采用三相法（TPP）分离纯化番薯过氧化物酶（SPP），以番薯过氧化物酶的酶活回收率和纯化倍数为优化目标，分别利用单因素法和响应面分析法对其影响因素进行考察。建立了最佳提取条件：以叔丁醇作溶剂、硫酸铵饱和度为66%、粗提物与叔丁醇体积比为1.0∶1.13、温度为26℃、pH=5.0，在最优提取条件下，其酶活回收率和纯化倍数分别达133.5%和3.5。SDS-PAGE电泳分析结果显示SPP酶蛋白主要为一条带，其分子质量约为40 kDa左右。三相法操作简便、耗时短、成本低，此外分离过程增强了SPP酶与底物的亲和力，提高了酶活回收率，有利于工业化应用。

Sweet potato peroxidase (SPP) is separated and purified by three-phase partitioning (TPP) method.The recovery rate and purification ratio of sweet potato peroxidase are optimized by single factor method and response surface analysis.The optimal extraction conditions are established as follows:t-butyl alcohol is used as solvent,the saturation of ammonium sulfate is 66%,the volumetric ratio of crude solution to t-butyl alcohol is 1.0:1.13,the temperature is set at 26℃ and pH is 5.0.Under these optimal conditions,the purification efficiency of sweet potato peroxidase can reach 3.5 times and the activity recovery can be 133.5%.It is shown by SDS-PAGE that SPP enzyme protein is a single band with a molecular weight of around 40 kDa.TPP is a simple,quick,low cost and very attractive process for primary purification of SPP and has a big potential use for industrial applications.

蔡燕(1983-),女,博士,副教授,研究方向为生物技术,yancai2010@ntu.edu.cn