HPLC测定人血浆乙酰水杨酸酯酶活性及其测定培养条件的优化选择

Determination of the Activity of Aspirin Esterase in Plasma by HPLC Method and Optimization of the Incubated Conditions in Vitro

目的建立测定血浆中乙酰水杨酸和水杨酸浓度的高效液相色谱法,采用水杨酸/（乙酰水杨酸＋水杨酸）表示乙酰水杨酸酯酶的活性,并对乙酰水杨酸酯酶体外培养条件进行了优化,测定血浆乙酰水杨酸酯酶活性可为临床合理用药提供科学依据。方法采用反相高效液相直接进样法测定血浆中乙酰水杨酸和水杨酸的浓度,并用正交实验设计分析法对其体外培养条件进行优化选择。结果乙酰水杨酸在0.08-5.12mg·L^-1内线性关系良好,水杨酸在0.05-4mg·L^-1内线性关系良好,体外最佳培养条件是A2B2C3D1,即以1mmol·L^-1乙酰水杨酸,200μL血浆,在pH为7.6的培养液中培养20min。结论本法操作简便、灵敏、快速,适用于血浆中乙酰水杨酸和水杨酸的浓度测定。

OBJECTIVE To develop a HPLC method for the determination of aspirin and salicylic acid in plasma, and use the ratio of salicylic acid/（ aspirin ＋ salicylic acid）to express the activity of aspirin esterase, and optimize the incubated conditions of aspirin esterase. METHODS The contents of aspirin and salicylic acid in the plasma were determined by RP-HPLC method. Orthogonal experimental design was used to optimize the incubated conditions of aspirin esterase. RESULTS Calibration curves of aspirin were linear in the range of 0. 08 - 5. 12 mg·L^-1（ r = 0. 999 5 ）. Calibration curves of salicylic acid were linear in the range of 0. 05 - 4 mg·L^-1 （ r = 0. 999 8 ）. The optimal incubated conditions were abtained that 1 mmol ·L^-1 aspirin and 200 μL plasma were incubated for 20 min in pH 7.6 incubation buffer. CONCLUSION The method is simple, accurate and rapid, suitable for the determination of aspirin and salicylic acid in plasma. The method can be used to assay the activities of aspirin esterase.