Methods in carbohydrate chemistry. Volume 4.

Shearing experiments were conducted in a stirred tank reactor with 0.1% lipase solutions of Candida cylindracea. Inactivation of the lipase solutions were observed at various shear rates from 50 to 150 s(-1) after continuous shearing for ca. 30-240 min under optimal pH and temperature conditions. However, there was no shear stress denaturation of the lipase when it was subjected to shear stresses of 0.72-109.2 kg/m/s(2) and shear rate of 100 s(-1). In the presence of polypropylene glycol, the rate of denaturation of the lipase decreased by 93%. When the lipase solution was filled to the brim, the rate of denaturation of the lipase decreased by 97% compared to that when reactor was half-filled. The rate of denaturation of the lipase decreased by 61% when probes in the fermentor were removed. There was no significant difference in the rate of denaturation of the lipase under ambient conditions compared with that in the absence of oxygen, or in the absence of free metal ions. Recovery of lipase activity from the first hour of shearing was observed at a shear rate of 150 s(-1). The native lipase and the lipase which had recovered its activity showed similar pH profiles, temperature profiles, and activation energies. Temperature was found to have no effect in the rate of shear-induced denaturation of the lipase in the range 20 to 30 degrees C during shearing at 100 s (-1)and optimal pH. Above 30 degrees C, the rate of denaturation of the lipase increased drastically as a function of temperature. The significance of the findings in the de sign of reactor systems for hydrolysis or esterification of oils by lipase will be discussed.

The kinetics and mechanism of shear inactivation of lipase from Candida cylindracea

The current invention relates to detergent compositions comprising one or more enzymes which are immobilized by a covalent binding on an activated polymer. Preferably the polymer is polyethylene glycol.

Detergent compositions comprising immobilized enzymes

Method of the removal of lignin and reuse of cellulases for a continuous saccharification of lignocelluloses were investigated. Only lignin could be separated from hydrolysates by differences in the settling velocity; it was removed from the saccharification process by flocculation with chitosan without loss of cellulases. The ultra-filtration membrane PM10 (Amicon) could be used for recovery of cellulases, but the membrane UH-1 (Toyo Roshi) was better for this purpose, because no cellulases leaked from the membrane, and the amount of cellulase adsorbed to the membrane was less. The cellulases were inactivated by vigorous agitation of the solution in an ultra-filtration device. The loss of cellulase activity by such agitation increased with agitation time, but could be controlled by recovery at a low speed of agitation, so the cellulases could be reused.

Removal of lignin and reuse of cellulases for continuous saccharification of lignocelluloses.

Immobilized cellulases for cellulose utilization

Aspergillus niger cellulase was imobilized on cyanogen bromide activated dextran of varying molecular weights. The effect of different concentrations of cyanogen bromide used for the activation process was also studied. About 50% conjugation and 70% retention activity was achieved in the immobilized cellulase. The pH activity of immobilized enzyme was unchanged, but exhibited more stable activity at acidic pH than the free enzyme. Higher resistance to heat inactivation was also observed.

Cellulase immobilized on a soluble polymer

Glucoamylase (α-1, 4 Glucan glucohydrolase, Exo1, 4, α-glucosidase E.C. 3.2.1.3.) from Rhizopus niveus was immobilized to cyanogen bromide-activated dextran. The glucoamylase-dextran conjugate was purified by gel filtration chromatography. It was found that 92% of the original protein of the enzyme was recovered in the conjugate and about 82% of the specific activity of the native enzyme retained. Some properties of the enzyme conjugate were investigated and compared with those of the free enzyme. The pH activity and pH stability of the immobilized enzyme were not much altered with the optimum pH at 5.5 as that of the free enzyme. Immobilized enzyme exhibited a wide range of temperature stability of 50-60°C as compared to the free enzyme. Heat inactivation at 55°C had less effect on the residual activity of the immobilized enzyme in which 60% activity was retained after prolonged heating for 60 minutes.

Chakamas Wongkhalaung

Papers

Cellulase immobilized on a soluble polymer

Properties of Glucoamylase Immobilized on Soluble Polymer