Study On The Synthesis Of Geranyl Butyrate In Nonaqueous Phase Using Candida Rugosa Lipase

As one of the most frequently used terpenoids,geranyl butyrate is widely used in food,medicine,perfume and cosmetic products.The chemical synthesis of geranyl butyrate,actually used in industry,has obvious shortcomings.The enzymatic synthesis has attractive prospect of application due to its specificity,its mild reaction conditions and its facility for the separation of reactants.The present work concerns the synthesis of geranyl butyrate catalyzed by Candida rugosa Lipase(CRL)in organic solvent.Firstly,the reactant was separated by liquid chromatography and identified as geranyl butyrate by FT-IR and NMR.The method for quantitative measuring of product was also established by gas chromatography.Then,the reaction conditions were optimized by single factor experiments.The highest esterification rate of 90.7%was obtained under optimal conditions:n-heptane as solvent,9mg/m L CRL,10 mg/m L of molecular sieves,1:1.5 molar ratio of butyric acid to geraniol and reaction time of 3.5 h.For further understanding on the reaction mechanism of the synthesis of geranyl butyrate catalyzed by CRL in organic phase,the kinetics of synthesis was studied.It was found that the reaction obeyed the Ping-Pong Bi-Bi mechanism with competitive inhibition by geraniol and butyric acid.The kinetic parameters were obtained by nonlinear simulation.For the practical application,CRL was immobilized.The weak-acidic macroporous cation exchange resin D151 was chosen as carrier.The optimal immobilization conditions were:15mg/m L CRL,pH of the phosphate buffer 6.5,temperature 45°C and immobilization time 8 h.An immobilization efficiency of 88.5%,a specific activity of 195.6×10~4 U/g and an activity recovery of 83.5%were achieved.The immobilized CRL had high catalytic ability and the highest esterification rate of 84.3%could be achieved.The properties of immobilized CRL were studied and it was found that the optimal temperature of 40°C was kept as same as that of free enzyme.On the other hand,the thermo stability was greatly improved.After being incubated under 80°C for 2 h,the immobilized CRL could achieve an esterification rate over 60%for the synthesis of geranyl butyrate while the free CRL could arrive only 16.4%.The immobilized CRL was used to catalyze the synthesis of geranyl butyrate for 7 times and the esterification ratio could be kept over 75%,showing a good stability.An experimental circulating fluidized bed reactor(CFBR)was designed to synthesize continuously geranyl butyrate catalyzed by immobilized CRL.The optimized operational conditions were:expansion ratio 1.6,feed flow rate 0.07 mL/min(corresponding average residence time of 4.7 h),butyric acid concentration 75 mmol/L,acid:alcohol molar ratio 1:1.5.The reaction was realized for 12 h under above conditions and a stable esterification rate of 77%was achieved.These results provide the basis for the scale-up and the industrial application of the process.Finally,the gas-liquid equilibrium was calculated by ASPEN PLUS software.It was proved that the relative volatilization between geraniol and geranyl butyrate was very high,which could be separated by conventional distillation.