Chronic toxicologic studies on isopropyl N-(3-chlorophenyl) carbamate (CIPC)

TOXICOLOGY

AND

APPLIED

Chronic

PHARMACOLOGY

Toxicologic

E. M. HENNIGAR,*

(1960)

Studies

N-(3Chlorophenyl) P. S. LARSON, GORDON R.

659-673

2,

on

Isopropyl

Cat-barn&e CRAWFORD,

H. B.

(CIPC)’

R.

BLACKWELL

HAAG,

AND

J.

SMITH, Ii.

Department of Pharmacology, Medical College of Virginia, Department of Pathology,* College of Medicine, State New York, Brooklyn 3 Received

June

JR.,

FINNEGAN? Richmond University

19 and of

24, 1960

Isopropyl N- (3-chlorophenyl) carbamate,” hereinafter referred to as CIPC, has been proposed for use as an herbicide in connection with the production of a number of food crops and for postharvest treatment of potatoes for control of sprouting. The present study was undertaken to ascertain what deleterious effects might be produced in rats and dogs through long-term consumption of CIPC in their diets, and to relate, if possible, the concentrations of this material in the diets with such effects as might be observed. Earlier toxicologic studies of CIPC made by Westrick et al. (1953) were concerned with the acute oral toxicity, and later studies by the same authors (1954) dealt with the effects of feeding this material to rats in their diets for a period of 90 days. They reported that the acute oral LDoo for rats lies between 5 and 7.5 g per kilogram body weight and for rabbits is approximately 5 g/kg. In their go-day study in which CIPC was administered to groups of 10 male rats at concentrations of 2.0, 0.5, 0.125, 0.031, and 0 ‘$ in a dry casein diet, they found that all the treated groups surpassed the control group in mean weight gain and food consumption. They further reported that the mean liver weights of the 2.0, 0.5, and 0.125 s groups were significantly increased over those of the control group, although histopathologic studies showed no liver abnor1 Supported under contract by the Market Marketing Service, United States Department 2 Deceased. 3 Furnished by the Columbia-Southern Pennsylvania. 659

Quality Research of Agriculture. Chemical

Division,

Corporation,

Agricultural

Pittsburgh

22,

660

P.

S.

LARSON

ET

AL.

malities; the mean kidney weights at each feeding level were not significantly different from that of the control group; and, finally, they observed minor kidney pathology in both experimental and control groups. More recently, van Esch ct al. (1958) have reported the production of skin tumors in mice by oral treatment with CIPC in combination with skin painting with croton oil. They noted that after a latency of about 90 days, there were many more tumors than in the control animals treated only with croton oil applications to the skin. They stated: ‘(The difference was in no case really significant according to the usual standards,” but added, “If the statistics are limited to the females, the difference is indeed significant (p = < 0.05 and > 0.02). . . . As, furthermore, many animals with CIPC had multiple tumors (one had even more than 20), we have the unmistakable impression that the combination of CIPC and croton oil is (also)-’ carcinogenic.” These authors found that most of the papillomas regressed completely when treatment of the skin with promoter (croton oil) was stopped but that a few of the remaining papillomas deteriorated into malignant tumors. One of 90 animals treated with CIPC and croton oil showed two malignant squamouscell carcinomas on the treated skin ,area as compared with none of 30 control animals treated only with croton oil. The authors concluded that this action of CIPC was weak as compared with that of urethan. METHODS

Two-Year Study in Rats Four groups of young albino rats consisting of 25 malesand 25 females with comparable littermates in each group were placed on diets containing these levels of CIPC for a period of two years: 0 (control), 0.02, 0.2, and 2.0 7’0. Finely ground Purina Dog Chow Kibbled Meal served as the basic diet. The 2.0 y0 CIPC diet was made by adding to 96 parts of meal 4 parts by weight of a 50 70 (w/v) solution of CIPC in corn oil. Using dilutions of this corn oil solution, the remaining diets were so made that in addition to the CIPC content, 2 parts by weight of corn oil were added. Thus all diets, including the control one, contained equal amounts of corn oil. The rats were individually caged and weighed once a week. Food consumption data on all rats for 3-day periods were collected at the end of 1, 3, 6, 12, and 24 months. 4 Refers to similar but currently with skin painting

more intense with croton

activity oil.

