphenyls to those of
various organochlorine pesticides, and to obtain preliminary information
about tissue storage trends for such materials.

The results contribute

to our understanding of molecular characteristics required for microsomal
enzyme induction and also allow some speculation about the consequences
of various types of chlorinated compounds found in biota.
METHODS
Groups of mala and female rats ware individually fed Purina Labora­
tory Chow diets containing 25, 50, and 100 ppm concentrations of one of
the several PCB and PCT materials tested.

The test materials, Aroclors#

provided by the Monsanto Co. , consist of crude products obtained by
chlorination of biphenyl, or Crlphenyl, to specified limits; each is
therefore a complex mixture.
cent chlorine.

The ten products ranged from 21 to 68 per­

Except for the negative controls, the diets were also

treated to contain 1 ppm dleldrin.

One group of rats in each experiment

was fed diet containing 50 ppm DDT and 1 ppm dleldrin.

The diets were

- 2 MOhS

G631G&

: '
fed for 15 days.

;

During Chet period test* of drug and insecticide metab-

olien In vivo were conducted.

Later, tests of microsomal enzyme activity

were carried out with liver homogenates.
Hexobarbltal sleeping time measurements were made on the tenth

day of treatment.

Hexobarbltal sodium was administered at the rate of

100 mg/kg l.p. In aqueous solution.

The duration of deep sleep ending

with the return of the enlsel's righting reflex was recorded as the
sleeping time.

The oxidative detoxification of EPN (0-ethyl-0-(p~nltro-

ph«ny1)-phenylphoephonoth1oate) and the O-demethylatlon of p»nltroanlsole (PNA) were measured by incubation of the respective substrates with
the 10,000 X g supernatant of liver homogenates following the method of
Heal and OuBols as modified by Klnoshita et al.

o

The ring hydroxylation

of aniline was measured by the incubation of aniline hydrochloride with
the 10,000 X g supernatant of liver homogenates following the method of
Kato and Gillette.^
Residual dieldrin in adipose tissue was determined by electron
capture gas chromatography on hexane extracts of the tissue.

The adipose

tissue extracts required special cleanup to eliminate PCB Interference in
the detection of dieldrin.

This was accomplished by passing the extract

over a 2-stage chromatography column consisting of 1:1 MgO-Celite (5V')
on the bottom and Florlsll containing 47. moisture (4") on the top.

After

eluting with hexane (200 ml) to bring out the PCB components, dieldrin
was recovered by eluting with 400 ml of a 1 + 4 dichloromethane + hexane
solution.
Qualitative and seml-quantltatlve evaluation of PCB storage in
adipose tissue was possible after careful evaluation of GLC data comparing
parent material and that recovered from tissue.

- 3 -

MCNS

063109

)
RESULTS
Short term feeding of polychlorinated biphenyls and triphenyls to
rats at rates up to 100 ppm In the diet had essentially no effects on
food consumption, weight gains or efficiency of food utilization (Table 1).
Liver enlargement was observed with all products tested, increasing in
dogree with the percentage chlorine in the product.

Products with 60

percent chlorine or more caused greater liver enlargement than equivalent
dot«e of the insecticide DDT*
Evidence of FCB induction of liver hydroxy1sting enzyme activity was
obtained with all enzyme testa employed.

Hexobarbital sleep times were

markedly reduced by the PCB materials at both 50 and 100 ppm In the diet
for 10 days (Fig. 1).

The rate of decrease was reasonably linear with

increasing degree of PCB chlorination.
Enzymatic aniline hydroxylation, EPN degradation and demethylation
of p-nitroanisole, as determined with liver homogenates, were increased
with preparations from PCB-treated rats (Fig. 2 and Table 2).

In general,

higher chlorination of the biphenyl resulted In greater enzymatic activ­
ities.

These enzyme activities were similarly affected in both sexes by

the treatment materials, although the male appeared to be more responsive
(Table 3).
Dleldrin metabolism In vivo, judging from its residue storage in
adipose tissue, was stimulated by the treatment materials, especially
those with high degrees of chlorination (Fig. 3).

The products contain­

ing 607. chlorine or greater reduced dleldrin storage to levels found in
untreated control animals in our laboratory.

This activity toward diel-

drin metabolism had been predicted by analogy to tha similar effects of
snzyme-lnducing drugs and organochlorine pesticides.

- 4 -

HONS

06311°

,

'

)

Higher dosage rates (100 ppm VS 50 ppm In the diet) produced greater
degrees of enzyme Induction.

