chloromethane; methyl chloride

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in mammalian cells

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline study with acceptable restrictions

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Data source

Reference

Reference 1

Reference Type:

publication

Title:

Assessment of the genotoxic effects of methyl chloride in human lymphoblasts

Author:

Fostel, J. et al.

Year:

1985

Bibliographic source:

Mutation research 155:75-81

Reference 2

Reference Type:

secondary source

Title:

Chloromethane CAS: 74-87-3

Author:

OECD SIDS

Year:

2004

Bibliographic source:

SIDS Initial Assessment Report for SIAM 15 ; www.iccahpv.com

Reference 3

Reference Type:

secondary source

Title:

Methyl Chloride

Author:

Löf, A. et al.

Year:

2000

Bibliographic source:

Concise International Chemical Assessment Document 28

Reference 4

Reference Type:

secondary source

Title:

Toxicological review of methyl chloride (CAS No. 74-87-3)

Author:

Greenberg, M. and Riddle, J.

Year:

2001

Bibliographic source:

U.S. Environmental Protection Agency, EPA/635/R01/003

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)

Deviations:

yes

Remarks:

no test with metabolic activation was performed

GLP compliance:

not specified

Type of assay:

mammalian cell gene mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Chloromethane

EC Number:

200-817-4

EC Name:

Chloromethane

Cas Number:

74-87-3

Molecular formula:

CH3Cl

IUPAC Name:

chloromethane

Details on test material:

- Name of test material (as cited in study report): methyl chloride
- Analytical purity: no data

Method

Target gene:

thymidine kinase

Species / strain

Species / strain / cell type:

human lymphoblastoid cells (TK6)

Details on mammalian cell type (if applicable):

- Type and identity of media: RPMI 1640 medium supplemented with 20% heat-inactivated horse serum and penicillin

Additional strain / cell type characteristics:

not specified

Metabolic activation:

without

Metabolic activation system:

no

Test concentrations with justification for top dose:

1, 2, 3, 4 or 5% chloromethane (1% chloromethane is equivalent to 0.73 mM (38.37 mg/L).)

Vehicle / solvent:

none

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

no

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: ethylmethanesulphonate (0.2 and 0.4 mM) and methylmethanesulphonate (0.025 and 0.05 mM)

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium
Between 2 and 2.5 x 10^7 TK6 cells were added to each of three 50 mL aliquots of complete medium in 75 cm2 flasks with lids that had been punctered with a 16-gauge needle. The flasks were sealed into 1.9 L Gas Pak chambers and 9 changes of an atmosphere containing a known concentration of chloromethane were pumped through each chamber. To correlate the percent of atmospheric chloromethane with chloromethane concentration in the culture medium, a small volume of [14C]chloromethane was added to pure chloromethane to generate a mixture of chloromethane with a known specific activity.

DURATION
- Exposure duration: 3 h
- Expression time (cells in growth medium): 3 d from the point where growth rate was normal again
- Selection time (if incubation with a selection agent): 12-14 d

SELECTION AGENT (mutation assays): 1 µg trifluorothymidine/ mL

NUMBER OF REPLICATIONS: 2 (4 experiments per replication)

NUMBER OF CELLS EVALUATED: 4 x 10^4 each experiment

DETERMINATION OF CYTOTOXICITY: extrapolation of the exponential portion of the growth curve back to the treatment time

Evaluation criteria:

The mutant fraction was calculated by dividing the efficiency of colony formation in the mutant dishes by the efficiency of colony formation on the dishes without selection.

Results and discussion

Test results

Species / strain:

human lymphoblastoid cells (TK6)

Metabolic activation:

without

Genotoxicity:

positive

Remarks:

dose-dependent increase

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at all concentrations there was a lag in the growth of the cultures

Vehicle controls validity:

not applicable

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Water solubility: according to measurements a linear relationship between atmospheric concentration of chloromethane and concentration in the medium was observed. 1% chloromethane is equivalent to 0.73 mM (38.37 mg/L).

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
positive without metabolic activation