Registration Dossier

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: well performed GLP and OECD guideline like study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1987
Report Date:
1987

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: Company guideline following the test method published by: B.N. Ames et al (1973), B.N. Ames et al (1976), M.H.L. Green et al (1976), A.P. Alvares et al (1973), J. McCann (1975) et al, which was similar to OECD 471.
Deviations:
no
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent

Method

Species / strainopen allclose all
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2
Species / strain / cell type:
S. typhimurium TA 1538
Metabolic activation:
with and without
Metabolic activation system:
Aroclor 1254 (TN) induced rat liver S9-mix
Test concentrations with justification for top dose:
First experiment: 0, 4, 20, 100, 500, 2500, 10000 µg/plate
Second experiment: 0, 0.16, 0.8, 4, 20, 100, 500, 2500, 5000 µg/plate
Vehicle / solvent:
DMSO
Controls
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
9-aminoacridine
2-nitrofluorene
sodium azide
benzo(a)pyrene
other: N-Methyl-N-nitro-N-nitrosoguanidine, 2-Aminoanthracene
Details on test system and experimental conditions:
METHOD OF APPLICATION: plate incorporation method

DURATION
- Incubation period: 48 to 72 hour at 37 °c in the dark

NUMBER OF REPLICATIONS: three plates per dose, two independent experiments


DETERMINATION OF CYTOTOXICITY
- Method: A reduced rate of spontaneously occuring colonies as well as visible thinning of the bacterial lawn were used as indicator

Evaluation criteria:
- dose dependent increase in the number of revertant colonies
- relevant increases in the number of revertant colonies
Statistics:
mean of three plates

Results and discussion

Test resultsopen allclose all
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2
Metabolic activation:
with and without
Genotoxicity:
positive
Remarks:
Without S9-Mix: dose a dependent increase of revertant colonies in Salmonella strainsTA 98, TA 100, TA 1537 and E. coli WP2uvrA. With S9-Mix: relevant increases of revertant colonies in Salmonella strains TA 98, TA 100, TA 1537 and E. coli WP.
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
at doses of 100 µg/plate
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Species / strain:
S. typhimurium TA 1538
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
at doses of 2500 µg/plate
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
positive

3-Nitro-4-methoxybenzoesäure is mutagenic with and without exogenous metabolic activation in this bacterial reverse mutation assay.
Executive summary:

3-Nitro-4-methoxybenzoesäure was tested for mutagenicity with the strains TA 100, TA 1535, TA 1537, TA 1538, TA 98 of Salmonella typhimurium and Escherichia coli WP2uvrA. The mutagenicity studies were conducted in the absence and in the presence of a metabolizing system derived from rat liver homogenate. A dose range of 8 different doses from 0.16 µg/plate to 5000 µg/plate was used. Control plates (solvent control) without test item showed that the number of spontaneous revertant colonies was similiar to that described in the literature. All the positive control compounds gave the expected increase in the number of revertant colonies. Toxicity: The test compound proved to be very toxic to the bacterial strains at 100 µg/plate. 5000 microgram/plate was chosen as top dose level for the mutagenicity study. Mutagenicity: In the absence of the metabolic activation system, the test compound gave a dose dependent increase in the number of revertant colonies with the bacterial strains Salmonella TA 98, TA 100, TA 1537, and E. coli WP2uvrA. Also in the presence of metabolic activation, treatment of the cells with 3-Nitro-4-methoxybenzoesäure results in relevant increases in the number of revertant colonies with the Salmonella strains TA 98, TA 100, TA 1537, and E. coli WP2uvrA. Summarizing, it can be stated that 3-Nitro-4-methoxybenzoesäure is mutagenic with and without exogenous metabolic activation in this bacterial reverse mutation assay.