Simultaneous determination of norisoboldine and its major metabolite in rat plasma by ultra‐performance liquid chromatography–mass spectrometry and its application in a pharmacokinetic study

Abstract

Norisoboldine (NIB) is one of the main bioactive isoquinoline alkaloids in Linderae Radix. A rapid, selective and sensitive method using UPLC-ESI/MS was first developed for simultaneous determination of NIB and norisoboldine-9-O-α-glucuronide (NIB-Glu), its major metabolite in rat plasma. A one-step protein precipitation with methanol was employed as sample preparation technique. Chromatographic separation was carried out on an Acquity UPLC BEH C(18) column (50 × 2.1 mm, i.d. 1.7 µm) with a gradient mobile phase consisting of acetonitrile and water containing 0.1% formic acid. Detection and quantification were performed using a quadrupole mass spectrometer by selective ion reaction-monitoring mode. Good linearity was achieved using weighted (1/x(2) ) least squares linear regression over the concentration ranges 0.01-2 µg/mL for NIB and 0.025-25 µg/mL for NIB-Glu. The lower limit of quantification of NIB and NIB-Glu was 0.01 and 0.025 µg/mL, respectively. The intra- and inter-day precisions (relative standard deviations) of the assay at all three quality control levels were 4.6-14.1% for NIB, and 5.0-12.2% for NIB-Glu. The accuracies (relative error) were -13.5-8.1% for NIB and -12.8-7.6% for NIB-Glu, respectively. This developed method was successfully applied to an in vivo pharmacokinetic study in rats after a single intravenous dose of 10 mg/kg NIB.