title: Esterase inhibitors as ester-containing drug stabilizers and their hydrolytic products: potential contributors to the matrix effects on bioanalysis by liquid chromatography/tandem mass spectrometry.
abstract: BACKGROUND
  Esterase inhibitors are widely used to stabilize ester-containing drugs in biological matrices for quantitative liquid chromatography/tandem mass spectrometry (LC/MS/MS) assays. These co-existing inhibitors could cause matrix effects on bioanalysis and jeopardize the assay performance. We therefore developed an LC/MS/MS methodology to monitor the fate of inhibitors and evaluate their matrix effects, which is described in this study.
  
  
  METHODS
  Human plasma containing 20 mM of diisopropylfluorophosphate (DFP), paraoxon, eserine, phenylmethylsulfonyl fluoride (PMSF) or 2-thenoyltrifluoroacetone (TTFA) was extracted by liquid-liquid extraction (LLE) and analyzed by an LC/MS/MS assay for BMS-068645 (a model drug) with additional pre-optimized selected reaction monitoring (SRM) transitions using positive/negative electrospray ionization (ESI) mode for each inhibitor. Hydrolytic products were characterized by product ion or neutral loss scan LC/MS/MS analysis. The matrix effect contribution from each inhibitor was evaluated by post-column infusion of BMS-068645.
  
  
  RESULTS
  In the extracted samples by LLE, SRM chromatograms revealed the presence of paraoxon, eserine and TTFA with peak intensity of >2.50E08. Three DFP hydrolytic products, diisopropyl phosphate (DP), triisopropyl phosphate (TP) and DP dimer, and one PMSF hydrolytic product, phenymethanesulfonic acid (PMSA), were identified in the extracted samples. In post-column infusion profiles, ion suppression or enhancement was observed in the retention time regions of eserine (~10% suppression), paraoxon (~70% enhancement) and DP dimer (~20% suppression).
  
  
  CONCLUSIONS
  The SRM transitions described here make it possible to directly monitor the inhibitors and their hydrolytic products. In combination with post-column infusion, this methodology provides a powerful tool to routinely monitor the matrix effects-causing inhibitors, so that their matrix effects on the bioanalysis can be evaluated and minimized.
authors.affiliation: Bioanalytical Sciences, Bristol-Myers Squibb Research & Development, Princeton, NJ 08543, USA.
authors.forename: Naiyu
authors.initials: N
authors.lastname: Zheng
authors.forename: Eliza N
authors.initials: EN
authors.lastname: Fung
authors.forename: Adela
authors.initials: A
authors.lastname: Buzescu
authors.forename: Mark E
authors.initials: ME
authors.lastname: Arnold
authors.forename: Jianing
authors.initials: J
authors.lastname: Zeng
mesh_terms: D000480:Alkynes / Q000097:blood / Q000737:chemistry; D001774:Blood Chemical Analysis / Q000379:methods* / Q000592:standards; D002853:Chromatography, Liquid / Q000379:methods*; D004355:Drug Stability; D004791:Enzyme Inhibitors / Q000097:blood / Q000737:chemistry* / Q000378:metabolism; D004950:Esterases / Q000037:antagonists & inhibitors*; D006801:Humans; D006868:Hydrolysis; D007531:Isoflurophate / Q000097:blood / Q000737:chemistry / Q000378:metabolism; D008956:Models, Chemical; D010261:Paraoxon / Q000097:blood / Q000737:chemistry / Q000378:metabolism; D010664:Phenylmethylsulfonyl Fluoride / Q000097:blood / Q000737:chemistry / Q000378:metabolism; D010830:Physostigmine / Q000097:blood / Q000737:chemistry / Q000378:metabolism; D011684:Purine Nucleosides / Q000097:blood / Q000737:chemistry; D053719:Tandem Mass Spectrometry / Q000379:methods*; D013804:Thenoyltrifluoroacetone / Q000032:analysis / Q000737:chemistry / Q000378:metabolism
chemical_list: D000480:Alkynes; C521541:BMS-068645; D004791:Enzyme Inhibitors; D011684:Purine Nucleosides; D007531:Isoflurophate; D013804:Thenoyltrifluoroacetone; D010664:Phenylmethylsulfonyl Fluoride; D010830:Physostigmine; D004950:Esterases; D010261:Paraoxon