Absolute quantitation of acetaminophen‐modified human serum albumin in acute liver failure patients by liquid chromatography/tandem mass spectrometry. (31st July 2018)

• Record Type: Journal Article
• Authors: Geib, Timon; LeBlanc, André; Shiao, Tze Chieh; Roy, René; Leslie, Elaine M.; Karvellas, Constantine J.; Sleno, Lekha
• Is Part Of: Rapid communications in mass spectrometry. Volume 32:Number 17(2018)
• Page Start: 1573
• Page End: 1582
• Publication Date: 2018-07-31
• Subjects: Mass spectrometry -- Periodicals; 543.65
• DOI: 10.1002/rcm.8206
• Languages: English
• ISSNs: 0951-4198

Abstract

Abstract : Rationale: Acetaminophen (APAP) is a well‐known analgesic, deemed a very safe over‐the‐counter medication. However, it is also the main cause of acute liver failure (ALF) in the Western world, via the formation of its reactive metabolite, N ‐acetyl p ‐benzoquinone imine (NAPQI), and its covalent attachment to liver proteins. The aim of this study was to develop a sensitive and robust quantitative assay to monitor APAP‐protein binding to human serum albumin (HSA) in patient samples. Methods: A combination of isotope dilution, peptic digestion and solid‐phase extraction coupled to liquid chromatography/multiple reaction monitoring (LC/MRM) was employed. An external calibration curve with surrogate modified protein spiked into blank serum was used for absolute quantitation. Samples were analyzed by LC/MRM to measure the modified active site peptide of HSA. The LC/MRM assay was validated and successfully applied to serum samples from patients suffering from APAP‐induced ALF. Results: Accuracy ranged from 83.8–113.3%, within‐run coefficient of variation (CV) ranged from 0.3–6.9%, and total CVs from 1.6–10.6%. Patient samples ranged from 0.12–3.91 nmol/mL NAPQI‐HSA; in‐between the assay dynamic range of 0.11–50.13 nmol/mL serum. In vivo median concentrations were found to be 0.62 nmol/mL and 0.91 nmol/mL for non‐spontaneous survivors ( n = 25) and individuals with irreversible liver damage ( n = 10), respectively ( p ‐value = 0.028), demonstrating significant potential as a biomarker for ALF outcome. Conclusions: A fast and sensitive assay was developed to accurately quantify NAPQI‐HSA as a biomarker for APAP‐related covalent binding in human serum.