[N,N,N',N',N'',N''-hexaethyl-29H,31H-phthalocyaninetrimethylaminato(2-)-N29,N30,N31,N32]copper

Endpoint summary

Additional information

Bacterial reverse mutation assay (Ames test)

In a reverse gene mutation assay in bacteria (BASF AG, 1999, OECD 471), strains TA1535, TA 1537, TA100, TA 98 of Salmonella typhimurium and WP2 uvr A of Escherichia coli were exposed to the test item (CAS number 28654 -73 -1) up to 5000 µg/plate in the presence and absence of mammalian metabolic activation S9 -mix. A slight decrease in revertants was observed as sign for bacterial toxicity in two independent experiments in concentrations of 500 -2500 µg/plate. In addition, precipitation from about 100 µg/plate onward was observed. The positive controls induced the appropriate responses in the corresponding strains.There was no evidence of induced mutant colonies over background. Therefore, the test item was considered to be non-mutagenic in this bacterial reverse mutation assay.

 

In a supporting gene mutation assay in bacteria (BASF AG, 2001, OECD 471) the Salmonella typhimurium strain 1537 (only one strain tested) was exposed to the test item (CAS number 28654 -73 -1) in the presence and absence of mammalian metabolic activation S9 -mix in concentrations up to 10000 µg/plate. Data about exposure duration or bacterial toxicity were not cited in the report. Precipitation occurred from about 100 µg/plate onwards. Some plates could not be evaluated because contaminations were observed. However, the authors mentioned that the result was not influenced by this confounding factor.

Under the experimental conditions reported, the test item did not induce genotoxicity as determined by this Salmonella typhimurum reverse mutation assay.

In vitro mammalian cell gene mutation test (HPRT test) (read across)

CAS number 75247 -18 -6 was assessed for its potential to induce gene mutations at the HPRT locus using V79 cells of the Chinese hamster according to OECD guideline 476 (BASF SE, 2012). The assay was performed in two independent experiments with identical experimental procedures, using two parallel cultures each. The first experiment was performed with and without liver microsomal activation and a treatment period of 4 hours. The second experiment was performed with a treatment period of 24 hours in the absence of metabolic activation and 4 hours in the presence of metabolic activation.

The cell cultures were evaluated at the following concentrations:

Experiment I:

without S9 mix: 0.8; 1.5; 3.0; 6.0 and 9.0 µg/mL

with S9 mix: 3.0; 6.0; 12.0; 24.0 and 48.0 µg/mL

Experiment II:

without S9 mix: 0.5; 1.0; 2.0; 3.0 and 4.0 µg/mL

with S9 mix: 0.8; 1.5; 3.0 and 6.0 µg/mL

 

Relevant cytotoxic effects indicated by a relative cloning efficiency or cell density below 50% in both parallel cultures occurred in the first experiment at 6.0μg/mL and above without metabolic activation and at 48.0μg/mL with metabolic activation. In the second experiment cytotoxic effects occurred at 3.0μg/mL and above with metabolic activation. Precipitation of the test item was noted at 6.0μg/mL and above in both experiments with metabolic activation. No relevant and reproducible increase in mutant colony numbers/10E6 cells was observed in both experiments up to and including the maximum concentration.

A linear regression analysis (least squares) was performed to assess a possible dose dependent increase of mutant frequencies. A significant dose dependent trend of the mutation frequency indicated by a probability value of <0.05 was solely detected in the first experiment with metabolic activation. This trend however, was judged as irrelevant as the mutation frequency remained within the historical range of solvent controls at all of the data points and there was no significant trend in the parallel culture under identical conditions.

In both experiments of this study (with and without S9 mix) the range of the solvent controls was from 10.8 up to 35.0 mutants per 10E6 cells; the range of the groups treated with the test item was from 4.9 up to 35.5 mutants per 10E6 cells. EMS (150μg/mL) and DMBA (1.1μg/mL) were used as positive controls and showed a distinct increase in the number of induced mutant colonies. Under the experimental conditions reported the test item did not induce gene mutations at the HPRT locus in V79 cells.

In vitro cytogenicity study in mammalian cells (Chromosome aberration) (waiving)

In accordance with column 2 of REACH Annex VIII, the in vitro cytogenicity study in mammalian cells does not need to be conducted if adequate data from an in vivo cytogenicity test are available. Since an in vivo mammalian erythrocyte micronucleus test is available for a structurally similar substance no test on cytogenicity on mammalian cells in vitro has to be conducted.

Mammalian erythrocyte micronucleus test (CAS number 81457 -65 -0) (read across)

The test item (CAS number 81457 -65 -0) was assessed in a nucleus anomaly test (BASF AG, 1982; equivalent to OECD 474) for its potential to induce clastogenic effects in the bone marrow of the Chinese hamster. The test item, dissolved in peanut oil, was administered orally to 3 Chinese hamsters/sex at dose levels of 116, 234 or 468 mg/kg bw once a day for 2 days. The post exposure period was 24 hours. Cyclophophamide was used as positive control of 128 mg/kg bw. After sacrifice, bone marrow cells were harvested, mixed and transferred on slides. 1000 bone marrow cells each were scored per animal and the following anomalies were registered: single jolly bodies, fragments of nuclei in erythrocytes, micronuclei in erythroblasts, micronuclei in leucopoietic cells and polyploid cells. In all dosage groups the percentage of cells displaying anomalies of nuclei did not differ significantly from the negative control. The positive control yielded a significant increase of the percentage of cells with anomalies. In conclusion of this experiment, no evidence of mutagenic effects was obtained in Chinese hamsters treated with the test substance.

Justification for selection of genetic toxicity endpoint
Two in vitro tests (Ames test, HPRT assay) and one in vivo test (micronucleus test) are used for assessment of genotoxicity. Since an in vivo mammalian erythrocyte micronucleus test is available no test on cytogenicity on mammalian cells in vitro has to be conducted. In a read across assessment the reliable data of the CAS number 75247-18-6 (HPRT assay) and CAS number 81457-65-0 (in vivo mammalian erythrocyte micronucleus test) was used . For detail please refer to chaper 13 of the IULCID, Assessment reports.

Short description of key information:
The absence of a mutagenic potential was demonstrated for gene mutations in bacteria (Ames test, target chemical) and in mammalian cells (HPRT assay, read across from CAS number 75247-18-6). A mammalian erythrocyte test with CAS number 81457-65-0 (read across) demonstrated the absence of clastogenicity in vivo.

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

Based on the results of the available genetic toxicity studies, the test item CAS number 28654 -73 -1 does not need to be subjected for classification and labelling for mutagenicity/germ cell mutagenicity according to Directive 67/548/EEC (DSD) and Regulation (EC) No 1272/2008(CLP).