Resolution Of Endo-Bicyclo[3.2.0]hept-2-en-6-ol By Immobilized Lipase Under Microwave

The resolution of endo-bicyclo [3.2.0] hept-2-en-6-ol catalyzed by lipase in organic solvents was studied.Firstly,the optimum lipase source was screened,and then the effects of organic reaction medium,water activity,acyl donors with different chain length,temperature,p H,ionic strength and substrate molar ratio on the catalytic activity and enantioselectivity of lipase have been investigated.The optimum reaction conditions were determined as follows: the enzyme source was Mucor Miehei lipase(MML,p H =7.8,ionic strength 0.2 m M),The organic solvent is n-hexane,the acyl donor is vinyl butyrate,the reaction temperature is 25 ?,the water activity is 0.43,and the molar ratio of substrate(vinyl butyrate: endo-bicyclo [3.2.0] hept-2-en-6-ol)= 1.5:1.Under the optimum reaction conditions,the highest enzyme activity of MML was 93 ?mol/g·h and the highest enantioselectivity is E = 63.Because the catalytic performance of lipase is not very ideal,this study hopes to further improve the catalytic performance of lipase by enzyme immobilization and microwave.Firstly,monodisperse silica nanospheres were successfully prepared.Lipase MML was immobilized on them and then used to catalyze the resolution of endo-bicyclo [3.2.0]hept-2-en-6-ol.Under the condition of temperature 15 ? and p H = 7.5,5 ml of lipase MML solution with concentration of 1 mg/ml and 10 mg monodisperse silica nanospheres were mixed together and stirred for 9 hours.The immobilization rate could reach 27%.The catalytic activity of immobilized MML can reach 172 ?mol/g·h at 30 ?and 0.57 of water activity,1.8 times higher than that of free enzyme.The enantioselectivity of the immobilized MML was almost unchanged compared with that of free MML.In addition,the immobilized MML showed good storage stability and reusability.On this basis,we further investigated the auxiliary effect of microwave on the catalytic resolution catalyzed by immobilized MML.The enzyme activity of immobilized MML can reach 625 ?mol/g·h when the microwave power is 240 W and the temperature is 20 ?,6.7 times higher than that of free enzyme.The highest enantioselectivity was E = 472.5,which was increased by 7.5 times.Under the optimum reaction conditions,the resolution reaction can be nearly completed after 16 hours,the conversion of the reaction can reach 53.1%,and the ee of the residual substrate can reach99.2%.