Calcination products of BaCO3, SrCO3, MgO, Al2O3, Eu2O3 and MnCO3

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

December 2003 - March 2004

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: The study has been performed according to OECD and/or EC guidelines and according to GLP principles. Only the preparation of the test solutions is not clearly described.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2004

Report date:

2004

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

(1997)

Deviations:

no

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Test material form:

solid: particulate/powder

Details on test material:

- Appearance: White powder

Specific details on test material used for the study:

- Specific gravity used: 3.83
- Solubility: Insoluble in water

Method

Target gene:

- S. typhimurium: Histidine gene
- E. coli: Tryptophan gene

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Species / strain 2

Species / strain / cell type:

E. coli WP2 uvr A

Metabolic activation:

with and without

Metabolic activation system:

Rat liver S9-mix induced by Aroclor 1254

Test concentrations with justification for top dose:

First, a suspension was obtained by extracting the test substance at 500 mg/mL vehicle at 45 degrees Celsius for at least 72 hours prior to each of the 2 experiments. Part of this suspension was diluted and used for 2 of the 5 test solutions in the experiments. Subsequently, part of the 500 mg/mL suspension was centrifuged for 10 minutes at 800 rpm. The extract of that suspension was also diluted and used for 3 of the 5 test solutions.

Experiment 1 and 2:
All strains: Without and with S9-mix: 1.67 and 5 mg/plate and 5.56, 16.67 and 50 mg extract after centrifugation/plate

As precipitation was observed, the test substance concentration was sufficiently high.

Vehicle / solvent:

- Vehicle used: phosphate buffered saline adjusted to pH 6.55
- Justification for choice of vehicle: the test substance is insoluble in water and soluble in inorganic acids. Since the use of acid is not possible, the test substance was extracted in phosphate buffered saline, adjusted to slightly acidic conditions

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

other: see section "Any other information on materials and methods incl. tables"

Details on test system and experimental conditions:

METHOD OF APPLICATION:
- Experiment 1: in agar (plate incorporation)
- Experiment 2: preincubation

DURATION
- Exposure duration: 48 to 72 hours

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

DETERMINATION OF CYTOTOXICITY
- Method: The reduction of the bacterial background lawn.

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

Evaluation criteria:

A test substance is considered positive if a concentration-related increase over the range tested and/or a reproducible increase in at least one or more concentrations in the number of revertant colonies per plate in at least one strain (with or without metabolic activation). Biological relevance of the results will be considered first. The increase in mean revertants at the peak of the dose response is at least three times the mean vehicle control value for strains TA1535 and TA1537 and at least twice the value for strains TA98, TA100 and WP2uvrA.
A test substance is considered negative (not mutagenic) in the test if a result does not meet the above criteria.

Statistics:

Dunnett's analysis was used.

Results and discussion

Test results

Test results 1

Key result

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, but tested up to precipitating concentrations

Vehicle controls validity:

valid

Positive controls validity:

valid

Test results 2

Key result

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Vehicle controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: Slight precipitation was observed at the 2 dose levels 1.67 and 5 mg/plate

RANGE-FINDING/SCREENING STUDIES:
Not applicable

COMPARISON WITH HISTORICAL CONTROL DATA:
- The 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.

ADDITIONAL INFORMATION ON CYTOTOXICITY:
- No toxicity or mutagenicity was observed up to and including the highest dose level

Applicant's summary and conclusion

Conclusions:

The mutagenic activity of the substance was evaluated in accordance with OECD 471 (1997) and according to GLP principles. Based on the results of this study it is concluded that the substance is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay with and without metabolic activation.

Executive summary:

The mutagenic activity of the substance was evaluated in accordance with OECD 471 (1997) and according to GLP principles. The test was performed in two independent experiments, both in the absence and presence of S9-mix. The substance was tested up to limit concentrations. No cytotoxicity was observed. Slight precipitation was observed at the 2 dose levels 1.67 and 5 mg/plate.

Adequate solvent and positive controls were included. The substance did not induce a significant dose-related increase in the number of revertant (His+) colonies in each of the four S. typhimurium tester strains (TA1535, TA1537, TA98 and TA100) and in the number of revertant (Trp+) colonies in tester strain WP2uvrA, both in the absence and presence of S9-metabolic activation. These results were confirmed in independently repeated experiments. Based on the results of this study it is concluded that the substance is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay with and without metabolic activation.