1H-Pyrazole-4-carboxylic acid, 3-(difluoromethyl)-1-methyl-, ethyl ester

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:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP compliant, guideline study, no restrictions, fully adequate for assessment

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

Principles of method if other than guideline:

Not applicable.

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

ethyl 3-(difluoromethyl)-1-methyl-1H-pyrazole-4-carboxylate

EC Number:

619-510-5

Cas Number:

141573-95-7

Molecular formula:

C8H10F2N2O2

IUPAC Name:

ethyl 3-(difluoromethyl)-1-methyl-1H-pyrazole-4-carboxylate

Details on test material:

- Name of test material (as cited in study report): DFMMP
- Lot/batch No.: Batch No. MKA13111
- Substance type: mono-constituent substance
- Physical state: powder
- Analytical purity: 99.0% (GC)

Method

Target gene:

his

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:

E. coli WP2 uvr A

Metabolic activation:

with and without

Metabolic activation system:

rat liver S9-mix induced by a combination of phenobarbital and ß-naphthoflavone

Test concentrations with justification for top dose:

100, 333, 1000, 3330 and 5000 µg/plate

Vehicle / solvent:

DMSO

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: Without metabolic activation: Sodium azide (TA1535), ICR-191 (TA1537), 2-Nitrofluorene (TA98), Methyl methanesulphonate (TA 100) and 4-nitroquinoline N-oxide (WP2uvrA); With metabolic activation: 2-aminoanthracene (all strains).

Details on test system and experimental conditions:

METHOD OF APPLICATION
Direct incorporation method: 0.1 ml of a fresh bacterial culture, 0.1 mL of the DMSO diluted substance and 0.5 ml S9-mix (in case of activation assay) or 0.5 ml phosphate buffer (in case of non-activation assays) were successively added to 3 ml molten top agar. After agitation the mix was poured onto a selective agar plate.
DURATION
- Preincubation period: not applicable
- Exposure duration: 48 hours at 37°C in the dark
NUMBER OF REPLICATES
- 3 plates/dose/strain.
- Two independent experiments were performed. In the first experiment DFMMP was tested both in the absence and presence of 5% (v/v) S9-mix in tester strains TA1535, TA1537, and TA98. In an independent repeat of the assay with additional parameters, the test substance was tested both in the absence and presence of 10% (v/v) S9-mix in all tester strains.
DETERMINATION OF TOXICITY
- Method: the reduction of the bacterial background lawn, the increase in the size of microcolonies and the reduction of revertant colonies were examined.

Evaluation criteria:

The mutagenicity study is considered valid if the mean colony counts of the vehicle control values of the strains are within the laboratory historical range, if the results of the positive controls meet the criteria for a positive response within the laboratory historical range and if the selected dose range includes a clearly toxic concentration or is extended to 5 mg/plate.

A test substance is considered positive (mutagenic) in the test if the total number of revertants in tester strain TA100 is greater than two (2) times the concurrent control, or the total number of revertants in tester strains TA1535, TA1537, TA98 or WP2uvrA is greater than three (3) times the concurrent vehicle control.
A positive or negative response should be reproducible in at least one independently repeated experiment.

Statistics:

No statistical analysis was performed.

Results and discussion

Test results

Species / strain:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

The test substance was not toxic to any strain, in both the absence and presence of S9-mix, as neither a decrease in the mean number of revertants nor a clearing of the background lawn of bacterial growth compared to the negative controls was observed.
In both the absence and presence of S9-mix in all strains, DFMMP did not induce a minimal 2-fold and/or dose related increase in the mean number of revertant colonies compared to the background spontaneous reversion rate observed with the negative control.
It is concluded that the results obtained with the test substance in Salmonella typhimurium strains TA 1535, TA 1537, TA 98 and TA 100, and in the Escherichia coli strain WP2 uvrA, in both the absence and the presence of the S9-mix, indicate that DFMMP is not mutagenic under the conditions employed in this study.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.