摘要
目的:采用中心组合设计法优化前阳离子脂质体的制备工艺,考察前阳离子脂质体的有关性质。方法:以薄膜分散膜挤压法制备前阳离子脂质体,以平均粒径、转染效率为指标,对鱼精蛋白(protamine)与质粒DNA的质量比、CHETA([2[[4-[羧甲基]二硫-1-亚氨丁基]氨基]乙基]氨基甲酸胆固醇酯)摩尔百分比浓度、CHETA与质粒DNA的质量比3个自变量的各个水平进行二项式拟合,通过响应面分析选取较佳工艺条件。以LacZ为报告基因转染HepG2细胞,定量测定β-半乳糖苷酶活性和总蛋白含量计算转染效率。结果:影响平均粒径、转染效率的3个主要因子的最佳取值分别为:protamine/DNA,2.5∶1;CHETA(mol%),45;CHETA/DNA,4∶1。优化条件制备的前阳离子脂质体形态近似于球体,平均粒径为(228.9±8)nm,多分散指数为0.122±0.02(n=3),Zeta电位为-(25.08±2.5)mV(n=3),载基因前阳离子脂质体转染HepG2肝癌细胞转染效率为(24.26±2.6)mUβ-半乳糖苷酶/mg protein。结论:中心组合设计应用简便、预测性好,制备的前阳离子脂质体符合设计要求。
Aim: To optimize the preparation of procationic liposomes by using central composite design and investigate the related characteristics of the procationic liposomes. Methods: Blank procationic liposomes were prepared by film dispersion-fiheration. The effects of protamine/DNA ratio ( w/w ), CHETA molar percent (mol % ) and CHETA/DNA ratio (w/w) on the mean particle size, and the transfection efficiency were investigated. A second-order polynomial equation was fitted to the data and the resultant model was used to predict the response in the optimal region. Estimation of the transfection efficiency was performed by galactosidase assay in HepG2 cells using LacZ as the reporter gene. Results:All the observed response variables were found to be highly dependent on the formulation variables. It was found that the optimized formulation consisted of protamine/DNA ratio of 2.5 : 1, CHETA molar percent of 45 and CHETA/DNA ratio of 4 : 1. The resultant procationic liposomes had a regular spherical surface with an average size of (228.9 ± 8)nm (PDI = 0. 122 ± 0.02, n = 3)and Zeta potential of - (25.08 ± 2.5)mV ( n = 3). The transfection efficiency of the procationic liposomes in HepG2 cells was (24.26 ± 2.6 ) mU β-galactosidase/rng protein. Conclusion: Central composite design was simple, predictable and the characteristics of the resultant procationic liposomes met with the design requirements.
出处
《中国药科大学学报》
CAS
CSCD
北大核心
2007年第2期115-119,共5页
Journal of China Pharmaceutical University
基金
国家自然科学基金资助项目(No.30371697)~~
关键词
中心组合设计
前阳离子脂质体
鱼精蛋白
转染效率
central composite design
procationic liposomes
protamine
transfection efficiency