Determination of urinary S-phenylmercapturic acid, a specific metabolite of benzene, by liquid chromatography/single quadrupole mass spectrometry

Abstract

A high-performance liquid chromatography/single quadrupole mass spectrometry (LC/MS) method is described for the determination of urinary S-phenylmercapturic acid (S-PMA), a specific metabolite of benzene. Urine samples were spiked with [13C6]S-PMA (used as the internal standard) and acidified; then they were purified by solid-phase extraction (SPE) on C18 cartridges. Analyses were conducted on a reversed-phase column by gradient runs with 1% aqueous acetic acid/methanol mixtures at different proportions as the mobile phase. The detector was used in electrospray negative ion mode (ESI-), the ions m/z 238 for S-PMA and 244 for [13C 6]S-PMA being recorded simultaneously. The detection limit (for a signal-to-noise ratio = 3) was 0.2 μg/L, thus allowing for the measurement of background excretion of S-PMA in the general population. The use of the internal standard allowed us to obtain good precision (CV% values <3%) and a linear calibration curve within the range of interest for monitoring occupational exposure to benzene (up to 500 μg/L). The method was applied to assay the metabolite concentration in a group of 299 workers (68 smokers and 231 non-smokers) occupationally exposed to relatively low levels of benzene (environmental concentration = 0.4-220 μg/m3, mean 11.4 μg/m3) and 236 non-exposed subjects (134 smokers and 102 non-smokers). The results clearly showed that smoking must be taken into account for the correct interpretation of the results of S-PMA measurements for the assessment of work-related benzene exposure. When only non-smokers were selected, the mean excretion of S-PMA was significantly higher in workers exposed to benzene (1.2 ± 0.9 μg/g creatinine) than in the control group (0.7 ± 0.6 μg/g creatinine) (p < 0.001), thus confirming the role of S-PMA as a biomarker of benzene on a group basis, even for relatively low exposure degrees. Copyright © 2005 John Wiley & Sons, Ltd.