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Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information
The substance is not mutagenic in bacteria. Data on related substances shows that it is not mutagenic in cultivated mammalian cells and not clastogenic.
Link to relevant study records
Reference
Endpoint:
in vitro gene mutation study in bacteria
Remarks:
Type of genotoxicity: gene mutation
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: guideline study; restriction: non-GLP
Qualifier:
according to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
GLP compliance:
no
Type of assay:
bacterial reverse mutation assay
Target gene:
S. typhimurium and E. coli
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2
Metabolic activation:
with and without
Metabolic activation system:
Aroclor-induced rat liver S-9 mix
Test concentrations with justification for top dose:
20 ug - 5000 ug/plate (Standard Plate Test = SPT)
4 ug - 2500 ug/plate (Preincubation Test = PIT)
Vehicle / solvent:
Due to the limited solubility of the test substance in water, DMSO was selected as the vehicle, which had been demonstrated to be suitable in bacterial reverse mutation tests and for which historical control data are available.
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
yes
Positive controls:
yes
Positive control substance:
other: with metabolic activation: 2-aminoanthracene; without metabolic activation: N-methyl-N'-nitro-N-nitrosoguanidine, 4-nitro-o-phenylendiamine, 9-aminoacridine and 4-nitroquinoline-N-oxide
Details on test system and experimental conditions:
Standard Plate Test and Preincubation Test
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
other: A weak bacteriotoxic effect was observed under all test conditions.
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
An increase in the number of his+ or trp+ revertants was not observed in the standard plate test or in the preincubation test either withaut S-9 mix or after the addition of a metabolizing system.
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.
Conclusions:
Interpretation of results (migrated information):
negative

Accarding to the results of the present study, the test substance Neozapon Blau 807 Typ neu is not mutagenic in the Salmonella typhimurium/Escherichia coli reverse mutation assay under the experimental cond itions chosen here.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Additional information

Additional information from genetic toxicity in vitro:

Solvent Blue 70 was tested for its mutagenic potential based on the ability to induce point mutations in selected Ioci of several bacterial strains,

(TA 1535, TA 100, TA 1537, TA 98 and E. coli WP2 uvrA), in a reverse mutation assay (OECD 471) (BASF 2003). Doses were for the standard plate test between 20 and 5000mg/plate and for the preincubation test between 4 and 2500mg/plate. Aroclor-induced rat liver S-9 mix was used for metabolic activation. Precipitation ofthe test substance was found from about 100mg/plate onward. A weak bacteriotoxic effect was observed under all test conditions. The substance did not induce an increase in mutant frequency and was found to be non mutagenic in the Ames test. The tested sample was a commercial product containing 100% of the UVCB substance.

Mutagenicity in mammalian cells in vitro (weight-of-evidence; see also attachment)

No mutagenicity was observed in bacteria. As this substance is of very low solubility in water, testing in mammalian cells in vitro can only be performed at very low doses. Bacteria, that are much less sensitive to the presence of precipitates, were tested with the limit concentration of 5000 µg/plate. Both the main and the remote analogue, which are both of higher water solubility, were tested in GLP-compliant HPRT tests (OECD 476, Klimisch 1) and found to be non mutagenic. The structures are representative for the target as they contain relevant functional groups. Alkylchains are not a structural feature known for association with mutagenic properties. Therefore, the slight difference in length of the alkyl side chain is not considered relevant for the outcome of the in-vitro gene mutation assay.

In addition, as outlined in the toxicokinetic section, the substance is considered too large and too insoluble for cellular uptake.

 

Solvent Blue 67: An OECD 476 study was performed to investigate the potential to induce gene mutations at the HPRT locus in V79 cells of the Chinese hamster (BASF 2012). The study was performed under GLP and is valid without restrictions.

The assay was performed in two independent experiments, using two parallel cultures each. The first main experiment was performed with and without liver microsomal activation and a treatment period of 4 hours.The second experiment was performed with a treatment time of 4 hours with and 24 hours without metabolic activation.

The highest concentration applied in the pre-experiment (2500.0 µg/mL) was limited by the solubility properties of the test item in DMSO and aqueous medium. The concentration range of the main experiments was limited by precipitation of the test item.No substantial and reproducible dose dependent increase of the mutation frequency was observed up to the maximum concentration with and without metabolic activation. Appropriate reference mutagens (EMS and DMBA), used as positive controls, induced a distinct increase in mutant colonies and thus, showed the sensitivity of the test system and the activity of the metabolic activation system.

Clastogenicity in vitro (weight of evidence, see also attachment)

The main analogue substance was found to be non clastogenic in a valid in vivo micronucleus study in Chinese hamster.

The substance is not skin sensitizing indicating that it lacks the ability to covalently bind to proteins. In addition, as outlined in the toxicokinetic section, the substance is considered too large and too insoluble for cellular uptake.

A study for clastogenicity in Chinese Hamster is available (Ciba-Geigy Ltd. 1982). The study is titled to be a nucleus anomaly test as the term micronucleus assay and its OECD testing guideline were not yet established. The design of the study is however comparable to this guideline with deviations that only 1000 bone marrow cells were scored and that evaluation criteria were given merely as a statistically significant increase in the number of cells with anomalies. The doses of 116, 234 and 468 mg/kg bw given by gavage in peanut oil were based on incidences of mortality in a range-finder study.

The experiment was performed to evaluate any mutagenic effect on somatic interphase cells in vivo. Mutagenic effects present themselves in interphase cells in form of nucleus anomalies of bone marrow cells. These anomalies occur in interphase cells as a consequence of damage during the mitotic process. The increase in anomalies shows a clear dose dependency, comparable to the occurrence of chromosome aberrations in metaphase preparations. Treatment consisted of one daily dose on two consecutive days. The animals were sacrificed 24 h after the second application. From the bone marrow smears were made.

The percentage of anomalies in bone marrow smears from animals treated with various doses of the test substance (range 0 - 0.3) showed no significant difference from the control (range 0.1- 0.2).

The incidence of bone marrow cells with anomalies of nuclei corresponds to the frequency observed in the control group. By contrast, a positive control experiment with cyclophosphamide

(128 mg/kg) yielded 5.6% cells with anomalies of nuclei. This is significantly different from the controls treated with the vehicle alone.

In conclusion, the test substance did not cause clastogenicity in Chinese Hamsters in vivo. Therefore, no experimental data on clastogenicity in vitro was retrieved.

Justification for classification or non-classification

Dangerous Substance Directive (67/548/EEC)

The available studies are considered reliable and suitable for classification purposes under 67/548/EEC. No mutagenicity was observed in bacteria and cultivated mammalian cells. No genotoxicity was observed in a micronucleus assay in vivo. As a result the substance is not considered to be classified for mutagenicity under Directive 67/548/EEC, as amended for the 28th time in Directive 2001/59/EC.

Classification, Labelling, and Packaging Regulation (EC) No. 1272/2008

The available experimental test data are reliable and suitable for classification purposes under Regulation 1272/2008.

No mutagenicity was observed in bacteria and cultivated mammalian cells. No genotoxicity was observed in a micronucleus assay in vivo.

As a result the substance is not considered to be classified for mutagenicity under Regulation (EC) No. 1272/2008, as amended for the fifth time in Directive EC 944/2013.

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