UPLC-MS/MS测定人血浆中长春西汀和阿朴长春胺酸的含量及其应用

Determination of vinpocetine and Apovincaminic acid in Human plasma by UPLC-MS/MS and its Application

关键词:  长春西汀  阿朴长春胺酸  血药浓度  超高效液相色谱串联质谱法

Key words:  vinpocetine  apovincaminic acid  plasma concentration  UPLC-MS/MS

摘要:

目的 建立和验证UPLC-MS/MS测定人血浆中长春西汀和阿朴长春胺酸的方法，并用于临床样品检测。方法 以长春西汀-d5、阿朴长春胺酸-d4为内标，人血浆样品经蛋白沉淀法沉淀处理，色谱柱为Waters ACQUITY UPLC^® BEH C₁₈(2.1 mm×50 mm，1.7 μm)，流动相为乙腈-水(含0.05%甲酸)，梯度洗脱，流速0.35 mL·min^-1；电喷雾离子化源正离子监测模式。结果 长春西汀和阿朴长春胺酸线性范围分别为0.04~20.0 ng·mL^-1(r=0.999 7)、0.50~250 ng·mL^-1(r=0.999 6)，检测限分别为0.01，0.10 ng·mL^-1，待测物与内标提取回收率均为94.81%~105.0％，基质效应为94.51%~105.0%，RSD均< 5%；批内、批间精密度RSD均<10%。结论 本法特异性强、快速、准确，重复性好，适用于长春西汀临床生物样品检测及生物等效性研究。

Abstract:

OBJECTIVE To establish and validate a UPLC-MS/MS for the determination of vinpocetine and apovincaminic acid in human plasma and apply to the determination of clinical samples. METHODS Vinpocetine-d5 and apovincaminic acid-d4 were used as internal standards. Human plasma samples were prepared with acetonitrile to precipitate to protein. The chromatographic column was Waters ACQUITY UPLC^® BEH C₁₈(2.1 mm×50 mm, 1.7 μm), the mobile phase was acetonitrile-water(containing 0.05% formic acid). Gradient elution with a flow rate of 0.35 mL·min^-1. Mass spectrometry conditions: Electrospray ionization source(ESI), positive ion detection mode. RESULTS The linear range were 0.04-20.0 ng·mL^-1(r=0.999 7) for vinpocetine and 0.50-250 ng·mL^-1(r=0.999 6) for apovincaminic acid, and the limit of quantification were 0.01, 0.10 ng·mL^-1, respectively. The recoveries of test sample and internal standard were in the range of 94.81%-105.0%, and the matrix effect were 94.51%-105.0%, RSDs were <5%. The RSDs of intra- and inter-batch precision were all <10%. CONCLUSION The method is specific, rapid, accurate and reproducible. Therefore, it is suitable for the determination of vinpocetine concentration in plasma and can applied to its bioequivalence study.