LC-MS/MS method for quantifying aescinate A and B and assessing their relationship with phlebitis.

The purpose of this study is to establish and validate a sensitive, robust and rapid liquid chromatography-tandem mass spectrometry method for quantifying the aescinate A and aescinate B in human plasma and assessing the association of phlebitis and aescinate A and aescinate B in vivo exposure. The chromatographic separation was completed on Agilent ZORBAX SB-C18 (2.1 mm × 100 mm, 3.5 µm, Agilent, USA) column with isocratic elution. The flow rate was 0.3 mL/min and the total run time was optimized within 5 min. The protein precipitation was applied to pretreat plasma sample using methanol as precipitant. The data acquisition was achieved with positive electrospray ionization in multi-reaction monitoring mode for both aescinate A and aescinate B. The calibration range of aescinate A and aescinate B are constructed in 100-2000 ng/mL, and their correlation coefficients are both >0.990. The intra-day and inter-day precision and accuracy of this method are less than 9.04% and within -13.75% and -0.93%. This analytical method has been successfully applied for the determination of plasma aescinate A and aescinate B concentrations in patients with cerebral infarction, and the results showed that the incidence and grade of phlebitis were not associated with the in vivo exposure of aescinate A and aescinate B.