EXTRACTION, PURIFICATION AND UTILIZATION OF PROTEOLYTIC ENZYME FROM THE LATEX OF Calotropis gigantea IN FRESH CHEESE MAKING

Abstract

The main objective of the present study was extraction, purification and utilization of proteolytic enzyme from the latex of Calotropis gigante in fresh cheese making. The crude extract was subjected to TPP in varying optimization parameters of ammonium salt concentration (30-80%), ratio of t-butanol to crude extract (1;0.5-1:2) and the extraction pH (4-9). Both intermediate phase and aqueous phase were analyzed using milk clotting activity, protein content and caseinolytic activity for maximum activity recovery (%) and purification fold. Furthermore, the optimum temperature and pH of milk in the cheesemaking, for maximum and minimum time of coagulation (TOC), were determined by response surface methodology. The physicochemical properties of prepared cheese were compared with rennet cheese. For the highest activity recovery and purification fold, the optimum condition parameters for TPP were found to be 60% ammonium sulphate precipitation, 1:1.25 ratio of crude extract to t-butanol (v/v) and extraction pH 6. Numerical optimization study revealed that the optimum temperature and pH for milk clotting in cheesemaking was found to be 55°C and pH 6 respectively. The optimized TOC and MCA were 12.91 s and 2779.65 units respectively. The analysis of rennet cheese and aank protease coagulated cheese revealed no significant difference (P>0.05) in fat, protein and ash content. There was also no significant difference in yield of both cheeses.