胰岛素反相制备色谱的方法开发

引用本文: 潘剑, 陶云国. 胰岛素反相制备色谱的方法开发[J]. 色谱, 2017, 35(8): 848-854. doi: 10.3724/SP.J.1123.2017.04014 shu

Citation: PAN Jian, TAO Yunguo. Method development of reversed-phase preparative liquid chromatography for insulin[J]. Chinese Journal of Chromatography, 2017, 35(8): 848-854. doi: 10.3724/SP.J.1123.2017.04014 shu

胰岛素反相制备色谱的方法开发

潘剑 ,
陶云国

海正药业(杭州)有限公司, 浙江杭州 311404

摘要: 以胰岛素反相制备色谱方法的开发和优化为目标，通过考察色谱保留参数、峰展宽及样品流出曲线的浓度分布等色谱参数，对流动相梯度、色谱填料、载样量等色谱条件进行了优化，并建立了胰岛素制备色谱峰参数的描述方法。结果表明，所建立的方法可快速筛选出最适于胰岛素分离的色谱条件（包括流动相梯度及分离填料），即流动相中的强洗脱溶剂（有机相）需采取缓梯度窄区间的变化条件，筛选出的分离填料需具备峰向两侧展宽且展宽程度较小、样品最高浓度居中分布的特点。将方法用于实际胰岛素粗品的纯化制备，获得了杂质去除效果好、胰岛素纯度高的产品。该法为胰岛素反相色谱纯化制备方法的快速建立提供了指导，具有较强的实用价值，同时为发展大分子化合物的制备色谱方法提供了参考。

关键词:

English

Method development of reversed-phase preparative liquid chromatography for insulin

PAN Jian ,
TAO Yunguo

Hisun Pharmaceutical(Hangzhou) Co. Ltd., Hangzhou 311404, China
Received Date: 13 April 2017

Abstract: The development and optimization of reversed-phase preparative liquid chromatography method for insulin were performed. The chromatographic retention, sample loading and peak broadening were investigated. The mobile phase gradient conditions and stationary phases were optimized. The parameters of the method contained the peak broadening levels under different amounts of sample loading and the concentration distribution of the target compound in the elution curves. The parameters of peak broadening levels were defined and expressed as a matrix, which consisted of sample loading, the forward broadening and the backward broadening levels. The most suitable chromatographic system (including stationary phases and mobile phase conditions) was selected. A gradient program with slow change of the strong elution solvent was used. The most suitable stationary phase should exhibit the narrower peak broadening and the peaks were best to broaden to both sides comparing to that under the analytical conditions. Besides, the concentration distribution of the target compound should be focused on the middle of the elution. The guide principles were validation by purification of crude insulin products. The preparative chromatographic system constructed through this method can remove impurities effectively. The method has good practical value. It can provide reference for the development of preparative chromatographic methods of other macromolecular compounds.

Key words:

reversed-phase preparative liquid chromatography
/ chromatographic parameters
/ peak broadening
/ concentration distribution
/ insulin

1. [1] Xiao Y J, Huang L, Cao C L, et al. Progress in Modern Biomedicine, 2013, 13(24):4785肖拥军, 黄亮, 曹春来, 等. 现代生物医学进展, 2013, 13(24):4785
2. [2] Jiang L M, Chen Y, Li H L, et al. Chinese Medicinal Biotechnology, 2013, 8(1):19蒋利敏, 陈勇, 李红亮, 等. 中国医药生物技术, 2013, 8(1):19
3. [3] Polez S, Origi D, Zahariev S, et al. PLoS One, 2016, 11(1):1
4. [4] Liu H F.[PhD Dissertation]. Shanghai:East China University of Science and Technology, 2013刘海峰.[博士学位论文]. 上海:华东理工大学, 2013
5. [5] Guo M L, Milton T W H, Reinhard I B. Acta Biologiae Experimentalis Sinica, 2000, 32(3):265郭敏亮, Milton T W H, Reinhard I B.生物化学与生物物理学报, 2000, 32(3):265
6. [6] Ding L, Guo Z M, Xiao Y S, et al. J Sep Sci, 2014, 37(18):2467
7. [7] Wang C R, Guo Z M, Long Z, et al. J Chromatogr A, 2013, 1281:60
8. [8] Sabharwal A P, Chase H A. Food Bioprod Process, 1999, 77(1):18
9. [9] Vajda P, Bocian S, Buszewski B, et al. J Sep Sci, 2010(23/24), 33:3644
10. [10] Kamarei F, Gritti F, Guiochon G, et al. J Chromatogr A, 2014, 1329:71
11. [11] Gritti F, Guionchon G. J Chromatogr A, 2014, 1356:96
12. [12] Gritti F, Guiochon G. J Chromatogr A, 2005, 1090(1/2):39
13. [13] Gritti F, Guiochon G. J Chromatogr A, 2013, 1282:58
14. [14] Shan Y C, Zhao R H, Zhang W B, et al. Chinese Journal of Chromatography, 2001, 19(3):256单亦初, 赵瑞环, 张维冰, 等. 色谱, 2001, 19(3):256
15. [15] Dai Y T, Jin G W, Qin X M, et al. Chinese Journal of Chromatography, 2009, 27(4):442代云桃, 金高娃, 秦雪梅, 等. 色谱, 2009, 27(4):442
16. [16] Zou H F, Zhang Y K, Lu P Z. High Performance Liquid Chromatography. Beijing:Science Press, 2001邹汉法, 张玉奎, 卢佩章. 高效液相色谱法. 北京:科学出版社, 2001
17. [17] Jin G W, Xue X Y, Zhang F F, et al. Anal Chim Acta, 2008, 628(1):95
18. [18] Jin G W, Guo Z M, Xiao Y S, et al. J Sep Sci, 2016, 39(20):3917
19. [19] Sabharwal A P, Chase H A. Food Bioprod Process, 1998, 76(1):47
20. [20] Shao C, Gao Y H. Chinese Journal of Chromatography, 2010, 28(2):128邵晨, 高友鹤. 色谱, 2010, 28(2):128
21. [21] Pharmacopoeia Commission of the People's Republic of China. Pharmacopoeia of the People's Republic of China, Part 2. Beijing:Chemical Industry Press, 2015:1166国家药典委员会. 中华人民共和国药典, 二部. 北京:化学工业出版社, 2015:1166

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胰岛素反相制备色谱的方法开发

English

Method development of reversed-phase preparative liquid chromatography for insulin

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中国化学会秘书处

胰岛素反相制备色谱的方法开发

English

Method development of reversed-phase preparative liquid chromatography for insulin

CCS Chemistry：嵌段共聚物自组装成 “三明治”二维有机材料，实现纳米级压力传感功能

CCS Chemistry：颜徐州、鲍哲南： 你的皮肤可能是一片SPMs

CCS Chemistry：工作高效不疲劳，只须让催化剂动起来

CCS Chemistry：静电组装导向聚合- 聚电解质纳米凝胶量化制备新策略

CCS Chemistry：金黄色葡萄球菌醛基脱氢酶是如何识别特异性底物的？

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