Carbon tetrachloride

Repeated dose toxicity: oral

Administrative data

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1986

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Scientifically sound study, but no individual data, limited biochemical and histological investigations.

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Data source

Reference

Reference Type:

publication

Title:

Oral toxicity of carbon tetrachloride: acute, subacute, and subchronic studies in rats.

Author:

Bruckner, J., V.; MacKenzie, W.,F.; Muralidhara, S.; Luthra, R.; Kyle, G., M.; Acosta, D.;

Year:

1986

Bibliographic source:

Fundamental and applied toxicology, vol. 6, p. 16-34

Materials and methods

Test guideline

Qualifier:

no guideline followed

Principles of method if other than guideline:

- rats (15 - 16 animals per dose, 4 doses) were treated with orally via gavage.
- dosing regime: 5 days a week
- blood samples were taken from 5 rats of each group at 2, 4, 6, 8, 10 and 12 weeks after study initiation, every rat was selected as blood donor 2 times with an interval of 6 weeks

GLP compliance:

no

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Carbon tetrachloride

EC Number:

200-262-8

EC Name:

Carbon tetrachloride

Cas Number:

56-23-5

Molecular formula:

CCl4

IUPAC Name:

tetrachloromethane

Details on test material:

- Name of test material (as cited in study report): carbon tetrachloride
- Analytical purity: analytical grade
- Supplier: Matheson, Colemn and Bell (Norwood, Ohio, U.S.A.)

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: TIMCO Breeding Laboratories (Houston)
- Weight at study initiation: 200-250 g (used for second experiment)
- Housing: in group of 5 or 6 animals in stainless-steel cages
- Diet (e.g. ad libitum): Ralston Purina Formulab chow, ad libidum
- Water (e.g. ad libitum): tap water, ad libidum
- Acclimation period: 2 weeks


ENVIRONMENTAL CONDITIONS
- Photoperiod (hrs dark / hrs light): 12 / 12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on oral exposure:

- Dose preparation: concentrations were adjusted so that required doses were applied at 1 mL/animal
- Vehicle: of Mazola corn oil from Best Foods CPC international (Engleufood Cliffs, N.J., U.S.A.)

Analytical verification of doses or concentrations:

no

Duration of treatment / exposure:

- 12 weeks

Frequency of treatment:

- 5 days a week

Doses / concentrations

Remarks:

Doses / Concentrations:
1, 10, 33 mg/kg bw
Basis:

No. of animals per sex per dose:

15-16, males only

Control animals:

yes, concurrent vehicle

Details on study design:

- blood samples were taken from 5 rats of each group at 2, 4, 6, 8, 10 and 12 weeks after study initiation, every rat was selected as blood donor 2 times with an interval of 6 weeks (see remark1 for details)
- 7 to 9 animals were killed after 12 weeks of treatment, the rest of the respective group (6-9 animals) were kept for additional 13 days recovery period (no treatment with substance or vehicle)
- liver and kidneys weighted at final necropsy
- histopathology of liver and kidney samples after final sacrifice (see remark 2)

Positive control:

- no

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: No data

DETAILED CLINICAL OBSERVATIONS: No data


BODY WEIGHT: Yes
Time schedule for examinations: twice weekly



FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No


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


OPHTHALMOSCOPIC EXAMINATION: No


HAEMATOLOGY: No


CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: after sacrifice
- Animals fasted: No
- How many animals: 5 per group
- Parameters checked in table 1 were examined.


URINALYSIS: Yes / No / No data
- Time schedule for collection of urine:
- Metabolism cages used for collection of urine: Yes / No / No data
- Animals fasted: Yes / No / No data
- Parameters checked in table [No.?] were examined.


