Inter-species variability in 4,4′-methylenedianiline metabolism: insights from human and rat precision-cut liver slices

Occupational exposure to 4,4’-methylenedianiline (MDA), a precursor in polyurethane foam production, has been linked to liver disease and cancer. Most evidence, however, comes from underpowered epidemiological studies, while empirical data are mainly from animal models, which point toward liver metabolism generating reactive metabolites. We thus used precision-cut liver slices (PCLS) from human and rat donors which we exposed to MDA at 10 and 100 μM for 24 hours, and supernatants were analyzed by liquid chromatography-‘SWATH’ mass spectrometry. The major metabolite detected in hPCLS and rPCLS was MDA’s known N-acetyl derivative, comprising approximately 80–90% of metabolites at 10 μM and 50-60% at 100 μM. Other metabolites were phase II conjugates, including putative N-formylation and urea-glutamine and urea-ethanolamine adducts. The latter accounted for ∼10% of metabolites at 10 μM, increasing to 50% at 100 μM in rPCLS but only reaching ∼25% in hPCLS. Cell viability remained at 100% across all human conditions and in rPCLS at 10 μM, but it dropped below 5% at 100 μM. In conclusion, this study confirmed MDA’s well-known N-acetyl metabolite across both species, while also unveiling rather unusual, previously unreported conjugates. Additionally, distinct interspecies differences were observed, underscoring the need for human-based studies. • Human and rat precision-cut liver slices showed distinct MDA metabolism patterns • Various unreported (putative) MDA conjugates were discovered • N-acetyl MDA was the dominant metabolite but decreased at higher MDA levels • Rat slices lost viability at 100 μM MDA, while human slices remained unaffected • Interspecies differences highlight need for human-based toxicity assessment