硫代反义寡核苷酸药物癌泰得纯化工艺的建立  

Purification of phosphorothioate antisense oligonucleotide Cantide

作　　者：张嵬[1] 梁乾德[1] 王晓博[1] 丁雨[1] 林汝仙[1] 王升启[1] 

机构地区：[1]军事医学科学院放射与辐射医学研究所,北京100850

出　　处：《中国药学杂志》2005年第15期1188-1192,共5页Chinese Pharmaceutical Journal

基　　金：国家重大科技专项(2002AA2Z3337)国家自然科学基金(3017111)

摘　　要：目的建立硫代反义寡核苷酸药物癌泰得的大规模纯化方法。方法利用DMT的疏水性,在SOURCE15Q为填料的离子柱上预先用高盐缓冲液洗脱不带DMT基团的合成失败序列,然后直接在柱上用0.4%TFA切割,脱DMT保护后,使用盐浓度梯度分离硫代不完全片断和少量的(n-x)片断,从而获得纯品。结果以290nm为检测波长,流速为382cm·h-1,载量为12mg·g-1填料的条件下,平均产品收率为83.40%,质量平衡回收率为98.20%,产品终纯度可达98.23%。结论建立的方法能够有效纯化硫代反义寡核苷酸药物癌泰得,并可进行大规模纯化。OBJECTIVE To develop a single step chromatographic method for large-scale purification of phosphorothioate antisense oligonucleotide. METHODS The hydrophobicity of the dimethoxytrityl （DMT） protecting group and the hydrophobicity of SOURCE 15Q media were used for purification .The oligonueleotide with DMT was highly retained by the SOURCE 15Q ，“Trityl off”failure sequences were separated from “trityl on” oligmer when high-salt buffer was used firstly.The trityl group was cleaved from the product by 0.4% trifluoraeetie acid.Finally, a linear gradient was used to separate full-length, full thioated phosphomthioate contents from synthetic failure, products. RESULTSU UV absorption wavelength was selected at 290 nm, linear flow rate was 382 cm·h^-1, sample loading was 12 mg· g^-1 media. Product yield for this .separation was 83.40% at a final purity of 98.23% and the mass balance recovery of all oligonueleotide was up to 98.20%. CONCLUSION A single step anion-exchange chromatographic protocol was developed for purifying phosphomthioate antisense oligonueleotide named“Cantide”.

关 键 词：硫代反义寡核苷酸 癌泰得 高效液相色谱法 

分 类 号：R943.4[医药卫生—药剂学]