Registration Dossier

Toxicological information

Developmental toxicity / teratogenicity

Currently viewing:

Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Study along OECD GL 414, 2 doses only, exposure gestation days 6 - 15, no evaluation details given.

Data source

Reference
Reference Type:
publication
Title:
Unnamed
Year:
1974

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
GLP compliance:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): carbon tetra chloride
- Lot/batch No.: Lot No. 9256, Burdick & Jackson Lab. Inc., Muskegon, Michigan

Test animals

Species:
rat
Strain:
Sprague-Dawley
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Spartan
- Age at study initiation:
- Weight at study initiation: average 250 g
- Fasting period before study: not reported
- Housing: wire bottom cages
- Diet (e.g. ad libitum): Purina Rat Chow, Ralston Purina Co. St. Louis, Missouri
- Water (e.g. ad libitum): ad libitum
- Acclimation period: not reported


ENVIRONMENTAL CONDITIONS
- Temperature (°C): controlled, temperature not reported
- Humidity (%): controlled, exact humidity not reported
- Air changes (per hr): not reported
- Photoperiod (hrs dark / hrs light): controlled, exact light cycle not reported


Administration / exposure

Route of administration:
inhalation: vapour
Type of inhalation exposure (if applicable):
whole body
Vehicle:
unchanged (no vehicle)
Details on exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: 3.7 m³ stainless steel, cubical dynamic exposure chamber
- Method of holding animals in test chamber: not reported
- Source and rate of air: not reported
- Method of conditioning air: not reported
- System of generating vapours: by metering the liquid at known rates into a temperature controlled evaporating flask and diluting this vapours with filtered room air at a rate calculated to give the desired concentration. The nominal concentration of each compound in the chamber atmosphere was calculated from the ratio of the material delivery rate to the rate of total air flow through the chamber.
- Temperature, humidity, pressure in air chamber: not reported
- Air flow rate: not reportes
- Air change rate: not reported
- Method of particle size determination: not applicable
- Treatment of exhaust air:


TEST ATMOSPHERE
- Brief description of analytical method used: 3 times daily using a Beckman IR10 infrared spectrophotometer and in addition by using combustion conductivity analysis
- Samples taken from breathing zone: yes


VEHICLE (if applicable)
- no vehicle used
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The analytical concentration was determined by analyzing the chamber atmosphere 3 times daily using a Beckman IR10 infrared spectrophotometer with a multipath gas cell having a variable path length of 1-10 m and a volume of 4.5 liters. In addition, the concentration in the chambers was continuously monitored using combustion conductivity analysis to assure the absence of significant deviations from the desired level.
Details on mating procedure:
- Impregnation procedure: cohoused
- If cohoused:
- M/F ratio per cage: not reported
- Length of cohabitation: not reported.
- Further matings after two unsuccessful attempts: not reported
- Verification of same strain and source of both sexes: not reported
- Proof of pregnancy: sperm in vaginal smear referred to as day 0 of pregnancy
- Any other deviations from standard protocol: none reported
Duration of treatment / exposure:
7 hours
Frequency of treatment:
daily from gestation day 6 to 15
Duration of test:
in total 20 days (pregnancy duration)
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
300 and 1000 ppm (2137 and 6425 mg/m3).
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
334 and 1004 ppm (2137 and 6425 mg/m3).
Basis:
analytical conc.
No. of animals per sex per dose:
litter size 22 -23
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: not reported
- Rationale for animal assignment (if not random): not reported

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: No data
- Time schedule: not reported


DETAILED CLINICAL OBSERVATIONS: No data



BODY WEIGHT: Yes
- Time schedule for examinations: not reported


FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
- Food consumption of each animal was measured at 2-day intervals throughout the experimental period.


WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No


POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 20
- Organs examined: in addition to the reproductive organs: livers, possibly others as well, but not reported

Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
Fetal examinations:
- External examinations: Yes, but not detailed
- Soft tissue examinations: Yes, but not detailed
- Skeletal examinations: Yes, but not detailed
- Head examinations: Yes, but not detailed
see table 2 for details
Statistics:
Fisher Exact Probability test and ANOVA + Dunnett's test
Indices:
not calculated
Historical control data:
not reported

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
- Feed consumption of rats at both dose levels was reduced throughout the 10 day exposure period (see Tables 3 and 4 of attached document).
Body weight gain was reduced in a dose dependent manner in both treated groups.
- Hepatotoxicity is reflected by significant increases in SGPT activities during all exposure days, but not thereafter (see Table 5 of attached document). Gross changes in the appearance of the liver characteristic of CTC intoxication (pale, mottled livers) were evident immediately following the last exposure but were not evident 6 days later. The relative liver weight of exposed rats was significantly increased at both doses while absolute liver weight was not always (see Table 6 of attached document).
- There were no gross observations during exposure on demeanor or or signs of toxicity.