NEUROBEHAVIOURAL EXAMINATION: Yes / No / No data
- Time schedule for examinations:
- Dose groups that were examined:
- Battery of functions tested: sensory activity / grip strength / motor activity / other:


OTHER: relative weight of organ/bodyweight determined for liver and kidneys

Sacrifice and pathology:

GROSS PATHOLOGY: No
HISTOPATHOLOGY: Yes, liver and kidney slices

Statistics:

- Students paired and unpaired t-test
- one way ANOVA

Results and discussion

Results of examinations

Clinical signs:

not examined

Mortality:

not examined

Body weight and weight changes:

effects observed, treatment-related

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

effects observed, treatment-related

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Histopathological findings: neoplastic:

not examined

Details on results:

- body weight development was reported only graphically: significant decrease only in the high dose group (becoming significant from day 30)

Effect levels

1

Dose descriptor:

LOAEL

Effect level:

10 mg/kg bw/day (nominal)

Sex:

male

Basis for effect level:

other: - biochemistry: moderate increase in SDH levels - histological findings: mild centrilobular vacuolization

2

Dose descriptor:

NOAEL

Effect level:

1 mg/kg bw/day (nominal)

Sex:

male

Basis for effect level:

other: - absence of significant effects

Target system / organ toxicity

Critical effects observed:

not specified

Any other information on results incl. tables

- table 2: Clinical Chemistry Data from Subchronic Study

Week of evaluation after study initiation	Daily dose of CCl4 (mg/kg)	Total dose of CCl4 (mg/kg)	OCT activity (mmol CO2/ml)	SDH activity (IU/ml)	GPT activity (IU/ml)	BUN (mg/dl)
2	0	0	43 ± 17	2.9 ± 0.7	23 ± 1	ND
	1	10	44 ± 13	8.3 ± 4.0	21 ± 1	ND
	10	100	31 ± 10	7.3 ± 1.5	25 ± 2	ND
	33	330	240 ± 56 ^d	111.5 ± 17.0 ^d	117 ± 28 ^d	ND
4	0	0	21 ± 6	2.6 ± 0.1	22 ± 1	15.6 ± 1.5
	1	20	18 ± 4	ND	23 ± 2	17.8 ± 1.2
	10	200	26 ± 3	2.9 ± 0.3	22 ± 1	18.5 ± 1.2
	33	660	140 ± 29 ^d	90.2 ± 14.7 ^d	75 ± 21 ^d	18.4 ± 0.5
6	0	0	55 ± 13	2.5 ± 0.5	10 ± 1	16.2 ± 0.2
	1	30	44 ± 5	2.4 ± 0.0	14 ± 2	15.4 ± 0.8
	10	300	40 ± 10	8.8 ± 0.8 d	16 ± 2	15.4 ±1.3
	33	990	251 ± 58 ^d	119.3 ± 16.9 ^d	82 ± 20 ^d	15.0± 0.8
8	0	0	18 ± 3	2.2 ± 0.2	21 ± 1	ND
	1	40	34 ± 9	1.6 ± 0.3	21 ± 1	ND
	10	400	30 ± 4	4.2 ± 1.5	19 ± 2	ND
	33	1320	151 ± 21 ^d	110.6 ± 10.0 ^d	130 ± 41 ^d	ND
10	0	0	28 ± 8	3.5 ± 0.4	18 ± 1	14.8 ± 0.5
	1	50	23 ± 3	2.3 ± 0.6	20 ± 1	16.6 ± 1.0
	10	500	55 ± 10	7.6 ± 2.5 d	23 ± 1	17.6 ± 0.5
	33	1650	148 ± 48 ^d	134.8 ± 15.0 ^d	617 ± 334 ^d	18.2 ± 2.5
12	0	0	45 ± 4	3.2 ± 0.4	20 ± 0.3	16.6 ± 0.7
	1	60	61 ± 12 d	1.9 ± 0.1	19 ± 1	17.1 ± 0.6
	10	600	69 ± 16	8.7 ± 2.0 d	27 ± 2 d	15.3 ± 1.2
	33	1980	247 ± 31 ^d	145.7 ± 57.9 ^d	502 ± 135 ^d	16.2 ± 0.8
14 ( 2weeks recovery phase)	0	0	169 ± 33	4.7 ± 0.5	20 ± 2	15.1 ± 0.5
	1	60	111 ± 25	3.4 ± 0.5	20 ± 1	16.8 ± 0.3
	10	600	169 ± 42	4.3± 0.8d	19 ± 1	16.7 ± 0.4
	33	1980	174 ± 24	6.0 ± 0.8 ^d	40 ± 7 ^d	14.6 ± 2.0

