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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:
experimental study
Adequacy of study:
key study
Study period:
2015
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study was conducted in compliance with OECD GLP (1997) and FDA GLP regulations. Study performed according to OECD 471 (1997).

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2015
Report Date:
2015

Materials and methods

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

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Test material form:
other: Viscous liquid
Details on test material:
- Name of test material (as cited in study report): MTDID 15655
- Substance type: Mono-constituent
- Physical state: Viscous liquid
- Analytical purity: 91.4%
- Purity test date: 21 April 2015
- Lot/batch No.: 565093
- Expiration date of the lot/batch: 27 August 2017
- Storage condition of test material: Room temperature, protected from light

Method

Target gene:
Histidine operon, tryptophan operon
Species / strain
Species / strain:
other: Salmonella typhimurium TA98, TA100, TA1535, TA 1537 and Escherichia coli WP2 uvrA
Metabolic activation:
with and without
Metabolic activation system:
Aroclor-induced rat liver S9
Test concentrations with justification for top dose:
50, 150, 500, 1500 and 5000 ug/plate
Vehicle:
- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: test article solubility
Controls
Negative controls:
no
Solvent controls:
yes
Remarks:
DMSO
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: With metabolic activation: All strains: 2-aminoanthracene. Without metabolic activation: TA98: 2-nitrofluorene, TA100, TA1535: sodium azide, TA1537: 9-aminoacridine, WP2 uvrA: methyl methanesulfonate
Details on test system and conditions:
METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Preincubation period: 12 hours
- Exposure duration: 48-72 hours
- Expression time (cells in growth medium): 60-84 hours
- Selection time (if incubation with a selection agent): 48-72 hours
- Fixation time (start of exposure up to fixation or harvest of cells): 48-72 hours

SELECTION AGENT (mutation assays): Minimal top agar

NUMBER OF CELLS EVALUATED: Colonies on all plates were counted either by hand or by Sorcerer Colony Counter.

DETERMINATION OF CYTOTOXICITY
- Method: The condition of the bacterial background lawn was evaluated for evidence of test article toxicity.
Evaluation criteria:
For the test article to be evaluated positive, it must cause a dose-related increase in the mean revertants per plate of at least one tester strain over a minimum of two increasing concentrations of the test article as specified below:
TA 1535 and TA1537: Data sets were judged positive if the increase in mean revertants at the peak of the dose response was equal to or greater than 3.0-times the mean vehicle control value.
TA98, TA100 and WP2 uvrA: Data sets were judged positive if the increase in mean revertants at the peak of the dose response was equal to or greater than 2.0-times the mean vehicle control value.

Results and discussion

Test results
Species / strain:
other: Salmonella typhimurium TA98, TA100, TA1535, TA1537 and Escherichia coli WP2 uvrA
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity:
no, but tested up to precipitating concentrations
Vehicle controls valid:
yes
Negative controls valid:
not applicable
Positive controls valid:
yes
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH:
- Effects of osmolality:
- Evaporation from medium:
- Water solubility:
- Precipitation:
- Other confounding effects:

RANGE-FINDING/SCREENING STUDIES: A range-finding assay found dose levels up to 5000 ug/plate were appropriate for testing

COMPARISON WITH HISTORICAL CONTROL DATA:

ADDITIONAL INFORMATION ON CYTOTOXICITY: No cytotoxicity was observed up to 5000 ug/plate but precipitation was observed at 1500 and 5000 ug/plate.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative

Under the conditions of this study, the test article is not mutagenic in the bacterial reverse mutation assay with or without metabolic activation.
Executive summary:

The mutagenic potential of the test article (Viscous yellow liquid, Lot 565093, Purity: 91.4% ) was evaluated in the Bacterial Reverse Mutation Assay with S. typhimurium strains TA98, TA100, TA1535 and TA1537 and E. coli strain WP2 uvrA in both the presence and absence of a metabolic activation system (S9-mix: Aroclor-induced rat liver). This study was performed in accordance with USFDA GLP and OECD GLP (1997). The study design was based on OECD 471 (1998). The test article was prepared in DMSO for addition to the test system. Doses were selected based on an initial toxicity-mutation assay. In the confirmatory assay, the test article was tested at 50, 150, 500, 1500 and 5000 ug per plate in the presence and absence of metabolic activation. Precipitate was observed beginning at 1500 ug/plate and no toxicity was observed. No increase in revertant colonies was observed in either the presence or absence of S9-mix for any strain. All criteria for a valid study were met as described in the protocol. Under the conditions of this study, the test article was not mutagenic in the Bacterial Reverse Mutation Assay in either the presence or absence of metabolic activation.