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Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
09.03.2018 to 26.04.2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2018
Report date:
2018

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Type of assay:
bacterial reverse mutation assay

Test material

Constituent 1
Chemical structure
Reference substance name:
[2,2,6,6-tetramethyl-4-(trimethylazaniumyl)piperidin-1-yl]oxidanyl chloride
EC Number:
814-375-0
Cas Number:
67036-09-3
Molecular formula:
C12H26N2OCl
IUPAC Name:
[2,2,6,6-tetramethyl-4-(trimethylazaniumyl)piperidin-1-yl]oxidanyl chloride
Test material form:
solid

Method

Species / strain
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
Metabolic activation:
with and without
Metabolic activation system:
S9 mix
Test concentrations with justification for top dose:
31.6, 100, 316, 1000, 2500 and 5000 μg/plate
Vehicle / solvent:
purified water
Controlsopen allclose all
Untreated negative controls:
yes
Remarks:
purified water
Negative solvent / vehicle controls:
yes
Remarks:
purified water
Positive controls:
yes
Remarks:
without metabolic activation
Positive control substance:
sodium azide
Remarks:
Strain TA100 (10µg/plate) and TA1535 (10µg/plate)
Positive controls:
yes
Remarks:
without metabolic activation
Positive control substance:
other: 4-NOPD; 4-nitro-o-phenylene-diamine
Remarks:
Strain TA98 (10µg/plate) and TA1537 (40µg/plate)
Positive controls:
yes
Remarks:
without metabolic activation
Positive control substance:
methylmethanesulfonate
Remarks:
Strain TA102 (1µg/plate)
Positive controls:
yes
Remarks:
with metabolic activation
Positive control substance:
other: 2-AA; 2-aminoanthracene
Remarks:
Strain TA98, TA100, TA1535, TA1537 (all 2.5µg/plate) and TA102 (10µg/plate)
Details on test system and experimental conditions:
The test item TMA-TEMPO was investigated for its potential to induce gene mutations according to the plate incorporation test (experiment I) and the pre-incubation test (experiment II) using Salmonella typhimurium strains TA98, TA100, TA1535, TA1537 and TA102.
In two independent experiments several concentrations of the test item were used. Each assay was conducted with and without metabolic activation. The concentrations, including the controls, were tested in triplicate.
Evaluation criteria:
The Mutation Factor is calculated by dividing the mean value of the revertant counts by the mean values of the solvent control (the exact and not the rounded values are used for calculation).
A test item is considered as mutagenic if:
- a clear and dose-related increase in the number of revertants occurs and/or
- a biologically relevant positive response for at least one of the dose groups occurs
in at least one tester strain with or without metabolic activation.

A biologically relevant increase is described as follows:
- if in tester strains TA98, TA100 and TA102 the number of reversions is at least twice as high
- if in tester strains TA1535 and TA1537 the number of reversions is at least three times higher
than the reversion rate of the solvent control

Results and discussion

Test results
Key result
Species / strain:
S. typhimurium, other: TA98, TA100, TA1535, TA1537, TA102
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:
No precipitation of the test item was observed in any tester strain used in experiment I and II (with and without metabolic activation).
No toxic effects of the test item were noted in any of the five tester strains used up to the highest dose group evaluated with and without metabolic activation in experiment I and II.

Applicant's summary and conclusion

Conclusions:
In conclusion, it can be stated that during the described mutagenicity test and under the experimental conditions reported, TMA-TEMPO did not cause gene mutations by base pair changes or frameshifts in the genome of the tester strains used.
Therefore, TMA-TEMPO is considered to be non-mutagenic in this bacterial reverse mutation assay.

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