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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
28-jul-2009 to 13-aug-2009
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The study has been performed according to OECD and/or EC guidelines and according to GLP principles.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2009
Report date:
2009

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Type of assay:
bacterial reverse mutation assay

Test material

Constituent 1
Chemical structure
Reference substance name:
Potassium chlorate
EC Number:
223-289-7
EC Name:
Potassium chlorate
Cas Number:
3811-04-9
Molecular formula:
ClHO3.K
IUPAC Name:
potassium chlorate
Details on test material:
- Name of test material (as cited in study report): Potassium chlorate
- Substance type: Fine white crystals
- Physical state: Solid
- Stability under test conditions: Stable
- Storage condition of test material: At room temperature in the dark

Method

Target gene:
- S. typhimurium: Histidine gene
- E. coli: Tryptophan gene
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 induced by a combination of phenobarbital and ß-naphthoflavone
Test concentrations with justification for top dose:
Experiment 1:
Preliminary test (without and with S9) TA100 and WP2uvrA: 3, 10, 33, 100, 333, 1000, 3330 and 5000 µg/plate

Main study: TA1535, TA1537 and TA98:
With and without S9-mix: 333, 1000, 3330 and 5000 µg/plate

Experiment 2:
TA1535, TA1537, TA98, TA100 and WP2uvrA:
With and without S9-mix: 333, 1000, 3330 and 5000 µg/plate
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: Milli-Q water
- Justification for choice of solvent/vehicle: Test substance forms dangerous reaction with ethanol and DMSO.
Controlsopen allclose all
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
Remarks:
, ethanol
Positive controls:
yes
Positive control substance:
methylmethanesulfonate
Remarks:
without S9

Migrated to IUCLID6: 650 µg/plate in DMSO for TA100
Positive controls:
yes
Positive control substance:
2-nitrofluorene
Remarks:
without S9

Migrated to IUCLID6: 10 µg/plate in DMSO for TA98
Positive controls:
yes
Positive control substance:
sodium azide
Remarks:
without S9

Migrated to IUCLID6: 5 µg/plate in saline for TA1535
Positive controls:
yes
Positive control substance:
9-aminoacridine
Remarks:
without S9

Migrated to IUCLID6: 60 µg/plate in milli-Q water for TA1537
Positive controls:
yes
Positive control substance:
4-nitroquinoline-N-oxide
Remarks:
without S9

Migrated to IUCLID6: 10 µg/plate in DMSO for WP2uvrA
Positive controls:
yes
Positive control substance:
other: 2-aminoanthracene in DMSO for all tester strains
Remarks:
with S9
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 48 hour

NUMBER OF REPLICATIONS:
- Doses of the test substance were tested in triplicate in each strain. Two independent experiments were conducted.

NUMBER OF CELLS EVALUATED: 10E8 per plate

DETERMINATION OF CYTOTOXICITY
- Method: The reduction of the bacterial background lawn, the increase in the size of the microcolonies and the reduction of the revertant colonies

OTHER EXAMINATIONS:
- The presence of precipitation of the test compound on the plates was observed.


Evaluation criteria:
A test substance is considered negative (not mutagenic) in the test if:
a) The total number of revertants in tester strain TA100 is not greater than two (2) times the concurrent control, and the total number of revertants in tester strains TA1535, TA1537, TA98 or WP2uvrA is not greater than three (3) times the concurrent control.
b) The negative response should be reproducible in at least one independently repeated experiment.

A test substance is considered positive if:
a) A two-fold (TA100) or more or a three-fold (TA1535, TA1537, TA98, WP2uvrA) or more increase above solvent control in the mean number of revertant colonies is observed in the test substance group.
b) The increase in the mean number of revertant colonies follows the concentration of test substance (dose-response relationship).

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
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
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: No precipitation was observed up to and including the top of 5000 µg/plate

RANGE-FINDING/SCREENING STUDIES:
- No toxicity or mutagenicity was observed up to and including the top of 5000 µg/plate

COMPARISON WITH HISTORICAL CONTROL DATA:
- The negative and strain-specific positive control values were within our laboratory historical control data ranges indicating that the test conditions were adequate and that the metabolic activation system functioned properly.

Applicant's summary and conclusion

Conclusions:
Under the test condition, the test substance is not mutagenic with and without metabolic activation in S. typhimurium (strains TA1535, TA1537, TA98 and TA100) and E.coli WP2 uvrA.
Executive summary:

In a reverse gene mutation assay performed according to the OECD test guideline No. 471 and in compliance with GLP, Salmonella typhimurium strains TA 1535, TA 1537, TA 98 and TA 100 and Escherichia coli strain WP2 uvrA were exposed to test substance, both with and without the addition of a rat liver homogenate metabolizing system. 


 


All bacterial strains showed negative responses over the entire dose range, i.e. no significant dose-related increase in the number of revertants in two independently repeated experiments.


The negative and strain-specific positive control values were within the laboratory historical control data ranges indicating that the test conditions were adequate and that the metabolic activation system functioned properly.


 


Under the test condition, test substance did not induce gene mutations by base pair changes or frameshifts in the genome of the strains used. It can be concluded that test substance is not mutagenic with and without metabolic activation in S. typhimurium (strains TA1535, TA1537, TA98 and TA100) and E.coli WP2 uvrA according to the criteria of the Annex VI of the Regulation (EC) No.1272/2008 (CLP) and to the GHS. This study is considered as acceptable and satisfies the requirement for reverse gene mutation endpoint.

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