A comparative study on immobilization of urease on different matrices

Abstract

Activity and stability of immobilized urease on different matrices (nylon-6, 6 beads, sepharose gel, silica gel and gelatin film coated on cellulose acetate membrane) have been investigated. Different covalent coupling methods using ascorbic acid or glutaraldehyde for polyamine matrices and periodate for polyhydroxy matrix and tosyl chloride for silica gel were used. A relatively less expensive source of urease (crude extract of jack bean meal) was used. Periodate oxidized sepharose CL 6B gel was able to retain 74% of the enzyme on immobilization while ascorbic acid coupling to gelatin film retained 66%. Immobilized urease on nylon beads and gelatin film stored in 50 mM phosphate buffer (pH 8) at 4°C showed practically no leaching of the enzyme retaining activity over a period of 30 days, while urease coupled to sepharose and silica gel stored under the same conditions was stable over a period of 10 days. Immobilized urease retained activity over a wide pH range and was more stable than the free enzyme at lower and higher pH values. Immobilized urease was also more stable than the free enzyme over a wide temperature range. Nylon bead immobilized jack bean urease had a higher Km (0.62 mM) than that of the soluble enzyme (0.55mM). Kinetic studies on the hydrolysis of urea using immobilized urease in batch and fixed bed flow reactor configurations have been carried out.