Anthraquinone

Endpoint summary

Additional information

Commercial anthraquinone (AQ) is produced by at least three different production methods:

- Oxidation of anthracene (AQ-OX)

 - Friedel-Crafts technology (AQ-FC)

- Diels-Alder chemistry (AQ-DA)

with the final product varying in color and purity.

AQ-OX begins with anthracene produced from coal tar and different lots can contain various contaminants, particularly the mutagenic isomers of nitroanthracene. AQ has been reported to be negative in a variety of genotoxicity tests including numerous Ames mutagenicity assays. In addition, it is reported that AQ-DA is negative in the Salmonella-Escherichia coli reverse mutation assays, the L5178Y mouse lymphoma forward mutation assay, for inducing chromosomal aberrations, polyploidy or endoreduplication in Chinese hamster ovary cells, and in the in vivo mouse micronucleus assay (Butterworth et al., 2001). Further, a previous 18 month bioassay conducted with AQ administered to male and female B6C3F1 and (C57BL/6 x AKR)F1 mice reported no induction of cancer. Thus, it was somewhat unexpected that in a long-term study conducted by the National Toxicology Program (NTP) AQ-OX induced a weak to modest increase in tumors in the kidney and bladder of male and female F344/N rats and a strong increase in the livers of male and female B6C3F1 mice. In the studies reported in Butterworth et al. (2001) a sample of the AQ-OX used in the NTP bioassay was shown to be mutagenic in the Ames tester strains TA98, TA100 and TA1537. Addition of an S9 metabolic activation system decreased or eliminated the mutagenic activity. In contrast, the purified NTP AQ-OX as well as the technical grade samples AQ-FC and AQ-DA were not mutagenic in the Ames test. The chemical structure of AQ does not suggest that the parent compound would be DNA reactive. Therefore, a mutagenic contaminant was present in the NTP bioassay sample that is either directly mutagenic or can be activated by bacterial metabolism. Analytical studies showed that the primary contaminant 9-nitroanthracene (9-NA) was present in the NTP AQ-OX at a concentration of 1200 ppm but not in the purified material. The 9-NA and any other contaminants that might have been present in the NTP AQ-OX induced measurable mutagenicity at 9-NA concentrations as low as 0.15 µg/plate in tester strain TA98, indicating potent mutagenic activity (Butterworth et al., 2001).

Therefore, many of the reported genotoxicity and carcinogenicity results in the literature for AQ and AQ derivative compounds must be viewed with caution.

 

Results:

- Genetic toxicity in vitro: Bacterial reverse mutation assay:

1.- Key study: Experimental results. Test method equivalent to OECD guideline 471. GLP study.

 

The substance AQ-FC did not show any mutagenic activity either with or without metabolic activation in the Ames tester strains (S. typhimurium TA1535, TA1537, TA98 and TA100 or in E. coli WP2uvrA).

 

2.- Key study: Experimental results. Test method equivalent to OECD guideline 471.

 

The substance anthraquinone (100% pure) did not show any mutagenic activity either with or without metabolic activation in theAmestester strains used (S. typhimurium TA98, TA100 and TA102).

 

3.- Key study: Experimental results. Test method equivalent to OECD guideline 471.

 

The substance anthraquinone (Friedel-Crafts process) did not show any mutagenic activity either with or without metabolic activation in theAmestester strains used (TA98 and TA100).

 

- Genetic toxicity in vitro: Chromosomal aberrations in mammalian cells:

Key study: Read-across from experimental data on the analogue CAS No. 518-82-1 (emodin). Test method equivalent to OECD guideline 473.

 

Chromosome aberrations were induced in cultured CHO cells in the absence of S9 activation and in the presence of S9; the response observed without S9 was stronger than with S9.

 

- Genetic toxicity in vitro: Mammalian cell micronucleus test:

Key study: Read-across from experimental data on the analogue CAS No. 518-82-1 (emodin). Test method equivalent to OECD guideline 487.

 

The test substance did not reveal any micronuclei inducing activity in either human lymphocytes or in Hep-G2.

 

- Genetic toxicity in vitro: Gene mutation in mammalian cells:

1.- Key study: Read-across from experimental data on the analogue CAS No. 518-82-1 (emodin). Test method equivalent to OECD guideline 476.

