Development of a sensitive method for the quantification of urinary 3-hydroxybenzo[a]pyrene by solid phase extraction, dansyl chloride derivatization and liquid chromatography-tandem mass spectrometry detection

Abstract

Analysis of the urinary metabolite of benzo[a]pyrene (BaP), 3-hydroxybenzo[a]pyrene (3-OHBaP), is useful for the biomonitoring of carcinogenic polycyclic aromatic hydrocarbons (PAHs) exposure. The determination of 3-OHBaP by liquid chromatography tandem mass spectrometry (LC-MS/MS) suffers from the poor sensitivity of detection in commonly used electrospray ionization (ESI) sources. In this study, a sensitive and selective LC-MS/MS method was developed for the determination of urinary 3-OHBaP. Following enzymatic hydrolysis of the glucuronide and sulfate conjugates, the metabolite was enriched and cleaned up by solid-phase extraction and then derivatized with dansyl chloride. The derivative was analyzed by LC-MS/MS with ESI in the positive ion mode. The derivatization of 3-OHBaP by the introduction of a dansyl group into the molecule greatly enhanced the detection sensitivity by improving both the efficiency of ESI in the positive ion mode and collision-induced dissociation in the collision cell. Good linearity was obtained in the range of 0.25–40.0 pg mL⁻¹ with a correlation coefficient (r²) of 0.9924. The limit of detection (LOD) and lower limit of quantification (LLOQ) were 0.1 pg mL⁻¹ and 0.25 pg mL⁻¹, respectively. Accuracy ranged from 87.7% to 107.5%. Intra- and inter-day relative standard deviations varied from 4.6% to 8.4% and 7.2% to 10.6%, respectively. Finally, this developed method was successfully applied for the analysis of urine samples from smokers and non-smokers to measure human exposure to PAHs.