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Simultaneous quantification of acetaminophen and structurally related compounds in human serum and plasma
Author(s) -
Bylda Caroline,
Thiele Roland,
Kobold Uwe,
Volmer Dietrich A.
Publication year - 2014
Publication title -
drug testing and analysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.065
H-Index - 54
eISSN - 1942-7611
pISSN - 1942-7603
DOI - 10.1002/dta.1527
Subject(s) - chemistry , chromatography , phenacetin , analyte , electrospray ionization , detection limit , acetaminophen , mass spectrometry , triple quadrupole mass spectrometer , selected reaction monitoring , tandem mass spectrometry , biochemistry
The method described in this study allows the simultaneous quantification of acetaminophen (APAP) and nine structurally related compounds, namely acetaminophen metabolites and structurally similar analogs (acetaminophen‐glucuronide [APG], ‐sulfate [APS], mercapturate [APM], ‐cysteine [APC], p ‐phenetidine, phenacetin), antidote (N‐acetylcysteine, NAC), and two tricyclic antidepressants (imipramine and amitryptiline). Due to the relatively high serum concentration levels in the µg/ml range, matrix effects were simply minimized by dilution. The samples were diluted with water and disulfide bonds between serum proteins and analytes reduced using tris(2‐carboxyethyl)phosphine. Chromatographic separation of the analytes was achieved by gradient elution using a pentafluorphenyl (PFP) column with subsequent detection by electrospray ionization (ESI) triple quadrupole mass spectrometry in positive and negative ionization multiple reaction monitoring (MRM) modes. Quantification was performed by means of deuterated analogues of the analytes as internal standards. Total run time of the assay was 19 min. The method was fully validated and allowed quantification of the analytes with lower limits of quantification between 50 and 0.5 ng/ml. The calibration curves were linear over the range 0.1–100 µg/ml for APAP, APG, NAC, p ‐phenetidine and phenacetin, 0.03‐50 µg/ml for APS, and 0.01‐10 µg/ml for APM, APC, imipramine and amitriptyline with correlation coefficients r 2 > 0.99. The intra‐assay precision was ≤5% for all analytes except NAC (CV < 10%). The inter‐day precision was ≤10% for all analytes except NAC (inter‐assay precision <11%). This method was used to analyze 77 patient and spiked samples and results were consistent with expected values from a round robin test. Copyright © 2013 John Wiley & Sons, Ltd.