 

The substance induced a moderate increase in mutant fraction (it was only tested without metabolic activation).

 

2.- Key study: Read-across from experimental data on the analogue CAS No. 518-82-1 (emodin). Test method equivalent to OECD guideline 476.

 

Treatment with the test substance did not result in an increase in the number of colonies resistant to 6 -TG. Exposure in suspension in the presence of liver homogenate was also negative.

 

3.- Key study: Read-across from experimental data on the analogue CAS No. 518-82-1 (emodin). Test method equivalent to OECD guideline 476.

 

In the V79-HGPRT mutation assay, the test substance emodin was highly mutagenic without metabolic activation. However, the increase in the induction of mutation was observed at highly toxic concentrations. Furthermore, a low plating efficiency was observed in the negative and positive controls.

 

- Genetic toxicity in vivo: Mammalian erythrocyte Micronucleus test:

1.- Key study: Read-across from experimental data on the analogue CAS No. 518-82-1 (emodin). Test according to OECD guideline 474.

 

There was no statistically significant enhancement in the frequency of micronucleated PCEs in comparison to the negative controls at both preparation intervals.

 

2.- Key study: Read-across from experimental data on the analogue CAS No. 518-82-1 (emodin). Test method equivalent to OECD guideline 474.

 

In peripheral blood samples from mice in the 14-week feed study, an increase in the frequency of micronucleated NCEs was seen in females, but not in males. The small increase in NCEs observed in the female mice was statistically significant (P=0.001), but no individual exposed group value differed significantly from the control value; the result in female mice was concluded to be weakly positive.

 

- Genetic toxicity in vivo: Mammalian bone marrow chromosome aberration test:

1.- Key study: Read-across from experimental data on the analogue CAS No. 518-82-1 (emodin). Test method equivalent to OECD guideline 475.

 

No increases in the frequencies of micronucleated erythrocytes were observed in any of the treatment groups (only males were used).

 

2.- Key study: Read-across from experimental data on the analogue CAS No. 518-82-1 (emodin). Test method equivalent to OECD guideline 475.

 

No increases in the frequencies of micronucleated erythrocytes were observed in any of the treatment groups (males and females were used).

 

 

Justification for selection of genetic toxicity endpoint
No study was selected, since the results obtained in the in vitro studies (Ames test and Mammalian cell gene mutation assay) and the results obtained in the in vivo studies (Chromosome aberrations tests) were negative.

Short description of key information:
- Genetic toxicity in vivo: Mammalian erythrocyte Micronucleus test:
1.- Key study: Read-across from experimental data on the analogue CAS No. 518-82-1 (emodin). Test according to OECD guideline 474.

There was no statistically significant enhancement in the frequency of micronucleated PCEs in comparison to the negative controls at both preparation intervals.

2.- Key study: Read-across from experimental data on the analogue CAS No. 518-82-1 (emodin). Test method equivalent to OECD guideline 474.

In peripheral blood samples from mice in the 14-week feed study, an increase in the frequency of micronucleated NCEs was seen in females, but not in males. The small increase in NCEs observed in the female mice was statistically significant (P=0.001), but no individual exposed group value differed significantly from the control value; the result in female mice was concluded to be weakly positive.

- Genetic toxicity in vivo: Mammalian bone marrow chromosome aberration test:
1.- Key study: Read-across from experimental data on the analogue CAS No. 518-82-1 (emodin). Test method equivalent to OECD guideline 475.

No increases in the frequencies of micronucleated erythrocytes were observed in any of the treatment groups (only males were used).

2.- Key study: Read-across from experimental data on the analogue CAS No. 518-82-1 (emodin). Test method equivalent to OECD guideline 475.

No increases in the frequencies of micronucleated erythrocytes were observed in any of the treatment groups (males and females were used).

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

The substance to be supported by this registration dossier is anthraquinone obtained by Friedel-Crafts technology (AQ-FC). Based on the negative results obtained from the in vitro studies and the in vivo genetic toxicity studies conducted with the analogue emodin, the substance anthraquinone is not classified for mutagenicity.