Registration Dossier

Administrative data

Key value for chemical safety assessment

Additional information

The test substance used in investigations supporting this endpoint was TBHP (aqueous 70% solution): results have been expressed in terms of dry TBHP.

The genotoxic potential of TBHP has been investigated in a number of GLP-compliant guideline studies peformed using bacterial and mammalian cells in vitro. It was clearly mutagenic toward Salmonella typhimurium tester strains (EG and G Mason Research Institute, 1981; Haworth et al., 1983) and L5178Y TK cells (EG and G Mason Research Institute, 1981) and induced structural chromosomal aberrations in CHO cells (TNO Study Director, 1982); similar results were obtained both in the absence and presence of S9 fraction. The results indicate that TBHP is genotoxic in bacterial and eukaryotic systems in vitro. The clastogenic potential of TBHP has also been investigated in vivo in a GLP-compliant guideline bone marrow micronucleus test (TNO Study Director, 1995). The incidence of micronucleated erythrocytes was unaffected, however the incidence of polychromatic erythrocytes was decreased only to a limited extent suggesting that the test substance had not reached the bone marrow. This observation is consistent with toxicokinetic data demonstrating the rapid conversion of TBHP to tert-butanol in vivo. A Comet Assay on lung tissue from rats exposed to severely irritating concentrations of TBHP was negative (BASF 2010) but was unable to make a determination in the nasal tissue. In a subsequent comet assay, advances in techniques have allowed for an assessment of the nasal tissues and were determined to be negative for genotoxicity (WIL 2016). In contrast, TBHP tested positive in a male mouse dominant lethal study following i.p. injection (Kumar and Muralidhara, 1999). The method used was broadly similar to EU method B.22 however the test was not conducted to GLP. These findings, together with supporting toxicokinetic information for TBHP, were considered by the EU Technical Committee for New and Existing Substances and the EU Classification and Labelling Working Group which concluded that direct exposure of the testis via the inguinal canal could not be ruled out i.e the results were indicative of a local mutagenic effect. In contrast, TBHP is unlikely to reach germ cells after oral, inhalation and dermal exposure and is therefore unlikely to result in inheritable genetic damage. It was concluded that the vivo mutagenicity of TBHP is confined to somatic cells in tissues of first contact with no potential for systemic exposure, however the results of the second Comet Assay indicate that site of contact genotoxicity is not observed following a relevant route of exposure (inhalation).


Short description of key information:
Several key and supporting studies have been identified that have assessed the mutagenic/genotoxic potential of TBHP in vitro and in vivo. Study design was generally consistent with the appropriate guideline, however not all investigations were GLP compliant. All studies used TBHP (70% solution, as manufactured and supplied) but results are expressed in terms of dryTBHP. TBHP is clearly mutagenic and genotoxic in bacterial and mammalian cell systems in vitro in the absence and presence of hepatic S9 fraction. It failed to show any clastogenic potential in an in vivo mouse micronucleus test after i.v. injection which is consistent with toxicokinetic data demonstrating rapid conversion to tert-butanol in vivo. However TBHP tested positive in a male mouse dominant lethal study following i.p. injection. Direct exposure of the testis via the inguinal canal could not be ruled out in this study leading the EU to conclude that the results were indicative of a local mutagenic effect with in vivo mutagenicity confined to somatic cells in tissues of first contact. An attempt to characterise the potential for local nasal epithelial cell genotoxicity using a Comet Assay method was unsuccessful due to methodological obstacles. However, TBHP did not cause DNA damage measured by Comet Assay in the rat lung following 5 days exposure to maximal tolerated concentrations.

Endpoint Conclusion: Adverse effect observed (positive)

Justification for classification or non-classification

TBHP is mutagenic and genotoxic in bacterial and mammalian cell systems in vitro; however it failed to show any clastogenic potential in an in vivo mouse micronucleus test after i.v. injection; this is consistent with the rapid conversion of TBHP to tert- butanol in the body. However it tested positive in a male mouse dominant lethal study following i.p. injection. When reviewing these findings the EU Classification and Labelling Working Group concluded that direct exposure of the testis via the inguinal canal could not be ruled out in this study, and that the results were indicative of a local mutagenic effect. The conclusion that it is a local effect is supported by toxicokinetic data which indicate that it is unlikely to reach germ cells after oral, inhalation and dermal exposure. In another in vivo study, a Comet assay, a relevant route of exposure was employed and the potential for localised genotoxicity on nasal epithelial tissue has been determined to be negative (WIL 2016). The data from the dominant lethal study indicates that the mutagenic potential of TBHP is confined to somatic cells in tissues of first contact. The weight of evidence for the in vivo data indicates that TBHP is unlikely to be a germ cell mutagen, however if the complete data set (in vitro and in vivo) is used in a weight of evidence assessment there is possibility for a weak mutagenic effects.

The available data support the following classification:

CLP regulation : Muta 2; H341 (Suspected of causing genetic defects)