of urethan

administered

orally

con-

TOXICITY

OF ISOPROPYL

N-

(3-CHLOROPHENYL)

CARBAMATE

661

Hematologic studies were made at 3-month intervals on 5, or occasionally 10, rats of each sex at each dietary level. Values measured were hemoglobin, hematocrit, total and differential white cells. Pooled urine samplesfrom 5 rats of each sex at each dietary level were collected at 3-month intervals and tested for sugar by the Morris Anthrone method and for protein with Albutest Reagent Tablets@. These were supplemented in the later stagesby tests with Uristixm paper. One-Year Study in Dogs Eight male and 8 female purebred beagle dogs were used in the study. They were immunized against distemper, infectious hepatitis, and leptospirosis and were treated for intestinal parasites as indicated. The diets used were the same as in the two-year rat study, 2 male and 2 female dogs being placed on each diet at an average age of 6 months. The food, mixed with 90 g of water per 100 g, was offered to each dog once a day for an approximately one half-hour period and the amount consumed was recorded. A bone was given each dog once a week. The animals were weighed weekly. Control blood studies were made prior to placing the dogs on the diet and thereafter at 2 weeks, 1, 3, 6, and 12 months. The values recorded represented averages of 2 or 3 determinations made on separate days at each study period. Urine samplescollected prior to the experimental period and at monthly intervals during the experiment were tested semiquantitatively for sugar and protein by the proceduresusedin the rat study. RESULTS

Effect of Adding CIPC to the Diet of Rats for Two Years Average weight changes are summarized in Table 1. Growth was depressedamong both male and female rats receiving 2 % CIPC. Individual weights compared with those of controls by analysis of variance at 1, 5, 10, 26, and 52 weeks gave P-values of 0.01 or less in all cases. A further analysis at 78 weeks gave a P-value of less than 0.01 for the females, but the value did not reach significance for the males. Mortality data expressedas cumulative death totals are given in Table 2. An analysis of these data by the life table technique described by Sachs (1959) with sexes combined is given in Table 3. A significant difference from control animals (t = 2.499) appears during the last 13 weeks of the study among the rats receiving 2 $X0CIPC. In Table 4

.

57

2.0

62

2 .o

79a

98 99

100

74a

88 87

86

1 Wk

107

137 134

137

97

116 114

113

2 Wk

134

168 164

164

112

130 132

131

No

analysis

206a

242 245 235

156n

174 168

170

5 Wk

was

made

306”

348 344 343

199a

225 224 217

342

374 373 371

217

241 237 234

2840 512 520 510

25w 452 4.50 456

104.week

476a

326 329 321

283 280 279

401a

52 Wk

data.

YEARS

26 Wk

FOR Two

13 Wk

(g)

DIET

of the

of rats

10 Wk

weight

TABLE 1 CIPC IN THEIR

Average

RECEIVING

3 Wk

DATA ON RATS

a Value differs from controls by P = 0.01 or less. b Value does not differ significantly from controls.

62 62 62

0.02 0.2

0

57 58

0.02 0.2

Male

58

0

Female

Start

(70)

sex

lPW=l *.. 1I-

Dietary

GROWTH

463b

508 495 476

293a

373 342 354

78 Wk

461

468 459 456

294

350 336

336

104 Wk

.F

2

g

c E

?

v ’

-

-

0 0.02 0.2 2.0

0 0.02 0.2 2.0

Male

1

6 Wk

Female

Sex

Dietary level (7%)

SURVNAL

2 3 2 1

1

1

2 1 2

-

-

1

28 Wk

19 Wk

Cumulative

5 5 6 2

2 2

1

36 Wk

death

6 8 9 2

2 3 2 2

DIET

19

19 20 24

14

14 20 20

12 11

18 11

8

15

18

21 22

9

18

104 Wk 15

16

91 Wk

10 15

11

10 8 5

78 Wk

YEARS

10

5 5 3 7

65 Wk

weeks

FOR Two

at indicated

52 Wk

totals

TABLE 2 DATA ON RATS RECEIVING CIPC IN THEIR

664

I’.

