1-chloro-1,1-difluoroethane

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian germ cell study: cytogenicity / chromosome aberration

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP-compliant near-guideline study, available as confidential study report and published in peer-reviewed literature, no restrictions, fully adequate for assessment (SIDS score: 1a)

Data source

Reference

Reference 1

Reference Type:

study report

Title:

Unnamed

Year:

1983

Report date:

1983

Reference 2

Reference Type:

publication

Title:

Toxicological evaluation of hydrochlorofluorocarbon 142b

Author:

Seckar JA, Trochimowicz HJ, HoganGK

Year:

1986

Bibliographic source:

Fd Chem. Toxic., 24, 237-240

Materials and methods

Principles of method if other than guideline:

Bone-marrow cytogenetic assay

GLP compliance:

yes

Type of assay:

chromosome aberration assay

Test material

Test material information

Constituent 1

Reference substance name:

1-chloro-1,1-difluoroethane

EC Number:

200-891-8

EC Name:

1-chloro-1,1-difluoroethane

Cas Number:

75-68-3

Molecular formula:

C2H3ClF2

IUPAC Name:

1-chloro-1,1-difluoroethane

Details on test material:

Name of test material: 1-chloro 1,1-difluoroethane
The test-material was presented as compressed form in 8 gas cylinders and has a purity level of 99.9%
Supplier: Pennwalt Corporation
Description: clear, colorless liquid

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Breeding Laboratories, Wilmington, Massachusetts
- Age at study initiation: 35 days
- Date of receipt: September 25, 1979
- Weight at study initiation: 226-281 g (mean 254 g)
- Assigned to test groups randomly: yes
- Housing: individually in stainless steel wire mesh cages
- Diet: Purina Rodent Laboratoriy Chow 5001, ad libitum
- Water: tap - city water (Elizabethtown Water Company) with automatic watering system, ad libitum

ENVIRONMENTAL CONDITIONS
- Photoperiod (hrs dark / hrs light): 12 / 12
- Air changes: 4 per minutes

Administration / exposure

Route of administration:

inhalation

Vehicle:

None

Details on exposure:

TYPE OF INHALATION EXPOSURE: whole body

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure chamber: chamber with total volume of 10 cubic meter, dynamic exposure
- Exposure generation: generated by volatilising the test material and directing the undiluted vapor in the main chamber air intake, where it was diluted with the chamber air supply to the desired concentration. The flow of the test material was monitored suing calibrated glass flow meters. The chamber concentration was regulated by adjusting the flow rate of the test material.
- Method of holding animals in test chamber: caged; exposure cages were rotated weekly to average each animals position in the chamber
- Air flow rate: 2.5 cubic meters per minute
- Air change rate: one complete air change/4 minutes, with a 99% equilibrium time of 18 minutes

TEST ATMOSPHERE
- Brief description of analytical method used: gas chromatogaphy

Duration of treatment / exposure:

13 weeks

Frequency of treatment:

6 h/day, 5 d/week, 13 weeks

Post exposure period:

< 24 hours

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
1000, 10000 and 20000 ppm (0.1, 1.0 and 2.0% v/v)
Basis:
nominal conc.

Doses / concentrations 2

Remarks:

Doses / Concentrations:
0.10, 1.04 and 1.98% v/v
Basis:
analytical conc.

No. of animals per sex per dose:

10 males/dose

Control animals:

yes, concurrent vehicle

Examinations

Tissues and cell types examined:

bone marrow and bone marrow cells

Details of tissue and slide preparation:

DETAILS OF SLIDE PREPARATION:
CRITERIA FOR DOSE SELECTION:
Animals were simultaneously exposed with animals in a two-year inhalation chronictoxicity/carcinogenicity study. The MTD was selected on the basis of a previous 90 day-sub-chronic toxicity sudy which had shown no treatment related effect at a concentration of 10,000 ppm (6h/d, 5d/w).

SAMPLING:
Within the 24h after the last exposure, Colcemid (4 mg/kg) was injected IP and bone marrow cells harvested 2 to 4 hours after the injection and processed for cytogenetic evaluation.

METHOD OF ANALYSIS:
At least 50 metaphases were scored per animal. The following parameters were recorded for each slide:
1. Mitotic index (metaphases/1000 cells)
2. Chromosome number and numer of popyploides
3. Chromatid and chromosome gaps.
4. Chromatid and chromosome breaks (including acentricfragments).
5. Exchanges andrearrangements (dicentrics, etc.)
6. Others (includes pulverisation, etc.)

CLINICAL OBSERVATIONS PERFORMED:  
mortality, clinical signs, body weights

ORGANS EXAMINED AT NECROPSY:
gross necropsy on main organs/tissues; microscopic examens on bone-marrow from femur

Evaluation criteria:

Statistical significant differences from the control value

Statistics:

Student's t-test and standard one way analysis of variance

Results and discussion

Test results

Sex:

male

Genotoxicity:

negative

Toxicity:

no effects

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

not applicable

Additional information on results:

MORTALITY:

All animals survived the duration of the treatment.

