毛细管区带电泳法测定艾塞那肽纯度及在稳定性研究中的应用

目的: 建立毛细管区带电泳(CZE)方法测定艾塞那肽纯度,并初步考察其稳定性. 方法: 采用未涂层熔融石英毛细管(48.5 cm×50 μm,有效长度40 cm),考察了运行缓冲液pH及浓度、分离电压、温度、添加剂乙腈浓度对分离效果的影响,最终确立电泳条件:运行缓冲液为200 mmol·L^-1 磷酸二氢钠-磷酸氢二钠溶液(pH 6.2;含15%乙腈),分离电压为20 kV,温度 25℃,检测波长214 nm,压力进样5 kPa×10 s. 结果: 艾塞那肽质量浓度在0.05~4.0 mg·mL^-1(r=0.999 9)范围内线性关系良好; 检测限为3.3 μg·mL^-1,定量限为8.0 μg·mL^-1;主峰迁移时间的日内和日间RSD分别为1.07%(n=6)和0.91%(n=9), 纯度的日内和日间RSD分别为0.08%(n=6)和0.10%(n=9).该方法与HPLC法比较,纯度测定结果基本一致,本法分离效果更好,分析时间缩短,能有效检查艾塞那肽的纯度. 结论: 该方法可用于艾塞那肽的质量控制和稳定性研究.

Objective: To develop a method of capillary zone electrophoresis (CZE) for determination of the purity of exenatide and conduct a preliminary study on its stability. Methods: An uncoated silica capillary (48.5 cm×50 μm,40 cm to the detector) was used.A number of separation parameters were investigated,including pH,concentration of the running buffer,separation voltage,temperature and concentration of additive acetonitrile.The final conditions were as follows.The running buffer was 200 mmol·L^-1 sodium dihydrogen phosphate-disodium hydrogen phosphate (adjusted to pH 6.2) containing 15% acetonitrile.The separation voltage was 20 kV and the temperature was 25℃.The detection was monitored at 214 nm.The sample was injected under pressure of 5 kPa for 10 s. Results: The calibration curve of exenatide was linear in the concentration range of 0.05-4.0 mg·mL^-1(r=0.999 9),and the limits of detection and quantitation were 3.3 μg·mL^-1 and 8.0 μg·mL^-1.The intra-and inter-day RSDs of the migration time of main peak were 1.07%(n=6) and 0.91%(n=9),and the intra-and inter-day RSDs of the purity were 0.08% (n=6) and 0.10% (n=9).The purity of exenatide was effectively determined by CZE and the results of the CZE method were in accord with the HPLC method.Moreover,the resolution was better and the migration time was shorter by CZE. Conclusion: The method can be applied in the quality control and stability study of exenatide.