ESTIMATED

PROPORTION

S.

LARSON

ET

TABLE

3

OF ORIGINAL

GROUPS

TO END Period (weeks)

0 ‘j% Diet

0.02

‘5% Diet

98.00 9600

iz 1.9Sb k 2.77

98.CO

2

1.98

96.00

i

2.77

39 52

88.00

-+ 4.59

84.00 72.00

& 5.18 & 6.35

86.00 80.00

91

58.00 40.00

zk 6.98 rt 6.92

104

26.00

t

a Calculated b Percent

6.19

at 13-week survival

(SEXSS

COMBINED)

SURVIVINI:

OF PERIODS

13 26

65 78

AL.

0.2 5% Diet

2.0 ‘$h Diet

98.00 96.00

& 1.98 & 2.77

98.00

AZ 1.98

98.00

2

AI 4.91 -r- 5.66

84.00

‘-c 5.18

78.00

c

5.86

96.00 94.00

Z 2.77 & 3.26

72.CO

& 6.35

68.00

2

6.60

70.00

I? 6.48

64.00

& 6.78

7.02 6.99

zb 7.06

k r

k k

56.00

44.00 26.00

56.00 42.00 18.00

k

5.44

32.00 s.00

Z 6.59 t 3.84

7.02 6.21

1.98

intervals.

?z standard

deviation.

similar data are presented according to sex. Here a significant difference (t = 2.065) appears during the last 13 weeks of the study only among male rats receiving 2 7. CIPC, the corresponding t for females being 1.441. Food consumption data collected at the stated intervals during the study are summarized in Table 5. While not uniform, there would appear to be some tendency for the rats receiving 2 s CIPC to have a somewhat increased food consumption per kilogram body weight. An estimate of the adequacy of this determination was arrived at by calculating the correlation coefficient between the weight of food that disappeared and body weight for each animal on each dietary level at the l-month point. These values in order of increasing dietary concentration were for females 0.823, 0.619, 0.600, and 0.520, and for males 0.688, 0.817, 0.538 and 0.614. At the degrees of freedom involved, all these values are significant to below the 1 $% level. In hematologic studies only the hemoglobin and hematocrit values showed effects possibly due to treatment. An indication of a relative depression in these values appeared to exist in the third month data for rats receiving 2 “/o CIPC, particularly male rats. This impression persisted, although at times in a somewhat erratic manner, throughout the remainder of the study. Studies of the urine samples collected revealed no effects of treatment throughout the feeding period with the possible exception that the rats receiving 2 ‘$ CIPC tended to show a less positive test for protein. Organ to body weight ratio data obtained on rats which survived the

13 26 39 52 65 78 91 104

Period (weeks)

100.00 96.00 92.00 80.00 60.00 44.00 28.00

?I -r-c A ? k L-

0.00 3.92 5.43 8.00 9.78 9.94 8.99

100.00 _c 0.00

0 70 Diet

lCO.00 396.00 -+: 92 .OO 2 88.00 2 84.00 -c 68.00 k 40.00 z!z 25.00 t

0.00 3.92 5.43 6.50 7.33 9.32 9.80 8.99

0.02 70 Diet

Female

ESTIMATED

_’ -c 2 k k k C &

0.00 3.92 5.43 5.43 5.43 8.00 9.60 7.33

0.2 % Diet 100.00 96.00 92.00 92 .OO 92.00 80.00 64.00 16.00

rats

PR~PORTIOK

100.00 100.00 100.00 96.00 72.00 56.00 40.00 12.00

2.0 s A & & -c ? 2 c 2

4 GROUPS

0.00 0.00 0.00 3.92 8.96 9.92 9.79 6.51

Di-t

TABLE OF ORIGINAL

96.00 92.00 80.00 75.79 61.58 53.88 42.33 23.09

-+& f. 2 t I?I -I 2

TO END

3.92 5.43 8.00 8.56 9.61 9.86 9.74 8.29

0 70 Diet

SURVIVING

96.00 96.00 80.00 72.00 60.00 60.00 48.00 24.00

0.02 s _’ 3~ -t zk 2 k -If

Male

3.92 3.92 8.00 9.00 9.78 9.78 9.98 8.78

Diet

OF PERIOD

96.00 92.00 76.00 64.00 44.00 32.00 20.00 20.00

-t e 2 k k k 2 _’

