Abstract
Three-phase partitioning (TPP) was used to recover proteases from the viscera of farmed giant catfish. Different ratios of crude enzyme extract to t-butanol (1.0:0.5, 1.0:1.0, 1.0:1.5, and 1.0:2.0; v/v) and concentrations of ammonium sulfate ((NH4)2SO4) (20, 30, 40, and 50%; w/v) were investigated. The best protease yield (163%) with a 5-fold purification was obtained in the interphase (I-ph) of the 2nd TPP system, which consisted of the crude extract to t-butanol ratio of 1.0:0.5 in the presence of 50% (NH4)2SO4. A higher concentration of (NH4)2SO4 increased the protease recovery at the interphase, whereas an increase in the t-butanol volume decreased the recovery. SDS-PAGE and zymography revealed a substantial level of isolation of proteases from the viscera of farmed giant catfish by TPP. The molecular weight of the major protease recovered was approximately 25kDa, corresponding to trypsin. The present study demonstrates desirable outcomes and could be used as a primary purification process.
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