Registration Dossier

Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1999
Report date:
1999

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 471 (Bacterial 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 (incl. QA statement)
Type of assay:
bacterial reverse mutation assay

Test material

Constituent 1
Chemical structure
Reference substance name:
Isopentyl salicylate
EC Number:
201-730-4
EC Name:
Isopentyl salicylate
Cas Number:
87-20-7
Molecular formula:
C12H16O3
IUPAC Name:
3-methylbutyl 2-hydroxybenzoate

Method

Target gene:
Histidine
Species / strain
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
Metabolic activation:
with and without
Metabolic activation system:
liver homogenate (S9) from Aroclor 1254
Test concentrations with justification for top dose:
5 to 5000 µg per plate.
Vehicle / solvent:
- Vehicle: DMSO
Controlsopen allclose all
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Remarks:
water
Positive controls:
yes
Positive control substance:
sodium azide
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Remarks:
DMSO
Positive controls:
yes
Positive control substance:
2-nitrofluorene
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Remarks:
DMSO
Positive controls:
yes
Positive control substance:
other: 2-Aminoanthracene
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Remarks:
water
Positive controls:
yes
Positive control substance:
mitomycin C
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Remarks:
water
Positive controls:
yes
Positive control substance:
9-aminoacridine
Details on test system and experimental conditions:
METHOD OF APPLICATION: standard plate-incorporation assay with and without liver homogenate activation.

DURATION
- Exposure duration: 48 to 72 h at 37°C in the dark

NUMBER OF REPLICATIONS: 3

EVALUATION
- Counting of the number of revertant colonies (his+ revertants)
- Examination normal background lawn
- Examination precipitates
- Microscopically: microcolony growth

OTHER
When there is any question about the nature of colonies scored as revertants and when positive mutagenic results are obtained, the genotype of revertant colonies are spot-checked by picking and streaking on histidine free plates.
To confirm the reversion properties and specificity of the bacterial strains as well as the activity of the liver homogenates used, positive mutagenesis controls were run simultaneously.
The genotypes of the tester strains (histidine requirement, spontaneous reversion frequency, sensitivity to UV-light, crystal violet and ampicillin) were checked in each experiment. The Vogel-Bonner minimal plates, the test compound, and the S9-mix viere checked for sterility.


Results and discussion

Test results
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
500 µg/plate (TA100, TA102, TA1535 and TA1537); 5000 µg/plate (TA98)
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
other: all strains/cell types tested

Applicant's summary and conclusion

Conclusions:
Interpretation of results: negative

The test item was not mutagenic to Salmonella typhimurium strains TA98, TA100, TA102, TA1535, and TA1537 in the presence and absence of a metabolizing system.
Executive summary:

The mutagenicity of the test item was studied with five mutant strains of Salmonella typhimurium (TA1535, TA1537, TA98, TA100, and TA102). The study has been performed according to OECD 471 and GLP.

The investigations were carried out using the standard plate incorporation assay with and without liver homogenate (S9) pretreated male rats as metabolic activation system.

The test item did not induce a significant increase in the mutation frequency of any of the tester strains in the absence and presence of a metabolic activation system. Therefore, the test item is under the experimental conditions described, not mutagenic.