N-Nitrosodimethylamine investigations in Muta™Mouse define point-of-departure values and demonstrate less-than-additive somatic mutant frequency accumulations.

The N-nitrosamine, N-nitrosodimethylamine (NDMA), is an environmental mutagen and rodent carcinogen. Small levels of NDMA have been identified as an impurity in some commonly used drugs, resulting in several product recalls. In this study, NDMA was evaluated in an OECD TG-488 compliant Muta™Mouse gene mutation assay (28-day oral dosing across seven daily doses of 0.02-4 mg/kg/day) using an integrated design that assessed mutation at the transgenic lacZ locus in various tissues and at the endogenous Pig-a gene-locus, along with micronucleus frequencies in peripheral blood. Liver pathology was determined together with NDMA exposure in blood and liver. The additivity of mutation induction was assessed by including two acute single-dose treatment groups (i.e. 5 and 10 mg/kg dose on Day 1), which represented the same total dose as two of the repeat dose treatment groups. NDMA did not induce statistically significant increases in mean lacZ mutant frequency (MF) in bone marrow, spleen, bladder, or stomach, nor in peripheral blood (Pig-a mutation or micronucleus induction) when tested up to 4 mg/kg/day. There were dose-dependent increases in mean lacZ MF in the liver, lung, and kidney following 28-day repeat dosing or in the liver and kidney after a single dose (10 mg/kg). No observed genotoxic effect levels (NOGEL) were determined for the positive repeat dose-response relationships. Mutagenicity did not exhibit simple additivity in the liver since there was a reduction in MF following NDMA repeat dosing compared with acute dosing for the same total dose. Benchmark dose modelling was used to estimate point of departure doses for NDMA mutagenicity in Muta™Mouse and rank order target organ tissue sensitivity (liver > kidney or lung). The BMD50 value for liver was 0.32 mg/kg/day following repeat dosing (confidence interval 0.21-0.46 mg/kg/day). In addition, liver toxicity was observed at doses of ≥ 1.1 mg/kg/day NDMA and correlated with systemic and target organ exposure. The integration of these results and their implications for risk assessment are discussed.

Mono-n-hexyl phthalate: exposure estimation and assessment of health risks based on levels found in human urine samples.

Mono-n-hexyl phthalate (MnHexP) is a primary metabolite of di-n-hexyl phthalate (DnHexP) and other mixed side-chain phthalates that was recently detected in urine samples from adults and children in Germany. DnHexP is classified as toxic for reproduction category 1B in Annex VI of Regulation (EC) 1272/2008 and listed in Annex XIV of the European chemical legislation REACH; thereby, its use requires an authorisation. Health-based guidance values for DnHexP are lacking and a full-scale risk assessment has not been carried out under REACH. The detection of MnHexP in urine samples raises questions about the sources of exposure and concerns of consumer safety. Here, we propose the calculation of a provisional oral tolerable daily intake value (TDI) of 63 µg/kg body weight/day for DnHexP and compare it to intake levels corresponding to levels of MnHexP found in urine. The resulting mean intake levels correspond to less than 0.2% of the TDI, and maximum levels to less than 5%. The TDI was derived by means of an approximate probabilistic analysis using the credible interval from benchmark dose modelling of published ex vivo data on reduced foetal testosterone production in rats. Thus, for the dose associated to a 20% reduction in testosterone production, a lower and upper credible interval of 14.9 and 30.0 mg/kg bw/day, respectively, was used. This is considered a conservative approach, since apical developmental endpoints (e.g. changed anogenital distance) were only observed at higher doses. In addition, we modelled various scenarios of the exposure to the precursor substance DnHexP from different consumer products, taking measured contamination levels into account, and estimated systemic exposure doses. Of the modelled scenarios including the application of sunscreen (as a lotion or pump spray), the use of lip balm, and the wearing of plastic sandals, and considering conservative assumptions, the use of DnHexP-contaminated sunscreen was highlighted as a major contributing factor. A hypothetical calculation using conservative assumptions for the latter resulted in a margin of safety in relation to the lower credible interval of 3267 and 1007 for adults and young children, respectively. Most importantly, it was found that only a fraction of the TDI is reached in all studied exposure scenarios. Thus, with regard to the reported DnHexP exposure, a health risk can be considered very unlikely.