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Toxicological information

Genetic toxicity: in vitro

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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
Study period:
18 January - 20 January 1983
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1983
Report date:
1983

Materials and methods

Test guideline
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Principles of method if other than guideline:
Salmonella typhimurium strains TA98, TA100, TA1535, TA1537 and TA1538 were tested in the Ames test.
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay

Test material

Constituent 1
Chemical structure
Reference substance name:
Reaction mass of 2-methylbutyl acetate and pentyl acetate
EC Number:
908-918-1
Molecular formula:
Unspecified
IUPAC Name:
Reaction mass of 2-methylbutyl acetate and pentyl acetate
Details on test material:
Primary amyl acetate consists of CH3COOC5H11 (mixed isomers: 60% amyl acetate, 35% 2-methylbutylacetate and 5% 3-methylbutylacetate)

Method

Target gene:
histidine gene
Species / strain
Species / strain / cell type:
S. typhimurium, other: TA98, TA100, TA1535, TA1537 and TA1538
Metabolic activation:
with and without
Metabolic activation system:
S9 liver homogenate prepared from Aroclor 1254-induced Sprague-Dawley male rats.
Test concentrations with justification for top dose:
0.01, 0.03, 0.1, 0.3 and 1.0 ul
Vehicle / solvent:
ethanol
Controls
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: 4-nitro-o-phenylenediamine, sodium azide, 9-aminoacridine and 2-aminoanthracene
Details on test system and experimental conditions:
Sample Preparation: The test substance was dissolved in absolute ethanol to a concentration of 6 ul/ml; all dilutions were made in the same solvent. Dilutions of the test substance were made fresh each day of testing.

Dose Selection: A preliminary toxicity test was performed using strain TA100 to determine the level of toxicity of the test substance. Ten doses were tested for toxicity with a plate assay performed in the manner used for mutagenicity determinations. Toxicity was assessed at 24 to 48 hours after treatment by either growth inhibition of the background lawn or a reduction in the number of spontaneous mutants.

Testing: The test chemical was tested in triplicate at five doses chosen to span a range which included moderately toxic to relatively nontoxic concentrations. If the substance was nontoxic in the preliminary toxicity test, 50 ul of liquid or 5000 ug of solid (or up to the limit of solubility) was used as the maximum dose. Testing was performed both with and without metabolic activation. Concurrent solvent and positive controls were run in each test.
Evaluation criteria:
The spontaneous reversion for the solvent controls must be within this laboratory's historical range. The positive controls must demonstrate that the test systems are responsive with know mutagens. A test chemical is considered a bacterial mutagen if the number of revertant colonies is at least twice the solvent control for at least one dose level and there is evidence of a dose-related increase in the number of revertant colonies. If a test chemical produces a marginal or weak response that cannot be reproduced, the initial positive results loses significance. If there is no evidence of a dose-related increase in the number of revertant colonies and the number of revertant colonies is not twice the solvent control, then the test chemical is not considered to be a bacterial mutagen.
Statistics:
Mean and standard deviations were calculated.

Results and discussion

Test results
Species / strain:
S. typhimurium, other: TA98, TA100, TA1535, TA1537 and TA1538
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
not specified
Positive controls validity:
valid
Additional information on results:
Primary amyl acetate was tested in a preliminary toxicity screen at 50, 10, 3, 1, 0.3, 0.1, 0.03, 0.01, 0.003 and 0.001 ul per plate with strain TA100 only. Because 100 ul of the solvent ethanol was toxic in a preliminary test, the test chemical was diluted so that 50 ul of the dilution would deliver the required dose. Phosphate-buffered saline was used to bring the total volume added per tube to 100 ul. The top four doses of 50, 10, 3 and 1 ul per plate inhibited all growth of the background lawn, while the 0.3 ul dose allowed some sparse growth of the background lawn. Based on these results, mutagenicity testing was done with 5 doses of 1.0, 0.3, 0.1, 0.03 and 0.01 ul per plate in triplicate.

No evidence of mutagenicity was observed at any of the tested doses, either by evidence of a dose-response relationship or a doubling of the number of colonies over the solvent control value.

Dose selection appeared to be in a suitable range in the mutagenicity tests because some toxicity was evident in both tests. In the tests with and without metabolic activation, toxicity was observed at 1 ul/plate with all strains as a slight reduction in the number of colonies per plate.

All strains exhibited a positive mutagenic response with the positive controls tested both with and without S9 metabolic activation. Negative (solvent) controls were also tested with each strain, and the average spontaneous reversion rates were within the historical range at this laboratory. All positive and negative controls were run concurrently with the test chemical. Concurrently run sterility checks showed that the S9 mix, PBS, the test chemical and all solvents and controls were sterile
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Any other information on results incl. tables

No additional information available.

Applicant's summary and conclusion

Executive summary:

Primary Amyl Acetate was tested for potential mutagenic activity using the Salmonella/microsome bacterial mutagenicity assay (Ames test). Primary Amyl Acetate was tested in triplicate at five dose levels, ranging from 0.01 microliters to 1.0 microliter per plate. Mutagenic activity was not observed with any of the five bacterial strains tested with or without metabolic activation. Primary Amyl Acetate was not mutagenic in this screening test.