Objective:To develop a high performance liquid chromatography-mass spectrometry(HPLC-MS/MS)method for the determination of vildagliptin in human anticoagulant plasma with ethylenediamine tetra acetic acid and apply it to the study of pharmacokinetics.Methods:13C-15N-vildagliptin was used as internal standard(IS).After extraction from human plasma by protein precipitation with acetonitrile,all components were separated by a Hypurity C18 column(150 mm ×2.1 mm,5 μm),using a gradient elution procedure consisting of methanol and 5 mmol·L-1 ammonium formate at a flow rate of 0.5 mL·min-1,and the column temperature was 40 ℃.Injection volume was just 2 μL Positive electrospray ionization was performed using multiple reaction monitoring(MRM)with transitions of m/z 304.3→154.2 for vildagliptin and m/z 310.3→160.3 for internal standard.Specificity,standard curve,lower limit of quantification,precision,recovery,matrix effect and stability were examined.Then this method was used to determine the plasma concentration of veragliptin in healthy subjects.Results:The calibration curve of vildagliptin in human plasma was linear over the concentration range of 1.11 to 534.0 ng·mL-1.The low-er limit of quantitation was 1.11 ng·mL-1.The intra-and inter-day precisions at four quality control levels were within 0.9％-8.5％,and the accuracy was within 99.8％-109.3％.The data of short-term stability at room temperature displayed that the accuracy percentage of LQC samples was 92.0％for 0.5 h exposure,87.6％for 1 h exposure,71.2％for 2 h exposure.These of LQC samples chilled on ice was 102.0％for 0.5 h exposure,94.5％for 1 h exposure,86.6％for 2 h exposure.These results showed a phenomenon that there was a possible degradation of vildagliptin in plasma.The results of extraction recovery and matrix effect and other stability met the requirements of biological sample analysis.The pharmacokinetic study results of 8 healthy subjects showed that t1/2 was(1.49± 0.37)h,tmaxwas(2.06±1.11)h,Cmax was(290.94±100.36)ng·mL-1,AUC0-24h was(1 343.46±186.89)ng·h·mL-1,AUC0-∞ was(1 351.31±188.79)ng·h·mL-1.Conclusion:This method is easy to operate,has high specificity,and sensitivity.It has been successfully applied to the pharmacokinetic study of 8 healthy subjects after oral administration of 50 mg vigagliptin tablets on an empty stomach.Therefore,it can be used as a reliable detection method for human pharmacokinetic research and therapeutic drug monitoring.
HPLC-MS/MSvildagliptindipeptidyl peptidase-4(DDP-4)stabilitypharmacokineticsdia-betestherapeutic drug monitoring(TDM)
维普
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