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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in mammalian cells
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
Oct 2016 - Apr 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2017
Report Date:
2017

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test using the Hprt and xprt genes)
Version / remarks:
2016
GLP compliance:
yes
Type of assay:
in vitro mammalian cell gene mutation test using the Hprt and xprt genes

Test material

Reference
Name:
Unnamed
Type:
Constituent

Method

Target gene:
HPRT
Species / strain
Species / strain / cell type:
Chinese hamster lung fibroblasts (V79)
Metabolic activation:
with and without
Metabolic activation system:
phenobarbital and ß-naphthoflavone induced rat liver S9 mix
Test concentrations with justification for top dose:
650.0 - 1600.0 µg/mL
The maximum concentration was chosen with respect to the current OECD Guideline 476 (2016) and a correction factor of 1.3 was used to correct for purity and the ratio of phenylguanidine to carbonate counter ion in the test substance.
Vehicle / solvent:
DMSO
Controls
Untreated negative controls:
yes
Remarks:
solvent control
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
7,12-dimethylbenzanthracene
ethylmethanesulphonate
Evaluation criteria:
A test chemical is considered to be clearly positive if, in any of the experimental conditions examined all of the following criteria are met:
a) at least one of the test concentrations exhibits a statistically significant increase compared with the concurrent solvent control in both parallel cultures,
b) the increase is concentration-related when evaluated with an appropriate trend test,
c) any of the results are outside the distribution of the historical solvent control data.
When all of these criteria are met, the test chemical is then considered able to induce gene mutations in cultured mammalian cells in this test system.
Providing that all acceptability criteria are fulfilled, a test chemical is considered clearly negative if, in all experimental conditions examined:
a) none of the test concentrations exhibits a statistically significant increase compared with the concurrent negative control,
b) there is no concentration-related increase when evaluated with an appropriate trend test,
c) all results are inside the distribution of the historical negative control data.
The test chemical is then considered unable to induce gene mutations in cultured mammalian cells in this test system.
Statistics:
The statistical analysis was performed in the mean values of culture I and II for all experiments.
A linear regression analysis was performed to assess a possible dose dependency of mutant frequencies.
A t-test was performed using a validated test script of "R", a language and environment for statistical computing and graphics, to evaluate an isolated increase of the mutation frequency at a test point exceeding the 95 % confidence interval and other if necessary.

Results and discussion

Test results
Species / strain:
Chinese hamster lung fibroblasts (V79)
Metabolic activation:
with and without
Genotoxicity:
negative
Vehicle controls validity:
valid
Untreated negative controls validity:
not applicable
True negative controls validity:
not applicable
Positive controls validity:
valid

Applicant's summary and conclusion

Conclusions:
The increases in the mutant frequency observed in the presence of metabolic activation were inconsistent between parallel cultures of the experiments and caused by increases in one culture only. None of those increases in MF were statistically significant increases and there was no relevant dose dependent trend. Based on these results, the test substance meets the criteria for being negative in the HPRT assay.
In conclusion it can be stated that under the experimental conditions reported the test item did not induce gene mutations at the HPRT locus in V79 cells of the Chinese hamster. Therefore, the test substance is considered to be non-mutagenic in this HPRT assay.