The dose-response curve (Fig. 4) obtained

with 627. Cl-PCB, In the range from 5 ppm to 100 ppm in the diet, was
generally similar to that of DDT or phenobarbltal yet that substance was
greater In potency at every dosage level.

Heptachlor epoxide, by contrast,

was considerably more potent than any of these compounds.
The two products containing chlorinated triphenyl components (one
42% Cl-PCT and the other a mixture of PCB:PCT, 60:40, containing 65%
chlorine) *-«*t*d

<n comparison to the PCB materials, less active In

terms of effect on the enzyme activities measured in vitro.

However,

hexobarbltal sleep time was more strongly Influenced by the 427. Cl-PCT
material than by the equivalent PCB, and dleldrln metabolism was highly
stimulated by both the PCT-contalnlng materials (Table 4).
The Arochlor products fed and the residual components recovered
from adipose tissue were all examined by electron capture gas-liquid
chromatography.

Some degree of storage of each product was observed al­

though the residues had greatly modified composition in comparison to the
parent materials administered.

Those products and individual components

having the least chlorine appeared to be stored the least in adipose
tissue.

That can be seen in the comparison illustrated (Pig. 5) for 62%

Cl-PCB and its tissue residue.

The aarly GLC peaks (components low in

chlorine) in the residue were greatly diminished relative to most of the
later peaks in the chromatogram,

peaks C and D In that residue match,

in retention time, two of the three major peaks reported by Risebrough
at al.^ in PCB residues from tissues and eggs of birds in the San Fran­
cisco bay area.

The tlasue residues for PCBs lower in chlorination were

much lower in proportion to chat of the highly chlorinated PCBs and

- 5 -

consisted chiefly of minor components of those materials.

Work is In

progress to quantitate these residues In terms of organic chlorine.
SUMMARY
To summarize all the responses evaluated, hepatic microsomal enzyme
induction was minimal with the PCBs of low chlorination (21-32% chlorine)
but Increased to very high values for those materials averaging over 60%
chlorine.

The latter exceeded DDT in potency of induction, but not hep-

tachlor epoxide.
,

Microsomal enzyme induction caused by PCB materials is probably

r

addlrlve with that produced by organochlorlne pesticides.^Hence, If

",

any of the toxicological aspects of organochlorlne pesticides In the
environment are related to their enzyme inducing action, the presence
of highly chlorinated biphenyls and triphenyls would intensify such
biological effects of the pesticides.
The very marked influence of the highly chlorinated PCB and PCT
materials on dieldrln metabolism (which, by analogy, would affect all
other cyclodiene Insecticides and DDT as well) could greatly confound the
Interpretation of exposures of animals and birds bearing mixed residues.
The toxicological Implications of such exposures would be equally con­
founded.
This work extends our previous investigations of the structureactivity aspects of microsomal enzyme inducing agents (presented In part
at the 156th ACS meeting, Agriculture and Food Chemistry Division) and
the results fit well with our working hypotheses regarding the activities
of various bridged and unbrldged diphenyl compounds.

6

MOMS

063112

LITERATURE CITED

1.

Jensen, S.

New Scl.

32:66(1966)

2.

Reynolds, L. M.

3.

Rlsebrough, R. W., F. Reiche and H. S, OlcotC.
Toxicol. 4:192-201(1969)

Bull. Envir. Cont, Toxicol.

4.

Von Oetcingen, W. F.

4:128-143(1969)
Bull: Envir. Cone.

U. S. Dept. of Health, Education and Welfare,

public Health Service Publication No. 414, 1955. The Halogenated
Aliphatic, Olefinic, Cyclic, Aromatic, and Aliphatic Aromatic
Hydrocarbons Including the Halogenated Insecticides, Their
Toxicity and Potential Dangers,
p. 306-307.

5.

Street, J. C.

Ann. New York Acad. Scl.
Nature

160:274-290(1969)

6.

peakall, D. B.

7.

F4««»t>r»t'gh, R. W., P. Reiche, D. B. Peakall, S. G. Herman, and
M. N. Kirvon. Nature 220:1098-1102(1968)

216:505(1967)

8.

Kinoahita, K. F., J. F. Frawley and K. P. DuBola.
Pharmacol. 9^505(1966)

9.

Kato, R. and J. R. Gillette.

Toxicol. Appl.

