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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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP compliant guideline study, available as unpublished report, no restrictions, fully adequate for assessment.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2012
Report Date:
2012

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Qualifier:
according to
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Qualifier:
according to
Guideline:
EPA OPPTS 870.5100 - Bacterial Reverse Mutation Test (August 1998)
GLP compliance:
yes (incl. certificate)
Remarks:
BASF SE, Experimental Toxicology and Ecology, 67056 Ludwigshafen, Germany
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
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 / strainopen allclose all
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 / 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
Controlsopen allclose all
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
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
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
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
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 resultsopen allclose all
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98 and 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
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
Additional information on results:
TOXICITY
No bacteriotoxic effect (reduced his- or trp- background growth, slight decrease in the number of his+ or trp+ revertants, slight reduction in the titer) was observed in the standard plate test up to the highest required concentration. In the preincubation assay bacteriotoxicity (slight decrease in the number of his+ revertants, slight reduction in the titer) was occasionally observed depending on the strain and test conditions at 2750 μg/plate only without S9 mix.

SOLUBILITY
No test substance precipitation was found with and without S9 mix.

DISCUSSION
According to the results of the present study, the test substance did not lead to a relevant increase in the number of revertant colonies either without S9 mix or after adding a metabolizing system in experiments carried out independently of each other (standard plate test and preincubation assay). Besides, the results of the negative as well as the positive controls performed in parallel corroborated the validity of this study, since the values fulfilled the acceptance criteria of this study. In this study with and without S9 mix, the number of revertant colonies in the negative controls was nearby the range of the historical negative control data for each tester strain. In addition, the positive control substances both with and without S9 mix induced a significant increase in the number of revertant colonies within the range of the historical positive control data or above.
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Applicant's summary and conclusion