Soybean oil, epoxidized, reaction products with methanol

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:

2012

Report date:

2012

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)

Test guideline 3

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes (incl. QA statement)

Remarks:

BASF SE, Experimental Toxicology and Ecology, 67056 Ludwigshafen, Germany

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Soybean oil, epoxidized, reaction products with methanol

EC Number:

287-837-7

EC Name:

Soybean oil, epoxidized, reaction products with methanol

Cas Number:

85586-35-2

Molecular formula:

C H4 O . Unspecified

IUPAC Name:

1-({13-[(1-{2,3-bis[(9,13-dihydroxy-10,12-dimethoxyoctadecanoyl)oxy]propoxy}-10-methoxy-1-oxooctadecan-9-yl)oxy]-9-hydroxy-10,12-dimethoxyoctadecanoyl}oxy)-3-[(9-hydroxy-10-methoxyoctadecanoyl)oxy]propan-2-yl 9,13-dihydroxy-10,12-dimethoxyoctadecanoate; 1-[(9,13-dihydroxy-10,12-dimethoxyoctadecanoyl)oxy]-3-[(9-hydroxy-10-methoxyoctadecanoyl)oxy]propan-2-yl 9,13-dihydroxy-10,12-dimethoxyoctadecanoate; 3-({8-[3-(3-hydroxy-2-methoxyoctyl)oxiran-2-yl]octanoyl}oxy)-2-[(8-{3-[(3-pentyloxiran-2-yl)methyl]oxiran-2-yl}octanoyl)oxy]propyl 9-hydroxy-10-methoxyoctadecanoate; 3-[(8-{3-[3-({1-[3-({8-[3-(3-hydroxy-2-methoxyoctyl)oxiran-2-yl]octanoyl}oxy)-2-[(8-{3-[(3-pentyloxiran-2-yl)methyl]oxiran-2-yl}octanoyl)oxy]propoxy]-10-methoxy-1-oxooctadecan-9-yl}oxy)-2-methoxyoctyl]oxiran-2-yl}octanoyl)oxy]-2-[(8-{3-[(3-pentyloxiran-2-yl)methyl]oxiran-2-yl}octanoyl)oxy]propyl 9-hydroxy-10-methoxyoctadecanoate

Details on test material:

- Name of test material: Soybean oil, epoxidized, reaction products with methanol
- Physical state: Liquid / yellow.
- Homogeneity: The homogeneity of the test substance was ensured by mixing before preparation of the test substance solutions.
- Storage condition of test material: Room temperature.
- Storage stability: The stability of the test substance under storage conditions throughout the study period is guaranteed until Dec 2013 as indicated by the sponsor, and the sponsor holds this responsibility.
- Expiration date of the batch: Dec 2013
- Batch identification: CE61910022
- Purity/composition: water: 0.1 g/100 g (see analytical report 12L00136)

Method

Target gene:

Histidine and tryptophan.

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Details on mammalian cell type (if applicable):

The Salmonella strains are checked for the following characteristics at regular intervals: deep rough character (rfa); UV sensitivity (Ä uvrB); ampicillin resistance (R factor plasmid). Histidine and tryptophan auxotrophy is checked in each experiment via the spontaneous rate.

Species / strain 2

Species / strain / cell type:

E. coli WP2 uvr A

Details on mammalian cell type (if applicable):

E. coli WP2 uvrA is checked for UV sensitivity. Histidine and tryptophan auxotrophy is checked in each experiment via the spontaneous rate.

Metabolic activation:

with and without

Metabolic activation system:

phenobarbital/β-naphthoflavone induced rat livers

Test concentrations with justification for top dose:

33, 100, 333, 1000, 2750 and 5500 μg/plate

Vehicle / solvent:

DMSO. Which had been demonstrated to be suitable in bacterial reverse mutation tests and for which historical control data are available

Controls

Controls 1

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

N-ethyl-N-nitro-N-nitrosoguanidine

Remarks:

No metabolic activation, Strains: TA 1535 and TA 100

Controls 2

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 4-nitro-o-phenylenediamine

Remarks:

No metabolic activation, Strains: TA 98

Controls 3

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

Remarks:

No metabolic activation, Strains: TA 1537

Controls 4

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

Remarks:

No metabolic activation, Strains: E. coli WP2 uvrA

Controls 5

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2-aminoanthracene.

Remarks:

With metabolic activation and for all strains

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation); preincubation

DURATION
Standard plate test
- Exposure duration: 48 - 72 hours at 37°C in the dark
Preincubation test
- Preincubation period: 20 minutes at 37°C
- Exposure duration: 48 - 72 hours at 37°C in the dark

NUMBER OF REPLICATIONS: 3 test plates per dose or per control

DETERMINATION OF CYTOTOXICITY
- Method: number of revertants, background lawn, determination of titer

Evaluation criteria:

ACCEPTANCE CRITERIA:
Generally, the experiment is considered valid if the following criteria are met: The number of revertant colonies in the negative controls was within the range of the historical negative control data for each tester strain; The sterility controls revealed no indication of bacterial contamination; The positive control substances both with and without S9 mix induced a distinct increase in the number of revertant colonies within the range of the historical positive control data or above; Fresh bacterial culture containing approximately 10^9 cells per mL were used. For approval the titer of viable bacteria was ≥ 10^8 colonies per mL.

ASSESSMENT CRITERIA:
The test substance is considered positive in this assay if the following criteria are met: A dose-related and reproducible increase in the number of revertant colonies, i.e. about doubling of the spontaneous mutation rate in at least one tester strain either without S9 mix or after adding a metabolizing system.
A test substance is generally considered non-mutagenic in this test if: The number of revertants for all tester strains were within the historical negative control range 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:

cytotoxicity

Remarks:

depending on the strain and test conditions at 2750 μg/plate only without S9 mix (slight decrease in the number of his+ revertants, slight reduction in the titer)

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

depending on the strain and test conditions at 2750 μg/plate only without S9 mix (slight decrease in the number of his+ revertants, slight reduction in the titer)

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 3

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

depending on the strain and test conditions at 2750 μg/plate only without S9 mix (slight decrease in the number of his+ revertants, slight reduction in the titer)

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 4

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

depending on the strain and test conditions at 2750 μg/plate only without S9 mix (slight decrease in the number of his+ revertants, slight reduction in the titer)

Vehicle controls validity:

valid

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

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Applicant's summary and conclusion

Conclusions:

Thus, under the experimental conditions of this study, the test substance is not mutagenic in the Salmonella typhimurium/Escherichia coli reverse mutation assay in the absence and the presence of metabolic activation.

Executive summary:

The test substance was tested for its mutagenic potential based on the ability to induce point mutations in selected loci of several bacterial strains, i.e. Salmonella typhimurium and Escherichia coli, in a reverse mutation assay.

STRAINS: TA 1535, TA 100, TA 1537, TA 98 and E. coli WP2 uvrA

DOSE RANGE: 33 μg - 5 500 μg/plate (SPT); 33 μg - 5 500 μg/plate (PIT)

TEST CONDITIONS: Standard plate test (SPT) and preincubation test (PIT) both with and without metabolic activation (liver S9 mix from induced rats).

SOLUBILITY: No precipitation of the testsubstance was found with and
without S9 mix.

TOXICITY: A weak bacteriotoxic effect was occasionally observed
depending on the strain and test conditions at 2 750 μg/plate

MUTAGENICITY:
A relevant increase in the number of his+ or trp+ revertants was not observed in the standard plate test or in the preincubation test either without S9 mix or after the addition of a metabolizing system.