Phenol, 2,4-bis[(dodecylthio)methyl]-6-methyl-

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in mammalian cells

Remarks:

Type of genotoxicity: gene mutation

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: GLP Guideline study. According to ECHA guidance, a study with a read-across substance can have no reliability of higher than 2. The study itself is valid without restriction.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1990

Report date:

1990

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes

Type of assay:

mammalian cell gene mutation assay

Test material

Test material information

Constituent 1

Details on test material:

- Analytical purity: 94.5/94.6%

Method

Target gene:

HPRT

Species / strain

Species / strain / cell type:

Chinese hamster lung fibroblasts (V79)

Metabolic activation:

with and without

Metabolic activation system:

S9-mix from rat livers induced with Aroclor 1254

Test concentrations with justification for top dose:

Original experiment:
Range with activation: 25 - 500 µg/mL
Range without activation: 50 - 1000 µg/mL

Confirmatory experiment:
Range with activation: 50 - 1000 µg/mL
Range without activation: 50 - 1000 µg/mL

Vehicle / solvent:

DMSO

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: with metabolic activation: 1.0 µL/mL of the promutagen N-nitroso-dimethylamine (DMN); without metabolic activation: 300 nL/mL of the ultimate mutagen ethylmethanesulphonate (EMS)

Details on test system and experimental conditions:

METHOD OF APPLICATION: in suspension

DURATION
- Preincubation period: 42 h
- Exposure duration: five hours by 22.5 mL treatment medium and 2.5 mL S9 activation mixture, or for 21 hours by 25 mL treatment medium alone
- Expression time (cells in growth medium): 5 days
- Selection time (if incubation with a selection agent): 7-8 days

SELECTION AGENT (mutation assays): 8 µg/mL 6-thioguanine
STAIN: Giemsa

DETERMINATION OF CYTOTOXICITY
- Method: cell viability

Evaluation criteria:

All mutant frequencies are normalized to a virtual cloning efficiency of 100% at the end of the expression period. If this cloning efficiency is lower than 15%, the corresponding mutant frequency is usually not calculated, owing to the high statistical insignificance of the result. For every concentration a mutant factor, which is defined as the ratio of the mutant frequency of the treated and the solvent control cultures, will be calculated.
The sensitivity of the test is restricted to a lower limit of the mutant frequency of 4x10e-6. If the mutant frequency measured is below this limit, it will be reported to be smaller than 4x10e-6 and the mutant factor will be calculated assuming a mutant frequency of 4x10e-6.

The test substance will be considered mutagenic in this test system, if either:
- The mutant frequency of the treated culture exceeds that of the solvent controls by a mutant factor of 2.5 and there is a dose-dependent increase of the mutant frequency; or:
- The mutant frequency in a treated culture exceeds that of the solvent control by a mutant factor of 3.0 at any concentration tested and reported and the absolute number of clones in the treated and untreated cultures differ by more than 20 clones per 10e6 cells plated.

Results and discussion

Test results

Species / strain:

Chinese hamster lung fibroblasts (V79)

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

other: A cytotoxicity test was performed on V79 cells as a preliminary test to determine the highest concentration of the test substance to be applied in the mutagenicity assay (causing about 90% reduction of viable cells in comparison with the solvent control.

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

Original mutagenicity test
In the original mutagenicity test with microsomal activation, (Summary)) after screening with 6-thioguanine the mutant frequencies in the solvent controls were both <4.00x10e-6. This value represents the lower limit of sensitivity of the test system. The mean value used for calculation is 4.00x10e-6.
At the five lowest concentrations tested, the calculated mutant frequency values were <4.00x10e-6. Comparison of this value with the mean of the two solvent controls revealed mutant factors of 1.00. At the higher concentration up to the highest, the calculated mutant-frequency values were 5.68x10e-6 (factor 1.42) and 5.21x10e-6, resulting in a mutant factor of 1.30.
The positive control treated with 1 µL DMN/mL medium revealed a mutant-frequency of 131.27x10e-6, giving a mutant factor of 32.82. The mutant-frequencies in the solvent controls of the cultures without microsomal activation were both <4.00x10e-6. At the six lowest concentrations tested, the calculated mutant frequency values were <4.00x10e-6. Comparison of this value with the mean of the two solvent controls revealed mutant factors of
1.00. At the highest concentration the calculated mutant-frequency value was 5.03x10e-6, resulting in a mutant factor of 1.26.
The positive control treated with 300 nL EMS/mL medium gave a mutant frequency of 1183.67x10e-6 and a corresponding mutant factor of 295.92.

Confirmatory mutagenicity test
In the confirmatory mutagenicity test with microsomal activation, after screening with 6-thioguanine the mutant-frequency values in the solvent controls were both <4.00x10e-6.
At the six lowest concentrations tested, the calculated mutant frequency values were <4.00x10e-6. Comparison of this value with the mean of the two solvent controls revealed mutant factors of 1.00. At the highest concentration the calculated mutant-frequency value was 10.83x10e-6, resulting in a mutant factor of 2.71.
The positive control treated with 1 nL DMN/mL medium revealed a mutant frequency of 268.34x10e-6, giving a mutant factor of 67.09. The mutant-frequencies in the solvent controls of the cultures without microsomal activation were both <4.00x10e-6. At all concentrations tested, the calculated mutant frequency values were <4.00x10e-6. Comparison of this value with the mean of the two solvent controls revealed mutant factors of 1.00.
The positive control treated with 300 nL EMS/mL medium gave a mutant frequency of 1179.96x10e-6 and a corresponding mutant factor of 294.99.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.