离子交换高效液相色谱法分析质粒pUDKH  

Analysis of Plasmid pUDKH by Anion-Exchange High Performance Liquid Chromatogaphy

作　　者：刘佳[1] 张庆林[1] 毕建进[1] 王晓娜[1] 龚萍[1] 哈小琴[1] 王正平 吴祖泽[1] 

机构地区：[1]军事医学科学院放射医学研究所,北京100850

出　　处：《色谱》2004年第1期65-67,共3页Chinese Journal of Chromatography

基　　金：国家"863"课题(课题号:2001AA217061).

摘　　要：采用阴离子交换高效液相色谱法监测重组质粒pUDKH的生产过程和该质粒产物中超螺旋pUDKH的比例。所用的色谱条件为:色谱柱TSKgelDNA NPR(4 6mmi d ×75mm);流动相体系由20mmol/LTris HCl(pH8 8)和1mol/LNaCl组成,梯度洗脱方式。分析结果表明:质粒产物中超螺旋pUDKH含量达95%(质量分数)以上时,检测不到残留的核糖核酸(RNA)。高效液相色谱法检测质粒pUDKH产物及其生产过程的灵敏度高、试样用量少,分辨率高。pUDKH, a new plasmid DNA comprising human hepatocyte growth factor (HGF), was developed as a potential genetherapeutic product for peripheral arterial disease. An anion-exchange high performance liquid chromatographic (HPLC) method was developed to determine the proportion of plasmid DNA in both opened and supercoiled forms. A TSK gel DNA-NPR column was equilibrated with 50% buffer A (20 mmol/L Tris-HCl, pH 8.8) and 50% buffer B (buffer A with 1 mol/L NaCl added). pUDKH was analyzed by using a gradient HPLC with a mobile phase from 50%B to 80%B and with a UV detection at 260 nm. The flow rate was 0.5 mL/min. The method was also used to monitor the production procedures.

关 键 词：重组质粒pUDKH 肝细胞生长因子 离子交换高效液相色谱法 超螺旋 生产过程 肢体动脉闭塞病 基因治疗制剂 质量控制 

分 类 号：R972.9[医药卫生—药品] R927[医药卫生—药学]