3.92 5.43 8.53 9.60 9.92 9.33 8.00 8.CO

0.2 70 Diet

rats

96.00 96.00 92.00 92.00 68.00 56.00 24.00 4.00

4 f. 51 & -c-t k f

3.92 3.92 5.43 5.43 9.32 9.91 8.54 3.92

2.0 70 Diet

TOXICITY

OF ISOPROPYL

iv-

(&CHLOROPHENYL)

667

CARBAMATE

full two years on the diet are summarized in Table 6. No trend is apparent at dietary levels through 0.2 %. The interpretation of the data with respect to the 2.0 5%dietary level is complicated by the small number of survivors in these groups. TABLE 6 ORGAN TO BODY WEIGHT RATIO DATA ON RATS RECEIVING IN THEZR DIET FOR Two YEARS

Sex Female

Male

Dietary level (%)

No. rats

0 0.02 0.2 2.0

7 7 4 3

33.0” 5.1 31.9 & 4.1 32.8 I+ 4.5 41.6 -r- 11.8

0 0.02 0.2 2.0

6 6 5

33.2 t 31.0 c 31.1 2 48.5

1

Organ Liver

to body Kidney

4.3 6.4 4.0

1.1

8.5 k 1.7 5.3 k 2.6 10.4 2 5.1 8.9 k 2.4 8.0 k

9.6

ratio

k S.D.

Heart

8.0 2 1.6 7.5 &

weight

1.1

3.5 3.5 3.7 4.1

-c 2 f k

CIPC

(X

Spleen 0.6 0.3 0.8 1.0

2.2 ” 0.4 2.0 2 0.3 3.3 IL 0.8 8.425.4

3.3 & 0.6 3.4 -c 0.6 3.3 2 0.3 4.0

2 .o ” 0.7 2.3 rt 0.6 2.6 -t 0.8 15.7

103) Testes 6.7 c 0.9 6.9 c 2.2 5.6 =!I 1.6 9.9

Histopathologic studies were done on animals dying during the study, in which autolysis did not preclude some interpretation, as well as on all survivors of the two-year period. The tissues studied included heart, lung, liver, kidney, gastroenteric, spleen, adrenal, pancreas, thyroid, gonad, bone marrow, bladder, skeletal muscle, skin, and brain. An unusually high incidence of infectious diseaseof the respiratory tract was evident. However, the distribution of these and the other lesions seen did not bear any apparent relationship to treatment. Tumor incidence is detailed in Table 7. Effect of Adding CIPC to the Diet of Dogs for One Year Weight change data are given in Table 8. All animals, the controls as well as those given the experimental diets, lost weight initially. At the end of the 12-month period, the animals on the control, 0.02, and 0.2 % CIPC diets showed group average weight gains of approximately 17, 15, and 6 $, respectively, over the starting group average weights. Those on the 2.0 % CIPC showed a group average weight loss of 7 70 after 12 months as compared with the starting group average for these animals. The maximum group average weight losses,all of which occurred within the first 4 weeks, were approximately 8, 9, 12, and 16 % for the groups fed diets containing 0, 0.02, 0.2, and 2 % CIPC, respectively.

668

P.

S. LARSON

ET

TABLE TUMOR

INCIDENCE

IN RATS

RECEIVING

AL.

7 CIPC

IN THEIR

DIET

FOR Two

YEARS

Sex

Dietary cont. (%)

No. of rats”

Female

0

22

Subcutaneous fibromas in 1 rat. Large fibromatosis of mammary gland in 1 rat. Subcutaneous fibroma in neck and in pelvic area in 1 rat. Malignant lymphoma of lungs, liver, kidney, and spleen in 1 rat

0.02

24

Fibrous adenomatosis of lung in 1 rat. fibroma in pelvic area in 1 rat

0.2

22

Adenoma of thyroid in 1 rat. Malignant lymphoma of liver and kidneys with infiltration into spleen, ovaries, and bone in 1 rat. Tumor in pelvic region (not diagnosed) in 1 rat

