Research On Preparation Of Fatty Acid Ethyl Ester From Semen Abutili Seed Oil Catalyzed By Lipase

Fatty acid ethyl ester(FAEE)can not only be used as renewable energy resource(biodiesel),but also as health care products.Besides,FAEE,a renewable and environment-friendly raw material,can be widely used in cosmetics,washing,textiles and other fields.Therefore,it is very interesting to develop and research FAEE.Currently,the feedstocks of FAEE are mainly from edible vegetable oils(accounting for?70%of raw materials),which can increase the cost of FAEE production and lead to competition in the edible oil market.Therefore,it is very interesting to develop new oil resources.Recently,free liquid lipase has attracted much attention because of its low cost and environmental friendliness.However,the acid value(AV)of free liquid lipase-catalyzed the transesterification of oil is usually out of specification(AV>4.0 mg KOH/g).Therefore,it is very urgent to explore the method to reduce the AV of FAEE.In addition,free liquid lipase also has the disadvantages of easy inactivation and difficult recovery.The immobilized enzyme technology can effectively improve the disadvantage of free liquid lipase.At present,the price of immobilized enzyme is high due to the foreign resin carrier,which limits its application.Therefore,it is very urgent to develop a new immobilized enzyme method.Based on the above,Semen Abutili seed oil(SASO)was used as a potential feedstock and low-priced free liquid lipase Eversa(?)Transform 2.0(ET)was used as a catalyst for FAEE production.In order to let AV of free liquid lipase-catalyzed FAEE meet the prescribed standard(AV?0.5 mg KOH/g),two methods of removing FFA from FAEE by lipase were compared.In addition,in order to improve the catalytic performance of free liquid lipase and reduce the cost of immobilized enzyme,lipase immobilized on low-cost domestic resin through covalent binding was studied.(1)Firstly,three free liquid lipases,including Eversa(?)Transform 2.0,Candia Antarctica Lipase A and Candia Antarctica Lipase B,were screened to produce FAEE from SASO in the solvent-free system.Among the tested lipases,free liquid lipase ET showed the highest catalytic performance and the fastest initial reaction rate.The conditions of FAEE production from SASO was optimized by single factor experiment and response surface methodology(RSM).Results showed that the maximum FAEE yield was 94.2±1.3%under the optimal conditions(lipase load 6%,water content 20%,7:1(mol/mol)ethanol to SASO,11 h,37?).Kinetic parameters(V_max and K'_m)of the transesterification of SASO were 4.16×10^-2 mol/(L·min)and 5.27×10^-1 mol/L,respectively.Arrhenius equation and Ea of ET-catalyzed the transesterification of SASO were ln V₀=4.8252-2382.3/T and 19.80k J/mol,respectively.(2)The acid value(AV)of free liquid lipase-catalyzed the transesterification of oil is usually out of prescribed standard(>4.0 mg KOH/g).In order to let AV of FAEE meet the prescribed standard(AV?0.5 mg KOH/g),two methods of removing FFA from FAEE by lipase were compared((i)the method of complex lipase removing FFA:complex lipase-catalyzed transesterification of SASO(AV=5.45±0.42 mg KOH/g)with esterification of residual FFA of SASO;(ii)the method of lipase powder removing FFA:lipase powder-catalyzed esterification of residual FFA of FAEE(AV=12.0±0.2 mg KOH/g)).The effect of reaction factors on the method of complex lipase removing FFA was optimized by single factor experiment,and the AV and yield of FAEE were 0.39±0.10 mg KOH/g and90.8±1.5%,respectively,under the optimal conditions(ET load 1%,CALB powder load0.8%,9:1(mol/mol)ethanol to SASO,4A-MS load 0.8 g/g,24 h,35?).The effect of reaction factors on the method of lipase powder removing FFA was optimized by single factor experiment and RSM.Results demonstrated that the yield and AV of FAEE were99.4±0.5%and 0.26±0.14 mg KOH/g,respectively,under the optimal conditions(CALB powder load 0.8%,6:1(mol/mol)ethanol to FFA,4A-MS load 0.8 g/g,8 h,35?).Kinetic parameters(V_max and K'_m)of the esterification of FFA were 3.95×10^-4 mol/(L·min)and1.85×10^-1 mol/L,respectively.The activation energy(Ea)was 28.7 k J/mol.(3)In order to improve the catalytic performance of free liquid lipase and reduce the cost of immobilized lipase,lipase ET and CALB were immobilized on domestic resin through covalent binding.The results showed that epoxy resin ES-108B was the best carrier,and the effect of conditions on immobilized complex lipase were optimized by RSM.The maximum lipase loading of ES-108B was 68.1±2.1 mg/g under the optimal conditions(p H 7,V_ET:V_CALB=7:3(mL/mL),21 h,25?).The preparation of FAEE from SASO catalyzed by immobilized complex lipase was optimized by RSM.Results showed that the maximum yield and AV of FAEE were 98.5±1.2%and 2.1±0.1 mg KOH/g,respectively,under the optimal conditions(lipase load 14%,8:1(mol/mol)ethanol to SASO,11 h,37?).(4)In order to study the performance of immobilized complex lipase-catalyzed esterification of residual FFA of SASO,the immobilized complex lipase-catalyzed the transesterification of low AV and high AV of SASO to produce FAEE(AV?0.5mg KOH/g)was studied.The conditions for preparation of FAEE from SASO with low acid value(AV=5.5±0.3 mg KOH/g)catalyzed by immobilized complex lipase were optimized.Results showed that the maximum yield and AV of FAEE were 93.3±1.5%and 0.43±0.07mg KOH/g,respectively,under the optimal conditions(lipase load 14%,ethanol to SASO11:1(mol/mol),4A-MS load 0.8 g/g,11 h,37?).The optimal conditions for the preparation of FAEE from SASO with high acid value(AV=20.4±0.1 mg KOH/g)catalyzed by immobilized complex lipase were optimized.Results showed that the maximum yield and AV of FAEE were 95.1±1.2%and 0.46±0.05 mg KOH/g,respectively,under the optimal conditions(lipase load 14%,ethanol to SASO 9:1(mol/mol),4A-MS load 1.2 g/g,11 h,37?).(5)After purified,the properties of FAEE from SASO were analyzed.The properties of FAEE from SASO were agreed with the quality standard of ASTM D6751 except oxidation stability,which can be improved by antioxidant.The purification of linoleic acid ethyl ester was pale yellow oily liquid,the physical and chemical properties of linoleic acid ethyl ester were agreed with the national standards.