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Administrative data

Key value for chemical safety assessment

Additional information

The substance was not mutagenic in a Bacterial Reverse Mutation (Ames) Test according to OECD TG 471 on Salmonella typhimurium TA 1535, TA 100, TA 1537, TA 98, and TA 102, tested without and with metabolic activation. As vehicle ethylene glycol dimethyl ether was used.

For assessing further genotoxicity endpoints a read across to a substance with a very similar chemical composition (EC No. 900 -066 -9; comparable allophanate-type HDI oligomerisation product) is applied.The read across is based on physicochemical and toxicological similarity of the two substances. Especially a comparative pulmonary irritant potency study according to TRGS 430 (Technical Rule for Hazardous Substances 430; published by the German Federal Ministry of Labour and Social Affairs, 2009), as the key study for justification of read-across, confirmed for both substances the same toxicological mode of action and nearly identical potency. For further justification of the grouping and read-across according to regulation (EC) No 1907/2006, Annex XI, 1.5 see document attached to chapter "Assessment Reports".

The read-across substance (EC No. 900 -066 -9) revealed no mutagenic potential in a mammalian cell gene mutation assay according to OECD 476 using V79 cells (HPRT). This assay was conducted in two independent experiments: 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 time of 4 hours with and 24 hours without metabolic activation. As vehicle ethylene glycol dimethyl ether was used.

Two in vitro micronucleus assays were performed with the read-across substance (EC No. 900 -066 -9). In one a positive and in the other a negative test result was obtained. In principle, both assays solely vary by the use of the solvent applied, ethylene glycol dimethylether (EGDE) versus dimethyl suloxide (DMSO). Indeed, the suitability of solvents used in genotoxicity tests with diisocyanates has already been a subject of scientific examination, especially in the case of aromatic diisocyanates (Gahlmann et. al., Zbl Arbeitsmed 43, 1993, 34-38; Herbold et al., Mutation Research 412, 1998, 167-175; Seel et. al., Mutation Research 438, 1999, 109-123). For aromatic diisocyanates like MDI (CAS number 26447-40-5), the products of aqueous hydrolysis, i.e. the aromatic amines, were suspected of causing positive results in Ames tests. It could be shown that if DMSO is used as solvent for MDI a rapid degradation of the isocyanate function occurs. One of the degradation products was found to be the corresponding aromatic amine 4,4’-methylenedianiline (MDA), which is a known genotoxin. If EGDE was used as a representative of a less polar solvent, analytical data revealed that MDI was quite stable and no formation of MDA could be detected.
EU risk assessment of MDI (final report, 2005) states that “It is evident from the results of studies .... that the solvent used for solubilisation of MDI in genotoxicity assays is crucial with respect to a valid and representative evaluation of MDI in such assays. There is adequate evidence to demonstrate that use of EDGE as solvent is appropriate whereas DMSO can give rise to false positives due to solvent-catalysed conversion to MDA.”

In fact, DMSO is not chemically inert. In synthetic organic chemistry it is known to react with certain polar groups to "activated dimethyl sulfoxide", which can further be used as reagent in organic synthesis (Tidwell, Synthesis, 1990, 857-870; Mancuso & Swern, Synthesis, 1981, 165-185). Moreover reactions of isocyanates and DMSO or at least reactions with catalytical activity of DMSO in chemical conversions of isocyanates are reported (Arbuzov et. al., Bulletin of the Academy of Sciences of the USSR, Division of Chemical Science (Engl. Translation) 24, 1975, 1113-1114; Sorenson, J. Org. Chem. 24, 1959, 978-980). Therefore, it is assumed, that single positive results of isocyanates in genotoxicity assays conducted with DMSO as solvent does not reflect a genotoxic property of the isocyanate, but reflects positive genotoxicity of a reaction product that is formed in this mixture of isocyanate, DMSO and possibly substances from liver homogenate also.

The in vitro micronucleus assay performed with EGDE (Bayer, 2012) can therefore be considered to be more reliable to assess the in vitro cytogenicity of the allophanate-type HDI oligomers in mammalian cells compared to the assay performed with DMSO (BASF, 2006), which is therefore marked as "disregarded study".

The micronucleus assay performed with EGDE was conducted in accordance to OECD 487 using Chinese Hamster V79 cells. Two independent assays were performed in this study, the first conducted with a treatment time of 4 hours, with and without a metabolizing system (S9 mix), the second assay with a treatment time of 24 hours (continuous treatment) without S9 mix and with a treatment time of 4 hours with S9 mix.

No biologically relevant increase in the frequencies of micronucleus containing V79 cells were found either in the absence (4 hours or 24 hours treatment) or in the presence (4 hours treatment) of S9 mix when tested up to cytotoxic or precipitating concentrations, thus no genotoxicity potential is concluded based on this in vitro MNT.

The negative genotoxicity is confirmed in an in vivo Mammalian Erythrocyte Micronucleus Test (OECD 474) in NMRI mice. In this test the substance, dissolved in corn oil, was administered twice orally, with a 24 -hour interval between administrations, to groups of 5 male animals at dose levels of 500 mg/kg, 1000 mg/kg and 2000 mg/kg body weight. Bone marrow was prepared 24 hours after the second administration.

The two oral substance administrations did not lead to any increase in the number of polychromatic erythrocytes containing either small or large micronuclei. The rate of micronuclei was always close to the same range as that of the concurrent negative control in all dose groups and within the range of the historical control data. A slight inhibition of erythropoiesis, determined from the ratio of polychromatic to normochromatic erythrocytes, was reported. Therefore, no indications for genotoxicity in vivo can be concluded from this study.


Justification for selection of genetic toxicity endpoint
No study was selected, since all available genotoxicity studies were assessed.

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

No classification required for genotoxicity according to Regulation (EC) No. 1272/2008, Annex I.