Optimization of Protease Enzyme Extraction from Calatropis Procera Using Aqueous Two-Phase Purification

Abstract

Though protease enzyme extraction has mainly been from microbes, plant proteases have been sought out as a supplement to further boost production and overcome the expensive and tedious nature of microbial protease extraction. At present, efficient and easily affordable processes for the partitioning and purification of enzymes that provide ample yield and high purity of the final products have been demanded by most industries. Thus, this study seeks to optimize conditions and experimental procedures to generate a pure, folded, and dimeric enzyme in its maximum yield from Calotropis procera. The aqueous two-phase purification system (ATPS) was used for the extraction process. The factors pH, NaCl, and temperature were confirmed as having a significant impact on the ATPS and therefore were selected for further optimization to maximize yield and purity. The study found that the maximum protease recovery for the leaves was 93%, 77%, and 89% at pH 6, 20°C, and a salt concentration of 0.0 M, respectively. The maximum protease recovery for the bark was 54%, 51%, and 47% at pH 6–8, 10–20°C, and a salt concentration of 0.0 M, respectively. Additionally, it showed that the leaves had the maximum protease purity at pH 8, temperature 10, and salt concentration of 0.2 M, respectively, while the bark had the highest protease purity at pH 8, temperature 30, and salt concentration of 0.2 M, respectively. Thus, the aqueous two-phase purification method is effectively optimized for maximum yield and purity of the protease enzyme extraction from the leaves and bark of the Calotropis procera plant at or around neutral pH, a temperature range of 10–30°C, and low salt concentrations of 0.0 M–0.1 M.