Simultaneous determination of bioactive components in essential oil of Xiang–Fu–Si–Wu Formula in Beagle dog plasma by UPLC–MS/MS and its application to pharmacokinetics

A highly sensitive and rapid ultraperformance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) has been developed and validated for simultaneous quantification of the three main bioactive compounds, i.e., ligustilide, dehydrocostuslactone and α-cyperone in dog plasma after oral administration of the essential oil of Xiang–Fu–Si–Wu Formula (XEO). Clarithromycin was used as an internal standard (IS). Plasma samples were processed by protein precipitation with methanol. The separation was performed on an Acquity BEH C18 column (100 mm × 2.1 mm, 1.7 μm) at a flow rate of 0.4 mL min−1, using 0.1% formic acid–acetonitrile as mobile phase. The MS/MS ion transit ions monitored were 190.5 → 90.9 for ligustilide, 231.1 → 185.1 for dehydrocostuslactone, 219.2 → 123.0 for α-cyperone and 748.5 → 158.1 for IS. Method validation was performed as per Food and Drug Administration guidelines and the results met the acceptance criteria. The lower limit of quantification (LLOQ) achieved was 5.12 ng/mL for ligustilide, 1.06 ng/mL for dehydrocostuslactone and 1.89 ng/mL for α-cyperone, respectively, and the calibration curves obtained were linear (r > 0.99) over the concentration range approximately 1–1000 ng/mL. The intra- and inter-day precision was less than 15% and the accuracy was within ±9.2%. After validation, this method was successfully applied to a pharmacokinetic study where dogs were orally given 0.3 g/kg XEO, equivalent to 183.6 mg/kg of ligustilide, 5.0 mg/kg of dehydrocostuslactone and 26.2 mg/kg of α-cyperone, respectively.