title: Integrated HPLC-MS and (1)H-NMR spectroscopic studies on acyl migration reaction kinetics of model drug ester glucuronides.
abstract: Acyl glucuronides (AGs) are common, chemically reactive metabolites of acidic xenobiotics. Concerns about the potential of this class of conjugate to cause toxicity in man require efficient methods for the determination of reactivity, and this is commonly done by measuring transacylation kinetics. High-performance liquid chromatography-mass spectrometry (HPLC-MS) and nuclear magnetic resonance (NMR) spectroscopy were applied to the kinetic analysis of AG isomerization and hydrolysis for the 1-beta-O-AGs of ibufenac, (R)- and (S)-ibuprofen, and an alpha,alpha-dimethylated ibuprofen analogue. Each AG was incubated in either aqueous buffer at pH 7.4 or human plasma at 37 degrees C. Aliquots of these samples, taken throughout the reaction time course, were analysed by HPLC-MS and (1)H-NMR spectroscopy and the results compared. For identification of the AGs incubated in pH 7.4 buffer and for analysis of kinetic rates, (1)H-NMR spectroscopy generally gave the most complete set of data, but for human plasma the use of (1)H-NMR spectroscopy was impractical and HPLC-MS was more suitable. HPLC-MS was more sensitive than (1)H-NMR spectroscopy, but the lack of suitable stable-isotope labelled internal standards, together with differences in response between glucuronides and aglycones, made quantification problematic. Using HPLC-MS a specific 1-beta-O-AG-related ion at m/z 193 (the glucuronate fragment) was noted enabling selective determination of these isomers. In buffer, transacylation reactions predominated, with relatively little hydrolysis to the free aglycone observed. In human plasma incubations the observed rates of reaction were much faster than for buffer, and hydrolysis to the free aglycone was the major route. These results illustrate the strengths and weaknesses of each analytical approach for this class of analyte.
authors.affiliation: Biomolecular Medicine, Department of Surgery and Cancer, Faculty of Medicine, Imperial College London, Sir Alexander Fleming Building, South Kensington, London SW7 2AZ, UK.
authors.forename: C H
authors.initials: CH
authors.lastname: Johnson
authors.forename: E
authors.initials: E
authors.lastname: Karlsson
authors.forename: S
authors.initials: S
authors.lastname: Sarda
authors.forename: L
authors.initials: L
authors.lastname: Iddon
authors.forename: M
authors.initials: M
authors.lastname: Iqbal
authors.forename: X
authors.initials: X
authors.lastname: Meng
authors.forename: J R
authors.initials: JR
authors.lastname: Harding
authors.forename: A V
authors.initials: AV
authors.lastname: Stachulski
authors.forename: J K
authors.initials: JK
authors.lastname: Nicholson
authors.forename: I D
authors.initials: ID
authors.lastname: Wilson
authors.forename: J C
authors.initials: JC
authors.lastname: Lindon
mesh_terms: D000215:Acylation; D002851:Chromatography, High Pressure Liquid; D020719:Glucuronides / Q000097:blood / Q000737:chemistry / Q000493:pharmacokinetics*; D006801:Humans; D006868:Hydrolysis; D007052:Ibuprofen / Q000097:blood / Q000737:chemistry / Q000493:pharmacokinetics; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D010648:Phenylacetates / Q000097:blood / Q000737:chemistry / Q000493:pharmacokinetics
chemical_list: D020719:Glucuronides; D010648:Phenylacetates; C100118:ibufenac; D007052:Ibuprofen