Department of biological science and engineering Korea advanced institute of science and technology

Abstract

Lipid hydrolysis reaction approached to complete conversion in the presence of isooctane in isooctane-water two phase. Although the lipase was unstable in organic solvent, the stability was improved by the substrate olive oil. The stabilizing effect of the substrate became less significant above 20\% olive iol. For the continuous lipid hydrolysis in a packed-bed reactor using isooctane, candida rugosa lipase was immobilized with DEAE-Sephadex A50, Sephadex G50, Sephadex LH20, Amberlite IRA94, and Amberlite XAD7. Among these, the ionic carrier gave higher activity yield (61\% with DEAE-Sephadex and 14\% with Amberlite IRA) than other carriers (less than 1\%) when assayed in olive oil emulsion substrate. The lipase immobilized on DEAE-Sephadex A50 showed an increased thermal stability. In order to carry out the continuous reaction in a glass column reactor, a strategy to pack the immobilized enzyme into the reactor was developed. By using rhe strategy, immiscible two phase (olive oil dissolved in isooctane and buffer solution) could be evenly distributed through the packed enzyme-carrier without any surfactant supplement. The reactor performance was best with DEAE-Sephadex. The operational stability of DEAE-Sephadex-immobilized enzyme in the reactor was influenced by initial water content of the support material. Also the types of buffer supplemented, ionic strength, pH, temprrature, and substrate concentration were important operating variables for the enzyme stability. Substrate extended the operational half life logarithmically as the concentration was increased. The immobilized enzyme was much unstable above 35$^\circ$C and at 25$^\circ$C. The best operating conditions found were 20\% olive oil, 25 mM triethanolamine buffer (pH 7.0) and 30$^\circ$C. Under these condition, the operational half life of the enzyme was 220 hrs at flow rate of 12.5ml/hr (volumetric ration of substrate to buffer was 1:0.28). Still the half life was under the effect of buffer flow r...