Solid-Phase Extraction Based on Captiva EMR-Lipid for Determination of 19 Aromatic Amine Antioxidants and Two p-Phenylenediamine Quinones in Human Plasma

A robust analytical method based on Captiva EMR-Lipid solid-phase extraction and HPLC-MS/MS was developed and validated for the simultaneous determination of 19 aromatic amine antioxidants (AAs) and two <i>p</i>-phenylenediamine-derived quinones (PPD-Qs) in human plasma. The optimized protocol effectively removed phospholipid interferences from complex blood matrix, significantly mitigating ion suppression and improving the recovery of hydrophobic AAs compared to conventional liquid-liquid extraction. Method validation demonstrated good accuracy (spike recoveries: 73.0-96.8%), precision (RSD < 11%), and sensitivity with method detection limits ranging from 0.81 to 21 pg/mL. The method was successfully applied to plasma samples from 20 adults, in which 11 AAs were detected at total concentrations of 240-710 pg/mL. Diphenylamine derivatives, particularly bis(4-tert-butylphenyl)amine (DBDPA) and diphenylamine (DPA), were identified as the predominant compounds, contributing over 69% of the total AA burden. No PPDs or PPD-Qs were detected, which may be attributed to their biotransformation and urinary excretion, as well as the limited sample size. This study provides a comprehensive biomonitoring tool for assessing combined human exposure to multiple AAs and establishes a foundation for further investigation into their health implications.