(2E)-2-methyl-3-(4-methylphenyl)prop-2-en-1-ol

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

Study period:

18-07-2011 to 04-08-2011

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study performed under GLP. All relevant validity criteria were met.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2011

Report date:

2011

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

JAPAN: Guidelines for Screening Mutagenicity Testing Of Chemicals

Version / remarks:

Guideline stipulated by the Japanese Ministry of Health, Labour and Welfare, Ministry of Economy, Trade and Industry and Ministry of the Environment (revised November, 2006)

Deviations:

no

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

(2E)-2-methyl-3-(4-methylphenyl)prop-2-en-1-ol

EC Number:

700-548-7

Cas Number:

56138-10-4

Molecular formula:

C11H14O

IUPAC Name:

(2E)-2-methyl-3-(4-methylphenyl)prop-2-en-1-ol

Details on test material:

- Physical state: Waxy solid at temperature < 23°C and liquid at temperature > 23°C
- Storage condition of test material: In refrigerator (2-8°C) in the dark under nitrogen
- Other: white waxy solid at temperature < 23°C and clear colourless liquid at temperature > 23°C

Method

Target gene:

histidine or tryptophan locus

Species / strain

Species / strain 1

Species / strain / cell type:

E. coli WP2 uvr A

Additional strain / cell type characteristics:

not applicable

Species / strain 2

Species / strain / cell type:

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

Additional strain / cell type characteristics:

not applicable

Metabolic activation:

with and without

Metabolic activation system:

Rat liver microsomal enzymes (S9 homogenate)

Test concentrations with justification for top dose:

Experiment 1: 10, 33, 100, 333, 1000, 5000 ug/plate
Experiment 2: 33, 100, 333, 1000, 2000, 3330 ug/plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: Dimethyl sulfoxide (DMSO)
- Justification for choice of solvent/vehicle: A solubility test was performed. The test substance could not be dissolved in water. The test substance was soluble at 50 mg/mL in dimethyl sulfoxide. Dimethyl sulfoxide was used in the assay. The test formulations were used within 4 hours for the assay.

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

2-nitrofluorene

sodium azide

benzo(a)pyrene

methylmethanesulfonate

other: ICR-191; 2-nitrofluorene

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: The plates were inverted and incubated in the dark at 37.0 ± 1.0 °C for 48 ± 4 h. After this period revertant colonies (histidine independent (His+) for Salmonella typhimurium bacteria and tryptophan independent (Trp+) for Escherichia coli were counted. The revertant colonies were counted automatically with a Colony Counter.

NUMBER OF REPLICATIONS: 3

DETERMINATION OF CYTOTOXICITY
- Method: Relative total growth (bacterial background lawn) and reduction in the number of revertants

Evaluation criteria:

See 'Any other information on materials and methods' for details on evaluation of the assay and positive criteria.

Statistics:

No formal hypothesis testing was done. See 'Any other information on materials and methods' for details on the acceptability and evaluation criteria of the assay.

Results and discussion

Test results

Test results 1

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at > 3330 μg/plate in all strains/cell types tested

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Test results 2

Species / strain:

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

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at > 3330 μg/plate in all strains/cell types tested

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: Precipitation of test substance on the plates was observed at the start of the incubation period at 3330 and 5000 µg/plate in the first mutation experiment. Precipitation was not observed at the end of the incubation period nor in the second incubation period.

RANGE-FINDING/SCREENING STUDIES: None.

COMPARISON WITH HISTORICAL CONTROL DATA:
The negative control values were within the laboratory historical control data ranges.

The positive control values were mostly but not all within the laboratory historical control data ranges, specifically TA1537 (first experiment, absence of S9-mix). However, this was more than three (3) times the concurrent control values and this deviation I the mean plate count had no effect on the results of the study. Indicating that the test conditions were adequate and that the metabolic activation system functioned properly. Historical control data from experiments was presented within the study report.

Applicant's summary and conclusion

Conclusions:

Interpretation of results:
negative
Under the conditions of this study the test item was considered to be non-mutagenic in the presence and absence of S9 activation.

Executive summary:

The study was performed to OECD TG 471, EU Method B.13/14, EPA OPPTS 870.5100 and the Japan Guidelines for Screening Mutagenicity of Chemicals in accordance with GLP; to evaluate the mutagenic activity of the test substance in the Salmonella typhimurium and the Escherichia coli in a reverse mutation assay (with independent repeat). In the first mutation assay, the test item was tested up to concentrations of 5000 μg/plate in the absence and presence of 5% (v/v) S9-mix. The test item did not precipitate on the plates at this dose level. Toxicity was observed in all tester strains. Cytotoxicity was observed in all tester strains in the absence and presence of S9-mix. In the second mutation assay, the test item was tested up to concentrations of 3330 μg/plate in the absence and presence of 10% (v/v) S9-mix. Cytotoxicity was observed in all tester strains.The negative and strain-specific positive control values were within the laboratory historical control data ranges indicating that the test conditions were adequate and that the metabolic activation system functioned properly, except the response for TA1537 in the absence of S9-mix (first experiment; positive control). The value of TA1537 was above the limit of the range. However, since more than a three-fold increase was observed compared to the solvent control, the validity of the test was considered to be not affected. The test item did not induce a significant dose-related increase in the number of revertant (His+) colonies in each of the four tester strains (TA1535, TA1537, TA98 and TA100) and in the number of revertant (Trp+) colonies in tester strain WP2uvrA both in the absence and presence of S9-metabolic activation. These results were confirmed in an independently repeated experiment. Under the conditions of this study, it is concluded that that the test item was not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.