尿刊酸偶联壳聚糖基因载体的合成和表征  

Synthesis and characterization of urocanic acid-coupled chitosan as gene vector

作　　者：李夷平[1] 姚静[1] 周建平[1] 王伟[1] 

机构地区：[1]中国药科大学,江苏南京210009

出　　处：《药学学报》2008年第12期1233-1238,共6页Acta Pharmaceutica Sinica

基　　金：国家自然科学基金资助项目(30672548)

摘　　要：本文将活化的尿刊酸(urocanic acid,UA)与壳聚糖(chitosan,CTS)上的氨基反应合成一种新型非病毒基因载体——尿刊酸偶联壳聚糖(urocanic acid-coupled chitosan,UAC),通过红外(FT-IR)、核磁共振(1H NMR)和元素分析对UAC结构进行确证。用正交实验对取代度的影响因素进行考察,结果表明,随着UA投料量的增加和活化时间的延长,UAC的取代度也随之增大;通过琼脂凝胶电泳阻滞实验分析UAC/质粒DNA(plasmid DNA,pDNA)的复合能力及对DNase I酶抗降解能力,结果证明UAC与pDNA有较好的复合能力和抵制DNase I酶降解能力;通过测定UAC/pDNA复合物粒径和对zeta电位的分析,结果证明复合物具有良好的稳定性和易于进入细胞的性质;体外细胞转染是通过荧光显微镜观察UAC介导pDNA在体外培养HepG2细胞中的表达,结果显示,UAC能介导pDNA转染HepG2细胞并在细胞中表达绿色荧光蛋白,其转染效果较CTS明显增强。因而UAC是一种制备工艺简单,具有应用前景的非病毒基因载体。A new nonviral gene vector—— the reaction of the activated urocanic acid （ structure of UAC was confirmed with FT-IR, substitution values were studied by orthogonal -- urocanic acid-coupled chitosan （UAC） was prepared by UA） with the amine group on the chitosan （CTS）. The ^1H NMR and element analysis. The influencing factors of test, and the substitution values of UAC increased with the prolongation of activating time of UA and the increasing ratio of UA to CTS. The condensation ability and the resistance to DNase I of UAC/pDNA were evaluated by agarose gel electrophoresis, and UAC showed good condensation ability with pDNA, well protecting pDNA from the degradation by DNase I. The particle size and zeta potential were evaluated by zetasizer, and the results showed that the UAC/pDNA complex was well stable and could easily enter into cells. The transfection studies were performed with HepG2 cells in vitro. It showed that the in vitro transfection of UAC/pDNA was efficient in HepG2 cells and could express more green fluorescent proteins than that of CTS. So the UAC is easy to prepare and a promising non-viral gene vector.

关 键 词：尿刊酸偶联壳聚糖 表征 凝胶电泳阻滞 UAC/pDNA 转染效率 

分 类 号：R943[医药卫生—药剂学]