Decolorization and purification of crude protease from Rhizopus oryzae by activated charcoal and its electrophoretic analysis

Activated charcoal decolorized and partially purified the protease from a c rude extract of solid state fermentation of wheat bran by Rhizopus oryzae. Treatment for 5 min was sufficient. Depending on the initial colour intensi ty of crude, the charcoal to crude extract ratio could be optimized to achi eve 90% decolorization, 85% enzyme recovery, and over a 3-fold purification , even up to 20-fold variation in batch size (from 1 ml to 20 ml crude extr act). Decolorization followed the Freundlich and the Langmuir models, the F reundlich constant, n, being 2.74. Partial purification was confirmed by na tive PAGE and the protease band identified by gelatin-PAGE. SDS-PAGE showed the protease consisted of two sub-units (about 22 and 24 kDa). List of symbols: c(o), initial solute concentration in liquid before adsorp tion; c*, equilibrium solute concentration in liquid after adsorption; k, e mpirical constant for Freundlich adsorption isotherm; U, unit of protease a ctivity; v, volume of solution per unit weight of adsorbent.