High throughput covalent immobilization process for improvement of shelf-life, operational cycles, relative activity in organic media and enzymatic kinetics of urease and its application for urea removal from water samples

摘要

High throughput covalent urease immobilization was performed through the amide bond formation between the urease and the amino-functional MNPs. The enzyme's performances, including shelf-life, reusability, enzymatic kinetics, and the enzyme relative activity in organic media was improved. At optimal conditions, the immobilization efficiency was calculated about 95.0% with keeping 94.7% of the urease initial specific activity. The optimal pH for maximum activity of the free and immobilized urease was calculated as 7.0 at 37.0 degrees C and 8.0 at 60.0 degrees C, respectively. The kinetics studies showed the K-m of 26.0 mM and 8.0 mM and the V-max of 5.31 mu mol mg(-1 )min(-1 )and 3.93 mu mol mg(-1) min(-1 )for the free and immobilized urease, respectively. The ratio K-cat/K(m )as a measure of catalytic efficiency and enzyme specificity was calculated as 0.09 mg(-1 )min(-1) and 0.22 mg(-1) min(-1 )for the free and immobilized urease, respectively, indicating an improvement in the enzymatic kinetics. The shelf-life and operational studies of immobilized urease indicated that approximately 97.7% and 88.5% of its initial activity was retained after 40 days and 17 operational cycles, respectively. The immobilized urease was utilized to urea removal from water samples with an efficiency between 91.5-95.0%.