title: Accurate quantification of the mercapturic acids of acrylonitrile and its genotoxic metabolite cyanoethylene-epoxide in human urine by isotope-dilution LC-ESI/MS/MS.
abstract: Acrylonitrile is a highly important industrial chemical with a high production volume worldwide, especially in the plastics industry. It is classified as a possible human carcinogen by the International Agency for Research on Cancer (IARC group 2B). During metabolism of acrylonitrile, the genotoxic metabolite cyanoethylene-epoxide is formed. The urinary mercapturic acids of acrylonitrile, namely N-acetyl-S-(2-cyanoethyl)-L-cysteine (CEMA) and cyanoethylene-epoxide, namely N-acetyl-S-(1-cyano-2-hydroxyethyl)-L-cysteine (CHEMA) are specific biomarkers for the determination of individual internal exposure to acrylonitrile and its highly reactive metabolite. We have developed and validated a sensitive method for the accurate determination of CEMA and CHEMA in human urine with a multidimensional LC/MS/MS-method using deuterium-labelled analogues for both analytes as internal standards. Analytes were stripped from urinary matrix by online extraction on a restricted access material, transferred to the analytical column and determined by tandem mass spectrometry. The limit of quantification (LOQ) for CEMA and CHEMA was 1 μg/L urine and allowed to quantify the background exposure of the (smoking) general population. Precision within and between series for CHEMA ranged from 2.6 to 8.0% at four concentrations ranging from 8.3 to 86 μg/L urine, mean accuracy was between 94 and 100%. For CEMA, precision within and between series ranged from 2.4 to 14.5% at four concentrations ranging from 15.1 to 196 μg/L urine, mean accuracy was between 91 and 104%. We applied the method to spot urine samples of 83 subjects of the general population with no known occupational exposure to acrylonitrile. Median levels (range) for CEMA and CHEMA in urine samples of non-smokers (n=47) were 1.9 μg/L (<1-16.4 μg/L) and<1 μg/L (<1-3 μg/L), while in urine samples of smokers (n=36), median levels were 184 μg/L (2-907 μg/L) and 29.3 μg/L (<1-147 μg/L), respectively. Smokers showed a significantly higher excretion of both acrylonitrile metabolites (p<0.001). Due to its automation and high sensitivity, our method is well suited for application in experimental studies on acrylonitrile metabolism or occupational studies.
authors.affiliation: Institute for Occupational and Social Medicine, Medical Faculty, RWTH Aachen University, Pauwelsstrasse 30, D-52074 Aachen, Germany. tschettgen@ukaachen.de
authors.forename: T
authors.initials: T
authors.lastname: Schettgen
authors.forename: J
authors.initials: J
authors.lastname: Bertram
authors.forename: T
authors.initials: T
authors.lastname: Kraus
mesh_terms: D000111:Acetylcysteine / Q000031:analogs & derivatives* / Q000652:urine*; D000181:Acrylonitrile / Q000652:urine*; D000328:Adult; D000368:Aged; D002853:Chromatography, Liquid; D005027:Ethylene Oxide / Q000031:analogs & derivatives* / Q000652:urine; D005260:Female; D006801:Humans; D007201:Indicator Dilution Techniques; D057230:Limit of Detection; D008297:Male; D008875:Middle Aged; D016273:Occupational Exposure; D012907:Smoking; D021241:Spectrometry, Mass, Electrospray Ionization; D053719:Tandem Mass Spectrometry
chemical_list: C033154:2-cyanoethylene oxide; C057647:N-acetyl-S-(1-cyano-2-hydroxyethyl)cysteine; D005027:Ethylene Oxide; D000181:Acrylonitrile; C025645:N-acetyl-S-(2-cyanoethyl)cysteine; D000111:Acetylcysteine