Immobilization Of Lipase For Catalyzing Reactions In Organic Solvent

The importance of chiral technology and the progress in the preparation of optical pure chemical compounds were reviewed. And the immobilization of enzyme was introduced, including conceptions, preparation methods and applications. The effect of immobilization on activity of enzyme was also referred.There are many limitations in application of free enzyme. So in this paper , immobilization of lipase QL was studied. Lipase QL can catalyze the transesterification of R-1-phenyl ethanol with high selectivity. To determine the effection of immobilization, the selectively transesterification of 1-phenyl ethanol, with vinyl acetate as both the acyl donor and the solvent, was taken as the model reaction.The reaction equation was as follows:The immobilization methods, carriers and conditions were studied. The results showed that lipase could be well aggradated onto diatomite, which was chosen as the carrier. The studies on immobilization conditions showed that the optimal ratio of carrier to lipase, the optimal pH, and the optimal concentration of buffer were 5:1, 9.2, and 0.03mol/L respectively. The transesterification activity of immobilized enzyme was proved to be 14.3 times higher than that of free lipase with same content of protein.The prepared immobilized lipase was applied in catalysis of transesterification of 1-phenyl ethanol. The catalytic, behaviors were investigated, including the influence of temperature, rotation speed and the concentration of substrate. It was found that the optimal temperature and rotate speed were at 40℃ and 150rpm respectively. The catalysis was inhibited at the concentration of substrate was 3.0mol/L. Compared to free lipase, it just cost 20% time for immobilized lipase to finish the transesterification reaction.The characteristics of immobilized lipase was investigated. The results showed it was more stable than free lipase in high temperature. After being reused for 10 cycles, the remaining activity of immobilized lipase was about 75.5%. After 56 days reserved in 4℃30℃, and 40℃, the residual activities of immobilized lipase were 70.4%, 56% and 45.7% respectively.With the same method and carrier, another lipase(from Alcaligenes sp.)was immobilized. The studies showed that the optimal ratio of carrier to enzyme was 13:1, ' the optimal pH was 8.5, and the optimal concentration of buffer was' 0.05mol/L. It was found that the immobilized lipase lost its activity seriously in organic solvent.Because S-1-phenyl ethanol did not react with vinyl acetate, the kinetics of R-1-phenyl ethanol was studied only. The reaction mechanism was hypothesized to be a ping-pong bi-bi reaction. The rate equation was:The five parameters of this equation were simulated from the experimental data. The model parameters were as follows: Kp=0.0003min-1 KA=0.9589mmol/L KB=0.0054mmol/L KQA=0.00002mmol/L KBQA=0.000011mmol/L.The equation was used to predict the conversion process at different concentrations of substrate and immobilized lipase. The model simulation curves were in good agreement with the experimental data.