运载双层抗原蛋白的CS-PLGA制剂制备及其表征鉴定  

The preparation and characterization of CS-PLGA preparation carrying bilayer antigen protein

作　　者：郑蕊 杨晓岚 王慧 罗德炎 Zheng Rui;Yang Xiaolan;Wang Hui;Luo Deyan(School of Basic Medical Science,Anhui Medical University,Hefei 230032;State Key Laboratory of Pathogen and Biosecurity,Institute of Microbiology and Epidemiology,Academy of Military Medicine,Academy of Military Science,Beijing 100071)

机构地区：[1]安徽医科大学基础医学院,合肥230032 [2]军事科学院军事医学研究院微生物流行病研究所病原微生物生物安全全国重点实验室,北京100071

出　　处：《安徽医科大学学报》2024年第7期1213-1218,共6页Acta Universitatis Medicinalis Anhui

基　　金：病原微生物生物安全国家重点实验室自主研究课题(编号:SKLPBS2220)。

摘　　要：目的制备运载双层抗原的CS-PLGA微球制剂,并对其进行相关表征鉴定。方法通过复乳法结合溶剂挥发法制备得到多孔微球后,在4℃水浴条件下装载抗原,利用抗原浓度梯度介导的物理扩散促进抗原进入微球内部,并通过静电作用结合在微球外表面,形成内外双层抗原运载。根据聚乳酸-羟基乙酸共聚物(PLGA)材料玻璃转化温度这一特性,促进多孔微球表面孔道在48℃条件下发生愈合,使其形成闭合的微球制剂,再将得到的微球制剂与壳聚糖溶液混悬镀层,进行阳离子修饰,逆转微球表面负性电位。通过扫描电子显微镜、动态光散射粒度仪等检测微球形态、粒径分布和电位变化,采用十二烷基硫酸钠聚丙烯酰胺凝胶电泳(SDS-PAGE)鉴定抗原是否装载至微球制剂中,采用荧光标记的BSA和荧光标记的PLGA材料进行激光共聚焦显微镜观察抗原装载后的分布情况,通过BCA法检测微球制剂的包封率和载药率。结果扫描电镜和光学显微镜结果显示多孔微球成孔良好,粒径大小为(73.94±0.81)nm,多分散性指数为0.038±0.004。Zeta电位由负转正说明壳聚糖已被成功镀层至微球表面。经过SDS-PAGE、激光共聚焦显微镜等证实BSA已被成功运载。经micro BCA试剂盒检测后多孔微球包封率为(3.01±0.04)%,载药率为(1.50±0.02)%。结论成功制备得到运载双层抗原的CS-PLGA制剂,为后续缓控释制剂研究提供新思路。Objective To prepare and identify CS-PLGA microspheres carrying bilayer antigen protein.Methods Porous microspheres were prepared by emulsion polymerization combined with solvent evaporation.The antigen was loaded in a water bath at 4℃,and the antigen was promoted to enter the microspheres by physical diffusion mediated by antigen concentration gradient and combined with the outer surface of the microspheres by electrostatic action,forming an inner and outer double-layer antigen carrier.Then,according to the glass transition temperature of PLGA material,the pores on the surface of porous microspheres were promoted to heal at 48℃to form a closed microsphere preparation,and then the obtained microsphere preparation was suspended with chitosan solution for cationic modification to reverse the negative potential on the surface of microspheres.In this study,the morphology,particle size distribution and potential changes of microspheres were detected by scanning electron microscope and dynamic light scattering particle size analyzer.In addition,sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE)was used to identify whether the antigen was loaded into the microsphere preparation.The fluorescent labeled BSA and fluorescent labeled PLGA materials were used to observe the distribution of the antigen after loading by laser confocal microscope.The encapsulation efficiency and drug loading rate of the microsphere preparation were detected by BCA method.Results The results of scanning electron microscope and optical microscope showed that the porous microspheres had good pore formation,the particle size was(73.94±0.81)nm,and the Polydispersity index was 0.038±0.004.Zeta potential changed from negative to positive,which indicated that chitosan had been successfully coated on the surface of microspheres.SDS-PAGE,laser confocal microscope and other detection methods confirmed that BSA had been successfully carried.The encapsulation rate of porous microspheres was(3.01±0.04)%and the drug loading rate was(1.5

关 键 词：聚乳酸-羟基乙酸共聚物 壳聚糖 多孔微球 抗原装载 自愈合 

分 类 号：R318.08[医药卫生—生物医学工程]