3'-(trifluoromethyl)acetophenone

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

The test item was tested for mutagenic effects in vitro according to OECD TG 471 in S. typhimurium TA98, TA 100, TA 102, TA 1535, TA 1537 and E. coli WP2 uvrA. The test was performed with and without the addition of rat-liver post mitochondrial supernatant (S9 fraction). The compound was dissolved in DMSO and tested at five concentrations in the range of 312.5 to 5000.0 µg/plate in the presence and absence of a metabolic activation system. In order to confirm the results, the experiments were repeated with and without metabolic activation at five concentrations in the range of 312.5 to 5000.0 ng/plate. Each strain was additionally tested in the presence and in the absence of a metabolic activation system with a suitable, known mutagen as positive control. The original experiment with and without metabolic activation and the confirmatory experiment without activation were performed as standard plate incorporation assay. The confirmatory experiments with metabolic activation were carried out as preincubation assay.

In both experiments, performed with and without metabolic activation, none of the tested concentrations of the test item led to an increase in the incidence of either histidine or tryptophan-prototrophic mutants by comparison with the negative control. Based on the results of these experiments, it is concluded that the test item and its metabolites did not induce gene mutations.

The test material was investigated forclastogenic (chromosome-damaging) effects on Chinese hamster ovary cells in vitro with and without extrinsic metabolic activation (S9) according OECD TG 473. The compound was dissolved in DMSO. In both experiments performed without and with metabolic activation no statistically significant increase in the number of metaphases containing specific chromosomal aberrations was observed. The incidence of aberrant cells was within the historical control range at all doses assessed. The positive control materials induced significant increases in the percentage of aberrant cells.

Link to relevant study records

Reference

Reference 1

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:

July - September 1998

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

1992

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

1983

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Specific details on test material used for the study:

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Solubility and stability of the test substance in the solvent/vehicle: To demonstrate that the test system was exposed to the intended concentrations of the test substance in the mutagenicity tests, the concentration of the substance in solution has been determined by the analytical unit. The analysis was performed with the lowest concentration, which was obtained by serial dilution of the highest concentration used.

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102

Species / strain / cell type:

E. coli WP2 uvr A

Metabolic activation:

with and without

Metabolic activation system:

rat liver S9 fraction

Test concentrations with justification for top dose:

20.58, 61.73, 185.19, 555.56, 1666.67, 5000 µg / plate in the range finding test
312.5 to 5000.0 ug/plate (original and conformatory experiment)

Vehicle / solvent:

DMSO

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: with :2-Aminoanthracene for TA 100, TA 102, TA 98, TA 1537, Cyclophosphamide for TA 1535, without : Sodium azide for TA 100 and TA 1535, Mitomycin-C for TA 102, 2-Nitrofluorene for TA 98, 9-Aminoacridine for TA 1537, 4-nitroquinoline for WP2 uvrA

Rationale for test conditions:

Range finding test
Six concentrations of the test item ranging from 20.6 to 5000.0 µg/plate were tested with strain Salmonella typhimurium TA100 and strain Escherichia coli WP2 uvrA to determine the highest concentration to be used in the mutagenicity assay. The experiments were performed with and without metabolic activation. Normal background growth was observed with both strains. The numbers of revertant colonies were not reduced. The test substance did not precipitate on thesurface of the agar plates.
From the results obtained the highest concentration suitable for the mutagenicity test was selected to be 5000.0 µg/plate with and without metabolic activation.

Evaluation criteria:

The test substance will be considered to be positive in the test system if one or both of thefollowing conditions are met:
• At least a reproducible doubling of the mean number of revertants per plate above that of the negative control at any concentration for one or more of the following strains: TA 98, TA 1535, TA 1537, E. coli WP2 uvrA.
• A reproducible increase of the mean number of revertants per plate for any concentration above that of the negative control by at least a factor of 1.5 for strains TA 100 or TA 102.
Generally a concentration-related effect should be demonstrable.

Statistics:

A statistical analysis was not performed. At present the use of statistical methods concerning this particular test system is not generally recommended.

