Lipase-Catalyzed Enantioselective Hydrolysis Of Ethyl Lactate In Organic Solvents

In this work,asymmetric hydrolysis of ethyl lactate catalyzed by lipase in organic systems has been studied.Lipase was used as the catalyst,organic solvents as the reaction medium, ethyl lactate as the optical resolution substrate. Studied the resolution process and optimized reaction condiyions. The kinetics of lipase-catalyzed enantioselective hydrolysis of ethyl lactate was investigated .Firstly, the feasibility of preparation of L-lactic acid by Lipase-catalyzed enantioselective hydrolysis of ethyl lactate in organic solvents was explored. Effects of enzyme sources and organic medium on the reaction were examined respectively. The results showed that Candida antarctic lipase B(N435)-had both the highest catalytic activity and enantioselectivity. The mixture of Tert-butanol and Isooctane (1:1 ,V/V) was the most suitable medium for the reaction, other optimum conditions for catalysis were 200 r/min for shaking rate, 1:5 for mass ratio of ethyl lactate to water, 0.27 g·mL-1 for ethyl lactate concentration, 0.8 g·mol-1 for enzyme concentration and 60℃for reaction temperature, respectively. Under these optimal conditions, a product enantiomeric excess (ee) of 90.02% and the yield of 28.69% were achieved after 16 h reaction.Secondly, based on the results of the above single factor experiments, significant factors were selected from the six factors researched in the single factor experiment by Plackett-Burman (PB) experiment. The result showed that enzymatic concentration, substrate concentration and reaction temperature were the significant impact factors. The above three major factors were further optimized by central composite experiment design and analyzed the data using SAS software. The optimum conditions of the reaction were obtained: 58℃for reaction temperature, 200 r/min for shaking rate, 0.82 g·mol-1 for enzyme concentration, 0.25 g·mL-1 for ethyl lactate concentration, 0.2 for mass ratio of ethyl lactate to water respectively. Under this condition, a product enantiomeric excess (ee) was 94.38% and the yield was 30.02%.Finally The kinetics of lipase-catalyzed enantioselective hydrolysis of ethyl lactate was investigated. The experimental results indicated that: the reaction could be described in terms of the Michaelis-Menten equation with a Ping-Pong Bi-Bi mechanism and competitive inhibition by both the substrates. The effect of D-lactic acid and D-ethyl lactate substrate concentration on the initial rate of asymmetric hydrolysis were studied. On this basis, the dynamic model was established.