J. Pharmacol. Exptl. Therap.

150:280(1965)

10. Street, J. C., D. J. Wagstaff, and F. Mayer. Proc. 6th Interamerlcan
Conf. on Toxicol, and Indust. Med., Univ. Miami School of Medicine,
1960.
(In presa)

*0N$

063113

Table 1,

Food consumpt1 h, uelght gain and efficiency,
p liver veigniv
of female rats treated with polychlorinated blpuenyls and DDT. 1
Food

Cons.

Height
Gain

S

Basel

226 t 5.6

Dleldrln, 1 ppm
+ DDT, 50 ppm

Treatment

g

Wt. Gain
t Food
g/100g

Liver Wt
g/l00g

45 t 1.6

20 1 0.8

4.1 t 0.44

233 t 3.3

49 t 3.3*

21 1 1.6

4.3 t 0.27

224 t 6.0

44 t 2.4

20 t 0.9

4.5 t 0.44*

Trial A

1- 48* Cl-PCB,

50 ppm
100 ppm

226 ± 4.7
225 ♦ 5.5

47 ± 3.2
44 ± 6.5

21 t 1.3
20 t 3.0

4.7 t 0.45**
4.7 t 0.46**

+ 54* Cl-PCB,

50 ppm
100 ppm

224 ± 5.4
231 t 9.0

47 t 2.0
48 ± 2.0*

21 t 0.7
21 * 1.7

4.7 t 0.39**
5.5 i 0.46**

+ 607, Cl-PCB,

50 ppm
100 ppm

226 t 7.1
225 * 5.5

50 ± 3.1*
59 t 2.2**

22 t 1.4*
26 i 3.0**

4.6 t 0.28**
5.3 ± 0.22**

+ 687. Cl-PCB,

50 ppm
100 ppm

224 t 9.6
227 t 6.3

50 t 5.0*
51 1 5.5*

22 ± 1.8*
22
1.8*

5.1 + 0.16**
5.1 t 0.46**

, 50 ppm
ppm

100

228 t 8.3
232 ♦ 7.9

43 i 3.0
49 t 1.7*

19 ♦ 0.9
21 t 1.1

4.8 t 0.31**
5.2 t 0.37**

50 ppm
100 ppm

218 + 4.6
211 ♦ 5.7

40 ± 4.3
48 t 5.8*

18 t 1.8
18 t L.l

4.4 + 0.21
4.7 ♦ 0.01**

39 ± 2.3

16 + 2.5

4.0 t 0.31
4.3 t 0.32*

(
+ 657. Cl; +

+ 427. Cl-PCT,

C
D

237 t

Basal

u>

Trial B

Dieldrln, 1 ppm

234 t 6.9

40 ± 8.0*

17 ♦ 3.0

+ DDT, 50 ppm

221 t 2.6*

32 ± 2.3

15 ± 1.6

4.5 t 0.21**

+ 217. Cl-PCB,

50 ppm
100 ppm

228 ± 6.6
228 1 6.9

33 t 1.3
33 ± 5.5

14 ± 1.7
14 ± 2.5

4.1 i 0.33
4.1 ± 0.56

+ 327. Cl-PCB,

50 ppm
100 ppm

225 t 4.5*
232 ± 4.3

38 1 1.7
39 1 4.7

17 1 1.9
17 ± 1.3

4.4 ♦ 0.52**
4.7 t 0.44**

+ 42S Cl-PCB,

50 ppm
100 ppm

226 t 7.7
211 t 5.0**

39 1 2.1
31 1 3.1

17 t 1.9
15 ± 3.8

4.5 t 0.26**
4.6 * 0.28**

+ 627. Cl-PCB,

50 ppm
100 ppm

216 t 5.4**
230 t 5.3

35 ± 2.5
48 1 3.6**

16 ± 1.1
20 ± 1.2**

4.8 ♦ 0.22**
5.3 ♦ 0.33**

All groupa except the baaal received 1 ppm dieldrln. Data are expressed
as means 1 S.E. for groups of five rats.
Asterisks, * and **, Indicate values judged significantly different from
the basal group by LSD analysis Kith P 5 .05 or .01, respectively.

HONS

063114

Table 2.

Enzyme respon. i and reduction in hexobarbltai leep time and
dieldrin storage in female rats created with polychlorinatedbiphenyls,-triphenyla or DDT. *

PNA
Demeth.
%

Aniline
Oxldn*
7.

94

98

104

100

100

431

Treatment

EPN
Detox.
7.

Basal
+ Dieldrin, 1 ppm
+ DDT, 50 ppm

Hexobarb.
Sleep Time
-7.

Dieldrin
Storage
-7.

