3-Pentanone, 1-hydroxy-2-methyl-

Genetic toxicity: in vitro

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:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2016

Report date:

2016

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.5100 - Bacterial Reverse Mutation Test (August 1998)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

1-hydroxy-2-methylpentan-3-one

EC Number:

939-893-5

Cas Number:

27970-79-2

Molecular formula:

C6 H12 O2

IUPAC Name:

1-hydroxy-2-methylpentan-3-one

Test material form:

liquid

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Test substance No.: 15/0439-1, Batch identification: J 1812
- Expiration date of the lot/batch: 08 Jul 2017
- Purity: 90.6 area-%; water content: 0.3 g/100 g

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature (protect against humidity; no direct
sunlight)
- Solubility and stability of the test substance in the solvent/vehicle: the test substance was administered immediately after preparation and is usually stable.

Method

Species / strain

Species / strain / cell type:

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

Metabolic activation:

with and without

Metabolic activation system:

S9 obtained from livers of rats treated with phenobarbital and β-naphthoflavone

Test concentrations with justification for top dose:

Duue to the purity of the test substance 5.5 mg/plate was used as top dose in all experiments.

Vehicle / solvent:

- Solvent used: DMSO
- Justification for choice of solvent/vehicle: Due to insolubility of the test substance in water, DMSO was used as vehicle

Controls

Untreated negative controls:

yes

Remarks:

Sterility control treated with soft agar, S9 mix, buffer, vehicle test substance without tester strains.

Negative solvent / vehicle controls:

yes

Remarks:

The vehicle control with and without S9 mix only contains the vehicle used for the test substance

True negative controls:

no

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

9-aminoacridine

other: 2-aminoanthracene (2-AA), N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), 4-nitro-o-phenylenediamine (NOPD)

Details on test system and experimental conditions:

METHOD OF APPLICATION: plate incorporation method and preincubation Test

DURATION
- Preincubation period: 20 minutes
- Exposure duration: 48 – 72 hours

SELECTION AGENT (mutation assays):
Salmonella typhimurium: amino acid solution (minimal amino acid solution for the determination of mutants: 0.5 mM histidine + 0.5 mM biotin)
Escherichia coli: amino acid solution (minimal amino acid solution for the determination of mutants: 0.5 mM tryptophan)

NUMBER OF REPLICATIONS:
1st Experiment (Standard plate test with and without S9 mix): 3 test plates per dose or per control
2nd Experiment (Preincubation test with and without S9 mix): 3 test plates per dose or per control
3rd Experiment (Preincubation test without S9 mix): 3 test plates per dose or per control

DETERMINATION OF CYTOTOXICITY
- Method: Toxicity detected by either a decrease in the number of revertants (factor ≤ 0.6)
or clearing or diminution of the background lawn (= reduced his- or trp- background growth).

Evaluation criteria:

The test substance was considered positive in this assay if the following criteria were met:

• A dose-related and reproducible increase in the number of revertant colonies, i.e. at least doubling (bacteria strains with high spontaneous mutation rate, like TA 98, TA 100 and E.coli WP2 uvrA) or tripling (bacteria strains with low spontaneous mutation rate, like TA 1535 and TA 1537) of the spontaneous mutation rate in at least one tester strain either without S9 mix or after adding a metabolizing system.

A test substance was generally considered non-mutagenic in this test if:
• The number of revertants for all tester strains were within the range of the historical negative control data under all experimental conditions in at least two experiments carried out independently of each other.

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Remarks:

A slight increase in the number of his+ revertants was observed in the preincubation test without S9 mix at 5500 μg/plate (Factor 2.0). This finding was not reproduced in a repeat experiment.

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 3

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 4

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Remarks:

A slight increase in the number of his+ revertants was observed in the preincubation test without S9 mix at 5500 μg/plate (Factor 1.9). This finding was not reproduced in a repeat experiment.

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 5

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Untreated negative controls validity:

valid

Positive controls validity:

valid

Applicant's summary and conclusion

Conclusions:

Under the experimental conditions chosen here, it is concluded that Hydroxymethylpentanon is not a mutagenic test substance in the bacterial reverse mutation test in the absence and the presence of metabolic activation.