Registration Dossier

Administrative data

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

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2016

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
GLP compliance:
yes (incl. certificate)
Type of assay:
bacterial forward mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
UVCB substancePhysical form: liquidsolubility/dispersability > 300 g/ltest item storage: room temperature, protected from lightStability: stable under storage conditionsExpiry date: 31 December 2018

Method

Species / strainopen allclose all
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Additional strain / cell type characteristics:
other: rfa wall mutation to increase permeability to certain classes of chemicals (crystal violet); uvrB mutation, deficient in a DNA excision repair system (UV irradiation sensitivity) TA98 and TA100 contain pKM101 plasmid for resistance to Ampicillin
Species / strain / cell type:
E. coli WP2 uvr A
Additional strain / cell type characteristics:
other: uvrA DNA repair deficiency, sensitivity to UV irradiation
Metabolic activation:
with and without
Metabolic activation system:
S9 tissue fraction from rat liver
Test concentrations with justification for top dose:
Main assay III concentrations (spacing of a factor of 0.5)TA1535 without activiation, 15.6 to 0.977 ug/plateTA1535 with activation 250 to 15.6 ug/plateTA1537 without activation 50 to 1.56 ug/plateTA1537 with activation 2000 to 31.3 ug/plateWP2 uvrA without activation 62 to 3.91 ug/plateWP2 uvrA with activation 2000 to 62.5 ug/plateTA98 without activation 31.3 to 0.977 ug/plateTA98 with activation 2000 to 62.5 ug/plateTA100 without activation 50 to 1.56 ug/plateTA100 with activation 2000 to 62.5 ug/plateThe highest dose was chosen as the maximum level to obtain a sufficient number of analysable concentrations
Controls
Positive controls:
yes
Positive control substance:
9-aminoacridine
2-nitrofluorene
sodium azide
methylmethanesulfonate
other: 2-aminoanthracene

Results and discussion

Test resultsopen allclose all
Species / strain:
E. coli WP2 uvr A pKM 101
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
at the (two) highest dose
Species / strain:
other: TA 1535, TA 1537, TA98 , TA100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
at the (two) highets dose level

Applicant's summary and conclusion

Conclusions:
The substance was tested following OECD 471 for genetic toxicity to bacteria. Under the experimental conditions the substance did not show amu mutagenic potential in all tested strains.
Executive summary:

The substance was examined for the ability to induce gene mutations in tester strains of Salmonella typhimurium and Escherichia coli, as measured by reversion of auxotrophic strains to prototrophy. The five tester strains TA1535, TA1537, TA98, TA100 and WP2 uvrA were used. Experiments were performed both in the absence and presence of metabolic activation, using liver S9 fraction from rats pre-treated with Phenobarbital and 5,6-Benzoflavone. The test item was used as a solution in sterile water for injection.

Toxocity test was performed as to establish correct doses for the main test, with plate incorporation method. In this first main assay toxicity was observed for all dose levels but TA1535 with activation system.

In the second main assay some dose levels were decresed and pre-incubation step was added since no relevant increase in the number of revertant was observed. However, severe to slight toxicity was again observed and a thrid main assya was performed with lower dose levels.

The substance did not induce reverse mutation of Salmonella T. or Escherichia Coli in the absence or presence of S9 metabolism under the experimental conditions