双水相萃取结合液相色谱法分离蛋白质  

Separation of Proteins by Aqueous Two-phase Extraction System Combined with Liquid Chromatography

作　　者：赵新颖[1,2] 屈锋[1] 董敏[1] 陈凡[1] 罗爱芹[1] 张经华[2] 

机构地区：[1]北京理工大学生命学院,北京100081 [2]北京市理化分析测试中心,北京100089

出　　处：《分析化学》2012年第1期38-42,共5页Chinese Journal of Analytical Chemistry

基　　金：国家重点基础研究发展计划(No.2007CB914101);国家自然科学基金(No.20875009)资助项目

摘　　要：建立了PEG/(NH4)2SO4双水相体系萃取富集,结合液相色谱分离分析多种蛋白质的方法。考察了无机盐种类和浓度、PEG分子量、pH值和温度等因素对双水相形成以及对细胞色素C、肌红蛋白、牛血清白蛋白、溶菌酶、胰蛋白酶分配行为的影响。结果表明,上述5种蛋白在室温、pH 3.5～9.0范围内,可在15%PEG-4000/10%(NH4)2SO4双水相体系中得到富集,且主要集中在下相。同样条件下,血清中的高丰度蛋白在上下相均有分配,下相分配量较大。通过双水相萃取分离蛋白质及对液相色谱一定时间段的色谱峰收集,可初步实现血清中高丰度蛋白质的分离去除。A combination of aqueous two-phase extraction system（ATPS） of polyethylene glycol（PEG）/（NH4）2SO4 with high performance liquid chromatography（HPLC） was proposed for enrichment and separation of proteins.The effects of inorganic salt,concentration,PEG molecular weight,pH,temperature and NaCl on the formation of two phases and partition of cytochrome C,lysozyme,trypsin,bovine serum albumin and myoglobin were investigated,respectively.The results showed that five proteins were enriched mainly in bottom phase in 15% PEG-4000/10%（NH4）2SO4 solution at room temperature and pH range from 3.5 to 9.0.High abundance proteins in serum also distributed in two phases and the bottom phase had larger amount at the same conditions,which could be extracted and separated by optimized ATPS.The proteins were collected based on their retention time in HPLC,and could be removed from serum samples.

关 键 词：双水相萃取 蛋白质分离 血清 

分 类 号：Q51[生物学—生物化学]