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

True negative controls validity:

not applicable

Positive controls validity:

valid

Species / strain:

other: S. typhimurium TA 98, TA 100, TA 102; TA 1535, TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

True negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

Due to a growth inhibiting effect of the test substance the number of revertant colonies was reduced in the original experiment with metabolic activation on strains TA 100, TA 102, TA 1537 (2500.0 and 5000.0 µg/plate), TA 1535 and TA 98 (5000.0 µg/plate). In the experiment without activation a reduction in the number of reverant colonies was observed with strains TA 100, TA 102, TA 98, TA 1537 (2500.0 and 5000.0 µg/plate) and TA 1535 (5000.0 µg/plate). Similar effects occurred in the confirmatory experiment with activation on strains TA 100, TA 1537 (2500.0 and 5000.0 µg/plate), TA 1535, TA 102, TA 98 and E. coli (5000.0 µg/plate). In the
experiment without activation a reduction in the number of reverant colonies was seen with strains TA 100, TA 1535, TA 102, TA 1537 (2500.0 and 5000.0 µg/plate) and TA 98 (5000.0 µg/plate). A reduction in the growth of the background lawn was observed in the experiments with activation on strains of S. typhimurium at the higher concentrations. In the experiments without activation also on strain E. coli a reduced growth was visible. The test substance did not precipitate on the surface of the agar plates.

Remarks on result:

other: negative

Executive summary:

The test item was tested for mutagenic effects in vitro according to OECD TG 471 in S. typhimurium TA98, TA 100, TA 102, TA 1535, TA 1537 and E. coli WP2 uvrA. The test was performed with and without the addition of rat-liver post mitochondrial supernatant (S9 fraction). The compound was dissolved in DMSO and tested at five concentrations in the range of 312.5 to 5000.0 µg/plate in the presence and absence of a metabolic activation system. In order to confirm the results, the experiments were repeated with and without metabolic activation at five concentrations in the range of 312.5 to 5000.0 ng/plate. Each strain was additionally tested in the presence and in the absence of a metabolic activation system with a suitable, known mutagen as positive control. The original experiment with and without metabolic activation and the confirmatory experiment without activation were performed as standard plate incorporation assay. The confirmatory experiments with metabolic activation were carried out as preincubation assay.

In both experiments, performed with and without metabolic activation, none of the tested concentrations of the test item led to an increase in the incidence of either histidine or tryptophan-prototrophic mutants by comparison with the negative control. Based on the results of these experiments, it is concluded that the test item and its metabolites did not induce gene mutations.

Reference 2

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Type of information:

experimental study

Adequacy of study:

key study

Study period:

August - November 1998

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EU Method B.10 (Mutagenicity - In Vitro Mammalian Chromosome Aberration Test)

Qualifier:

according to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.5375 - In vitro Mammalian Chromosome Aberration Test

GLP compliance:

yes (incl. QA statement)

Type of assay:

in vitro mammalian chromosome aberration test

Specific details on test material used for the study:

Stability (in the vehicle used, under the conditions of the test): To confirm that the cells were actually exposed to the intended test concentrations and to confirm the stability of the test substance in the vehicle used, determination of the concentration of the test substance in solution was performed by the analytical unit. This determination was performed with the lowest concentrations of the stock solutions used in the first and last segment of the mutagenicity test.
The concentration values found were in agreement with the nominal concentrations, indicating sufficient stability of the test substance in the vehicle used.

The test substance was dissolved in DMSO at room temperature. The highest soluble concentration was about 557 mg/ml. This solution caused the formation of strong precipitates after 100 fold dilution with culture medium. The highest concentration in DMSO (stock solution) resulting in a tolerable homogeneous turbidity after 100 fold dilution with culture medium was 150 mg/ml. Lower concentrations were prepared by serial dilution of the stock solution with DMSO. The respective solutions were added (1:100) to the cell cultures. The final concentration of the vehicle DMSO in the culture medium was 1%. The addition of the stock solutions to the cell cultures produced turbidity in the culture medium at
the final concentrations of 1500 down to 750 µg/ml in the experiments without and with activation.

