Registration Dossier

Diss Factsheets

Toxicological information

Genetic toxicity: in vitro

Currently viewing:

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:
comparable to guideline study

Data source

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

Materials and methods

Test guideline
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay

Test material

Constituent 1
Chemical structure
Reference substance name:
3-(4-hydroxy-4-methylpentyl)cyclohex-3-ene-1-carbaldehyde; 4-(4-hydroxy-4-methylpentyl)cyclohex-3-ene-1-carbaldehyde
EC Number:
915-617-9
Molecular formula:
C13H22O2
IUPAC Name:
3-(4-hydroxy-4-methylpentyl)cyclohex-3-ene-1-carbaldehyde; 4-(4-hydroxy-4-methylpentyl)cyclohex-3-ene-1-carbaldehyde
Test material form:
liquid

Method

Target gene:
S. typhimurium TA1535: hisG46 rfa uvrB; S. typhimurium TA1537: hisC3076 rfa uvrB; S. typhimurium TA 1538: hisD3052 rfa uvrB; S. typhimurium TA98: hisD3052 rfa uvrB pKMl01; S. typhimurium TA100: hisG46 rfa uvrB pKM101; E. coli WP2 uvrA: trp
Species / strainopen allclose all
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Species / strain / cell type:
E. coli WP2 uvr A
Metabolic activation:
with and without
Metabolic activation system:
Rat S9
Test concentrations with justification for top dose:
In the preliminary cytotoxicity assay: 0, 6.7, 10, 33, 67, 100, 333, 667, 1000, 3333 and 5000 μg/plate
In the main study: 0, 75, 200, 600, 1800 and 5000 μg/plate
Vehicle / solvent:
- Solvent used: DMSO
- Justification for choice of solvent:based on the solubility of the test substance and compatibility with the target cells
- Solubility in DMSO: the test article was soluble in dimethyl sulfoxide at approximately 500 mg/mL, the maximum concentration tested
Controls
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
9-aminoacridine
2-nitrofluorene
sodium azide
methylmethanesulfonate
other: 2-aminoanthracene (with S9)
Details on test system and experimental conditions:
METHOD OF APPLICATION:
The assay was performed in two phases, using the plate incorporation method. The first phase, the preliminary toxicity assay, was used to establish the dose range for the mutagenicity assay. The second phase, the mutagenicity assay, was used to evaluate the mutagenic potential of the test article.

INCUBATION CONDITIONS:
- Duration: 48 - 72 hr
- Temperature: 37 °C

NUMBER OF REPLICATIONS:
- triplicate

DETERMINATION OF CYTOTOXICITY
- Method: relative total growth

Evaluation criteria:
The test article was considered to give a positive result, if it caused a dose-related increase in the mean revertants per plate of at least one tester strain with a minimum of two increasing concentrations of test article. Data sets for strains TA1535 and TA1537 were judged positive if the increase in mean revertants at the peak of the dose response is equal to or greater than 3 times the mean vehicle control value. Data sets for strains TA98, TA100 and WP2 uvrA were judged positive if the increase in mean revertants at the peak of the dose response is equal to or greater than two times the mean vehicle control value.

Results and discussion

Test resultsopen allclose all
Key result
Species / strain:
S. typhimurium TA 1535
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
True negative controls validity:
not applicable
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium TA 1537
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
True negative controls validity:
not applicable
Positive controls validity:
valid
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:
valid
True negative controls validity:
not applicable
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
True negative controls validity:
not applicable
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:
cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
True negative controls validity:
not applicable
Positive controls validity:
valid
Additional information on results:
RANGE-FINDING/SCREENING STUDIES:
- In the preliminary cytotoxicity assay, no precipitation was observed, but toxicity was observed at 5000 μg/plate with some test conditions. Based on the results of the study, 5000 μg/plate was selected as a maximum concentration for the main study.

MAIN STUDY:
- In the mutagenicity assay, no precipitate was observed but toxicity was generally observed at 5000 µg per plate with test strains TA98 and TA1537 in the absence of S9 activation. Non-dose responsive increases were observed in the tester strains TA98 (1.8-fold, maximum increase) and TA100 (1.5-fold, maximum increase) in the absence of S9 activation. These two strains/activation combinations were retested to clarify the responses observed. A non-dose responsive increase was observed again with tester strain TA98 (2.4-fold, maximum increase) in the absence of S9 activation. The maximum revertant count was within the normal historical vehicle control range and is not believed to be biologically relevant.

Applicant's summary and conclusion

Conclusions:
The test substance gave negative results in the Ames test with S. typhimurium strains TA98, TA100, TA1535 and TA1537 and E. coli strain WP2 uvr A with and without metabolic activation at concentration levels up to and including 5000 μg/plate.
Executive summary:

The ability of the test substance to cause gene mutations in prokaryotic cells was investigated in a study, comparable to OECD guideline 471 in compliance with GLP. S. typhimurium strains TA98, TA100, TA1535 and TA1537 and E. coli strain WP2 uvr A were treated with 0 (vehicle control), 75, 200, 600, 1800 and 5000 μg/plate of the test substance in DMSO in the presence and absence of metabolic activation, using a plate incorporation method. The doses were selected based on the results of a preliminary range-finding study. Cytotoxicity was observed at the highest concentration levels in strains TA98 and TA1537. No statistically significant increase in the number of revertants was noted in any cases, indicating that the substance gave a negative result in the Ames test.