蕹菜过氧化物酶的分离纯化及理化性质  

Isolation, Purification and Characterization of Peroxidase from Ipomoea aquatica Forsk.

作　　者：王红扬[1] 王洁[1] 李星[1] 唐云明[1] 

机构地区：[1]西南大学生命科学学院重庆市甘薯工程研究中心三峡库区生态环境教育部重点实验室,重庆400715

出　　处：《食品科学》2015年第3期166-170,共5页Food Science

基　　金：重庆市科委重点攻关项目(CSTC2011AB1027)

摘　　要：新鲜蕹菜经硫酸铵分级沉淀、DEAE-Sepharose离子交换层析和Superdex-200凝胶过滤层析后获得电泳纯的过氧化物酶(peroxidase,POD),该酶的比活力、回收率、纯化倍数和产率分别为35 972.96 U/mg、12.21%、168.48和211.07 U/g。该酶的亚基分子质量为42.7 kD,最适温度为40 ℃,最适pH值为 6,并且在20～50 ℃及pH 5～8的范围内具有良好的稳定性。以不同浓度的H2O2为底物,测得该酶的Km值为18.32 mmol/L。NaCl、尿素(Urea)、Zn2+、Mg2+对该酶都具有较强的激活作用,十二烷基硫酸钠(sodium dodecyl sulfate,SDS)、硫氰化钾(potassium thiocyanate,KSCN)、抗坏血酸(ascorbic acid,AsA)、Ba2+、Mn2+ 、Fe2+、甲醇、乙醇、正丁醇和异丙醇对蕹菜POD活力均有抑制作用,其中抗坏血酸对POD有极强的抑制作用,当浓度为0.01 mol/L时,酶活力接近于0。Electrophoresis-purity peroxidase (POD) was extracted and purified from Ipomoea aquatica Forsk. through ammonium sulfate precipitation, DEAE-Sepharose and Superdex-200 chromatography. The specific activity of the purified peroxidase was 35 972.96 U/mg and the yield was 211.07 U/g with a recovery rate of 12.21% and a purification fold of 168.48. Its molecular mass was around 42.7 kD as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE). Besides, the POD with the optimal temperature and pH of 40 ℃ and 6, respectively, was relatively stable in the temperature range of 20–50 ℃ and pH range of 5–8. This enzyme showed a Km value of 18.32 mmol/L towards the substrate hydrogen peroxide. In addition, the POD was found to be activated by NaCl, urea, Zn2+and Mg2+ but inhibited by SDS, KSCN, ascorbic acid (AsA), Ba2+, Mn2+, Fe2+, methanol, ethanol, n-butyl alcohol and isopropanol, and completely inactivated by 0.01 mmol/L AsA.

关 键 词：蕹菜 过氧化物酶 分离纯化 酶活力 

分 类 号：Q946.5[生物学—植物学]