响应面法优化米曲霉蛋白酶的双水相萃取条件  

Optimization of Aqueous Two-phase System for Protease Extraction from Aspergillus oryzae by Response Surface Methodology

作　　者：黄福如[1] 罗颖飞 张敏[1] 杨海龙[1] 

机构地区：[1]温州大学生命与环境科学学院,浙江温州325035

出　　处：《核农学报》2014年第10期1860-1866,共7页Journal of Nuclear Agricultural Sciences

基　　金：浙江省科技计划项目(2012C22028)

摘　　要：为获得一种快速纯化米曲霉(Aspergillus oryzae,A.oryzae)蛋白酶的方法,采用聚乙二醇(polyethylene glycol,PEG)/盐双水相系统,从米曲霉固态发酵粗提液中萃取蛋白酶,考察PEG分子质量、盐、PEG浓度、(NH4)2SO4浓度、pH值、粗酶液加入量对蛋白酶萃取的影响,并以Box-Behnken试验设计结合响应面分析法优化了萃取条件。结果表明,选用PEG/(NH4)2SO4双水相系统的最佳提取条件为:PEG-600浓度25%,(NH4)2SO4浓度24.82%,pH值6.97,粗酶液加入量为24.0%,在此条件下蛋白酶的提取率94.74%。双水相法为纯化米曲霉A.oryzae蛋白酶提供了一种有效的方法。To obtain a convenient method to purify protease, protease was extracted by polyethylene glycol （PEG）/salt aqueous two-phase system from the crude extracting solution of Aspergillus oryzae, which was cultured by solid-state fermentation. The effects of several parameters, such as phase forming polymer molecular weight, the types of salt, concentration of phase forming components [ （PEG, （NH4） 2SO4, crude enzyme solution] , pH of the buffer solution on the extraction rate, partition coefficient and purification factor of protease were studied, and the extracting condition was optimized by response surface methodology based on Box-behnken experiment design. The results showed that the optimal extracting conditions were constituted by 25% of PEG - 600, 24.82% of （ NH4 ） 2SO4, 24.0% of crude enzyme solution and pH 6.97. The extraction rate was 94.74% under the optimal conditions. Thus the aqueous two-phase system [ PEG/（ NH4） 2SO4 ] provides an effective method for separation of protease from A. oryzae.

关 键 词：双水相 萃取 米曲霉 蛋白酶 响应面 

分 类 号：TS201.3[轻工技术与工程—食品科学] TQ028.32[轻工技术与工程—食品科学与工程]