[2,9,16,23-tetrachloro-29H,31H-phthalocyaninato(2-)-N29,N30,N31,N32]copper

Endpoint summary

Additional information

A category assessment is performed, which is based on the hypothesis that the copper phthalocyanine pigments are too insoluble in water or fat / octanol for cellular uptake. A data matrix and further information are provided in the chapter on toxicokinetic properties. All available data has consistently shown absence of genotoxic properties. The full set of in-vitro/in-vivo studies is available for the basic copper phthalocyanine, which has the lowest molecular weight. Experimental data is also available for the halogenated derivatives.

Data on category members is summarized below:

CAS No. 147-14-8:

GENOTOXICITY IN-VITRO:

Valid experimental data were available to assess the genetic toxicity in-vitro.

Gene mutation in bacteria

Copper phthalocyanine was not mutagenic in a standard plate Ames test with and without metabolic activation (tested up to 10000 μg/plate in Salmonella typhimurium TA1535, TA1537, TA98 and TA100; metabolic activation: S9 fraction from the liver of male Sprague-Dawley rats, treated with a single dose of 500 mg/kg bw Aroclor 1254 five days before sacrifice and mixed with a series of cofactors. Cytotoxicity was not observed with or without metabolic activation.

In another Ames test a pre-incubation assay was conducted with and without metabolic activation (tested up to 5000 μg/plate in Salmonella typhimurium TA100, TA 102, TA 98 and TA97; metabolic activation: S9 fraction from the liver of rats, treated with KC-400, equivalent to PCB), copper phthalocyanine was also not mutagenic (JETOC 1995). No cytotoxicity was observed.

Several other studies provided also negative results for copper phthalocyanine in the Ames test (Hayatsu 1983, Val. 4; Manandhar 1982, Val. 4). In another Ames test, a positive result was obtained. However, the mutagenicity test was conducted in two S. typhimurium strains (TA1535 and TA1538) only. No data were available in terms of cytotoxicity and validity of positive, solvent and negative controls (Milvy 1978, Val. 3).

Gene mutation in mammalian cells

The test substance was negative for genotoxicity in a mouse lymphoma assay using L5178Y cells at levels up to 1000 µg/plate (with and without metabolic activation) (Manandhar, Val. 4).

Cytogenicity in mammalian cells

CHL cells were used in an in-vitro chromosomal aberration test (acc. Japanese Guidelines for Screening Mutagenicity Testing of Chemicals, JETOC 1995), with and without metabolic activation. The test substance was found to be negative for causing cytogenicity at dose levels from 750 µg/ml up to 3000 µg/ml. Cytotoxicity was observed at > 3000 µg/ml.

Genome mutation in mammalian cells

A cell transformation test using C3H/10T1/2 was found to be negative for causing genome mutations at dose levels from 10 µg/plate up to 1000 µg/plate (Manandhar 1982, Val. 4).

DNA damage and/or repair

An UDS assay with rat hepatocytes was found to be negative for DNA damage/or repair at dose levels from 10 µg/plate up to 1000 µg/plate (Manandhar 1982, Val. 4).

It should be noted that there were several weakly positive Ames test results and one weakly positive HPRT test, when crude copper phthalocyanine was investigated for mutagenic effects in bacteria as well as in V79 cells (Ciba-Geigy 1986 (3x) and 1988 (1x), all Val. 3). However, today it is known that these crude copper phthalocyanines were manufactured by using nitrobenzene and chlorobenzene as solvents in the past. Therefore the test material contained residues of these solvents and these impurities were considered to be responsible for the weakly mutagenic effects observed in-vitro. Meanwhile this manufacturing process was replaced by other manufacturing processes which all use alternative solvents.

GENOTOXICITY IN-VIVO:

Valid experimental data were also available to assess the genetic toxicity in-vivo.

