A new method for the analysis of styrene mercapturic acids by liquid chromatography/electrospray tandem mass spectrometry.
A new method based on liquid chromatography/tandem mass spectrometry has been developed for the direct determination of specific urinary mercapturic acids arising from the conjugation of (R)-and (S)-enantiomers of styrene 7,8-oxide with glutathione (GSH), i.e. (R,R)- and (S,R)-N-acetyl-S-(1-phenyl-2-hydroxyethyl)cysteine (R,R-M1 and S,R-M1) and (R,R)- and (S,R)-N-acetyl-S-(2-phenyl-2-hydroxyethyl)-cysteine (R,R-M2 and S,R-M2). The four diastereoisomers were separated on a C18-DB (7.5 cm, 3 microm) column using variable proportions of 20 mM aqueous ammonium formate buffer and methanol at a flow-rate of 0.5 mL/min. The analytes were ionized by electrospray, in negative-ion mode. Operating in selected-reaction monitoring mode, linearity of the MS response versus analyte concentration was established over 4 orders of magnitude, the detection limits being 0.7-1.0 microg/L for all the mercapturates. Precision of the method determined at 50 microg/L (n = 12), expressed as relative standard deviation, was respectively 3.1, 4.8 and 6.9% within the run, intra-day and inter-day. The corresponding figures at 1.0 mg/L (n = 12) were respectively 2.0, 3.6 and 5.5%. The method was applied to the quantitative analysis of conjugated metabolites in urine samples from workers occupationally exposed to styrene. The diastereoisomers R,R-M1 and S,R-M2 accounted respectively for 50 and 40% of total mercapturates, whereas the proportion of R,R-M2 was 7% and only minor amounts of S,R-M1 were detectable. Styrene mercapturates represented a minor fraction of total styrene metabolites, less than 1% on average. The ratio mercapturates/main metabolites (mandelic + phenylglyoxylic acid) showed a bimodal distribution, the medians of the two subgroups being 0.2 and 1%, respectively. Such subgroups are probably characterized by the genetic polymorphisms of the drug-metabolizing enzymes to be identified.
Manini
P
P
Dipartimento di Clinica Medica, Nefrologia e Scienze della Prevenzione, Laboratorio di Tossicologia Industriale, Università degli Studi di Parma, Italy. medlav@unipr.it
Andreoli
R
R
Bergamaschi
E
E
De Palma
G
G
Mutti
A
A
Niessen
W M
WM
D000111:Acetylcysteine / Q000031:analogs & derivatives* / Q000378:metabolism / Q000652:urine*; D002853:Chromatography, Liquid / Q000379:methods*; D004785:Environmental Pollutants / Q000378:metabolism / Q000652:urine; D004852:Epoxy Compounds / Q000378:metabolism; D005978:Glutathione / Q000378:metabolism; D006038:Glyoxylates / Q000378:metabolism / Q000652:urine; D006801:Humans; D008333:Mandelic Acids / Q000378:metabolism / Q000652:urine; D015394:Molecular Structure; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D013237:Stereoisomerism; D020058:Styrene / Q000378:metabolism* / Q000652:urine*
D004785:Environmental Pollutants; D004852:Epoxy Compounds; D006038:Glyoxylates; D008333:Mandelic Acids; C012482:phenylglyoxylic acid; D020058:Styrene; C013690:styrene oxide; D005978:Glutathione; C037938:mandelic acid; D000111:Acetylcysteine