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Simultaneous determination of t,t-muconic, S-phenylmercapturic and S-benzylmercapturic acids in urine by a rapid and sensitive liquid chromatography/electrospray tandem mass spectrometry method.

We describe a rapid and sensitive high-performance liquid chromatography/electrospray tandem mass spectrometry (HPLC/ESI-MS/MS) method for simultaneous determination of the most relevant metabolites of benzene and toluene, t,t-muconic acid (t,t-MA), S-phenylmercapturic acid (S-PMA), and S-benzylmercapturic acid (S-BMA). Urine samples were purified before analysis by solid-phase microextraction (SPE) on SAX cartridges with 50 mg sorbent mass. The developed method fulfils all the standard requirements of precision and accuracy. Calibration curves were linear within the concentration range of the standards (0-80 microg/L(urine) for t,t-MA, and 0-25 microg/L(urine) for S-PMA and S-BMA), and had correlation coefficients > or =0.997. Limits of detection were 6.0 microg/L for t,t-MA, 0.3 microg/L for S-PMA, and 0.4 microg/L for S-BMA. The method was used to determine t,t-MA, S-PMA and S-BMA levels in urine of 31 gasoline-station workers, with personal monitoring data obtained from radial symmetry passive diffusive samplers. In the context of mean work-shift exposures of 75.9 microg/m(3) (range 9.4-220.2) for benzene and 331.9 microg/m(3) (78.2-932.1) for toluene, metabolite concentrations in end-of-shift urine samples ranged from 23.5-275.3 microg/g(creatinine) for t,t-MA, non-detectable to 0.9 microg/g(creatinine) for S-PMA, and 3.8-74.8 microg/g(creatinine) for S-BMA. No significant correlation was found between the environmental concentrations and urinary metabolites (p > 0.05 for all cases); the ratios of benzene metabolites could be influenced by exposure levels and co-exposure to xylenes and toluene. The high throughput of this procedure should facilitate exploration of the metabolic effects of benzene-related co-exposure to toluene and alkylbenzenes in large populations of subjects exposed to gasoline.