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:
2014-06-16 to 2014-07-03
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study conducted according to OECD method and in accordance with GLP.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2014
Report Date:
2014

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)
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
other: Liquid
Details on test material:
- Name of test material (as cited in study report): Bis-Tes (dried)
- Molecular formula (if other than submission substance): C16H34OSi2
- Molecular weight (if other than submission substance): 298
- Physical state: Liquid
- Stability under test conditions: Unknown
- Storage condition of test material: At room temperature in the dark under nitrogen

Method

Target gene:
Four histidine-requiring strains (Salmonella typhimurium (TA1535, TA1537, TA98 and TA100)) and one tryptophan-requiring strain (Escherichia coli (WP2uvrA).
Species / strain
Species / strain / cell type:
other: Salmonella typhimurium: TA1535, TA1537, TA98, TA100. Escherchia coli WP2uvrA.
Metabolic activation:
with and without
Metabolic activation system:
S-9 mix (rat liver S9-mix induced by Aroclor 1254)
Test concentrations with justification for top dose:
For genotoxicity experiment concentrations (with & without metabolic activation) used: 5000 ug/plate for plate incorporation method.
Vehicle / solvent:
Solvent- ethanol
Controlsopen allclose all
Untreated negative controls:
yes
Remarks:
Ethanol
Negative solvent / vehicle controls:
no
True negative controls:
no
Positive controls:
yes
Remarks:
With metabolic activation
Positive control substance:
other: 2-aminoanthracene
Positive controls:
yes
Positive control substance:
sodium azide
Remarks:
Without metabolic activation
Positive controls:
yes
Positive control substance:
2-nitrofluorene
Remarks:
Without metabolic activation

Migrated to IUCLID6: 2.5 ug/plate (TA98) without s-9
Positive controls:
yes
Positive control substance:
4-nitroquinoline-N-oxide
Remarks:
Without metabolic activation
Positive controls:
yes
Positive control substance:
methylmethanesulfonate
Remarks:
Without metabolic activation
Details on test system and experimental conditions:
Precipitation of Bis-Tes (dried) on the plates was observed at the start and at the end of the incubation
period at concentrations of 1600 and 5000 µg/plate. Except in the tester strains TA1535, TA1537 and
TA100 in the absence of S9-mix, in which no precipitation of the test substance was observed at
1600 µg/plate.

There was no reduction of the bacterial background lawn and no
biologically relevant decrease in the number of revertants at any of the concentrations tested in all
tester strains in the absence and presence of S9-mix.

In strain TA1537 (presence of S9-mix), a fluctuation in the number of revertant colonies below the
laboratory historical control data range was observed at the mid dose of 164 µg/plate. However, since
no dose-relationship was observed, the reduction is not considered to be caused by toxicity of the test
substance
Evaluation criteria:
A test substance is considered negative (not mutagenic) in the test if:
a) The total number of revertants in tester strain TA100 is not greater than two (2) times the
concurrent control, and the total number of revertants in tester strains TA1535, TA1537, TA98 or
WP2uvrA is not greater than three (3) times the concurrent vehicle control.
b) The negative response should be reproducible in at least one independently repeated experiment.
A test substance is considered positive (mutagenic) in the test if:
a) The total number of revertants in tester strain TA100 is greater than two (2) times the concurrent
control, or the total number of revertants in tester strains TA1535, TA1537, TA98 or WP2uvrA is
greater than three (3) times the concurrent vehicle control.
b) In case a repeat experiment is performed when a positive response is observed in one of the tester
strains, the positive response should be reproducible in at least one independently repeated
experiment.
Statistics:
Mean and standard deviation of the plate counts for each treatment were determined.

Results and discussion

Test results
Species / strain:
other: S. typhimurium TA 1535, TA 1537, TA 98, TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
other: yes. Negative control values were within the laboratory historical control data ranges, except for TA1535 in the presence of S9-mix (1st experiment). Since this value was just outside the limit of the range, the validity of the test was not affected.
Positive controls validity:
valid
Additional information on results:
There were no deviations from standard operating procedures that affected the integrity of the study.

Bis-Tes (dried) did not induce a significant dose-related increase in the number of revertant (His+
)
colonies in each of the four tester strains (TA1535, TA1537, TA98 and TA100) and in the number of
revertant (Trp+
) colonies in tester strain WP2uvrA both in the absence and presence of S9-metabolic
activation. These results were confirmed in an independently repeated experiment.
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative

Based on the results of this study it is concluded that Bis-Tes (dried) is not mutagenic in the
Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.
Executive summary:

No increase in the number of revertants was observed upon treatment with Bis-Tes (dried) under all

conditions tested. Bis-Tes (dried) is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.