Chemistry Department, Faculty of Science and Arts, Adnan Menderes University, Aydin, Turkey.
Appl Biochem Biotechnol. 2012 Jun;167(3):552-63. doi: 10.1007/s12010-012-9707-7. Epub 2012 May 10.
Supermacroporous poly(2-hydroxyethyl methacrylate) [poly(HEMA)] monolithic cryogel was prepared by radical cryocopolymerization of HEMA with N,N'-methylene bisacrylamide as crosslinker. Reactive Green 5 dye was immobilized to the cryogel with nucleophilic substitution reaction, and this dye attached cryogel column was used for affinity purification of papain from Carica papaya latex. Reactive Green 5-immobilized poly(HEMA) cryogel was characterized by swelling studies, Fourier transform infrared spectroscopy, scanning electron microscopy, and energy dispersive X-ray analysis. Maximum papain adsorption capacity was found to be 68.5 mg/g polymer while nonspecific papain adsorption onto plain cryogel was negligible (3.07 mg/g polymer). Papain from C. papaya was purified 42-fold in single step with dye attached cryogel, and purity of papain was shown by silver-stained sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
采用自由基冻共聚的方法，以 N,N'-亚甲基双丙烯酰胺为交联剂，制备了大孔聚（2-羟乙基甲基丙烯酸酯）[聚（HEMA）]整体冰晶。通过亲核取代反应将活性绿 5 染料固定在冰晶上，并将该染料固定的冰晶柱用于从木瓜乳中亲和纯化木瓜蛋白酶。通过溶胀研究、傅里叶变换红外光谱、扫描电子显微镜和能量色散 X 射线分析对负载活性绿 5 的聚（HEMA）冰晶进行了表征。最大的木瓜蛋白酶吸附容量为 68.5mg/g 聚合物，而普通冰晶对木瓜蛋白酶的非特异性吸附可以忽略不计（3.07mg/g 聚合物）。用负载染料的冰晶在单步纯化中对木瓜蛋白酶进行了 42 倍的纯化，通过银染的十二烷基硫酸钠-聚丙烯酰胺凝胶电泳显示了木瓜蛋白酶的纯度。