Effect levels (maternal animals)

Dose descriptor:
LOAEC
Effect level:
2.11 mg/L air (analytical)
Basis for effect level:
other: maternal toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
- Reproductive parameters (see Table 1 of attached document):
Exposure to Carbon terachloride had no effect on reproductive parameters except for reduced foetal body weight and crown-rump length.
- Foetal examinations (see Table 2 of attached document):
A significant incidence of subcutanous edema was observed at 300 ppm but not at 1000 ppm. The incidence of sternebral anomalies was significantly increased in the foetuses exposed to 1000 ppm.

Effect levels (fetuses)

open allclose all
Dose descriptor:
NOAEC
Effect level:
2.11 mg/L air (analytical)
Basis for effect level:
other: teratogenicity
Dose descriptor:
LOAEC
Effect level:
2.11 mg/L air (analytical)
Basis for effect level:
other: fetotoxicity

Fetal abnormalities

Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Any other information on results incl. tables

- Analytical concentration: in inhalation chambers: mean of 334 ± 48 and 1004 ± 90 ppm for the 300 and 1000 ppm groups, respectively.

- Table 1: Effect of inhaled carbon tetrachloride on observations made at Cesarean Section

Statistic

Air control

Carbon

tetrachloride

CTC concentration

--

334 ppm

1004 ppm

No. of litters

43

22

23

% pregnancy

91(43/47)

92(22/24)

82(22/28

Corp. lutea/dam

14±2

14±2

14±3

Implantation sites /litterb

11±3

13±2

10±4

Live fetuses/litterb

11±3

12±2

9±4

% resorptions/impl. sites

7(34/492)

6(16/284)

10(25/239)

% litters with resorption

44(19/43)

41(9/22)

48(11/23)

Litters totally resorbed

0/43

0/22

1/23

Resorptions/litter with resorptions

1.8(34/19)

1.8(16/9)

2.3(25/11)

Sex ratio, M/F

56:44

55:45

55:45

Fetal body weight (g)c

5.64±0.34

5.29±0.34e

4.96±0.68e

Fetal crown-rump length (mm)c

43.7±1.0

42.2±1.0c

41.8±2.2e

b Mean ± SD

c Mean of Litter means ±SD

e Significantly different from control by an analysis of variance and Dunnett’s test, p<0.05

- Table 2: Effect of inhaled carbon tetrachloride on the incidence of fetal anomalies among rat litters

Air control

Carbon

334 ppm

Tetrachloride

1004 ppm

Number of litters examined

22

22

22

% of Litters affected (No. of litters)

Gross

(0)

(0)

(0)

Skeletal

Skull anomalies (delayed ossific.)

12(5)

9(2)

9(2)

Lumbar ribs or spurs

24(10)

41(9)

27(6)

Vertebral anomalies (bipartite centra)

21(9)

27(6)

14(3)

Sternebral anomalies (unpooled controls)

61(14)

68(15)

(bipartite; delayed ossific.) (unp. cont.)

11(2)

59(13)c

Total skeletal

58(25)

91(20)c

68(15)

Soft tissue

Subcuatneous oedema

33(14)

59(13)c

50(11)

Dilated Ureters

12(5)

14(3)

5(1)

Total soft tissue

51(22)

68(15

59(13)

c Incidence significantly different from control by the Fisher Exact Probability test, p<0.05

- Table 3: Effect of Carbon tetrachloride on rat maternal feed consumption (g/rat/day ± SD)

Feed consumption on day

n

4-5

6-7

8-9

10-11

12-13

14-15

16-17

18-19

Control

43

20±3

20±3

20±3

23±2

23±2

23±3

25±5

27±3

300 ppm

22

18±2

12±3c

13±3c

13±3c

16±4c

18±4c

23±3

NDb

1000 ppm

23

19±3

9±3c

13±3c

14±3c

12±4c

12±5c

23±7

28±7

ND not determined

c Significantly different from control by Dunnett’s test p<0.05

- Table 4: Effect of Carbon tetrachloride on maternal rat body weight (g, mean ± SD)

Control

334 ppm

1004 ppm

No. of dams

43

22

23

Gestation day

6

283±17

290±21

275±19

13

317±18b

281±25b

253±22b

21

397±24b

368±33b

336±57b

b Significantly different from control by Dunnett’s test, p<0.05

- Table 5: Effect of inhaled Carbon tetrachloride on SGPT activiy in rats (SE units ±SD)

Day of exp.

