Registration Dossier

Toxicological information

Genetic toxicity: in vitro

Currently viewing:

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:
From 15 DEC 2004 to 10 JAN 2005
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study (OECD 471)

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2005
Report Date:
2005

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
in accordance to German Chemikaliengesetz and Directive 88/320/EEC
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent

Method

Species / strainopen allclose all
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Additional strain / cell type characteristics:
not specified
Species / strain / cell type:
E. coli WP2 uvr A
Additional strain / cell type characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
induced rat liver S9 (phenobarbital/ß-naphtoflavone used for induction)
Test concentrations with justification for top dose:
Experiment I (plate incorporation): 3, 10, 33, 100, 333, 1000, 2500, and 5000 µg/plate
Experiment II (preincubation test): 33, 100, 333, 1000, 2500, and 5000 µg/plate
Vehicle / solvent:
- Vehicle used: DMSO
Controlsopen allclose all
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: sodium azide (TA 1535, TA 100), 4-nitro-o-phenylene-diamine (TA1537, TA 98), methyl methan sulfonate (WP2uvrA)
Remarks:
without metabolic activation
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: 2-aminoanthracene (all strains)
Remarks:
with metabolic activation
Details on test system and experimental conditions:
METHOD OF APPLICATION:
Experiment I: plate incorporation
Experiment II: preincubation

DURATION
- Preincubation period: only Experiment II: 60 minutes at 37 °C
- Exposure duration: 48 hours


NUMBER OF REPLICATIONS: 3 plates per strain and dose level, including controls
Evaluation criteria:
test item considerer as a mutagen, if:
- number of revertants exceeding a threshold of twice (TA 98, TA 100 and WP2uvrA) or
- number of revertants exceeding a threshold of thrice (TA 1535, TA 1537)
the colony count of the corresponding solvent control

- a dose dependent increase is considered relevant if the threshold is exceeded more than once

- increase in threshold at only one concentration is judged relevant if reproduced in a second independent experiment

-dose dependent increase below the threshold is regarded as indication of a mutagenic potential if reproduced in a second independent experiment, whenever the colony counts remain within the historical range of negative and solvent controls such an increase is not considered relevant

Results and discussion

Test results
Species / strain:
other: S. typhimurium TA 1535, TA 98, TA 100 and TA 1537, E. coli WP2uvrA
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Remarks:
minor effect in plate incorporation assay at 5000 µg/plate in TA1537 (reduction in the number of revertants to ~50% of control plates)
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: visible in Experiment II without S9 at a concentration of 5000 µg/plate and with S9-Mix at concentrations of 1000 µg/plate and above

COMPARISON WITH HISTORICAL CONTROL DATA:
- historical range of positive controls was exceeded with metabolic activation in strains TA 98 and TA 1537 (Experiment I) and in strain TA 100 (Experiment/ and II), this effect indicates the sensitivity of the strains rather than compromising the assay
- Strain WP2uvrA (ExperimentI) with metabolic activation historical control range of negative and solvent controls was slightly exceeded
- Strain WP2uvrA and TA 1535 (ExperimentII) with and without metabolic activation historical control range of negative and solvent controls was slightly exceeded
- Strain TA 1535 and TA 98 (ExperimentII) with metabolic activation historical control range of negative and solvent controls was slightly exceeded

ADDITIONAL INFORMATION ON CYTOTOXICITY:
- TA 1537: plate incorporation assay without metabolic activation minor effect at 5000 µg/plate
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Any other information on results incl. tables

The test item showed no mutagenic activity in both experiments (plate incorporation assay, preincubation assay) each with and without metabolic activation.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative with metabolic activation
negative without metabolic activation

The test item did not exert mutagenic activity in the reverse bacterial mutation assay (plate incorporation assay and preincubation assay) with and without metabolic activation.
Executive summary:

Mutagenic activity of the test item was investigated in Salmonella typhimurium strains TA 1535, TA 1537, TA98, TA100 and Escherichia coli strain WP2uvrA with (induced rat liver S9 mix) and without metabolic activation at concentrations of 3, 10, 33, 100, 333, 1000, 2500, and 5000 µg/plate using the plate incorporation assay. Additionally, a preincubation assay with or without metabolic activation was performed using the concentrations 33, 100, 333, 1000, 2500, and 5000 µg/plate.

The test item did not reveal any mutagenic activity under the conditions tested. The appropriate reference mutagenes showed distinct positive mutagenic effects.