2.0

25

Granulosa

0

24

Lymphosarcoma cystic reticulum in 1 rat

0.02

23

Xanthogranulomatous

0.2

24

Subcutaneous tumor (not diagnosed) in 1 rat. Malignant lymphoma of mediastinum with lung invasion in 1 rat

2 .o

24

Large lymphosarcoma attached to the spleen in 1 rat. Leiomyosarcomata filling the abdominal cavity with invasion of heart muscle in 1 rat. Inflammatory papillary adenomatosis of lung involving the mediastinum in 1 rat

Male

n In the remaining

Tumor

rats, gross examination

cell tumor of

revealed

incidence

of ovary

Subcutaneous

in 1 rat

lung in 1 rat. Undifferentiated cell sarcoma ( ?) in neck region mass in mediastinum

in 1 rat

no tumors.

Food consumption data are summarized in Table 9. Amounts consumed by the animals on the 2.0 % CIPC diet reflect the weight change data given above. No fatalities occurred and all dogs appeared to be in general good health throughout the experimental period. The only positive hematologic findings, relatively lower hemoglobin and hematocrit values noted in the l-month through the 6-month observations on the dogs receiving the 2 $ CIPC diet, were not apparent in the observations made at the end of the twelfth month. The urine samples collected at monthly intervals did not reveal any

littermates.

6.95

10.6 7.6

7.1

11.6

8.2

M-61”

M-12”

8.8 8.5 9.3

F-7”

0 Superscripts

9.0

7.6 10.4

M-622 M-134

indicate

7.6 10.4

Averages

8.55

F-19: F-83

2 .o

M-154

M-592

7.6

F-18”

8.1

7.15 8.85

6.6 9.8

7.5

7.6 7.5 8.45

6.5

6.7

6.25

7.5

F-81’

F-63”

6.7

7.4

7.8

7.9

7.2 8.75

6.5 9.3

7.8

7.95 7.85 9.0

6.35

7.45

10.35

6.6

6.8 6.05

7.2

8.15 8.10

8.0 8.0

M-144

M-602

8.6 9.0

2 Wk

TABLE RECEIVING

6.35 6.15

6.7 7.0

F-801 F-10”

1 Wk

DATA ON Dots

6.35 6.35

Start

Sex and no.a

Averages

0.2

Averages

0.02

Averages

0

Dietary level (%)

BODY WEIGHT weight

8.2

8.05

8.3

8.9

6.85 9.3

8.5

7.6

6.3

5.95

8.2 8.55

8.2

8.0

9.6

8.6

8.45 9.85

7.8 8.4

9.6

10.55

9.45 10.3

8.45 8.85

8.4 9.2

9.4

9.4

8.05

9.3

12.35

8.4

9.1 7.3

9.3

8.9

9.7

9.65 7.95

8.15

8.4

7.75 9.8

7.9s

7.95

9.1

10.5

10.8 9.5

8.9 9.8

7.35

9.4

13.65

8.35

7.1

8.45

9.1

9.95 9.8

9.1 7.65

52 Wk

9.55 10.3

7.4

9.7

13.5

8.45

8.85 7.9

9.6

10.55

Wk

10.15

39 9.65 8.05 10.02

Wk 9.2 7.85 9.85

26

YEAR

6.95

8.2

7.5 10.75

6.45

8.0

8.3

6.95 8.75

8.0 8.4

7.0

7.7

10.1

7.0

7.8

6.1

7.9

7.85 8.35

6.45

7.8

10.40

7.05

7.65 6.15

7.85

8.7

9.25

9.15

8.9

8.75

13 Wk

(kg)

FOR ONE

1 7.95 7.0

8 Wk

DIET

7.3 6.6

Body

IN THEIR

7.15 6.65 8.85

4 Wk

8 CIPC

^%

?

5

s s

% ;: %

2

;;

670

P.

S.

LARSON

ET

T.4BLE FOOD

Dietary level

(%I 0

CONSUMPTION

Sex and no.

