ATWLPPR配体修饰的载多肽药物PEDF免疫脂质体的制备和表征初步研究

Prelimanary study of the preparation and character of immunoliposome loading polypeptide PEDF with ATWLPPR as ligand modified on the surface

目的:研究表面键合ATWLPPR配体的载多肽药物PEDF免疫脂质体的制备方法,并对其性质进行初步研究。方法:采用逆向蒸发法制备空白脂质体,通过羧基活泼酯化,利用带末端羧基的磷脂酰乙醇胺(PE)的聚乙二醇衍生物(DSPE-PEG-COOH)将配体寡肽ATWLPPR共价键合到脂质体上,制备空白免疫脂质体,采用pH梯度法装载PEDF;HPLC测量配体结合率;用PEDF试剂盒间接测定PEDF免疫脂质体的包封率;并采用相应方法分析免疫脂质体的形态、平均粒径、表面电荷及体外稳定性等药剂学性质。结果:制备了表面键合ATWLPPR的载多肽药物PEDF免疫脂质体,平均配体结合率59.85%;平均PEDF包封率74.6%;平均粒径357.5nm;表面电荷-46.6mV;平均24h体外PEDF漏出率0.99%。结论:尝试采用pH梯度法成功制备了载多肽药物PEDF免疫脂质体,其药剂学性质稳定,为实现PEDF体内以新途径主动靶向传输到视网膜脉络膜新生血管发挥临床作用奠定了实验基础。

OBJECTIVE To study the preparation method and the character of immunoliposome loading polypeptide with ATWLPPR as ligand linked on the Surface. METHODS Use reverse-phase evaporation method to prepare blank liposome. To prepare blank immunoliposome, oligopeptide ATWLPPR was covalently linked to the blank liposome using DSPE-PEG-COOH as spacer. And PEDF was loaded into the blank immunoliposome by pH gradient methods. HPLC was used to determine ligand binding efficiency. PEDF kit was used to indirectly determine the envelop efficiency of PEDF immunoliposome. Using corresponding methods to analyze the appearance , the mean diameter, the surface charge, and the in vitro stability of the immunoli posome. RESULTS PEDF loading immunoliposome with ATWLPPR linked on the surface was successfully prepared. The average ligand binding efficiency was 59. 85%. The average envelop efficiency was 74. 60%. The mean diameter was 357. 5 nm. The average surface charge was - 46. 6 mV. The 24 h leakage of PEDF in vitro was 0. 99%. CONCLUSION The pH gradient method was tried, and the polypeptide loading immunoliposome was prepared successfully, and the pharmaceutical character was stable. This experiment settled a fundation for the objective of using novel method to active targeting deliver PEDF to retina choroidal neovascularization.