Simultaneous determination of a novel M3 muscarinic receptor antagonist and its active 5-OH metabolite in human plasma using liquid chromatography/tandem mass spectrometry.
A sensitive, specific, and robust liquid chromatography (LC)/mass spectrometry (MS)/MS method has been developed and validated for a novel M(3) muscarinic receptor antagonist (I) and its active 5-OH metabolite (II) in human plasma. The assay involves a two-step liquid-liquid extraction of the compounds from human plasma, high performance liquid chromatography (HPLC) separation, and MS/MS for the detection of the analytes. The method provides a linear response from a quantitation limit of 0.05-20 ng/ml for I and 0.1-20 ng/ml for II using 1 ml of plasma. The mean absolute recovery was 85.4% for I and 80.8% for II, respectively. The intra-assay accuracy of I and II averaged from 95.0 to 105.3% with coefficient of variation (CV) values <or=6.5% over the standard curve range. The stability study showed that I and II are stable in the plasma matrix over a period of 11 months at -70 degrees C. The accuracy, ruggedness, and reproducibility of this method were demonstrated by analyzing over 5000 plasma samples in clinical pharmacokinetics studies over a 6-month period.
Yan
Kerri X
KX
Department of Drug Metabolism, Merck Research Laboratories, WP75A-303, West Point, PA 19486, USA. yan@merck.com
Song
Hengchang
H
Riffel
Kerry
K
Lo
Man-Wai
MW
D002853:Chromatography, Liquid / Q000379:methods; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods; D018727:Muscarinic Antagonists / Q000032:analysis* / Q000097:blood / Q000737:chemistry / Q000378:metabolism*; D043587:Receptor, Muscarinic M3; D011976:Receptors, Muscarinic / Q000097:blood
D018727:Muscarinic Antagonists; D043587:Receptor, Muscarinic M3; D011976:Receptors, Muscarinic