IN THEIR

DIET

by end of indicated

week

(kg) 11

10 Wk

20 Wk

30 Wk

40 Wk

7.7

15.4

31.8

49.0

67.4

97.2

2.5

4.0

16.4

34.4

51.4

88.9

M-60

3.3

9.6

19.0

38.9

104.7

M-14

3.0

8.4

16.4

34.0

59.8 50.7

63.1 78.7 67.0

88.7

2.83

8.30

16.8

34.8

52.8

69.6

94.9

2.7 2.0

8.4 7.0

17.1 15.8

35.4

56.4

75.0

55.1

73.5

102.6 96.4

54.5

72.7

9i.7

63.2

88.5

122.4

F-81 F-63

52 Wk

M-61

2 .o

7.2

14.9

M-12

2.9

8.0

17.3

35.6 34.5 39.5

2.40

7.65

16.3

36.3

j7.3

77.4

101.8

F-18

1.7

5.3

10.6

23.5

37.6

6?.2

F-7 M-59 M-15

1.9 2.5

7.6 8.2

15.3 16.2

33.9

51.0

48.5 65 3

35.4

70.6

88.6 94.1

2.8

9.4

18.5

37.3

53.9 54.4

71.0

95.6

2.23

7.63

15.2

32.5

49.2

63.9

8i.l

F-19

1.3

F-8

1.9

3.9 5.2

8.9 10.2

21.6 22.9

35.5 40.5

47.3 62.1

64.8 93.1

M-62 M-13

1.4 1.9

5.0

12.5 14.1

25.4 30.4

35.4 48.3

50.0

65.1

6.1

64.0

89.;

1.69

5 .OS

11.4

23.1

40.7

55.9

78.9

Average

Average u Corrected

consumption

CIPC

5 Wk

Average

2 .o

food

9 RECEIVING

2.5

.4veraae

0.2

Cumulative

ON Dots

2 Wk

F-80 F-10

0.02

DATA

AL.

for added

water.

difference as regards protein and sugar content among the control and the treated groups. Organ to body weight ratio data collected on sacrifice of the dogs after 12 months on diet are given in Table 10. The animals were anesthetized with pentobarbital i.v., exsanguinated via a carotid artery, and the chest opened. Liver ratios for the dogs receiving the 2.0 s diet appear to be moderately increased and the spleen ratios definitely so. Tissues taken for histopathologic examination corresponded to the recommendations of the Armed Forces Institute of Pathology. The findings as regards lesions possibly related to CIPC intake were negative except for splenic congestion in 2 dogs fed the 2.0 s CIPC diet. This is consistent with the spleen to body weight ratio data noted above.

Average

Average -2.0

.4verage 0.2

0.02

Average

__

8.1

8.2

~

8.8

M-62

M-13

8.S

F-8

F-19

.___-__-

8.1

M-15

7.9 7.1

7.5 8.0 8.7 7.7 7.0

M-12

F-18 F-7 M-59

7.2 8.7 8.6

_____

8.4 7.8

F-81 F-63 M-61

M-14

7.7

8.6

Heart

F-10 M-60

.-~

6.6

___

F-80

(%)

0

WEIGHT

Sex and no.

