Simultaneous determination of triptolide, wilforlide A and triptonide in human plasma by high-performance liquid chromatography-electrospray ionization mass spectrometry.

A simple and sensitive method for the determination of triptolide, wilforlide A and triptonide in human plasma was developed and validated, using high-performance liquid chromatography-electrospray ionization mass spectrometry (LC-ESI-MS). Plasma samples were purified using solid-phase extraction (SPE) columns. The HPLC separation of the analytes was performed on a MACHEREY-NAGEL C(18)column (2.0 mm x 125 mm, 3 microm), using 2.7 mM formic acid containing 10 mM ammonium acetate-acetonitrile (55:45, v/v) as mobile phase, with a flow-rate of 0.25 ml/min. The compounds were ionized in the electrospray ionization (ESI) ion source of the mass spectrometer and were detected in the selected ion recording (SIR) mode. The calibration curves were linear in the 0.80-300 ng/ml range for all the three analytes, and the limits of detection were 0.25, 0.40, and 0.35 ng/ml for triptolide, wilforlide A, and triptonide, respectively. The average absolute recoveries for all the three analytes were above 81%. The methodology recoveries were greater than 91% and the relative standard deviations (RSD) of intra-day and inter-day were less than 15%. The developed method was successfully applied to the determination of triptolide, wilforlide A and triptonide concentration in patients' plasma after taking the medicament containing Tripterygium wilfordii HOOK. F.