Dataset Information

Identification of alcohol-dependent clopidogrel metabolites using conventional liquid chromatography/triple quadrupole mass spectrometry.

ABSTRACT: Clopidogrel (CLO) is a prodrug used to prevent ischemic events in patients undergoing percutaneous coronary intervention or with myocardial infarction. A previous study found ethyl clopidogrel (ECLO) is formed by transesterification of CLO when incubated with alcohol in human liver microsomes. We hypothesize that ECLO will be subject to further metabolism and developed an assay to identify its metabolites.A liquid chromatography/triple quadrupole mass spectrometry (LC/MS/MS) method was developed to identify metabolites of ECLO. According to the predicted metabolic pathway of ECLO, precursor-product ion pairs were used to screen the possible metabolites of ECLO in human liver S9 fractions. Subsequently, the detected metabolites were characterized by the results of product ion scan.In the presence of alcohol, CLO was tranesterified to ECLO, which was further oxidized to form ethylated 2-oxo-clopidogrel and several ethylated thiol metabolites including the ethylated form of the H4 active metabolite.The ECLO formed by transesterification with alcohol is subject to metabolism by CYP450 enzymes producing ethylated forms of 2-oxo-clopidogrel and the active H4 thiol metabolite.

SUBMITTER: Hu ZY

PROVIDER: S-EPMC4034390 | biostudies-literature | 2014 Jun

REPOSITORIES: biostudies-literature

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Identification of alcohol-dependent clopidogrel metabolites using conventional liquid chromatography/triple quadrupole mass spectrometry.

Hu Zhe-Yi ZY Laizure S Casey SC Herring Vanessa L VL Parker Robert B RB

Rapid communications in mass spectrometry : RCM 20140601 11

<h4>Rationale</h4>Clopidogrel (CLO) is a prodrug used to prevent ischemic events in patients undergoing percutaneous coronary intervention or with myocardial infarction. A previous study found ethyl clopidogrel (ECLO) is formed by transesterification of CLO when incubated with alcohol in human liver microsomes. We hypothesize that ECLO will be subject to further metabolism and developed an assay to identify its metabolites.<h4>Methods</h4>A liquid chromatography/triple quadrupole mass spectromet ...[more]

PMID: 24760569

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Project description:Accumulating evidence indicates that the health impact of dietary phenolic compounds, including the principal grape-derived polyphenols, (+)‑catechin and (-)‑epicatechin, is exerted by not only the parent compounds but also their phenolic metabolites generated by the gut microbiota. In this work, a new high-throughput, sensitive and reproducible analytical method was developed employing ultra-high performance liquid chromatography coupled with triple quadrupole tandem mass spectrometry (UHPLC-QqQ-MS/MS) for the simultaneous analysis of 16 microbial-generated phenolic acid metabolites (PAMs) along with their precursors, catechin and epicatechin. Following optimizing the solvent system, LC conditions and MS parameters, method validation was carried out to evaluate the sensitivity, selectivity, accuracy and precision of the proposed method, and to ensure promising recovery of all analytes extracted from the matrix prior to bioanalysis. Results showed that the optimized analytical method allowed successful confirmation and quantitation of all analytes under dynamic multiple reaction monitoring mode using trans‑cinnamic acid‑d7 as an internal standard (I.S.). Excellent sensitivity and linearity were obtained for all analytes, with lower limits of detection (LLODs) and lower limits of quantification (LLOQs) in the ranges of 0.225-2.053 ng/mL and 0.698-8.116 ng/mL, respectively. By examining blank matrix spiked with standard mixture at different concentration levels, promising recoveries at two spiking levels (low level, 91.2-115%; high level 90.2-121%), and excellent precision (RSD < 10%) were obtained. This method was then successfully applied to an in vitro study where catechin/epicatechin-enriched broth samples were anaerobically fermented with gut microbes procured from healthy human donors. All sources of bacteria employed showed remarkable activity in metabolizing grape polyphenols and distinct variations in the production of PAMs. The successful application of this method in the in vitro fermentation assays demonstrates its suitability for high-throughput analysis of polyphenol metabolites, particularly catechin/epicatechin-derived PAMs, in biological studies.

Dataset Information

Identification of alcohol-dependent clopidogrel metabolites using conventional liquid chromatography/triple quadrupole mass spectrometry.

Publications

Identification of alcohol-dependent clopidogrel metabolites using conventional liquid chromatography/triple quadrupole mass spectrometry.

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