TO Born

Dietary level

ORGAN

39

40 38

38

30 40

23 28 33 28 30 29

32 29 28

30.5

30

30 30

32

Liver

RATIO

__

-

7.5

13.5 8.8

3.7

2.8 4.1

2.6 3.1 3.1

2.1 2.3

1.8

2.1 2.5 2 .o

2.7

2.4

3.1 2.2

3.0

TABLE

-~

Organ

2 .5 3.7

2.1

2.7

3.0 2.9

2.8

2.2

2.4

2.5

2.4 2.3

2.0 2.7 2.5

2.7 2.8 3.2

2.75

2.7

10

to body

CIPC

2.75

3.0 2.9

2.8

2.45 2.3

2.1 2.2 2.7

2.1 2.7 2.8

2.5 2.9 3.4

2.8 2.7

3.5

2.5

2.1

L. Kidney

RECEIVING

R. Kidney

ON DOGS

Spleen

DATA

x

-

1.2 1.3

0.042

0.056

1.4

0.036 0.99

0.049

0.86

1.5

0.104 0.050

1.4

0.088 1.2

0.084

R. Gonad

weight

IN THEIR

-

1.0

1.3

0.052

0.048

1.3

1.1

0.048

0.064

0.84

0.078 0.070 1.5

1.4

0.118 1.3

0.056

0.071

,081

X60

,081

.C6 7 ,061

.087 ,056 .06 1

,070 ,062

,060

,077 .083 .C60

.067

,061

.C67

.C65

0.075

R. Adrenal

FOR 12 MONTHS

L. Gonad

___.__

103

DIET

.___

0.070

,081

.056

.080

,063 ,061

.060 ,070 _____ ,060 ,087 ,048 ,058

,059

.083 .077

,065

.05 7

,064

,059

0.080

L. Adrenal

-

ii

&

E G

s -

2

z

g

6 s

&

;

z

g

z

8 Y

z e

2

672

P. S. LARSON

ET

AL.

DISCUSSION

The studies by Westrick et al. (1953, 1954) indicate that CIPC has a low order of acute toxicity when given orally to rats and rabbits, LD;,,‘s of 5-7 g/kg and 5 g/kg, respectively, being observed. When fed CIPC for 90 days at dietary levels of 0.125 5& and higher, the experimental rats exhibited significantly increased mean liver weights as compared with controls; kidney weights were not altered. Treatment-related pathologic changes were not seen. The present extended feeding studies in rats (two years) and dogs (one year) indicate similarly a relatively low chronic oral toxicity. In rats, weight gains were significantly depressed by dietary levels of 2.0 ‘j% but not by the lower levels fed. Mortality was significantly increased, as compared with controls, only among the male rats on the 2.0 % level during the last 13 weeks of feeding. Relatively lower hemoglobin and hematocrit values likewise were observed only among rats on the 2.0 % level; and urine studies were negative save for a tendency toward a less positive test for protein at this level. Relatively greater liver- and spleen-body weight ratios were observed only among the 2.0 ‘/( animals. Histopathologic studies revealed no treatment-related lesions at any feeding level. In dogs, adverse effects on body weight, as compared with the controls, were seen only at the 2.0 $% level. All animals showed initial weight loss, but only the 2.0 7” animals failed to surpass the average starting weight for the group during the experimental period. There was no mortality or apparent morbidity. Urine studies were negative. Hematologic studies showed relatively lower hemoglobin and hematocrit values through the sixth month on the 2 yO, but not at the end of the 12-month feeding period. As in the rats, increased liver- and spleen-body weight ratios were seen at the 2.0 y0 level, the latter being in keeping with the observation of splenic congestion at autopsy. No other pathologic changes were seen. The report of van Esch et al. (1958) that CIPC and croton oil administered concurrently in mice. may produce skin tumors is interesting; but the relationship of this findin, e to dietary carcinogenesis, if any, is obscure. In the present study, the extended feeding of CIPC produced no tumors in rats or dogs. SUMMARY The feeding of isopropyl N-(3-chlorophenyl) carbamate to rats for two years and to dogs for one year at dietary concentrations of 0.2 % or below produced no At a dietary concentration of 2.0 %, adverse effects discernible adverse effects. occurred in both rats and dogs.

TOXICITY

OF ISOPROPYL

A;- (3-CHLOROPHENYL)

CARBAMATE

673

REFERENCES SACHS, R. (1959). Life table technique in the analysis of response-time data from laboratory experiments on animals. Toxicol. Appl. Pharmacol. 1, 203-227. VAN ESCH, G. J., VAN GENDEREN, H., and VINX, H. H. (1958). The production of skin tumors in mice by oral treatment with urethane, isopropyl-l\r-phenyl carbamate or isopropyl-N-chlorophenyl carbamate, in combination with skin painting with croton oil and Tween 60. Brit. /. Cancer 12, 355-362. WESTRICK, M. L., GROSS, P., and SCKRENK, H. H. (1953). Acute oral toxicity of isopropyl N-(3chlorophenyl)-carbamate. Unpublished data. Mellon Institute, Pittsburgh. WESTRICK, M. L., GROSS, P., and SCHRENX, H. H. (1954). Toxicity of isopropyl N-(3-chlorophenyl) -carbamate: ninety-day feeding experiments. Unpublished data.