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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Remarks:
Type of genotoxicity: gene mutation
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
2008
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: GLP study according to OECD guideline 471 (adopted 1997). Study was originally judged Klimisch 1. However, according to the "Practical guide 6: How to report read-across and categories" the maximum score for read-across is 2.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2008
Report Date:
2008

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
Qualifier:
according to
Guideline:
other: Commision Directive 2000/32/EEC
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
-Name of test material (as cited in study report): Biofert Plusz
Details are presented in "Confidential details on test material"

Method

Target gene:
The Salmonella typhimurium histidine (his) and the E. coli tryptophan (trp) reversion system measures his- → his+ and trp- → trp+ reversions, respectively.
Species / strainopen allclose all
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Species / strain / cell type:
E. coli WP2 uvr A
Metabolic activation:
with and without
Metabolic activation system:
Rat liver S9-mix
Test concentrations with justification for top dose:
Pre-experiment/Experiment I: 3, 10, 33, 100, 333, 1000, 2500 and 5000 ug/plate (related to dry mass)
Experiment II: 156, 312, 625, 1250, 2500 and 5000 ug/plate (related to dry mass)

The tested concentrations were adjusted based on the dry mass of the material (e.g. 5000 ug/plate corresponds to 17793 ug liquid test item/plate)
Vehicle / solvent:
-Vehicle(s)/solvent(s) used: Deionized water
-Justification for choice of solvent/vehicle: According to the solubility properties of the test item
Controls
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
other: Without metabolic activation: sodium azide (TA1535 and TA100); 4-nitro-o-phenylene-diamine (TA98 and TA1537); methyl methane sulfonate (E. coli); With metabolic activation: 2-aminoanthracene (S. typhimurium and E. coli)
Details on test system and experimental conditions:
METHOD OF APPLICATION: In agar (plate incorporation)

DURATION: Preincubation period: 4 hours; Exposure duration: 48 hours

SELECTION AGENT: Minimal histidine agar

NUMBER OF REPLICATIONS: 3 plates per dose level in each experiment; 2 independent experiments

DETERMINATION OF CYTOTOXICITY: To evaluate the cytotoxicity of the test item, a pre-experiment was performed. Eight concentrations were tested for toxicity and mutation induction with three plates each.
Evaluation criteria:
According to guideline
Statistics:
According to OECD Guideline 471, a statistical analysis of data is not mandatory

Results and discussion

Test resultsopen allclose all
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
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
Species / strain:
E. coli WP2 uvr A
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:
TEST-SPECIFIC CONFOUNDING FACTORS
-Effects of pH: pH value was specified as 4.4. No effects to this value were noted
-Water solubility: completely soluble in water
-Precipitation: none

RANGE-FINDING/SCREENING STUDIES: In a pre-experiment (Experiment I), strains TA 1535, TA 1537, TA 98, TA 100, and WP2 uvrA were tested at 8 concentrations with each 3 plates. Concentrations of the dry mass were between 3 and 5000 ug/plate. There were no toxic effects and no mutagenic activity. Same results in experiment II.

COMPARISON WITH HISTORICAL CONTROL DATA: yes
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Any other information on results incl. tables

The plates incubated with the test item showed normal background growth up to 5000 ug/plate of the dry mass with and without metabolic activation in both independent experiments. No toxic effects, evident as a reduction in the number of revertants (below the indication factor of 0.5), occurred in the test groups with and without metabolic activation. No substantial increase in revertant colony numbers in any of the five tester strains at any concentration level, neither in the presence nor absence of metabolic activation. There was also no tendency of higher mutation rates with increasing concentrations in the range below the generally acknowledged border of biological relevance. Appropriate reference mutagens were used as positive controls. They showed a distinct increase in induced revertant colonies. 

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative

The test item did not induce gene mutations in S. typhimurium TA 1535, TA 100, TA 1537, TA 98 and E. coli WP2uvrA with and without metabolic activation at concentrations up to 5 mg/plate (related to dry mass).