N

Control

334 ppm

1004 ppm

2nd

10

63±3

159±20b

NDc

4th

10

75±2

154±21b

81±5b

7th

10

57±

218±51b

ND

10th

10

56±2

241±68b

154±11b

6 days after last exp.

Non-pregnant

6

57±3

52±5

46±4

Pregnant

10

72±3

62±3

ND

b Significantly different from control by Tukey’s test, p0.05

ND not determined

- Table 6: Effect of inhaled Carbon tetrachloride on absolute and relative liver weights

Parameter

Control

334 ppm

1004 ppm

Following last exp., nonpregnant (n04)

Abs. liver weight, g, mean±SD

9.0 ±0.6

10.0 ±1.4

10.4 ±1.5

Six days after last Exposure, nonpregnant, (n=6)

Abs. liver weight, g, mean±SD

9.3 ±0.8

9.9 ±1.9

11.4 ±1.4

Six days after last Exposure, pregnant, (n=)

Abs. liver weight, g, mean±SD

14.7 ±1.2
(43)

14.9 ±2.1
(22)

14.8 ±2.4
(23)

Following last exposure, non pregnant (n=4)

Rel. liver weight, mg liver/g bw, mean±SD

34 ±1

43 ±3b

49 ±4b

Six days after last exposure, nonpregnant, (n=6)

Rel. liver weight, mg liver/g bw, mean±SD

34 ±3

39 ±5

48 ±3b

Six days after last exposure,
(n=
  )

Rel. liver weight, mg liver/g bw, mean±SD

37 ±3
(43)

40 ±3b
(22)

45 ±6b
(23)

b Significantly different from control by Dunnett’s test, p< 0.05

- Table 7: Summary of the toxicity of inhaled Carbon tetrachloride to pregnant rats

334 ppm

1004 ppm

Embryo- and fetotoxicity

Resorptions

--

--

Fetal body measurements

dec

dec

Incidence of:

Gross anomalies

--

--

Skeletal anomalies

inc

inc

Soft tissue anomalies

inc

--

Maternal toxicity

Weight gain during gestation

dec

dec

Feed consumption

dec

dec

Conception rate

--

--

No. of implantations

--

--

Litter size

--

--

SGPT activity

inc

inc

Absolute liver weight

--

--

Relative liver weight

inc

inc

Gross liver pathology

inc

inc

Demeanor

--

--

Applicant's summary and conclusion

Conclusions:
Inhalation exposure of pregnant rats to carbon tetra chloride at levels of 300 and 1000 ppm resulted in maternal toxicity and retarded foetal development. No teratogenic effects were reported.
Executive summary:

Sprague Dawley rats were exposed by inhalation to 300 or 1000 ppm carbon tetrachloride for 7h/day on days 6-15 of pregnancy. During the dosing period, food consumption by the dams was markedly reduced, resulting in a mean decrease in body weight of 7% (300 ppm) and 15% (1000 ppm) by day 21 in comparison with controls. Evidence of maternal hepatotoxicity was seen in both groups; serum glutamic-pyruvic transaminase (SGPT) was significantly elevated during exposure but had returned to normal by day 21 when relative liver weights were significantly increased but absolute weights unchanged. There was no statistically significant effect on resorptions though 1/23 litters was fully resorbed in the 1000 ppm group. No gross external abnormalities were seen in any group. The data on internal and skeletal anomalies is estimated difficult to evaluate: only information on the number and percentage of litters affected is given, with no data on the numbers of fetuses affected. However, no significant increases in anomalies are reported, except for subcutaneous oedema in the 300 ppm group and sternebral anomalies in the 1000 ppm group. These increases are unlikely to be of any biological significance since oedema was not significantly elevated in the 1000 ppm group and the incidence of sternebral anomalies varied considerably in the two control groups. Fetal body weight and crown-rump length were significantly decreased in a dose related manner but this is not unexpected in view of the severe effect on food consumption in the dams.