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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
24 May 2000 - 30 May 2000
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2000
Report date:
2001

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay

Test material

1
Chemical structure
Reference substance name:
3a,4,5,6,7,7a-hexahydro-4,7-methano-1H-indenyl propionate
EC Number:
272-805-7
EC Name:
3a,4,5,6,7,7a-hexahydro-4,7-methano-1H-indenyl propionate
Cas Number:
68912-13-0
Molecular formula:
C13H18O2
IUPAC Name:
3a,4,5,6,7,7a-hexahydro-4,7-methano-1H-inden-6-yl propionate
Test material form:
liquid

Method

Target gene:
His-gene and Trp-Gene
Species / strain
Species / strain / cell type:
S. typhimurium, other: TA97a, TA98, TA100, TA102, TA1535
Details on mammalian cell type (if applicable):
Not applicable
Additional strain / cell type characteristics:
not applicable
Metabolic activation:
with and without
Metabolic activation system:
S9
Test concentrations with justification for top dose:
Study 1
Without metabolic activation:
- TA 97a, TA 98, TA 100, TA 1535, TA 102: 0.0016 - 0.005 - 0.016 - 0.05 - 0.16 mg/plate
With metabolic activation:
- TA 100, TA 1535: 0.0016 - 0.005 - 0.016 - 0.05 - 0.16 mg/plate
- TA 97a: 0.005 - 0.016 - 0.05 - 0.16 - 0.5 - 1.6 mg/plate
- TA 98, TA 102: 0.016 - 0.05 - 0.16 - 0.5 - 1.6 mg/plate

Study 2:
Without metabolic activation:
- TA 97a, TA 1535: 0.0016 - 0.005 - 0.016 - 0.05 - 0.16
- TA 98, TA 100, TA 102: 0.005 - 0.016 - 0.05 - 0.16 - 0.5
With metabolic activation:
- TA 97a: 0.0016 - 0.005 - 0.016 - 0.05 - 0.16
- TA 100, TA 1535: 0.005 - 0.016 - 0.05 - 0.16 - 0.5
- TA 98, TA 102: 0.016 - 0.05 - 0.16 - 0.5 - 1.6
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: dimethylsulfoxide (DMSO)
Controls
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
sodium azide
cumene hydroperoxide
other: ICR 191, 4-Nitro-1,2-phenylenediamine, Nitrofurantoine, 2-Aminoanthraceen, Danthron
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 48 hours (at 37C)

SELECTION AGENT (mutation assays): agar containing Histidine or Tryptophan

NUMBER OF REPLICATIONS: Three replicates per concentration. Titer control replicates in 2-fold.

DETERMINATION OF CYTOTOXICITY: reduced background lawn was regarded to be a cytotoxic effect. Plates with reduced background lawn were not included into evaluation procedures.

VALIDITY CRITERIA: the following genotypes of the tested strains had to be confirmed:
- Histidine auxotrophy
- Ampicillin resistance
- Tetracycline resistance (only TA 102)
- UV-sensitivity (except TA 102)
- Growth inhibition with crytal violet

Titer of the overnight culture >= 1x10^8 cells/ml.
Spontaneous revertant (negative controls) had to be within the following ranges:
- TA 97a: 60 - 300
- TA 87: 15 - 50
- TA 100: 60 -200
- TA 102: 300 -600
- TA 1535: 5 - 30
Evaluation criteria:
Plates with reduced background lawn were not included into evaluation procedures. The induction rate of mean valiues was calculates as revertant colonies of test item/revertant colonies of the corresponding control. The test item is to be interpretated mutagenic if there is a concentration effect relationship and the induction rate is >= 2.
Statistics:
Arithmetic mean values and standard deviations were calculated from colonies per plate of three replicates.

Results and discussion

Test results
Key result
Species / strain:
S. typhimurium, other: TA97a, TA98, TA100, TA102, TA1535
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
ADDITIONAL INFORMATION ON CYTOTOXICITY: Cytotoxicity was observed in the highest concentrations of all dose levels, and therefore these concentrations were not included in the evaluation process.

Applicant's summary and conclusion

Conclusions:
In a bacterial reverse mutation assay (Ames test), cyclaprop did not show mutagenic properties, under the conditions of this test.
Executive summary:

The mutagenic effects of the test item Cyclaprop were determined in a bacterial reverse mutation assay according to OECD guideline 471 in two independent studies. Test systems were the Salomonella Typhimurium strains TA 97a, TA 98, TA 100, TA 102 and TA 1535 with and without the metabolic actication system S9 (from male Wistar rats) each. Positive and negative controls were included in each study. The duration of each study was 48 hours. The test item was dissolved in DMSO and applied once at test initiation with concentrations ranging from 0.0016 mg/plate - 1.6 mg/plate. The validity criteria of the test were fulfilled.

Cyclaprop did not induce any mutagenicity. Cytotoxicity occured at the highest concentrations tested, therefore these concentrations were not included in the evaluation. Cyclaprop was found to have no mutagenic effects on Salmonella Typhimurium strains with and without S9, under the conditions of this test. According to the criteria outlined in Annex VI of 67/548/EEC (DSD) and Annex I of 1272/2008/EEC (CLP), Cyclaprop does not have to be classified as mutagenic.