Accurate quantification of mercapturic acids of styrene (PHEMAs) in human urine with direct sample injection using automated column-switching high-performance liquid chromatography coupled with tandem mass spectrometry.
Styrene is one of the most important industrial chemicals, with an enormously high production volume worldwide. The urinary mercapturic acids of its metabolite styrene-7,8-oxide, namely N-acetyl-S-(2-hydroxy-1-phenylethyl)-L-cysteine (PHEMA 1) and N-acetyl-S-(2-hydroxy-2-phenylethyl)-L-cysteine (PHEMA 2), are specific biomarkers for the determination of individual internal exposure to this highly reactive intermediate of styrene. We have developed and validated a fast, specific and very sensitive method for the accurate determination of the sum of phenylhydroxyethyl mercapturic acids (PHEMAs) in human urine with an automated multidimensional liquid chromatography-tandem mass spectrometry method using (13)C(6)-labelled PHEMAs as internal standards. Analytes were stripped from the urinary matrix by online extraction on a restricted access material, transferred to the analytical column and subsequently determined by tandem mass spectrometry. The limit of quantification (LOQ) for the sum of PHEMAs was 0.3 microg/L urine and allowed us to quantify the background exposure of the (smoking) general population. Precision within series and between series ranged from 1.5 to 6.8% at three concentrations ranging from 3 to 30 microg/L urine; the mean accuracy was between 104 and 110%. We applied the method to spot urine samples from 40 subjects of the general population with no known occupational exposure to styrene. The median levels (range) for the sum of PHEMAs in urine of non-smokers (n = 22) were less than 0.3 microg/L (less than 0.3 to 1.1 microg/L), whereas in urine of smokers (n = 18), the median levels were 0.46 microg/L (less than 0.3 to 2.8 microg/L). Smokers showed a significantly higher excretion of the sum of PHEMAs (p = 0.02). Owing to its automation and high sensitivity, our method is well suited for application in occupational or environmental studies.
Reska
M
M
Institute for Occupational and Social Medicine, Medical Faculty, RWTH Aachen University, Pauwelsstrasse 30, 52074 Aachen, Germany.
Ochsmann
E
E
Kraus
T
T
Schettgen
T
T
D000111:Acetylcysteine / Q000378:metabolism / Q000652:urine*; D000328:Adult; D002851:Chromatography, High Pressure Liquid / Q000295:instrumentation / Q000379:methods*; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D016273:Occupational Exposure; D012680:Sensitivity and Specificity; D020058:Styrene / Q000378:metabolism*; D053719:Tandem Mass Spectrometry / Q000379:methods*
D020058:Styrene; D000111:Acetylcysteine