色谱

色谱

• 研究论文 • 上一篇    下一篇

硫代反义寡核苷酸药物FT19有关物质的分析

张宏燕,鲁丹丹,吴利霞,周喆,王升启   

  1. Beijing Institute of Radiation Medicine, Beijing 100850, China
  • 收稿日期:2008-10-27 修回日期:2008-10-27 出版日期:2008-10-27 发布日期:1983-09-25
  • 通讯作者: 王升启
  • 基金资助:
    国家科技攻关项目(2004BA519A35).

Analysis of the related substances in phosphorothioate antisense oligonucleotide FT19

ZHANG Hongyan, LU Dandan, WU Lixia, ZHOU Zhe, WANG Shengqi   

  1. Beijing Institute of Radiation Medicine, Beijing 100850, China
  • Received:2008-10-27 Revised:2008-10-27 Online:2008-10-27 Published:1983-09-25
  • Contact: WANG Shengqi

PDF

652

被引次数

6

摘要： 为了进行硫代反义寡核苷酸药物FT19的质量控制研究,建立了用阴离子交换高效液相色谱(AX-HPLC)和毛细管凝胶电泳(CGE)分析自行合成的FT19有关物质的方法。设计合成了FT19的硫代不完全序列(P=O)1和短序列(n-x)并将它们作为已知杂质。在AX-HPLC上,使用的分析柱为DNA Pac PA-100 (4 mm×250 mm);流动相A为10 mmol/L NaOH-0.1 mol/L NaCl,流动相B为10 mmol/L NaOH-3 mol/L NaCl;梯度洗脱条件为流动相B液在8 min内从60%升至100%;流速1 mL/min;柱温40 ℃;检测波长为260 nm。CGE所用毛细管规格为内径100 μm,总长度为31 cm,有效长度为20 cm;电泳缓冲溶液为三羟甲基氨基甲烷（Tris）-硼酸-7 mol/L尿素,pH 8.5;采用电动进样,进样电压-10 kV;分离胶为250 g/L的聚丙烯酰胺;检测波长为254 nm。结果表明,FT19与硫代不完全的(P=O)1序列在AX-HPLC上能够达到基线分离,与短序列(n-1)在CGE上能达到基线分离。说明AX-HPLC和CGE联合应用能够很好地分析FT19中的有关物质。

关键词: 硫代反义寡核苷酸, 毛细管凝胶电泳, 阴离子交换高效液相色谱, 有关物质

Abstract: A method using anion exchange high performance liquid chromatography (AX-HPLC) and capillary gel electrophoresis (CGE) was established for the analysis of the related substances in phosphorothioate antisense oligonucleotide FT19 for quality control. The FT19 and its analogs of partial phosphodiester compounds (P=O)1 as well as deletion sequences (n-1) were analyzed. AX-HPLC with the DNA Pac PA-100 column (4 mm×250 mm) was used for the separation of phosphorothioate modifications. The size of the capillary column used in the CGE was 31 cm × 100 μm with an effective length of 20 cm. It was found that full phosphorothioate and partial phosphodiester oligonucleotides of the same length were successfully separated, and the deletion sequences (n-1) and the full-length sequence n were successfully separated in CGE. The results demonstrated that the related substances of phosphorothioate oligonucleotides can be well analyzed by AX-HPLC and CGE.

Key words: capillary gel electrophoresis (CGE), phosphorothioate oligonucleotide, related substances , anion exchange high performance liquid chromatography (AX-HPLC)