Improvement of an ultra-high liquid chromatography-tandem mass spectrometry method for the simultaneous quantification of eleven amino-polycyclic aromatic hydrocarbons in human urine

Abstract

Urinary amino-polycyclic aromatic hydrocarbons (amino-PAHs) are metabolites of nitro-polycyclic aromatic hydrocarbons (nitro-PAHs) and serve as biomarkers for assessing occupational exposure to nitro-PAHs, particularly in populations exposed to diesel exhaust. Current methods for detecting urinary amino-PAHs mainly rely on liquid-liquid extraction, which has limitations in covering a wide range of nitro-PAHs and often struggles to resolve interference from isomeric compounds. This study aimed to develop a reliable UPLC-MS/MS method incorporating solid-phase extraction (SPE) pretreatment for detecting 11 amino-PAHs in urine samples from workers exposed to nitro-PAHs. Briefly, urine samples were enzymatically hydrolyzed, purified using SPE extraction, and analyzed with an optimized UPLC-MS/MS method. Effective separation of the 11 amino-PAHs was achieved using reversed-phase HSS PFP columns with a gradient elution of 0.1 % formic acid in water and acetonitrile. Improved MS/MS conditions were established to achieve sufficient sensitivity using positive ion electrospray ionization (ESI), with limits of quantification (LOQs) ranging from 0.009 to 0.378 μg L⁻¹. The method was optimized for linearity, accuracy, precision, and matrix effects, utilizing two isotopically labeled internal standards (1-Aminonaphthalene-d7 and 1-Aminopyrene-d9). This method enabled precise and accurate quantification of the 11 amino-PAHs, with a coefficient of variation (CV) of less than 11 % and recovery rates between 82.0 % and 106.9 %. The improved method provides a unique analytical approach that can be used for biomonitoring workers exposed to diesel exhaust.