Crude Copper Phthalocyanine was administered by gavage to Chinese hamsters (Cricetulus griseus) of either sex. Treatment consisted of one daily dose of 1250, 2500 or 5000 mg/kg on each of two consecutive days. The animals were sacrificed 24 h after the second application. From the bone marrow smears were made. The experiment was performed to evaluate any mutagenic effect on somatic interphase cells in vivo (Ciba Geigy 1986, Val. 2). The bone marrow smears from animals treated with the various doses of the test material showed no significant difference from the control. The incidence of bone marrow cells with anomalies of nuclei corresponded to the frequency observed in the control group. By contrast, a "positive control" experiment with cyclophosphamide (128 mg/kg bw) yielded 9.48 % cells with anomalies of nuclei. This is significantly different from the controls (0.1 %) treated with the vehicle (0.5 % CMC) alone.

In a second in-vivo experiment, crude copper phthalocyanine was administered in a single intraperitoneal injection to pregnant female mice (C57 Bl/6) on the 10th day after conception. Doses of 1250, 2500 and 5000 mg/kg were given. The experiment was performed to ascertain whether the substance might have a mutagenic effect on somatic cells in vivo. The mouse spot test system permits the detection of induced point mutations and other genetic events in the melanoblasts of embryos exposed in utero. The induction of mutation is monitored post-natally by examination of the fur of young mice for recessive spots (RS) resulting from expression of recessive genes involved in coat-colour determination (Ciba Geigy 1986, Val. 2). The average litter size was not markedly affected by any of the doses administered. The survival rate of the young animals at the beginning of the observation period (approx. the 12 th day) was decreased with increasing doses. Altogether 899 animals were examined for spots. 0.29 % of the control animals showed RS. The frequencies of RS in the group treated with 1250, 2500 and 5000 mg/kg were respectively 0.93, 0 and 0 %. Thus, the relative incidence of RS among the animals treated with the various doses of the test material did not differ significantly from that of the control (sesame oil). By contrast, the positive control experiment with ethylnitrosourea (50 mg/kg) performed simultaneously yielded an statistically significant average RS frequency of 4.75 %.

In both of these in-vivo tests, also the test material TK 13143 (crude copper phthalocyanine) was used which had produced slightly positive effects in in-vitro tests (see above). However, in these in-vivo experiments no evidence for mutagenic effects were obtained.

CAS No. 1328-53-6:

GENOTOXICITY IN-VITRO:

Valid experimental data were available to assess the genetic toxicity in-vitro.

Gene mutation in bacteria

Polychloro copper phthalocyanine was not mutagenic in a preincubation Ames test (tested up to 5000 μg/plate in Salmonella typhimurium TA1535, TA1537, TA98, TA100 and E. coli WP2 uvrA with metabolic activation, up to 625 µg/plate in S. typhimurium TA100, TA1535, TA98 and TA1537 as well as up to 5000 µg/plate in E. coli WP2 uvrA without metabolic activation; metabolic activation: Rat liver, induced with phenobarbital and 5,6-benzoflavone. Cytotoxicity was observed at 313 µg/plate or more (TA100 and TA1535) and at 625 µg/plate (TA98 and TA1537) without metabolic activation, and at 5000 µg/plate (TA100, TA1535, TA98 and TA1537) with metabolic activation (acc. Guidelines for Screening Mutagenicity Testing of Chemicals Japan, JETOC 2001).

Polychloro copper phthalocyanine was also not mutagenic in a preincubation Ames test with and without metabolic activation (tested up to 5000 μg/plate in Salmonella typhimurium TA1535, TA1537, TA98, TA100 and E. coli WP2 uvrA with and without metabolic activation; metabolic activation: Rat liver, induced with phenobarbital and 5,6-benzoflavone. Cytotoxicity was not observed.

