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Additional information

Green liquor sludge has not been tested for mutagenicity or genotoxicity earlier. For this report in vitro gene mutation study in bacteria (Ames test), in vitro cytogenicity study in mammalian cells (chromosome aberration test in human lymphocytes) and in vitro gene mutation study in mammalian cells (cell mutation assay at the TK locus in mouse lymphoma L5178Y cells) were performed to elucidate genotoxicity of the GLS.

According to the results obtained in the in vitro gene mutation study in bacteria (Ames test) GLS is non-mutagenic in various S. typhimurium strains (TA97a, TA98, TA100, TA102 and TA1535) with and without an external metabolising system up to precipitating concentration (5000 µg/plate). The study was conducted in accordance with the OECD-guideline 471 and the Council Regulation (EC) No 440/2008, Method B.13/14. In this study, the test substance was not soluble in water or DMSO, but a homogeneous suspension was prepared with DMSO.

In in vitro cytogenicity study in mammalian cells GLS did not induce structural chromosomal aberrations in human lymphocytes under the experimental conditions reported, and was considered to be non-clastogenic when tested up to precipitating concentrations in the absence and presence of metabolic activation. The study was conducted according to Regulation (EC) No 440/2008 method B.10. and to OECD Guideline 473.

In in vitro gene mutation study in mammalian cells Green liquor sludge did not induce mutations in the mouse lymphoma thymidine kinase locus assay using the cell line L5178Y in the absence and presence of metabolic activation under the experimental conditions reported. Therefore, GLS was considered to be non-mutagenic up to precipitating and cytotoxic concentrations in this mouse lymphoma assay. The study was conducted according to the OECD Guideline 476 and Commission Regulation (EC) No. 440/2008 method B.17.

In conclusion, GLS was non-mutagenic and non-clastogenic based on the negative results in in vitro gene mutation study in bacteria (REACH Annex VII, section 8.4.1) and in vitro cytogenicity study in mammalian cells (REACH Annex VIII, section 8.4.2). Negative results in Annex VII, section 8.4.1. and Annex VIII, section 8.4.2. were confirmed by a third negative result in in vitro mutagenicity study, a gene mutation study in mammalian cells (REACH Annex VIII, section 8.4.3). The data was done according to the EU/OECD guidelines and assessed to be of good quality and relevant. Therefore, the genotoxicity test results can be taken as indicators for non-mutagenic and non-genotoxic effects of (tested) GLS. According to Reach Annex VIII (8.4.3) in column 2 of specific rules for adaptation standard information requirements, further in vivo mutagenicity studies need not to be considered in this case, since positive result were not detected for GLS in any of the genotoxicity studies in Annex VII or VIII. Further, GLS can be concluded to be not genotoxic and not mutagenic (European Chemicals Agency, Guidance on information requirements and chemical safety assessment, Chapter R.7a: Endpoint specific guidance, Table R.7.7-5, row 4). In addition, major Green liquor sludge components are common inorganic substances or ions and naturally present in the human body (See Chapter 5.1, Toxicokinetics) and therefore they are not likely to be mutagenic or genotoxic.

 

However, due to the lack of in vivo and human data available, a certain level of uncertainty remains when extrapolating these testing data to the effect in humans. The composition of GLS can also vary, and GLS includes substances that may have carcinogenic effects, such as nickel, cadmium, carbon black and possibly sulphides and polycyclic aromatic hydrocarbons (PAHs) attached on soot particle surfaces. However, in the analysis results for this report nickel and cadmium were detected intrace amounts (< 0.1 %) and no sulphides were detected (See Chapter 1.2, Composition of the substance). Carbon black is listed possibly carcinogenic to humans by IARC, but not listed as a carcinogen by European Union. PAH amounts in GLS were not determined for this report, but their concentration is likely to be very low and they are not likely to be released from carbon particles in biological fluids (See chapters Composition of the substance and Toxicokinetics). In addition, organic solvent (DMSO) was used in the in vitro gene mutation study in bacteria, that should extract PAHs from carbon particles. However, no mutagenicity was detected in the study.


Short description of key information:
Green liquor sludge is not mutagenic based on three negative in vitro test results (1. Ames test) (2. in vitrocytogenicity study in mammalian cells) (3. in vitro gene mutation study in mammalian cells ). The tested GLS material was not mutagenic in the tests. The tests followed the information requirements of the REACH annexes VII-VIII, and standardised OECD/EU testing guidelines.

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

Mutagenicity

According to the information requirements in REACH annexes VII-VIII, and based on three negative in vitro test results, it can be concluded that the tested Green liquor sludge material was not mutagenic and classification is not needed.

Carcinogenicity

No indication of carcinogenicity was detected in the Green liquor sludge combined repeated dose toxicity study with the reproduction / developmental toxicity screening test. Based on the existing mutagenicity studies it cannot be classified as a mutagen, and there is no indication of carsinogenicity in the repeated dose study. Therefore there are no basis or need for any carcinogenicity classification and no studies are proposed by the registrant. Some uncertainties of carcinogenicity potensy of Green liquor sludge still exists (related to trace impurities), but GLS has no widespread use and no direct exposure to consumers or general population and long-term worker exposure can be kept under control by risk management measures.

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