An Evidence‐Based Biomarker Framework for n‐Hexane‐Induced Neuropathy: Urinary 2,5‐Hexanedione, Pyrrole Adducts, and Neurofilament Light Chain

Neuropathy induced by n-hexane is a classical toxic axonopathy caused primarily by metabolic activation of n-hexane to 2,5-hexanedione (2,5-HD). Although urinary 2,5-HD is widely used for biological monitoring, its interpretation remains complicated by analytical heterogeneity, particularly the distinction between free 2,5-HD and acid-hydrolyzed total 2,5-HD. This review integrates mechanistic toxicology and analytical biomonitoring evidence to clarify how different biomarkers reflect different stages of n-hexane neurotoxicity. 2,5-HD reacts with lysine residues to form pyrrole adducts, promotes neurofilament cross-linking, disrupts axonal transport, and contributes to distal axonal degeneration. Urinary 2,5-HD is an established occupational exposure biomarker, although free and acid-hydrolyzed total 2,5-HD are not interchangeable. Pyrrole adducts are candidate toxicodynamic biomarkers supported predominantly by animal studies and limited worker evidence. Neurofilament light chain (Nf-L) is an exploratory, nonspecific axonal-injury biomarker with 2,5-HD evidence confined to animal models and no human occupational validation for n-hexane. Accordingly, we present the three analytes as a research framework with unequal validation maturity rather than as co-equal, clinically actionable layers. Future longitudinal occupational studies should evaluate them together with air monitoring, electrophysiology, and clinical outcomes to determine whether the candidate pyrrole-adduct and exploratory Nf-L layers add temporal or practical value beyond established exposure monitoring.