4-methylpent-3-en-2-one

Genetic toxicity: in vitro

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:

June 2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: fully GLP compliance

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2010

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Mesityl Oxide

IUPAC Name:

Mesityl Oxide

Constituent 2

Reference substance name:

4-methylpent-3-en-2-one

EC Number:

205-502-5

EC Name:

4-methylpent-3-en-2-one

Cas Number:

141-79-7

Molecular formula:

C6H10O

IUPAC Name:

4-methylpent-3-en-2-one

Details on test material:

- Name of test material (as cited in study report): Mesityl Oxide
- Physical state: liqid
- Composition of test material, percentage of components: 99.87%
- Purity test date: 16-Apr-2010
- Lot/batch No.: A1YB3N010101
- Expiration date of the lot/batch: 16-Apr-2012
- Storage condition of test material: Ambient condition

Method

Target gene:

S. typhimurium tester strains: histidine operon - E. coli: tryptophan operon

Species / strain

Species / strain / cell type:

other: Salmonella typhimurium TA1535, TA1537, TA98, TA100 and Escherichia coli WP2 uvrA

Additional strain / cell type characteristics:

other: uvrA, uvrB: Sensitivity to UV irradiation. rfa: Sensitivity to Crystal Violet. pKM101: Resistance to Ampicillin.

Metabolic activation:

with and without

Metabolic activation system:

S9 rat liver tissue fraction - Inducing Agents :Phenobarbital – 5,6-Benzoflavone

Test concentrations with justification for top dose:

Preliminary toxicity test: 5000, 2500, 500, 158 and 50.0 µg/ml
Main Assay I (plate incorporation): 5000, 2500, 1250, 625 and 313 µg/plate
Main Assay II (pre-incubation): 5000, 2500, 1250, 625 and 313 µg/plate

Vehicle / solvent:

dimethylsulfoxide (DMSO)

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: Sodium azide: TA1535, TA100 without S9 metabolism; 9-amino-acridine TA1537 without S9 metabolism; methylmethanesulphonate WP2 uvrA without S9 metabolism; 2-aminoanthracene all tester strains with S9 metabolism

Details on test system and experimental conditions:

METHOD OF APPLICATION: preliminary toxicity test and main assay I in agar plate incorporation; main assay II preincubation

DURATION
- Preincubation period: 30 minutes at 37 °C
- Incubation time: approximately 72 hours at 37 °C

NUMBER OF REPLICATIONS: three replicate plates for each experimental point

NUMBER OF CELLS EVALUATED: 100 - 500 million of viable bacteria/plate (titre) for each strain.

DETERMINATION OF CYTOTOXICITY
Reduction of revertant numbers and/or thinning of the background lawn

Evaluation criteria:

For the test item to be considered mutagenic, two-fold (or more) increases in mean revertant numbers must be observed at two consecutive dose levels or at the highest practicable dose level only. In addition, there must be evidence of a dose-response relationship showing increasing numbers of mutant colonies with increasing dose levels.

Statistics:

Regression lines, after square root transformation, are calculated using a minimum of the three lowest dose levels and then including the further dose levels in turn. The correlation co-efficient (r), the value of students "t" statistic and the p-value for the regression lines are also calculated.

Results and discussion

Test results

Species / strain:

other: Salmonella typhimurium TA1535, TA1537, TA98, TA100 and Escherichia coli WP2 uvrA

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
Precipitation: No precipitation of the test item was observed at the end of the incubation period at any concentration.

RANGE-FINDING/SCREENING STUDIES:
No toxicity was observed up to 5000 µg/plate
COMPARISON WITH HISTORICAL CONTROL DATA: that mean plate counts for untreated and positive control plates fell within the RTC historical control range
ADDITIONAL INFORMATION ON CYTOTOXICITY:
Main Assay I (plate incorporation method): No toxicity was observed with any tester strain at any dose level, in the absence or presence of S9 metabolism.
Main II (pre-incubation method): Toxicity was observed with all tester strains at the two or three highest concentrations, in the absence and presence of S9 metabolism

Any other information on results incl. tables

No two-fold increases in the number of revertant colonies were observed after treatment at any dose level, in any tester strain, in the absence or presence of S9 metabolism. It must be concluded that the test item Mesityl oxide is not mutagenic to S. typhimurium or E. coli under the reported experimental conditions.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative

It is concluded that the test item Mesityl oxide does not induce reverse mutation in Salmonella typhimurium or Escherichia coli in the absence or presence of S9 metabolism, under the reported experimental conditions.

Executive summary:

The test item Mesityl oxide was examined for the ability to induce gene mutations in tester strains of Salmonella typhimurium and Escherichia coli,as measured by reversion of auxotrophic strains to prototrophy. The five tester strains TA1535, TA1537, TA98, TA100 and WP2uvrA were used. Experiments were performed both in the absence and presence of metabolic activation, using liver S9 fraction from rats pre-treated with phenobarbitone and betanaphthoflavone.  The test item was used as a solution in dimethylsulfoxide (DMSO). Mesityl oxide was assayed in the toxicity test at a maximum concentration of 5000 µg/plate and at four lower concentrations spaced at approximately half-log intervals: 1580, 500, 158 and 50.0 µg/plate. No precipitation of the test item was observed at the end of the incubation period at any concentration. No toxicity was observed with any tester strain at any dose level in the absence or presence of S9 metabolism.   In Main Assay I, using the plate incorporation method, the test item was assayed at the maximum dose level of 5000 µg/plate and at four lower dose levels spaced by two-fold dilutions: 2500, 1250, 625 and 313 µg/plate. No toxicity was observed with any tester strain at any dose level, in the absence or presence of S9 metabolism. No precipitation of the test item was observed at the end of the incubation period at any concentration. As no increases in revertant numbers were observed at any concentration tested, a pre-incubation step was included for all treatments of Main Assay II. The test item was assayed at the same dose levels employed inToxicity was observed with all tester strains at the two or three highest concentrations, in the absence and presence of S9 metabolism. No precipitation of the test item was observed at the end of the incubation period at any concentration. The test item did not induce two-fold increases in the number of revertant colonies in the plate incorporation or pre-incubation assay, at any dose level, in any tester strain, in the absence or presence of S9 metabolism. It is concluded that the test item Mesityl oxide does not induce reverse mutation in Salmonella typhimurium or Escherichia coli in the absence or presence of S9 metabolism, under the reported experimental conditions.