title: Determination of the urinary benzene metabolites S-phenylmercapturic acid and trans,trans-muconic acid by liquid chromatography-tandem mass spectrometry.
abstract: To investigate how various levels of exposure affect the metabolic activation pathways of benzene in humans and to examine the relationship between urinary metabolites and other biological markers, we have developed a sensitive and specific liquid chromatographic-tandem mass spectrometric assay for simultaneous quantitation of urinary S-phenylmercapturic acid (S-PMA) and trans,trans-muconic acid (t,t-MA). The assay involves spiking urine samples with [13C6]S-PMA and [13C6]t,t-MA as internal standards and clean up of samples by solid-phase extraction with subsequent analysis by liquid chromatography coupled with electrospray-tandem mass spectrometry-selected reaction monitoring (LC-ES-MS/MS-SRM) in the negative ionization mode. The efficacy of this assay was evaluated in human urine specimens from smokers and non-smokers as the benzene-exposed and non-exposed groups. The coefficient of variation of runs on different days (n = 8) for S-PMA was 7% for the sample containing 9.4 microg S-PMA/l urine, that for t,t-MA was 10% for samples containing 0.07 mg t,t-MA/l urine. The mean levels of urinary S-PMA and t,t-MA in smokers were 1.9-fold (P = 0.02) and 2.1-fold (P = 0.03) higher than those in non-smokers. The mean urinary concentration (+/-SE) was 9.1 +/- 1.7 microg S-PMA/g creatinine [median 5.8 microg/g, ranging from not detectable (1 out of 28) to 33.4 microg/g] among smokers. In non-smokers' urine the mean concentration was 4.8 +/- 1.1 microg S-PMA/g creatinine (median 3.6 microg/g, ranging from 1.0 to 19.6 microg/g). For t,t-MA in smokers' urine the mean (+/-SE) was 0.15 +/- 0.03 mg/g creatinine (median 0.11 mg/ g, ranging from 0.005 to 0.34 mg/g); the corresponding mean value for t,t-MA concentration in non-smokers' urine was 0.07 +/- 0.02 mg/g creatinine [median 0.03 mg/g, ranging from undetectable (1 out of 18) to 0.48 mg/g]. There was a correlation between S-PMA and t,t-MA after logarithmic transformation (r = 0.41, P = 0.005, n = 46).
authors.affiliation: Naylor Dana Institute, American Health Foundation, Valhalla, NY 10595, USA.
authors.forename: A A
authors.initials: AA
authors.lastname: Melikian
authors.forename: R
authors.initials: R
authors.lastname: O'Connor
authors.forename: A K
authors.initials: AK
authors.lastname: Prahalad
authors.forename: P
authors.initials: P
authors.lastname: Hu
authors.forename: H
authors.initials: H
authors.lastname: Li
authors.forename: M
authors.initials: M
authors.lastname: Kagan
authors.forename: S
authors.initials: S
authors.lastname: Thompson
mesh_terms: D000111:Acetylcysteine / Q000031:analogs & derivatives* / Q000652:urine; D000818:Animals; D001554:Benzene / Q000008:administration & dosage / Q000493:pharmacokinetics*; D015415:Biomarkers; D001711:Biotransformation; D002138:Calibration; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004781:Environmental Exposure; D005069:Evaluation Studies as Topic; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D051381:Rats; D011916:Rats, Inbred F344; D012015:Reference Standards; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D012907:Smoking / Q000652:urine*; D013011:Sorbic Acid / Q000031:analogs & derivatives* / Q000032:analysis; D016482:Urinalysis / Q000295:instrumentation / Q000379:methods*
chemical_list: D015415:Biomarkers; C005939:muconic acid; C053003:S-phenyl-N-acetylcysteine; D001554:Benzene; D000111:Acetylcysteine; D013011:Sorbic Acid