In another Ames test a pre-incubation assay and a standard plate test were conducted, both with and without metabolic activation (tested up to 5000 μg/plate in Salmonella typhimurium strains TA100, TA98, TA1535 and TA1537; metabolic activation: S9 fraction from the liver of male Sprague-Dawley rats (treated with a single dose of 500 mg/kg bw Aroclor 1254 five days before sacrifice) and mixed with a series of cofactors. Polychloro copper phthalocyanine was also not mutagenic in this assay (comp. OECD 471, BASF AG 1988). No cytotoxicity was observed.

In another Ames test, a weakly positive result was obtained at concentrations >= 333 µg/plate in S. typhimurium TA98 only with metabolic activation (comp. OECD 471, Zeiger 1988). The mutagenicity potential of the test substance was tested up to 10000 μg/plate in two Salmonella typhimurium strains TA100 and TA98; metabolic activation: S9-mix, prepared from the liver of either male SD rats or male Syrian hamsters, induced with Aroclor 1254.Without metabolic activation in the strain TA98 as well as in the strain TA100 (with or without metabolic activation), no increase of the number of his+ revertants was observed. Since the analyzed purity of the test substance used in this study was indicated to be 94 % and no further information on the remaining content was given, this result may be due to impurities of the test substance.

Another study provided also negative results for polychloro copper phthalocyanine in the Ames test (Milvy 1978, Val. 4).

Cytogenicity in mammalian cells

In a Mammalian Cell Micronucleus Test (acc. OECD 487 draft proposal, BASF AG 2001) V79 cells were used, with and without metabolic activation. The test substance was found to be negative for causing cytogenicity at dose levels from 0.78 µg/ml up to 75 µg/ml. Cytotoxicity was not observed.

CHL/IU cells were used in two in-vitro chromosomal aberration tests (acc. Japanese Guidelines for Screening Mutagenicity Testing of Chemicals, JETOC 1995 and 2001), with and without metabolic activation. The test substance was found to be negative for causing cytogenicity at dose levels up to 5000 µg/ml. Cytotoxicity was not observed.

 

CAS No. 574-93-6:

GENOTOXICITY IN-VITRO:

Valid experimental data were available to assess the genetic toxicity in-vitro.

Gene mutation in bacteria

Heliogen Blue MFA was not mutagenic in a preincubation in a standard plate Ames test (tested up to 5000 μg/plate in Salmonella typhimurium TA1535, TA1537, TA98, TA100 and E. coli WP2 uvrA with and without metabolic activation (S9 fraction from the liver of male Sprague-Dawley rats (treated i.p. with 500 mg/kg bw Aroclor 1254, 5 days before sacrifice), mixed with a series of cofactors). Cytotoxicity was not observed (acc. OECD guideline 471, BASF AG 2000).

Heliogen Blue MFA was also not mutagenic in a preincubation test with and without metabolic activation (tested up to 5000 μg/plate in Salmonella typhimurium TA98 and TA100 with and without metabolic activation; metabolic activation: Rat liver, induced with phenobarbital and 5,6-benzoflavone. Cytotoxicity was not observed.

In two further tests, standard plate tests were conducted, both with and without metabolic activation, tested up to 5000 μg/plate in Salmonella typhimurium strains TA100, and TA98. Polychloro copper phthalocyanine was also not mutagenic in this assays (comp. OECD 471, BASF AG 1993). No cytotoxicity was observed.

In another Ames test, a weakly positive result was obtained. Without S9-mix, no increase in the number of his+ revertants was seen in all strains tested. With S9-mix, no increase in the number of his+ revertants was seen for the strains S. typhimurium TA 100. Weakly positive reactions were seen with a maximum increase in the number of mutant colonies at 6000µg/plate for TA 98. Since the analyzed purity of the test substance used in this study was unknown, this result may be due to impurities of the test substance (comp. OECD 471, BASF AG 1993).

Another study with limited validity also provided negative results for polychloro copper phthalocyanine in the Ames test (BASF AG 1993, Val. 4).

