title: HPLC-MS/MS methods for the quantitative analysis of ophthalmic acid in rodent plasma and hepatic cell line culture medium.
abstract: Ophthalmic acid (OA), an endogenous tripeptide analogue of glutathione, has been suggested as a potential biomarker for paracetamol/acetaminophen hepatotoxicity. Here HPLC-MS/MS methods have been developed for the precise, sensitive and specific detection and quantification of OA in in vitro cell culture medium and plasma. For the cell culture medium the LLOQ was found to be 1 ng/ml, with less than 1% between sample carry over at all concentrations and precision below 15% for within day and below 9% for between day analyses. For rat plasma the presence of endogenous OA resulted in the LLOQ being 25 ng/ml (defined as the lowest concentration on the calibration curve where the base peak was less than 20% of the LLOQ). For the plasma assay the percentage carry over was less than 1% for all concentrations and within and between batch precision was below 21%. The methods were linear for both sample types from the LLOQ up to 5 μg/ml. The method was successfully applied to the determination of OA in samples obtained following the chronic administration of the rat hepatotoxin methapyrilene, where plasma OA concentrations were observed to show a weak negative correlation with those of established liver injury biomarkers such as aspartate aminotransferase (AST).
authors.affiliation: Manchester Centre for Integrative Systems Biology and Doctoral Training Centre ISBML, Manchester Interdisciplinary Biocentre, University of Manchester, 131 Princess Street, Manchester M1 7DN, UK.
authors.forename: Suzanne
authors.initials: S
authors.lastname: Geenen
authors.forename: Filippos
authors.initials: F
authors.lastname: Michopoulos
authors.forename: J Gerry
authors.initials: JG
authors.lastname: Kenna
authors.forename: Kyle L
authors.initials: KL
authors.lastname: Kolaja
authors.forename: Hans V
authors.initials: HV
authors.lastname: Westerhoff
authors.forename: Ian
authors.initials: I
authors.lastname: Wilson
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000097:blood; D002460:Cell Line; D056486:Chemical and Drug Induced Liver Injury / Q000097:blood / Q000209:etiology; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003470:Culture Media / Q000032:analysis*; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D057230:Limit of Detection; D008099:Liver / Q000166:cytology / Q000378:metabolism*; D008297:Male; D008701:Methapyrilene / Q000633:toxicity; D009842:Oligopeptides / Q000032:analysis / Q000097:blood*; D051381:Rats; D015203:Reproducibility of Results; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D015415:Biomarkers; D003470:Culture Media; D009842:Oligopeptides; C016632:ophthalmic acid; D008701:Methapyrilene