地特胰岛素高分子蛋白质定性和定量分析

目的： 探索和建立地特胰岛素高分子蛋白质的定性和定量分析测定方法。 方法： 采用Tricine SDS-PAGE凝胶电泳有效地分离地特胰岛素中的高分子蛋白质组分，确定高分子蛋白质的主要组成形式；采用分子排阻色谱法分离地特胰岛素单体以及高分子蛋白质，并对高分子蛋白质进行准确定量。 结果： 确定了地特胰岛素中高分子蛋白质的主要形式为二聚体和寡聚体，建立的分子排阻色谱法可准确地测定地特胰岛素中高分子蛋白质的含量，并考察了高分子蛋白质产生的主要因素以及国内外产品的高分子蛋白质含量。 结论： 建立的方法可快速有效地对地特胰岛素高分子蛋白质进行定性和定量分析，可作为地特胰岛素的质量研究和质控分析方法。

Objective: To explore and establish the method of qualitation and quantitation of high molecular weight proteins in insulin detemir. Methods: The components of high molecular weight proteins were well separated and the form of the proteins was determined by Tricine SDS-PAGE gel electrophoresis.The high molecular weight proteins were separated with insulin detemir monomer and accurately determined using size exclusion chromatography. Results: The main forms of high molecular weight proteins in insulin detemir were determined to be dimer and oligomer.The content of high molecular weight proteins was determined accurately using the established size exclusion chromatography.The main factor of high molecular weight proteins and the difference of quality between domestic and imported products were also investigated. Conclusion: The established methods are fast and effective for qualitative and quantitative analysis of high molecular weight proteins in insulin detemir,and can facilitate specification research and quality control of insulin detemir.

[1] Pavli-Renar,Praek M,Djoi M.Insulin detemir—a novel basal insulin[J].Diabetologia Croatica,2003,32(4):163
[2] ChP 2010.Vol Ⅱ(中国药典2010年版.二部)[S].2010:566,568
[3] Donald DK,Kelly MW,Elva A.Resolution of high molecular weight glutenin subunits by a new SDS-PAGE system incorporating a neutral pH buffer[J].Cereal Chem,1998,75(1):70
[4] CAO Zuo-wu(曹佐武).An effective method of tricine SDS-PAGE for separating the 1 kDa peptide(有效分离1 kDa小肽的Tricine SDS-PAGE方法)[J]. China Biotechnol(中国生物工程杂志),2004,24(1):74
[5] Katakam M,Banga AK.Aggregation of insulin and its prevention by carbohydrate excipients[J].PDA J Pharm Sci Technol,1995,49(4):160
[6] HOU Yu-rong(侯玉荣),YUAN Yao-zuo(袁耀佐),ZHANG Mei(张玫),et al.Determination of high molecular mass impurities in cefuroxime axetil and its preparation by polystyrene-based gel filtration chromatography(高效凝胶色谱法测定头孢呋辛酯及制剂中高分子杂质的含量)[J].Chin J Pharm Anal (药物分析杂志),2012,32(2):267
[7] Robbins DC,Hirshman M,Wardzala LJ,et al.High-molecular-weight aggregates of therapeutic insulin:In vitro measurements of receptor binding and bioactivity[J].Diabetes,1988,37(1):56