血小板细胞膜色谱构建及柱温研究  

Establishment of Platelet Membrane Chromatography and Column Temperature Study

作　　者：李欧 付真真 刘康妮 雷雨甜 李佳瑞 高小康 LI Ou;FU Zhen-zhen;LIU Kang-ni;LEI Yu-tian;LI Jia-rui;GAO Xiao-kang(College of Pharmacy,Hubei Universily of Medicine,Shiyan 442000,China;The Open Project of Hubei Key labortory of Wudang local Chinese Medicine Research,Hubei Universily of Medicine,Shiyan 442000,China;Taihe Hospital Stem Cell Research Center,Shigyun 442000,China)

机构地区：[1]湖北医药学院药学院,湖北十堰442000 [2]湖北医药学院武当特色中药研究湖北省重点实验室,湖北十堰442000 [3]太和医院干细胞研究中心,湖北十堰442000

出　　处：《中国药学杂志》2020年第15期1289-1294,共6页Chinese Pharmaceutical Journal

基　　金：湖北省自然科学基金项目资助(2018CFC868);湖北省教育厅科研计划指导性项H资助(B2017113);湖北省十堰市科技局引导性项目资助(17Y05)。

摘　　要：目的建立血小板细胞膜色谱模型,考察不同温度下抗血小板聚集药物在该色谱柱上的保留行为,模拟药物在正常和发热病理状态下与血小板的相互作用。方法通过物理吸附法.构建血小板细胞膜色谱固定相,湿法装柱,制成血小板细胞膜色谱柱,BCA蛋白浓度测定试剂盒测定蛋白含量,Na^+、K^+-ATPase检测试剂盒测定生物活性,应用该色谱模型,考察色谱柱的特异性及在35.0~42.0℃温度范围下药物的保留特性。结果构建的血小板细胞膜色谱模型其血小板ATP酶活性值为0.214,固定前血小板膜蛋白质量浓度为0.3409 mg·mL^-1,固定后血小板膜蛋白质量浓度为0.0805 mg·mL^-1。3个抗血小板聚集药物氯吡格雷,双嘧达莫和西洛他唑在血小板细胞膜色谱柱和空白硅胶柱上保留特征有较大差异,36.0℃时3个药物在血小板细胞膜色谱柱上的保留时间均为最大值,然后随温度的升高,保留时间均呈下降的趋势。结论成功构建了血小板细胞膜色谱模型,并首次研究了不同温度下抗血小板聚集药物在该色谱模型上的保留特性,模拟了正常和发热体温时抗血小板聚集药物的色谱保留行为。OBJECTIVE To establish a platelet cell membrane chromatographie model and investigate the retention behaviors of anti-platelet aggregation drugs on chromatographie column at different temperatures,and simulate the interactions between drugs and platelets under normal and febrile pathological conditions.METHODS The platelet cell membrane chromatographie stationary phase was constructed by physical adsorption method.The column was packed with wet method.The protein content was determined by BCA proltein concentration assy kit.The biological activity was determined by Na^+、K^+-ATP ase assay kit.The chromatographie model was used to investigate the speeifieity of the column and the retention characleristics of drugs in the lemperalure range of 35.0-42.0℃.RESULTS The activity of platelet ATPase was 0.214,and the concentration of platelet membrane protein was 0.3409 mg·mL^-1 before bonding and 0.0805 mg·mL^-1 after bonding.The retention characteristics of clopidogrel,dipyridamole and cilostazole on platelet cell membrane chromalographie column and blank silica gel column were quite diferent.The retention time of the three drugs on platelet cell membrane chromatographie column was the maximum at 36.0℃,and then decreased with the increase of temperature.CONCLUSION A platelet cell membrane chromatographic model is scessfully constructed,and the retention characteristics of an-tiplatelet aggregales at diferent temperatures are studied for the first time.The chromatographie retention behaviors of antiplatelet ag gregates at normal and febrile body temperalures are simulated.

关 键 词：细胞膜色谱 血小板细胞膜色谱固定相 高效液相色谱法 抗血小板药物 

分 类 号：R917[医药卫生—药物分析学]