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Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

In a study according to OECD TG 471 under GLP concidtions, the substance was not mutagenic in five strains (TA97, TA98, TA100, TA102, TA1535) with and without metabolic activation.

Moreover, based on the results of an analogue approach (read-across), the substance is considered to be not genotoxic in an in vitro micronucleus assay (OECD 487, GLP) and in an in vitro mammalian gene mutation assay (OECD 476, HPRT test, GLP).

Link to relevant study records

Referenceopen allclose all

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Version / remarks:
July 21, 1997
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
GLP compliance:
yes (incl. certificate)
Type of assay:
bacterial reverse mutation assay
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Batch no. 1190567/006 provide by Raschig GmbH
- Expiration date of the lot/batch: 31.08.2001
- Purity test date: not available

RADIOLABELLING INFORMATION (if applicable): not available

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: stable
- Stability under test conditions: stable
- Solubility and stability of the test substance in the solvent/vehicle: soluble (in water)
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: none

TREATMENT OF TEST MATERIAL PRIOR TO TESTING: none

Species / strain / cell type:
S. typhimurium TA 97
Species / strain / cell type:
S. typhimurium TA 98
Species / strain / cell type:
S. typhimurium TA 100
Species / strain / cell type:
S. typhimurium TA 102
Species / strain / cell type:
S. typhimurium TA 1535
Metabolic activation:
with and without
Metabolic activation system:
S9
Test concentrations with justification for top dose:
First experiment: 0.05.; 0.15; 0.5; 1.5; 5.0 mg/plate, where 5 mg/plate is the recommended maximum test concentration for soluble non-cytotoxic substances
Second experiment: 1.25; 2.5; 5.0 mg/plate, where 5 mg/plate is the recommended maximum test concentration for soluble non-cytotoxic substances
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: water
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
sodium azide
benzo(a)pyrene
other: 4-Nitro-1,2-phenylene diamine and 2-Amino-anthrcene
Details on test system and experimental conditions:
METHOD OF APPLICATION: 1st experiment: in agar (plate incorporation); 2nd experiment: preincubation

DURATION
- Preincubation period: 20 min
- Exposure duration: 48 h (both experiments)
- Expression time (cells in growth medium): not available
- Selection time (if incubation with a selection agent): not applicable
- Fixation time (start of exposure up to fixation or harvest of cells): not applicable

SELECTION AGENT (mutation assays): not applicable

NUMBER OF REPLICATIONS: 4 plates per strain and dose

METHODS OF SLIDE PREPARATION AND STAINING TECHNIQUE USED: not applicable

NUMBER OF CELLS EVALUATED: not applicable

DETERMINATION OF CYTOTOXICITY
- Method: other: titre ratio
- Any supplementary information relevant to cytotoxicity: none

OTHER EXAMINATIONS: none

- OTHER: none
Evaluation criteria:
reproducible increase of revertant colonies per plate (factor >=2)
Statistics:
none
Key result
Species / strain:
S. typhimurium TA 97
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
Key result
Species / strain:
S. typhimurium TA 98
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
Key result
Species / strain:
S. typhimurium 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
Key result
Species / strain:
S. typhimurium TA 102
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
Key result
Species / strain:
S. typhimurium TA 1535
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: not available

RANGE-FINDING/SCREENING STUDIES: none

HISTORICAL CONTROL DATA (with ranges, means and standard deviation and confidence interval (e.g. 95%): not available

ADDITIONAL INFORMATION ON CYTOTOXICITY: none

In a study according to OECD TG 471 under GLP concidtions, the substance was not mutagenic in five strains (TA97, TA98, TA100, TA102, TA1535) with and without metabolic activation.

Executive summary:

The mutagenic potential of the substance was investigated in a bacterial reverse mutation assay according to OECD TG 471 under GLP. The substance was soluble and non-cytotoxic up to 5 mg/plate, i.e. the recommended maximum concentration. The substance was tested in five strains (TA97, TA98, TA100, TA102, TA1535) with and without metabolic activation (S9).

In a first experiment, five concentrations (0.05; 0.15; 0.5; 1.5; 5.0 mg/plate) were tested for 48 hours in the plate incorporation method. In the second experiment the concentrations 1.25, 2.5 and 5.0 mg/plate were tested (48 h) in the pre-incubation method. The number of colonies were evaluated by eye.

While the positive controls were clearly mutagenic, the substance did not increase the number of colonies. Therefore, the substance is considered to be not mutagenic.

Endpoint:
in vitro gene mutation study in mammalian cells
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
For this endpoint a one-to-one read across was performed to a chemical similar compound of the same chemical class with a comparable phys. chem. profile and similar response in biological assays. The relevant study was performed according to GLP and the methods applied are fully compliant with OECD Guideline 476. A read across justification is provided in the attached document (Chapter 13).
Reason / purpose:
read-across source
GLP compliance:
yes (incl. certificate)
Type of assay:
in vitro mammalian cell gene mutation test using the Hprt and xprt genes
Key result
Species / strain:
Chinese hamster lung fibroblasts (V79)
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Positive controls validity:
valid
Endpoint:
in vitro cytogenicity / micronucleus study
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
For this endpoint a one-to-one read across was performed to a chemical similar compound of the same chemical class with a comparable phys. chem. profile and similar response in biological assays. The relevant study was performed according to GLP and the methods applied are fully compliant with OECD Guideline 487. A read across justification is provided in the attached document (Chapter 13).
Reason / purpose:
read-across source
GLP compliance:
yes (incl. certificate)
Type of assay:
in vitro mammalian cell micronucleus test
Key result
Species / strain:
lymphocytes: human
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Additional information

In a study according to OECD TG 471 under GLP concidtions, the substance was not mutagenic in five strains (TA97, TA98, TA100, TA102, TA1535) with and without metabolic activation.

Using the analogue approach (read-across) the genotoxicity of a very similar compound (source substance) was investigated in an in vitro micronucleus test according to OECD test guideline 487 under GLP-conditions. As the non-cytotoxic substance (up to the recommended maximum concentration of 10 mM, corresponding to 1.95 mg/mL) did not increase the proportion of binucleated cells with micronuclei as compared to control for the concentrations of 0.49, 0.98 and 1.95 mg/mL, the source substance was considered as not genotoxic under the test conditions. This can also be assumed for the target compound (see read across justification).

Applying the analogue approach (read-across) the mutagenicity of a very similar compoun (source substance) was investigated in an in vitro mammalian cell gene mutation test using the HPRT genes according to OECD test guideline 476 under GLP-conditions using the Chinese hamster cells V79. As the substance was not cytotoxic in a pre-test, it was tested up to the recommended maximum concentration of 10 mM in two experiments (experiment 1: 4 hour treatment with and without metabolic activation; experiment 2: 14 hour treatment without metabolic activation). After an expression time of at least 168 hours and further incubation of 7 days, the substance did not increase mutant colony numbers in a substantial and reproducible dose-dependent manner. Therefore, the source substance was considered as not mutagenic under the conditions of this test. This can also be assumed for the target compound.

Justification for classification or non-classification

As the substance did show any mutagenic potential, it does not need to be classifed for mutagenicity.