α-chlorotoluene

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

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: see 'Remark'

Remarks:

The authors tested the mutagenicity and recombinogenicity of benzylchloride following a methodology similar to the OECD guideline 471 (Bacterial Reverse Mutation Test). While GLP standards are not specified, materials and methodology as well as results are very well described and few minor deviations were observed. Thus the study can be considered a Klimisch 2.c, comparable to a guideline study with acceptable restrictions.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1976

Report date:

1976

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

yes

Remarks:

maximum concentration tested is 10 µl/plate, no E. coli strain tested and four tested concentrations instead of five

GLP compliance:

not specified

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

chloromethylbenzene

IUPAC Name:

chloromethylbenzene

Constituent 2

Reference substance name:

α-chlorotoluene

EC Number:

202-853-6

EC Name:

α-chlorotoluene

Cas Number:

100-44-7

Molecular formula:

C7H7Cl

IUPAC Name:

(chloromethyl)benzene

Details on test material:

- Name of test material (as cited in study report): BIO-76-16, Benzyl Chlorine, Analytical reagent grade
- Physical state: colourless liquid

No more data available

Method

Target gene:

Histidine for Salmonella typhimurium

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Species / strain 2

Species / strain / cell type:

S. typhimurium TA 1538

Species / strain 3

Species / strain / cell type:

Saccharomyces cerevisiae

Metabolic activation:

with and without

Metabolic activation system:

rat liver (pretreatment with Aroclor 1254)

Test concentrations with justification for top dose:

0.1 µL/plate, 1 µL/plate, 5 µL/plate, 10 µL/plate

Vehicle / solvent:

No data

Controls

Controls 1

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: methylnitrosoguanidine (MNNG)

Remarks:

solvent water or saline, without metabolic activation

Controls 2

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

2-nitrofluorene

Remarks:

solvent dimethylsulfoxide, without metabolic activation

Migrated to IUCLID6: (NF)

Controls 3

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: quinacrine mustard (QM)

Remarks:

solvent water or saline, without metabolic activation

Controls 4

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2-anthramine (ANTH)

Remarks:

solvent dimethylsulfoxide, with metabolic activation

Controls 5

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

2-acetylaminofluorene

Remarks:

solvent dimethylsulfoxide, with metabolic activation

Migrated to IUCLID6: (AAF)

Controls 6

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 8-aminoquinoline (AMQ)

Remarks:

solvent dimethylsulfoxide, with metabolic activation

Controls 7

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: dimethylnitrosamine (DMNA)

Remarks:

solvent saline, with metabolic activation

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 48 to 72h

NUMBER OF CELLS EVALUATED: 10^9 cells from a log phase culture

No further data available

Evaluation criteria:

Amino-acid requiring strains of bacteria are used to detect reverse gene mutations. Based on the number of revertants, the mutagenicity of the test compound is assessed. Indeed, if a clear relation is established between the number of revertants and the tested non toxic concentrations (in presence or not of metabolic activativation), the test is interpretated to be positive.

Statistics:

No data

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Metabolic activation:

with and without

Genotoxicity:

positive

Remarks:

for strain TA 100 with and without metabolic activation, strain TA 1535 weak response at highest dose with metabolic activation

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

valid

Untreated negative controls validity:

not specified

Positive controls validity:

valid

Test results 2

Species / strain:

S. typhimurium TA 1538

Metabolic activation:

with and without

Genotoxicity:

not specified

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

valid

Untreated negative controls validity:

not specified

Positive controls validity:

valid

Test results 3

Species / strain:

Saccharomyces cerevisiae

Metabolic activation:

with and without

Genotoxicity:

positive

Remarks:

with and without metabolic activation

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

valid

Untreated negative controls validity:

not specified

Positive controls validity:

valid

Additional information on results:

No data

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

Table 1: Result for the mutagenicity (Salmonella strains) and recombinogenicity (Saccharomyces) experiment with benzylchloride without metabolic activation for all tested strains, including controls.

		S. typhimurium	S. typhimurium	S. typhimurium	S. typhimurium	S. typhimurium	S. cerevisiae
		TA-1535	TA-1537	TA-1538	TA-98	TA-100	D4
Solvent control		37	38	19	17	170	49		*	TA-1535	MNNG	10 µL/plate
Positive control*		>10^4	>10^4	821	>10^3	>10^4	460			TA-1537	QM	10 µL/plate
Test compound	0.1 µL/plate	50	44		24	192	37			TA-1538	NF	100 µg/plate
	1 µL/plate	31	38	18	28	214	34			TA-98	NF	100 µg/plate
	5 µL/plate	38	31	15	265	248	56			TA-100	MNNG	10 µL/plate
	10 µL/plate	42	25	18	38	674	333			D4	MNNG	10 µL/plate

Table 2: Result for the mutagenicity (Salmonella strains) and recombinogenicity (Saccharomyces) experiment with benzylchloride with metabolic activation for all tested strains, including controls.

		S. typhimurium	S. typhimurium	S. typhimurium	S. typhimurium	S. typhimurium	S. cerevisiae
		TA-1535	TA-1537	TA-1538	TA-98	TA-100	D4
Solvent control		29	68	48	35	97	37		**	TA-1535	ANTH	100 µg/plate
Positive control**		>10^3	>10^3	>10^3	>10^3	>10^3	44			TA-1537	AMQ	100 µg/plate
Test compound	0.1 µL/plate	31	35	45	23	155	40			TA-1538	AAF	100 µg/plate
	1 µL/plate	30	33	53	20	121	38			TA-98	AAF	100 µg/plate
	5 µL/plate	11	19	47	23	174	44			TA-100	ANTH	100 µg/plate
	10 µL/plate	69	27	30	22	391	134			D4	DMNA	100 µmoles/plate

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
positive with and without metabolizing system

The authors tested mutagenicity and recombinogenicity of benzylchloride in the absence and presence of a metabolizing system with a methodology similar to the OECD guideline 471. Under these test conditions, the test substance was considered mutagenic for Salmonella typhimurium and recombinogenic for Saccharomyces cerevisiae with and without metabolizing system.

Executive summary:

The authors tested the mutagenicity and recombinogenicity of benzylchloride (CAS n° 100-44-7) in the absence and presence of a metabolizing system with a methodology similar to the OECD guideline 471. The Salmonella typhimurium strains TA 1535, TA 1537, TA 1538, TA 98 and TA 100 and the Saccharomyces cerevisiae strain D4 were used and the metabolizing system was obtained from rat liver homogenates. These strains were exposed to 0.1, 1, 5 and 10 µL/plate with and without metabolic activation for 48 to 72 h. Solvent controls and positive controls were performed.

The test showed that benzylchloride was mutagenic for Salmonella typhimurium strain TA 100 and recombinogenic for Saccharomyces cerevisiae strain D4. This respons was observed in absence and presence of the metabolic activation system, demonstrating that metabolic activation is not essential for mutagenicity or recombinogenicity. Furthermore, there was also an indication of a weak response at the high dose of the

Salmonella typhimurium strain TA 1535. Effects were only seen at relatively high doses and dose-related responses at the lower end of the dose range were not observed. Hence, based on this information, further in vitro testing would be required with other systems to clarify if in vivo testing would be necessary.

No data was available on the GLP status of this study. However this study was similar to the OECD guideline 471 with minor deviations as following, the maximum concentration tested was 10 µl/plate, as no E. coli strain was tested and as four concentrations were tested instead of five.Therefore, this study should be considered reliable with restictions, a Klimisch 2.c study comparable to a guideline study with acceptable restrictions.