High performance liquid chromatography – tandem mass spectrometric determination of cyclovirobuxine D in human plasma

A sensitive, specific and rapid high performance liquid chromatography – tandem mass spectrometric (LC/MS/MS) method for the determination of cyclovirobuxine D in human plasma was developed and validated. The triple–quadrupole tandem mass spectrometric detector with an electrospray interface (ESI) was operated under the selected reaction monitoring (SRM) mode. After the addition of citalopram as an internal standard (IS), plasma samples were extracted with ethyl acetate. Chromatographic separation of the analytes was performed on a Kromasil CN column with a mobile phase of methanol/water (88/12, v/v) containing 0.4% formic acid. Linearity was established for the range of concentration 0.2–40 ng/ml. Under optimized conditions, the mean recovery was 86.6%. The intra-day precision ranged from 4.56% to 7.81%, while the intra-day accuracy ranged from 2.75% to 11.0%. The inter-day precision was in the range 3.87–10.7%, and the inter-day accuracy was in the range −4.00% to 2.50%. The cyclovirobuxine D was stable in human plasma after three freeze–thaw cycles, under storage at room temperature for 12 h, in a freezer at −20 °C for 15 days and during processing (in autosampler) at 10 °C for 24 h. The validated method is suitable for quantitative determination of cyclovirobuxine D in human plasma in pharmacokinetics study and has been successfully applied to the analysis of clinical samples.