 

CAS No. 14302-13-7:

GENOTOXICITY IN-VITRO:

Valid experimental data were available to assess the genetic toxicity in-vitro.

Gene mutation in bacteria

Heliogen Green 8GA was not mutagenic in a preincubation in a standard plate Ames test (tested up to 5000 μg/plate in Salmonella typhimurium TA1535, TA1537, TA98, TA100 and E. coli WP2 uvrA with and without metabolic activation (S9 fraction from the liver of male Sprague-Dawley rats (treated i.p. with 500 mg/kg bw Aroclor 1254, 5 days before sacrifice), mixed with a series of cofactors). Cytotoxicity was occasionally observed depending on the strain and test conditions from about 500 µg - 2500 µg/plate onward. (acc. OECD guideline 471, BASF AG 2000).

 

CAS No. 27614-71-7:

GENOTOXICITY IN-VITRO:

Valid experimental data were available to assess the genetic toxicity in-vitro.

Gene mutation in bacteria

Heliogen Blue K 6915 was not mutagenic in a preincubation and in a standard plate Ames test (tested up to 5000 μg/plate in Salmonella typhimurium TA1535, TA1537, TA98, TA100 and E. coli WP2 uvrA with and without metabolic activation (S9 fraction from the liver of male Sprague-Dawley rats (treated i.p. with 500 mg/kg bw Aroclor 1254, 5 days before sacrifice), mixed with a series of cofactors). Cytotoxicity was not observed (acc. OECD guideline 471, BASF AG 2000).

 

CAS No. 68987-63-3:

Genotoxicity in vitro:

Gene mutation in bacteria

The substance was tested in a GLP and OECD guideline 471 compliant study up to the limit dose (Safepharm 1994). The plate incorporation method was used for both experiments. No cytotoxicity, but precipitation was observed. The pigment was not mutagenic in Salmonella typhimurium TA1535, TA1537, TA98, TA100 and E. coli WP2 uvrA with and without metabolic activation.

Clastogenicity in vitro

The effects on chromosomal structure of exposure to the substance were investigated in cultured CHL (Chinese hamster lung) cells (DIC 1997). Tests were conducted with and without the inclusion of a rat liver-derived metabolic activation system (S-9 mix):

without S-9 mix cells were exposed continuously for 6, 24 or 48 hours, with S-9 mix exposure was limited to 6 hours; cells exposed for 6 hours were cultured in fresh medium for a further 18 hours before harvesting.

Treatments were established by the addition oftest suspensions (in culture medium) to 24-hour cultures. Cell division was arrested by the addition ofthe spindle poison, Colcemid, two hours before the cells were harvested; slides were then prepared for microscopic analysis.

All slides were scored for chromosomal aberrations. The highest concentration scored for chromosomal aberrations (5000 μg/ml) is the highest required by EC, OECD, US and Japanese regulatory test guidelines.

One hundred metaphases were analysed from all selected cultures. Treatment with the test substance did not produce biologically or statistically significant increases in the frequency of metaphases with aberrant chromosomes at any concentration tested,

compared to vehicle control values (p>0.05 both including and excluding gap-type aberrations), at any sampling time, either in the presence or absence of S-9 mix.

The known clastogens, Mitomycin C and cyclophosphamide, induced significant increases in the frequency of metaphases with aberrant chromosomes, compared to the vehicle control values, at both sampling times in both cytogenetic tests (p<0.001 in all cases), thus demonstrating the sensitivity ofthe test procedure, and the metabolic activity of the S-9 mix employed.

Short description of key information:
Copper phthalocyanine based pigments have shown to be non genotoxic both in-vitro and vivo.

Endpoint Conclusion: No adverse effect observed (negative)

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 genotoxicity was observed in-vitro or in vivo. As a result the substance is not considered to be classified for mutagenicity under Directive 67/548/EEC, as amended for the 31st time in Directive2009/2/EG.

 

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 genotoxicity was observed in vitro or 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 EC944/2013.