Species / strain / cell type:

Chinese hamster Ovary (CHO)

Details on mammalian cell type (if applicable):

Cell line: ATCC (American Type Culture Collection) CCL 61 (ovary, Chinese hamster, CHO Kl).

Metabolic activation:

with and without

Metabolic activation system:

S9 fraction from Aroclor 1254 induces rat liver

Test concentrations with justification for top dose:

Experiments without metabolic activation:
- original experiment:
3 h treatment/18 h recovery: 93.75, 187.50 and 375.00 µg/ml
confirmatory experiment:
21 h treatment: 93.75, 187.50 and 375.00 µg/ml
- 45 h treatment: 187.50, 375.00 and 750.00 µg/ml
Higher concentrations could not be scored due to cytotoxicity.
Experiments with metabolic activation:
- original experiment:
3 h treatment/18 h recovery: 93.75, 187.50 and 375.00 µg/ml
- confirmatory experiment:
3 h treatment/18 h recovery: 93.75, 187.50 and 375.00 µg/ml
3 h treatment/42 h recovery: 93.75, 187.50 and 375.00 µg/ml
Higher concentrations could not be scored due to cytotoxicity.

Vehicle / solvent:

The test substance was dissolved in DMSO at room temperature. The highest soluble concentration was about 557 mg/ml. This solution caused the formation of strong precipitates after 100 fold dilution with culture medium. The highest concentration in DMSO (stock solution) resulting in a tolerable homogeneous turbidity after 100 fold dilution with culture medium was 150 mg/ml. Lower concentrations were prepared by serial dilution of the stock solution with DMSO. The respective solutions were added (1:100) to the cell cultures. The final concentration of the vehicle DMSO in the culture medium was 1%. The addition of the stock solutions to the cell cultures produced turbidity in the culture medium at
the final concentrations of 1500 down to 750 µg/ml in the experiments without and with activation.

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

yes

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

cyclophosphamide

mitomycin C

Evaluation criteria:

- Criteria for a positive response:
Under the standard conditions of our laboratories, the test substance is generally considered to be active in the Chinese Hamster cells if the following conditions are met:
- The percentage of metaphases containing specific aberrations in a treatment group is higher than 6.0 (based on historical negative control range) and differs statistically significant from the respective value of the negative control.
- A concentration-related response should be demonstrable.
Criteria for a negative response:
Under the standard conditions of our laboratories, the test substance is generally considered to be inactive in the Chinese Hamster cells if the following conditions are met:
- The percentage of metaphases containing specific aberrations in all treatment groups is less than or equal to 6.0 (based on historical negative control range) and does not differ statistically significant from the respective value of the negative control.

Statistics:

The evaluated numbers of specific aberrations were subjected to statistical analysis. In the preliminary tests the data were assessed for flask effects (dependence of cells within each culture) using a chi-squared test. The nonsignificant result of this test means there is no substantial evidence to conclude a flask effect (although a flask effect still might exist). Accordingly, a chi-squared test for trend was performed modelling all cells in a given experiment as independent. That is, the individual cell is taken as the experimental unit. Consequently the power of the test is substantially increased, resulting in a rather safe judgement of the observed effects.

Species / strain:

Chinese hamster Ovary (CHO)

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not specified

True negative controls validity:

not specified

Positive controls validity:

valid

It is concluded that under the given experimental conditions no evidence of clastogenic effects was obtained in Chinese hamster ovary cells in vitro treated with the test material.

Conclusions:

It is concluded that under the given experimental conditions no evidence of clastogenic effects was obtained in Chinese hamster ovary cells in vitro treated with the test material.

Executive summary:

The test material was investigated forclastogenic (chromosome-damaging) effects on Chinese hamster ovary cells in vitro with and without extrinsic metabolic activation (S9) according OECD TG 473. The compound was dissolved in DMSO. In both experiments performed without and with metabolic activation no statistically significant increase in the number of metaphases containing specific chromosomal aberrations was observed. The incidence of aberrant cells was within the historical control range at all doses assessed. The positive control materials induced significant increases in the percentage of aberrant cells.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

Based on the study results no classification according to Regulation (EC) No. 1272/2008 (CLP), ANNEX I, is warranted.