A rapid and sensitive bioassay based on liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the simultaneous determination of four isomeric escin saponins (escin Ia, escin Ib, isoescin Ia and isoescin Ib) in human plasma has been developed and validated. Sample preparation of plasma after addition of telmisartan as internal standard (I.S.) involved solid-phase extraction (SPE) on C18 cartridges. Separation was based on reversed phase chromatography using gradient elution with methanol-acetonitrile (50:50, v/v) and 10 mM ammonium acetate solution (pH 6.8). MS/MS detection in the positive ion mode used multiple reaction monitoring of the transition at m/z 1113.8-->807.6. Stability issues with the four saponins required the addition of formic acid to plasma samples prior to storage at -80 degrees C and analysis within 30 days. The method was linear at concentrations up to 10 ng/mL with correlation coefficients>0.996 for all analytes. The lower limit of quantitation (LLOQ) for all four saponins was 33 pg/mL. Intra- and inter-day precisions (as relative standard deviation) were all <15% and accuracies (as relative error) in the range -5.3% to 6.1%. The method was successfully applied to a pharmacokinetic study of escins in healthy volunteers after oral administration of sodium aescinate tablets containing 60 mg escin saponins.
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