ND: not determined

d: significantly different from control at p0.05, paired I test

e: significant increase with increasing dose at p0.05, analysis of variance

- table 3: Severity of Liver Lesions in the Subchronic Studies

Time point of sacrifice after study initiation	Daily dose of CCl4 (mg/kg)	Lipid vacuolization	Nuclear and cellular polymorphism	Bile duct hyperplasia	Periportal fibrosis
12 weeks	0	0.0 + 0.0	0.0 + 0.0	0.0 + 0.0	0.0 + 0.0
	1	0.0 + 0.0	0.0 + 0.0	0.0 + 0.0	0.0 + 0.0
	10	3.7 + 0.7	3.5 + 0.3	0.0 + 0.0	0.0 + 0.0
	33	4.0 + 0.4	5.7 + 0.7	4.0 + 0.6	3.7 + 0.7
14 weeks	0	0.0 + 0.0	0.0 + 0.0	0.0 + 0.0	0.0 + 0.0
	1	0.0 + 0.0	0.0 + 0.0	0.3 + 0.3	0.0 + 0.0
	10	2.9 + 1.0	0.0 + 0.0	0.0 + 0.0	0.0 + 0.0
	33	1.4 + 0.7	2.6 + 0.7	3.4 + 0.8	2.9 + 0.8

Extent of lesions was graded on a scale of 0 (absent) to 8 (mean values of 7 – 9 rats)

Applicant's summary and conclusion

Conclusions:

The present study (Bruckner 1986) presents for CTC a LOAEL of 10 mg/kg bw in rats for subchronic repeated dose toxicity via oral exposure (12 weeks, treatment 5 d/week)

Executive summary:

The present study (Bruckner 1986) presents for CTC a LOAEL of 10 mg/kg bw in rats for subchronic repeated dose toxicity via oral exposure (12 weeks, treatment 5 d/week). Basis are biochemical parameters indicative for liver and kidney injury (moderate increase in SDH levels) and histological findings (mild centrilobular vacuolization). The corresponding NOAEL for subchronic oral toxicity is 1 mg/kg bw. The repeated dose toxicity of CTC (cited as carbon tertrachloride in the report) was evaluated in a 12 week oral toxicity study in rats following no official guideline. Male Sprague Dawley rats were treated with the test substance by oral gavage at 5 days a week. Approximately half of the animals of each dose group were killed at the end of the treatment period while the other half was kept for a recovery period of 13 days without any treatment before sacrifice. Blood samples were taken from 5 rats of each group at 2, 4, 6, 8, 10 and 12 weeks after study initiation, every rat was selected as blood donor 2 times throughout the study with an interval of 6 weeks. Three parameters of liver injury (OCT activity, SDH activity and GPT activity) and one parameter for kidney injury (blood urea nitrogen (BUN)) were determined. Liver and kidney lesions were analyzed histologically and graded with a scoring system. Body weights were recorded and relative organ weights of liver and kidney determined. No mortality was reported. Liver toxicity and nephrotoxicity parameters were not elevated for the two lower dose groups except for the SDH value in the mid dose group after 6 weeks and at the end of the treatment phase (fully reversible after the recovery period). In the high dose group lipid vacuolization, nuclear and cellular pleomorphism, bile duct hyperplasia and periportal fibrosis were apparent after the treatment period and only partly reversible within the recovery period). In the mid dose group (10 mg/kg bw) only lipid vacuolization was apparent and partly reversible within the recovery period. Based on these findings a LOAEL of 10 mg/kg bw (rat, subschronic (12 weeks), oral) can be derived. The corresponding NOAEL is 1 mg/kg bw.