Abstract
A specific and simultaneous assay of morphine, morphine-3-glucuronide (M-3-G) and morphine-6-glucuronide (M-6-G) in monkey and dog plasma has been developed. These methods are based on rapid isolation using solid phase extraction cartridge, and high-performance liquid chromatography (HPLC)–electrospray ionization (ESI)-tandem mass spectrometric (MSMS) detection. Analytes were separated on a semi-micro ODS column in acetonitrile–formic (or acetic) acid mixed solution. The selected reaction monitoring for assay in monkey and dog plasma, as precursor→product ion combinations of m/z 286→286 for morphine, m/z 462→286 for glucuronides and m/z 312→312 for internal standard (IS, nalorphine) were used. The linearity of morphine, M-3-G and M-6-G was confirmed in the concentration range of 0.5–50, 25–2500, 2.5–250 ng/ml in monkey plasma, 0.5–100, 25–5000, 2.5–500 ng/ml in dog plasma, respectively. The precision of this assay method, expressed as CV, was less than 15% over the entire concentration range with adequate assay accuracy. Therefore, the HPLC–ESI–MSMS method is useful for the determination of morphine, M-3-G and M-6-G with sufficient sensitivity and specificity in pharmacokinetic studies.
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