Thiophanate-methyl

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

March 8, 1990 - March 19, 1990

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1990

Report date:

1990

Materials and methods

Test guideline

Test guideline 1

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: EPA OPP 88-2

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Thiophanate-methyl

EC Number:

245-740-7

EC Name:

Thiophanate-methyl

Cas Number:

23564-05-8

Molecular formula:

C12H14N4O4S2

IUPAC Name:

methyl N-{[2-({[(methoxycarbonyl)amino]methanethioyl}amino)phenyl]carbamothioyl}carbamate

Test material form:

solid: particulate/powder

Method

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:

Type and composition of metabolic activation system:
- source of S9 : 9000 xg supernatant prepared from Sprague-Dawley adult male rat liver induced by phenobarbital and 5,6-benzoflavone was purchased from ORIENTAL YEAST CO., LTD.

Test concentrations with justification for top dose:

5000, 2500, 1250, 625, 312.5, 156.3, 78.1, 39.1 µg/plate (with and without S9 Mix)
The limit concentration required in the guideline was used for the tests.

Vehicle / solvent:

DMSO

Controls

Controls 1

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2-(2-furyl)-3-(5-nitro-2-furyl)acrylamide

Remarks:

without S9 mix, TA100, TA98, WPuvrA

Controls 2

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 1,2-benzpyrene

Remarks:

with S9 mix, TA100, TA98, TA1537

Controls 3

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

N-ethyl-N-nitro-N-nitrosoguanidine

Remarks:

without S9 mix, TA1535

Controls 4

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2-aminoanthracene

Remarks:

with S9 mix, TA1535, WP2uvrA

Controls 5

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

Remarks:

with S9 mix, TA1537

Details on test system and experimental conditions:

NUMBER OF REPLICATIONS:
- Number of cultures per concentration: triplicate
- Number of independent experiments : 2

METHOD OF TREATMENT/ EXPOSURE:
- Cell density at seeding: 2x10E9 cells/mL
- Test substance added: preincubation

TREATMENT AND HARVEST SCHEDULE:
- Preincubation period: 20 min
- Exposure duration/duration of treatment: 65 hours at 37°C

METHODS FOR MEASUREMENT OF CYTOTOXICITY
- Method: background growth inhibition

Evaluation criteria:

If mean number of revertants on triplicate plates of the test substance exceed two-fold compared to mean number of revertants on solvent control, and its dose relative appearance and reproducibility are observed, the test substance is judged a mutagen.

Results and discussion

Test results

Test results 1

Key result

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

from 625 µg/plate without S9 mix

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

Test results 2

Key result

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at 5000 µg/plate without S9 mix

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

Test results 3

Key result

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

Test results 4

Key result

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

from 1250 µg/plate without S9 mix

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

Test results 5

Key result

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation and time of the determination: The test compound precipitated at higher concentrations (final concentrations : 5000, 2500, 1250, 625 µg/plate) with and without presence of S9 mix.

STUDY RESULTS
- Concurrent vehicle negative and positive control data : see attachments

- Signs of toxicity : In the case of metabolic activation test, the test compound did not inhibit any bacterial growth. In the test without metabolic activation, the test substance showed inhibition of the bacterial growth at concentration of 1250 µg/plate or higher. E. coli was not inhibited at all.

- Individual plate counts : see attachments

- Mean number of revertant colonies per plate and standard deviation : see attachments

HISTORICAL CONTROL DATA
- no data

Applicant's summary and conclusion

Conclusions:

The substance was tested for mutagenic activity in the Ames-test with the bacterial strains S. typhimurium TA98, TA100,TA1535, TA1537, TA1538 and E.coli WP2 uvrA with and without metabolic activation in two independent studies. No increase of the number of revertants per plate was observed.

Executive summary:

A study similar to OECD TG 471 was performed using Salmonella typhimurium (TA100, TA1535, TA98 and TA1537) and Escherichia coli (WP2 uvrA) with and without metabolic activation. This study consisted of two independent preincubation tests. Dimethylsulfoxide was used as solvent for preparation of the test substance. Eight concentrations of the test substance (5000, 2500, 1250, 625, 312.5, 156.3, 78.1, 39.1 µg/plate) were used in the tests. The limit concentration required in the guideline was used as the highest concentration. No increases in the numbers of revertant colonies of bacteria were recorded for any of the strains of bacteria used, at any concentration, either with or without metabolic activation.