Quantification of the O- and N-demethylated metabolites of hydrocodone and oxycodone in human liver microsomes using liquid chromatography with ultraviolet absorbance detection

Andrew Menelaou, Mark R. Hutchinson, Ingvild Quinn, Anders Christensen, Andrew A. Somogyi

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    High-performance liquid chromatographic assays for the O- and N-demethylated oxidative metabolites of hydrocodone and oxycodone formed in human liver microsomes are described. A solvent-solvent extraction/re-extraction procedure followed by reversed-phase HPLC with UV detection at 210 nm allows for the quantification of hydromorphone, norhydrocodone, oxymorphone and noroxycodone. Calibration curve concentration ranges were 0.63-400 μM (0.18-114 μg/ml) and 1.25-400 μM (0.36-114 μg/ml) for hydromorphone and norhydrocodone, respectively and 0.13-20 μM (0.04-6.03 μg/ml) and 1-200 μM (0.30-60 μg/ml) for oxymorphone and noroxycodone, respectively. Assay performance was determined by intra- and inter-assay precision and inaccuracies for quality control samples and was <15% for all metabolites at each quality control concentration. These methods provide good precision, accuracy and sensitivity for use in in vitro kinetic studies investigating the oxidative metabolism of hydrocodone and oxycodone in human liver microsomes.

    Original languageEnglish
    Pages (from-to)81-88
    Number of pages8
    JournalJournal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences
    Issue number1
    Publication statusPublished or Issued - 25 Feb 2003


    • Hydromorphone
    • Norhydrocodone
    • Noroxycodone
    • Oxymorphone

    ASJC Scopus subject areas

    • Analytical Chemistry
    • Biochemistry
    • Clinical Biochemistry
    • Cell Biology

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