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A highly specific and sensitive liquid chromatography–tandem mass spectrometry method has been developed and validated for the determination of ponicidin in dog plasma. The plasma samples were prepared using liquid–liquid extraction with ethyl acetate as the extraction solvent. Chromatographic separation was accomplished on a Waters XTerra MS C18 column. The extracted ponicidin and the internal standard, oridonin, were detected by tandem mass spectrometry in positive electrospray ionization mode with multiple reaction monitoring. The optimized mass transition ion pairs ( m/z ) for quantitation were 363.08–345.08 for ponicidin and  m/z  365.10–347.06 for the internal standard. The lower limit of quantification was 5 ng/mL. The linear range of the method was from 5 to 5,000 ng/mL. The intra-day and inter-day precision measurements were lower than 5.3 and 6.0% in terms of relative standard deviation and the accuracy was within ±8.4% in terms of relative error. Additionally, no significant matrix ionization suppression or enhancement was observed. The validated method was successfully applied in a pharmacokinetic study of ponicidin in dogs. The primary pharmacokinetic parameters in dogs were: terminal elimination half-life, 8.14 ± 1.35 h; mean residence time, 12.30 ± 2.08 h; area under the plasma concentration–time curve from time zero to the last measurable concentration, 14.34 ± 1.37 µg/h/mL; area under the plasma concentration–time curve from time zero to infinity, 15.75 ± 1.44 µg/h/mL; apparent volume of distribution, 4.79± 1.68 L/kg; total body clearance, 0.41 ± 0.08 L/kg/h.

LC–MS-MS for the Determination of Ponicidin in Dog Plasma