Oxacycloheptadecan-2-one

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:

22 August 1997 - 9 September 1997

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 guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

26 May 1983

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

OECD Guideline 472 (Genetic Toxicology: Escherichia coli, Reverse Mutation Assay)

Version / remarks:

26 May 1983

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)

Version / remarks:

29 December 1992

Deviations:

no

Test guideline 4

Qualifier:

according to guideline

Guideline:

other: US EPA, Method: HG-GEne Muta - S. typhimurium: The Salmonella typhimurium revere mutation assay

Version / remarks:

1984

Deviations:

no

Test guideline 5

Qualifier:

according to guideline

Guideline:

other: US EPA, Method: HG-Gene Muta - E. coli: The Escherichia coli reverse utation assay, 1984

Test guideline 6

Qualifier:

according to guideline

Guideline:

JAPAN: Guidelines for Screening Mutagenicity Testing Of Chemicals

Version / remarks:

28 January 1985, 11 September 1989, 31 March 1987, 16 June 1987

Deviations:

no

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Oxacycloheptadecan-2-one

EC Number:

203-662-0

EC Name:

Oxacycloheptadecan-2-one

Cas Number:

109-29-5

Molecular formula:

C16H30O2

IUPAC Name:

oxacycloheptadecan-2-one

Test material form:

solid: crystalline

Method

Target gene:

S. typhimurium: his
E. coli: trp

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2

Species / strain 2

Species / strain / cell type:

S. typhimurium TA 1538

Metabolic activation:

with and without

Metabolic activation system:

Aroclor 1254-induced rat liver S9

Test concentrations with justification for top dose:

Preliminary toxicity test: 5, 50, 500 and 5000 µg/plate
Mutation test 1: 39.1, 78.1, 156.3, 312.5, 625, 1250, 2500 and 5000 µg/plate
Mutation test 2: 156.3, 312.5, 625, 1250, 2500 and 5000 µg/plate
The top dose corresponded to the maximal solubility of the test substance in the chosen solvent.

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: based on the solubility of the test substance (insoluble in water)

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

2-nitrofluorene

N-ethyl-N-nitro-N-nitrosoguanidine

other: 2-aminoanthracene

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 3 days

NUMBER OF REPLICATIONS: two independent experiments, each dose in each experiment tested in triplicate

DETERMINATION OF CYTOTOXICITY
- Method: reduction in revertant colony counts or absence of a complete background bacterial lawn

Evaluation criteria:

If treatment with a test substance produces an increase in revertant colony numbers of at least twice the concurrent solvent controls, with some evidence of a positive dose-relationship, in two separate experiments, with any bacterial strain either in the presence or absence of S09 mix, it is considered to show evidence of mutagenic activity in this test system.
If treatment with a test substance does not produce reproducible increases of at least 1.5 times the concurrent solvent controls, at any dose level with any bacterial strain, it is considered to show no evidence of mutagenic activity in this test system. No statistical analysis is performed.
If the results obtained fail to satisfy the criteria for a clear "positive" or "negative" response, the following approach is taken in order to resolve the issue of the substance's mutagenic activity in this test system:
1) Repeat tests may be performed using modifications of the experimental method. These modifications include (but are not restricted to), the use of a narrower dose range than that alrady tested; the use of different levels of liver homogenate S-9 fraction in the S-9 mix. Should an increase in revertant colony numbers be observed which satisfies the first paragraph, the substance is considered to show evidence of mutagenic activity in this test system. No statistical analysis is performed.
2) IF no clear "positive" response is obtained, the test data may be subjected to analysis to determine the statistical significance of any observed increases in revertant colony numbers. The statistical procedures used will be those described by Mahon et al. (1989) and will usually be analysis of variance followed by Dunnett's test.

Statistics:

No statistical analysis was performed.

Results and discussion

Test results

Test results 1

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 applicable

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

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Test results 3

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:

not applicable

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

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Test results 5

Key result

Species / strain:

S. typhimurium TA 1538

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Test results 6

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:

not applicable

Positive controls validity:

valid

Additional information on results:

RANGE-FINDING/SCREENING STUDIES:
Four concentrations of test substance were assessed for toxiciyt using the six tester strains. The highest concentration was 50 mg/mL of test substance in the chosen solvent, which provided a final concentration of 5000 µg/plate. Three 10-fold serial diluions of the highest concentration were also tested. The chosen solvent, dimethyl sulphoxide, was used as the negative control. Any toxic effects of the test substance can be detected by a substantial reduction in revertant colony counts or by the absence of a complete background bacterial lawn.

HISTORICAL CONTROL DATA (with ranges, means and standard deviation and confidence interval (e.g. 95%)
- Positive historical control data, mean (minimum-maximum):
TA 1535:
ENNG, -S9: 351.9 (68-2849)
AA, +S9: 151.3 (44-2171)
TA1537:
9-AC, -S9: > 3500 (> 3500- > 3500)
AA, +S9: 82.4 (29-714)
TA1538:
NF, -S9: 403.2 (79-767)
AA, +S9: 174.3 (40-1725)
TA98:
NF, -S9: 277.7 (68-782)
AA, +S9: 175.3 (57-1617)
TA 100:
ENNG, -S9:
472.3 (178-2564)
AA, +S9: 443.0 (159-2498)
WP2 uvrA:
ENNG, - S9: 853.4 (373-2331)
AA, +S9: 394.5 (143-2345)

- Negative (solvent/vehicle) historical control data:
TA1535: -S9: 15.8 (8-20), +S9: 16.3 (7-21)
TA1537: -S9: 10.0 (4-16), +S9: 11.1 (2-19)
TA1538: -S9: 13.0 (7-21), +S9: 14.5 (8-23)
TA98: -S9: 24.5 (16-34), +S9: 26.9 (17-37)
TA100: -S9: 125.4 (81-168), +S9: 129.7 (80-165)
WP2 uvrA: -S9: 68.1 (43-88); +S9: 67.9 (46-88)

ADDITIONAL INFORMATION ON CYTOTOXICITY:
The test substance was toxic towards all the tester strains at 5000 µg/plate. 5000 µg/plate was chosen as the top dose elvel in the first mutation test, but a total of eight dose levels were included to ensure that sufficient non-toxic concentrations were assessed.
No other toxicity except at the highest dose level was observed in the first mutation test, therefore a total of only six dose levels were chosen for the second test.

Applicant's summary and conclusion

Conclusions:

The test substance oxacycloheptadecan-2-one was not mutagenic in the Ames test performed according to OECD guidelines 471 and 472, both with and without metabolic activation, when tested up to cytotoxic concentrations.

Executive summary:

In a GLP-compliant OECD Guideline 472 and 472 study, oxacycloheptadecan-2 -one was tested in S. typhimurium TA98, TA100, TA1535, TA1537 and TA1538 strains and E. coli WP2 uvrA strain both in the absence and presence of metabolic activation, up to the maximal concentration of 5000 µg/plate. Cytotoxicity was observed at the highest concentration. Two independent mutation tests were performed, and each concentration was tested in triplicate. No evidence of mutagenic activity was seen at any dose level in either mutation test. The concurrent positive controls produced satisfactory results. Therefore oxacycloheptadecan-2 -one was concluded to be non-mutagenic in bacterial cells under the conditions of the study.