100

0

0

228

205

75.0

93.4

Trial A

+ 4B7. Cl-PCR,

50 ppm
100 ppm

225
362

408
568

193
295

34.6
57.7

60.7
77.5

+ 547. Cl-PCB,

50 ppm
100 ppm

444
475

555
672

309
346

65.4
80.0

85.8
91.6

+ 607. Cl-PCB,

50 ppm
100 ppm

475
544

291
368

231
273

78.8
80.0

93.4
96.5

+ 68% Cl-PCB,

50 ppm
100 ppm

600
525

272
242

266
240

83.7
80.0

96.5
96.0

Trial B
Baaal

92

114

114

+ Dieldrin, 1 ppm

100

100

100

0

0

+ DDT, 50 ppm

450

300

248

78.6

92.2

+ 217. Cl-PCB, 50 ppm
100 ppm

117
117

148
128

138
. 154

11.3
31.0

7.8
10.9

+ 327. Cl-PCB,

50 ppm
100 ppm

183
233

257
411

194
225

48.8
55.4

29.7
45.3

+ 42% Cl-PCB,

50 ppm
100 ppm

200
275

386
474

250
305

37.5.
65.0

37.5
64.7

+ 627. Cl-PCB,

50 ppm
100 PP«

625
717

403
474

293
312

85.7
87.5

94.5
96.5

1 Each value is Che mean percentage increase (or decrease) in the measured
response for groups of five rats compared to the groups that received the
basal diet plus only 1 ppm dieldrin.

HONS

063115

Table 3

Sex coi.. irisons in selected response oi Its to
various polychlorinated biphenyl materials. 1

PCB Treatment

Sex

BPS
Detox.
7.

PNA
Demeth.
%

Dleldrin
Storage
-7.

48% Cl, SO ppm
25 ppm

P
21

193
155

416
265

46.0
46.7

547. Cl, 50 ppm
25 ppm

F
M

293
225

497
341

74.7
76.6

60% Cl, 50 ppm
25 ppm

F
M

413
340

316
331

91.8
85.0

627. Cl t 50 ppm
25 ppm

F
H

433
408

286
363

91.5
90.0

687. Cl, 50 ppm
25 ppm

F
M

467
430

254
359

92.7
88.7

Data from rats fed diets containing 1 ppm dleldrin plus
each treatment at the indicated level. Each value is the
mean percentage Increase (or decrease) in the measured re­
sponse of groups of five rats compared to groups that receiv­
ed Che basal diet containing only 1 ppm dleldrin.

Table 4.

Comparisons of responses from polychlorinated biphenyls to poly­
chlorinated triphenyl materials equivalent in chlorination,*

EPN
Detox.
%

PNA
Demeth.
7.

Aniline
Oxidn.
%

Kexobarb.
Sleep Time
-7.

Dleldrin
Storage
-7.

427. Cl-PCB
427. Cl-PCT

200
238

386
208

250
145

37.5
46.2

37.5
73.6

68% Cl-PCB
62% Cl-PCB
65% Cl-iPCB'
I +
. PCT

600
625
519

272
403
249

266
293
231

85.6
85.7
66.3

96.4
95.0
95.5

Treatment

Data from female rats fed diets containing 1 ppm dleldrin plus each
treatment at a level of SO ppm. Each value la the mean percentage in­
crease (or decrease) in the measured response of groups of five rats com­
pared to groups Chat received the basal diet containing only l ppm
dleldrin.

MQNS

G63116

I

I

)

Figure 1. Reduction In hexobarbltal sleep
time In female rats after receiving PCB
materials in the diet (50 ppm) for 10 days.

Figure 2.
Increases In activity of liver
homogenates from female rets In degrading
EPN and p-nitroaniaole after receiving PCB
materials In the diet (50 ppm) for 15 days,
Data are presented as percentage increase
In activity relative to rata receiving the
basal diet containing only 1 ppm dieldrln.

HONS

063117

Figure 3. Reduction in dieldrin storage
in adipose tissue of female rats fed
diets containing I ppra dieldrin and
various PCB materials (SO ppm) for 15 days.

Figure 4. Dose response curves for various
microsomal enzyme inducing agents in terms
of stimulation of the in vitro degradation
of EPN by liver mitochondrial supernatant preparations. The Inducing agents were ad­
ministered in the dlete of female rats for
15 days.

MOMS

063118

Figure 5.
Electron capture gas chromatograms
of 627* Cl-PCB material and its residue recovered
from rat adipose tissue.

MQNS

063119