Registration Dossier

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:
2009
Reliability:
1 (reliable without restriction)

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2010

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent

Method

Target gene:
Salmonella typhimurium LT2
Species / strainopen allclose all
Species / strain / cell type:
S. typhimurium TA 1535
Additional strain / cell type characteristics:
other: hisG46, uvrB, rfa
Species / strain / cell type:
S. typhimurium, other: TA97a
Additional strain / cell type characteristics:
other: hisD6610,uvrB, pKM 101, rfa
Species / strain / cell type:
S. typhimurium TA 98
Additional strain / cell type characteristics:
other: hisD3052, uvrB, pKM 101, rfa
Species / strain / cell type:
S. typhimurium TA 100
Additional strain / cell type characteristics:
other: hisG46, uvrB, pKM 101, rfa
Species / strain / cell type:
S. typhimurium TA 102
Additional strain / cell type characteristics:
other: hisG428, pKM 101, rfa
Metabolic activation:
with and without
Metabolic activation system:
S9 mix
Test concentrations with justification for top dose:
The test item was dissolved in dimethyl sulfoxide (DMSO). The stock solution containing 50g/L was prepared.
First experiment (plate incorporation method): test concentrations: 5020 / 1506 / 502 / 151 / 50 µg/plate
Second experiment (pre-incubation method): test concentrations: 5051 /2526 / 1263 µg/plate
Vehicle / solvent:
DMSO
Controlsopen allclose all
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Remarks:
H2O, DMSO
True negative controls:
other: comparison with historical data
Positive controls:
yes
Positive control substance:
benzo(a)pyrene
Remarks:
strain 98, with metabolic activation system

Migrated to IUCLID6: concentration per plate: 40 µg
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Remarks:
H2O, DMSO
True negative controls:
other: comparison with historical data
Positive controls:
yes
Positive control substance:
other: 2-Aminoanthracene (1-AA), 3 µg per plate
Remarks:
strains 97a, 100, 102, 1535, with metabolic activation system
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Remarks:
H2O, DMSO
True negative controls:
other: comparison with historical data
Positive controls:
yes
Positive control substance:
other: 4-Nitro-1,2-phenylene diamine, 80 µg per plate
Remarks:
strains 97 a, 98, 102, without metabolic activation system
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Remarks:
H2O, DMSO
True negative controls:
other: comparison with historical data
Positive controls:
yes
Positive control substance:
sodium azide
Remarks:
strains 100, 1535, without metabolic activation system

Migrated to IUCLID6: concentration per plate: 6 µg

Results and discussion

Test resultsopen allclose all
Species / strain:
S. typhimurium TA 1535
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
Species / strain:
S. typhimurium, other: TA 97a
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
Species / strain:
S. typhimurium TA 98
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
Species / strain:
S. typhimurium TA 100
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
Species / strain:
S. typhimurium TA 102
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
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Any other information on results incl. tables

All results including tables and figures are given in the attached full study report.

Applicant's summary and conclusion

Conclusions:
The test item B 1061 did not show mutagenic effects in both experiments. The number of revertant colonies was not increased in comparison with
the spontaneous revertants (solvent only). Cytotoxicity of the test item was not detected. The background lawn was visible and the number of rever- tants was not significantly decreased.
Therefore it can be stated, that under the test conditions, the test item B 1061 is not mutagenic in the Bacterial Reverse Mutation Test using
Salmonella typhimurium, strains TA 97a, TA 98, TA 100, TA 102 and TA 1535.
Executive summary:

Two valid experiments were performed.

First experiment: Five concentrations of the test item B 1061, dissolved in DMSO (ranging from 5020 to 50 µg/plate) were used. Five genetically manipulated strains of Salmonella typhimurium (TA 97a, TA 98, TA 100, TA 102, TA 1535) were exposed to the test item both in the presence and in the absence of a metabolic activation system (S9) for 48 hours, using the plate incorporation method.

None of the concentrations caused a significant increase in the number of revertant colonies in the tested strains. The test item didn't show any mutagenic effects in the first experiment.

No signs of toxicity towards the bacteria could be observed.

The sterility control and the determination of the titre didn't show any inconsistencies. The determined values for the spontaneous revertants of the negative controls were in the normal range. All positive controls showed mutagenic effects with and without metabolic activation.

Second experiment: To verify the results of the first experiment, a second experiment was performed, using three concentrations of the test item (ranging from 5051 to 1263 µg/plate) and a modification in study performance (pre-incubation method).

The test item B 1061 didn't show mutagenic effects in the second experiment, either.

No signs of toxicity towards the bacteria could be observed.

The sterility control and the determination of the titre didn't show any inconsistencies. The determined values for the spontaneous revertants of the negative controls were in the normal range. All positive controls showed mutagenic effects with and without metabolic activation.

Under the conditions of the test, the test item B 1061 didn't show mutagenic effects towards Salmonella typhimurium, strains TA 97a, TA 98, TA 100, TA 102 and TA 1535. Therefore, no concentration-effect relationship could be determined.

The test item B 1061 is considered as "not mutagenic under the conditions of the test".