Purification of Papain Using Reactive Green 5 Attached Supermacroporous Monolithic Cryogel

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?Cpolyacrylamide gel electrophoresis.