Determination of sec-O-glucosylhamaudol in rat plasma by gradient elution liquid chromatography–mass spectrometry

• A simple and selective liquid chromatography mass spectrometry (LC–MS) method for determination of sec-O-glucosylhamaudol in rat plasma was developed.
• After addition of carbamazepine as internal standard (IS), simple one-step protein precipitation by acetonitrile–methanol (9:1, v/v) was used as sample preparation.
• The LC–MS method successfully applied to a pharmacokinetic study of sec-O-glucosylhamaudol after intravenous administration of single dosage 2.5 mg/kg to rats.

Sec-O-glucosylhamaudol is one of the major bioactive compounds of the Saposhnikoviae Radix. A simple and selective liquid chromatography–mass spectrometry (LC–MS) method for determination of sec-O-glucosylhamaudol in rat plasma was developed. After addition of carbamazepine as internal standard (IS), protein precipitation with acetonitrile–methanol (9:1, v/v) was used as sample preparation. Chromatographic separation was achieved on a Zorbax SB-C18 (2.1 mm × 150 mm, 5 μm) column with acetonitrile–0.1% formic acid in water as mobile phase with gradient elution. Electrospray ionization (ESI) source was applied and operated in positive ion mode; selective ion monitoring (SIM) mode was used for quantification using target fragment ions m/z 439 for sec-O-glucosylhamaudol and m/z 237 for the IS. Calibration plots were linear over the range of 50–8000 ng/mL for sec-O-glucosylhamaudol in rat plasma. Mean recovery of sec-O-glucosylhamaudol in plasma was in the range of 74.8–83.7%. Intra-day and inter-day precision were both <15%. This method was successfully applied in pharmacokinetic study after intravenous administration of 2.5 mg/kg sec-O-glucosylhamaudol in rats.