High throughput liquid chromatography-tandem mass spectrometry assay for mercapturic acids of acrolein and crotonaldehyde in cigarette smokers' urine.
3-Hydroxypropylmercapturic acid (3-HPMA) and 3-hydroxy-1-methylpropylmercapturic acid (HMPMA) are urinary metabolites of the toxicants acrolein and crotonaldehyde, respectively. Virtually all human urine samples contain these metabolites, resulting from the action of glutathione-S-transferases on acrolein and crotonaldehyde, which are lipid peroxidation products, environmental and dietary contaminants, and constituents of cigarette smoke. We have developed a high throughput liquid chromatography-tandem mass spectrometry method for quantitative analysis of 3-HPMA and HMPMA in large numbers of small urine samples, as would be required in molecular epidemiology and clinical studies relating levels of these metabolites to cancer risk. Solid-phase extraction on mixed mode reverse phase-anion exchange 96-well plates provided sufficient purification for LC-MS/MS analysis, which was performed by auto-injection using a 96-well format, and resulted in clean, readily interpretable chromatograms, with detection limits of 4.5pmol/mL urine for 3-HPMA and 3.5pmol/mL urine for HMPMA. Accuracy was 92% for 3-HPMA and 97% for HMPMA while inter-day precision was 9.1% (coefficient of variation) for 3-HPMA and 11.0% for HMPMA. The method was applied to more than 2600 urine samples from smokers; mean levels of 3-HPMA and HMPMA were 4800±5358 (S.D.)pmol/mL and 3302±3341pmol/mL, respectively.
Masonic Cancer Center, University of Minnesota, 420 Delaware Street SE, MMC 806, Minneapolis, MN 55455, USA. carme001@umn.edu
Steven G
SG
Carmella
Menglan
M
Chen
Adam
A
Zarth
Stephen S
SS
Hecht
D000111:Acetylcysteine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000652:urine; D000171:Acrolein / Q000737:chemistry / Q000378:metabolism / Q000652:urine*; D000447:Aldehydes / Q000737:chemistry / Q000378:metabolism / Q000652:urine*; D002853:Chromatography, Liquid / Q000379:methods; D057166:High-Throughput Screening Assays / Q000379:methods; D006801:Humans; D057230:Limit of Detection; D015203:Reproducibility of Results; D012907:Smoking / Q000378:metabolism / Q000652:urine*; D052616:Solid Phase Extraction; D053719:Tandem Mass Spectrometry / Q000379:methods
C518631:3-hydroxy-1-methylpropylmercapturic acid; D000447:Aldehydes; D000171:Acrolein; C012796:2-butenal; C001423:S-(3-hydroxypropyl)cysteine N-acetate; D000111:Acetylcysteine
Acrolein; Crotonaldehyde; High throughput; Liquid chromatography–tandem mass spectrometry; Mercapturic acids