Studies on Production, Optimization and Purification of Uricase from Gliocladium viride

Five different fungal strains belonging to Gliocladium and Gliomastix species wereinitially screened for their uricase producing capability, among which Gliocladium virideMTCC 3835 was identified to produce highly active uricase. Statistical designs were appliedto optimize uricase production. Using Plackett–Burman design, peptone, yeast extract andCuSO4 were found to have significant effect on enzyme activity. Box-Behnken design wasused to find the optimal concentrations of significant variables, which were as follows,Peptone-12.71g/L, Yeast Extract-10.57g/L, CuSO4-0.0762g/L. Maximum activity of 84.92 U/mlwas observed experimentally, which is 1.344 times higher than the activity got in basalmedium. Crude uricase was further purified using three-phase partitioning (TPP), and thesignificant factors like inorganic phase saturation, ratio between organic phase and crudeenzyme, operating temperature were optimized. Crude enzyme saturated with 50%(w/v) withammonium sulphate and at crude enzyme to t-butanol ratio of 1:1(v/v) at 30°C resulted in80.109% yield of uricase with 1.44 fold purification.