2-fluoro-6-trifluoromethylpyridine

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Link to relevant study records

Reference

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:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 472 (Genetic Toxicology: Escherichia coli, Reverse Mutation Assay)

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Specific details on test material used for the study:

Substance ID: F6TF
Lot #: 8922/16
Purity: 99.7%

Target gene:

histidine synthesis

Species / strain / cell type:

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

Species / strain / cell type:

E. coli WP2

Remarks:

WP2P

Species / strain / cell type:

E. coli WP2 uvr A

Metabolic activation:

with and without

Metabolic activation system:

Type and composition of metabolic activation system: S9 fraction (3 mL), Sucrose-Tris-EDTA buffer (S9 buffer) (7 mL) and Cofactor solution (20 mL)

Test concentrations with justification for top dose:

100, 200, 500, 1000, 2500 and 5000 µg/plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO

Untreated negative controls:

yes

Remarks:

DMSO

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

Positive controls:

yes

Positive control substance:

other: acridine mutagen IRC 191, 2-aminoanthracene (2AA), Daunomycin HCl (DR), N-ethyl-N'-nitro-N-nitrosoguanidine (ENNG), Mitomycin C (MMC), and sodium azide (NaZ)

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Preincubation period: 60 minutes at 37ºC (only for Trial 2 in the presence of S9 mix)
- Exposure duration: 3 days in the dark at 37ºC
- Fixation time (start of exposure up to fixation or harvest of cells): Following the total incubation period (3 days), the plates were examined for the lack of microbial contamination and evidence that the test was valid: i.e., there was a background lawn on the solvent control plates and on the plates for (at least) the lower concentrations of test substance, and that the positive controls had responded as expected.

NUMBER OF REPLICATIONS: 3 plates per concentration in each of two trials

Evaluation criteria:

Test data from individual experiments are considered valid if
a) the concurrent solvent control data are acceptable;
b) the positive control data show unequivocal positive responses;
Failure of one or more tester strainlS9 combinations does not invalidate the data for the remainder of a concurrent experiment.

A positive response in a (valid) individual experiment is achieved when one or both of the following criteria are met:
a) a statistically significant dose-related increase in the mean number of revertant colonies is obtained;
b) a two-fold or greater increase in the mean number of revertant colonies (over that observed for the concurrent solvent control plates) which is statistically significant, is observed at one or more concentrations.

A negative result in a (valid) individual experiment is achieved when:
a) there is no statistically significant dose-related increase in the mean number of revertant colonies per plate observed for the test substance; and
b) in the absence of any such dose response, no increase in colony numbers is observed (at any test concentration) which exceeds 2x the concurrent solvent control.

For a positive response in an individual experiment to be considered indicative of an unequivocal positive, i.e. mutagenic, result for that strain/S9 combination, then the observed effect(s) must be consistently reproducible.

Statistics:

An assessment of statistical significance was carried out using a one-tailed Student's t-test. The corresponding probability for each dose level was derived by computer using the appropriate degrees of freedom.

Key result

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not determined

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not determined

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not determined

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not determined

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

E. coli WP2

Remarks:

WP2P

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not determined

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not determined

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: Negative for mutagenicity

In two separate assays, the test substance did not induce any significant, reproducible increases in the observed number of revertant colonies in any of the tester strains in the absence of S9-mix, or in strains TA1535, TA1537, TA98, TA100 and WP2P in the presence of S9-mix.

With strain WP2P uvrA in the presence of S9-mix, increases in revertant colony numbers were observed in both experiments conducted. There was, however, no dose response in either experiment and the highest increase in revertant colony numbers (over negative control values) was 2-fold, observed in only one of the two experiments.

The positive controls for each experiment induced the expected responses indicating the strains were responding satisfactorily in each case.

Conclusions:

Under the conditions of this assay, the test substance gave a negative response to all strains tested in both the presence and absence of S9-mix.

Executive summary:

The test substance was evaluated in a bacterial mutagenicity assay over a range of concentrations using 4 strains of Salmonella typhimurium (TA1535, TA1537, TA98, TA100) and two strains of Escherichia coli (WP2P and WP2PuvrA) in the presence and absence of a rat liver-derived metabolic activation system (S9-mix).

 

In two separate assays, the test substance did not induce any significant, reproducible increases in the observed number of revertant colonies in any of the tester strains in the absence of S9-mix, or in strains TA1535, TA1537, TA98, TA100 and WP2P in the presence of S9-mix. With strain WP2P uvrA in the presence of S9-mix, increases in revertant colony numbers were observed in both experiments conducted. There was, however, no dose response in either experiment and the highest increase in revertant colony numbers (over negative control values) was 2-fold, observed in only one of the two experiments. The positive controls for each experiment induced the expected responses indicating the strains were responding satisfactorily in each case. Under the conditions of this assay, the test substance gave a negative response to all strains tested in both the presence and absence of S9-mix.

Additional information

In an in vitro clastogenicity study with human lymphocytes, the results were weakly positive with and without activation. In a second in vitro clastogenicity study with human lymphocytes, negative results were observed without activation, but weakly positive results were observed with activation in 1 of 2 trials.  Negative results were observed in an in vitro study in bacterial cells (Salmonella typhimurium and E. coli) and an in vivo micronucleus (mouse bone marrow) study. Overall, the test substance is considered negative for genotoxicity.

Justification for classification or non-classification:

Overall, the test substance did not produce mutagenicity or clastogenicity when evaluated in laboratory animals. Based on an assessment of the robust genetic toxicity data for this substance, the substance does not need to be classified for mutagenicity according the EU Directive 67/548/EEC and EU Classification, Labelling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008.

Short description of key information:
In Vitro (Mutagenic effects - bacterial): OECD 471 and 472; Bacterial reverse mutation assay. Negative. Reliability = 1..
In Vitro (Clastogenic effects - mammalian): OECD 473; Chromosome aberrations in human lymphocytes. Weakly positive with and without activation. Reliability = 1
In Vitro (Clastogenic effects - mammalian): OECD 473; Chromosome aberrations in human lymphocytes. Negative without activation, but was weakly positive with activation in 1 of 2 trials. Reliability = 1
In Vivo (Clastogenic effects - mammalian): OECD 474; in vivo rat micronucleus study; Negative at doses up to 800 ppm. Reliability = 1.

Endpoint Conclusion: No adverse effect observed (negative)