Characterization ofin vitrometabolites from human liver microsomes using directly coupled hplc-nmr: application to a phenoxathiin monoamine oxidase-A inhibitor

Abstract

1. The metabolism of 1-ethylphenoxathiin-10,10-dioxide (BW1370U87), an experimental compound designed as an inhibitor of monoamine oxidase-A for use as a possible anti-depression agent, has been studied in a human liver microsome preparation. 2. The identities of the metabolites have been determined using directly coupled hplc-1H nmr at 600 MHz in the stop-flow mode in this first study of in vitro xenobiotic metabolism using hplc-nmr. 3. The xenobiotic substances that were identified comprised the parent compound BW1370U87 (with -CH2CH3 at C1) together with sidechain-oxidized metabolites with C1 substituents of -CHOH.CH3, -CH2.CH2OH, -CHOH.CH2OH and -CH2.COOH, plus the unsubstituted phenoxathiin-10,10-dioxide.

Magnetic Resonance Methods and Applications in Pharmaceutical Research

“…The different types of samples that can be analyzed (e.g., urine, liver and kidney tissues, microsomal fractions 122,130 ), together with the combination of NMR and MS with separation techniques such as high performance liquid chromatography, [130][131][132] illustrate the usefulness of the approach for drug discovery and testing. Recent studies with exhaled breath condensates 133,134 demonstrate applications to the field of biomarkers of response reflecting underlying pathological change caused by disease or chemical exposure (Fig.…”

High‐throughput screening for stability and inhibitory activity of compounds toward cytochrome P450‐mediated metabolism

“…To solve this problem, complex mixtures of metabolites can first be separated by HPLC and then directly introduced into an NMR spectrometer. LC‐NMR hardware has been commercially available since the early 1990s,71–77 followed by directly coupled LC‐NMR‐MS in 1996,78–82 and has proven useful in eliminating some of the costly and time‐consuming isolation procedures required by NMR alone, and furthermore has increased the speed and ease with which a drug metabolite can be identified by NMR. For example, Holt et al82 described the advantage of combining LC‐MS with LC‐NMR for elucidating the structures of individual peptides from a commercial mixture.…”

Isolation of circulating metabolites in drug discovery using high-performance liquid chromatography, and their identification by liquid chromatography coupled with tandem mass spectrometry and nuclear magnetic resonance spectroscopy

“…Data generated from these MS/MS techniques can be combined with MS n data from a quadrupole ion trap and accurate mass measurements from a quadrupole-time-of-flight instrument to provide a detailed assessment of metabolite structure [2]. There have been a number of publications using LC-MS and LC-NMR to identify drug metabolites in human and rat urine [6 -8], liver microsomal incubations [9], and cell culture media [10]. NMR coupled to liquid chromatography is a very powerful technique for the separation and structural elucidation of minute amounts of organic molecules, and this technique is now being actively applied to drug meta-bolism studies [4,5].…”