PHYSICAL OBSERVATIONS:

During the study, several animals in all groups exhibited mucoid nasal discharge dry rales and moist rales. Observations of dry rales were scatterd throughout treatment and control groups and a treatment related response pattern was not evident. Observations of mucoid nasal discharge and moist rales were typically more numerous in Groups III (mid level) and IV (high level) which may indicate a slight dose-response relationship. A limited number of animals in all groups also exhibited fresh or dry red nasal discharge and soft stool which were not considered related to treatment with the test substance. Other physical observations not considered related to treatment include the following: excessive lacrimation (Groups I (control), III and IV up to two observations), excessive salivation (Groups III and IV - up to two observations), chromodacryorrhea (Group II (low level) - two observations), corneal opacity (Group I - eight observations) and hair loss (Groups III and IV (one animal from each group for up to eight observations). Singular observations of yellow staining of the anogenital gur (Group IV), dehydration (Group II), orange staining around the nose (Group IV) and dry material around the eye (Group IV) were also recorded.

BODY WEIGHTS:

Mean body weights for all treated male rats were similar to the controls throughout the study with the exception of week 2. At the week 2 interval, mean body weights were signifiantly lower in Group II (low level; p <=0.01) and Group IV (high level, p <=0.05). In view of the fact that body weights of the treated and control groups were comparable for all remaining intervals, these differences were not considered treatment-related.

GROSS POST MORTEM OBSERVATIONS

Necropsy observations of abnormalities consisted primarily of lung discoloration (mottling and foci). This observation was recorded for four of ten control animals, four of ten low level animals, two of ten mid level animals, and four of ten high level animals. The findings were not considered treatment-related.

CYTOGENETIC EVALUATION:

A slight increase in the percent mitotic rate in the mid and high level exposure groups was observed. This was not attributed to the treatment. No correlation of aneuploidy or gaps with exposure was seen. No induction of rearrangements was observed.
An increased number of open breaks was observed in all dose groups when compared to the control group, with the highest number of breaks being observed in the Group IV (high level) animals. Differences from control were not statistically significant (p <= 0.05) when analyzed both by the Student's t-test and by a standard one way analysis of variance (with an arcsin transformation applied to the dependent variable). However, the validity of the increased number of open breaks noted in the high dose group is confirmed by the increased number of acentric fragments also noted for this group. There were large differences in the numbers of open breaks observed in individual animals. For example, two animals in the high level exposure group contributed 50% of the breaks observed. However, each animal in this group had at least one break, while 4 of 10 in the control group showed no breaks.
Although these data suggest that the substance caused a slight increase in chromosomal breakage at the high exposure level, the difference from control was not statistically significant. As the sample size was regarded as small for a definitive evaluation of cytogenetic potential of the substance, the chronic toxicity and carcinogenciity bioassay with a large number of animals initiated in parallel with this study provides a more definitive indication of the effects of the substance: i.e. no treatment related effects upon histopathological evaluation of selected tissues nor statistical analysis of neoplasm data.

Any other information on results incl. tables

Table 1. A summary of cytogenetic findings

	Control	Low dose	Mid dose	High dose
Mitotic rate	1.38	1.29	1.63	1.75
(Time)	(3.03)	(3.10)	(3.32)	(3.34)
% Hypoploid	0.098	0.062	0/076	0.074
% Diploid	0.892	0.924	0.912	0.914
%Hyperploid	0.010	0.014	0.012	0.012
# Gaps	50	56	39	56
% Gaps	0.100	0.112	0.078	0.112
# Breaks	18	27	27	38
% Breaks	0.036	0.054	0.054	0.076
# Ace	-	-	1	4
# R	1	1	1	1

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): negative
1-chloro 1,1-difluoroethane did not demonstrate statistically significant chromosomal aberration activity in this 90 day-inhalation bone marrow cytogenic study in male rats.

Executive summary:

The exposure of 10 male rats to 4 different target concentrations of 1-chloro-1,1-difluoroethane (0, 0.1, 1.0 and 2%, v/v) 6 h/d, 5 d/weeks for 13 weeks did not produce statistically significant chromosome aberration activity. No mortality was observed. Body weights for all treated animals were comparable to controls. Gross necropsy observations did not reveal evidence of a treatment-related effect. A slight increase in the percent mitotic rate was observed in the mid and high level exposure group; however, this was not attributed to treatment. No correlation of aneuploidy or gaps with exposure level was observed. No induction of rearrangements was observed. An increased number of open breaks were observed in all dose groups when compared to the control groups, with the highest number observed in the high dose animals; however, these differences were not statistically significant (p = 0.05).