Registration Dossier

Toxicological information

Genetic toxicity: in vitro

Currently viewing:

Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
21 . Dec. 2018 - 11 Apr. 2019
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2012
Report Date:
2019

Materials and methods

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

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid

Method

Species / strain
Species / strain / cell type:
other: Salmonella typhimurium LT2 Strains TA97a, TA98, TA100, TA102 and TA1535
Metabolic activation:
with and without
Metabolic activation system:
S9-Mix
Test concentrations with justification for top dose:
Exp. I: 50, 150, 500, 1500 and 5000 μg/plate (with and without metabolic activation)
Exp. II: 156, 313, 625, 1250, 2500 and 5000 μg/plate (with and without metabolic activation)
Vehicle / solvent:
In a non-GLP pre-test, the solubility of the test item was tested in a concentration of 50 g/L in demineralized water, dimethyl sulfoxide (DMSO), tetrahydrofuran and acetone. The test item is not sufficiently soluble in all solvents. Based on the non-GLP pre-test, acetone was chosen as vehicle, because it showed the best suspension and this solvent does not have any effects on the viability of the bacteria or the number of spontaneous revertants in the tested concentrations.
Controlsopen allclose all
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
sodium azide
other: 4-Nitro-1,2-phenylene diamine; CAS-No.: 99-56-9
Remarks:
without metabolic activation
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
benzo(a)pyrene
other: 2-aminoanthracene
Remarks:
with metabolic activation
Evaluation criteria:
The colonies were counted visually and the numbers were recorded. A validated spreadsheet software (Microsoft Excel®) was used to calculate mean values and standard deviations of each treatment, solvent control and positive control. The mean values and standard deviations of each threefold determination was calculated as well as the increase factor f(l) of revertant induction (mean revertants divided by mean spontaneous revertants) of the test item solutions and the positive controls. Additionally, the absolute number of revertants (Rev. Abs.) (mean revertants less mean spontaneous revertants) was given. A substance is considered to have mutagenic potential, if a reproducible increase of revertant colonies per plate exceeding an increase factor of 2 in at least one strain can be observed. A concentration-related increase over the range tested is also taken as a sign of mutagenic activity.

Results and discussion

Test results
Key result
Species / strain:
other: Salmonella typhimurium LT2 Strains TA97a, TA98, TA100, TA102 and TA1535
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

Applicant's summary and conclusion

Conclusions:
All validity criteria are fulfilled. The study can be considered valid. Based on the results of this study it is concluded that Bisamid is not mutagenic in the Salmonella typhimurium strains TA97a, TA98, TA100, TA102 and TA1535 in the absence and presence of metabolic activation under the experimental conditions in this study.
Executive summary:

Three valid experiments were performed. The study procedures described in this report were based on the most recent OECD and EC guidelines. The test item Bisamid was tested in the Salmonella typhimurium reverse mutation assay with five strains of Salmonella typhimurium (TA97a, TA98, TA100, TA102 and TA1535). The test was performed in three experiments in the presence and absence of metabolic activation, with +S9 standing for presence of metabolic activation, and –S9 standing for absence of metabolic activation. Experiment 1a: In the experiment 1a, the test item (suspended in acetone) was tested up to concentrations of 5000 μg/plate in the absence and presence of S9-mix in the strains TA97a, TA98, TA100, TA102 and TA1535 using the plate incorporation method. The test item showed no precipitates on the plates at any of the concentrations. The bacterial background lawn was not reduced at any of the concentrations and no relevant decrease in the number of revertants was observed in all bacteria strains. The test item showed no signs of toxicity towards the bacteria strains in both the absence and presence of metabolic activation. The results of this experiment showed that none of the tested concentrations showed a significant increase in the number of revertants in all tested strains, in the presence and the absence of metabolic activation. Due to contamination of bacteria strain TA97a, the evaluation of the bacteria strain TA97a was not possible, therefore a repetition of the experiment for this bacteria strain was performed (Experiment 1b). Experiment 1b: Based on the contamination of the bacteria strain TA97a in experiment 1a, the experiment was repeated under the same conditions with this bacteria strain. The test item (suspended in acetone) was tested up to concentrations of 5000 μg/plate in the absence and presence of S9-mix for the bacteria strain TA97a. The test item showed no precipitates on the plates at any of the concentrations. The bacterial background lawn was not reduced at any of the concentrations and no relevant decrease in the number of revertants was observed in all bacteria strains. The test item showed no signs of toxicity towards the bacteria strains in both the absence and presence of metabolic activation. The results of this experiment showed that none of the tested concentrations showed an increase in the number of revertants in the tested strain, in the presence and the absence of metabolic activation.

Experiment 2: Based on the first experiments, the test item was tested up to concentrations of 5000 μg/plate in the absence and presence of S9-mix in all bacteria strains using the pre-incubation method. The test item showed no precipitates on the plates at any of the concentrations. The bacterial background lawn was not reduced at any of the concentrations and no relevant decrease in the number of revertants was observed in all bacteria strains. The test item showed no signs of toxicity towards the bacteria strains in both the absence and presence of metabolic activation. The results of this experiments showed that the test item caused no increase in the number of revertants in all bacteria strains compared to the solvent control, in both the absence and presence of metabolic activation. The test item did not induce a dose-related increase in the number of revertant colonies in all strains, in the presence and absence of metabolic activation.


Route: .live2