Coupled Kinetic Resolution Of (R, S)-2-Methylbutyric Acid And Its Ester By Lipases

2-methylbutyric acid and its short chain esters are typical important flavor material with the apple,pineapple and pineapple taste.Its two kinds of enantiomer,(S)-and(R)-configuration, have the different flavor and application separately.The enzyme catalysis kinetic resolution is easy to enlarge the scale,friendly to the environment,the product of which satisfies "the green",security and food grade requests.Therefore it is widely used to resolute many chiral flavor materials.The resolution of 2-methylbutyric acid,one of the typical flavor compounds,is difficult by enzymatic kinetic method,because the steric differences between the methyl and ethyl groups attached to its chiral carbon atom are too small to allow efficient stereochemical recognition by enzyme.In order to resolute the hard-resolute 2-methylbutyric acid and its ester,a novel method should be considered—coupled kinetic resolution method,that is,coupling the lipase catalyzed selective esterification with the selective hydrolysis.The kinetic model was established,and 4 different configuration isomers with high optical purity were obtained.The detailed contents of this dissertation follow:1) In order to obtain high optically pure and yield product,the selective esterification and the selective hydrolysis reaction system are separately established and optimized, screening lipases that have both high optical selectivity and high catalytic activity in each system.The esterification reaction system is established and optimized as follows:lipase from Rhizopus chinensis(RCL) has both high optical selectivity and high esterification active,the enzyme dosage 16 mg/ml,the substrate concentration 0.1 mol/L,4℃As for the hydrolysis system,the reacting system is established and optimized as follows:lipase RCL,the enzyme dosage 15 mg/ml,the substrate concentration 0.2 mol/L,4℃.2) In order to enhance the yield of product of coupled kinetic resolution,the model of coupled kinetic resolution reaction is establishing to guide theoretically the reaction.Under the requirement of desired ee_p,the conversion of the two steps reaction were controlled to obtain the highest product yield.Under this model instruction,the conversion of the first step hydrolysis reaction is controlled at 45%and the second step esterification reaction is 38%; and ee value of 90.64%of(S)-ethyl 2-methylbutyrate was obtained finally.As for the coupled kinetic resolution with firstly esterification followed by hydrolysis,the conversion of the first and second step were respectively controlled at 34.8%and 40.2%;and ee value of 92.3%of (S)-2-methylbutyric acid was obtained.3) The research discovered that RCL showed higher hydrolysis enantioselectivity in aqueous phase;and ee value of 92.4%of(R)-ethyl 2-methylbutyrate was obtained after 10 h hydrolysis reaction at 40℃through first hydrolysis(S)-ethyl 2-methylbutyrate.Furthermore, the influence of temperature on the esterification and hydrolysis resolution catalyzed by RCL was investigated,occurred with higher enantioselectivity of reaction under 4℃than 40℃; and ee value of(R)-ethyl 2-methylbutyrate obtained through the hydrolysis resolution was enhanced to 95.0%while 4℃.Extract the remaining substrate(R)-ethyl 2-methylbutyrate, using the lipase Novozym 435 with weak(S)-selectivity but high hydrolysis activity to catalysis the second step hydrolysis reaction,then obtained the ee value of 92.2%of (R)-2-methylbutyric acid.Thus,by two step hydrolysis as well as two kind of different sequence coupled kinetic resolution reactions,these materials have been obtained:the final ee value of 95.0%of (R)-ethyl 2-methylbutyrate,92.2%of(R)-2-methylbutyric acid,90.6%of(S)-ethyl 2-methylbutyrate as well as 92.3%of(S)-2-methylbutyric acid.