Response surface methodology as a tool to study the lipase‐catalyzed synthesis of betulinic acid ester

BACKGROUND: The synthesis of betulinic acid ester using betulinic acid and oleyl alcohol catalyzed by Novozym 435 (immobilized Candida antarctica lipase) was carried out. Response surface methodology (RSM) based on a five‐level, three‐variable, central composite rotatable design (CCRD) was employed to evaluate the interactive effects of various parameters. The parameters were reaction time (8–16 h), temperature (20–60 °C) and enzyme amount (120–160 mg). RESULTS: Simultaneously increasing reaction time, temperature and amount of enzyme increased the yields of betulinic acid ester produced. CONCLUSION: The optimum conditions derived via RSM for the reaction were reaction time of 10.2 h, temperature of 53.1 °C and enzyme amount of 138 mg. The actual experimental yield was 48.5% under optimum conditions, which compared well with the maximum predicted value of 47.6%. Copyright © 2008 Society of Chemical Industry     

Modeling and optimization of lipase‐catalyzed synthesis of dilauryl adipate ester by response surface methodology

BACKGROUND: Adipate esters are used as low‐temperature and low‐viscosity plasticizers for polyvinyl chloride and its copolymers. In this work, optimization of lipase‐catalyzed production of dilauryl adipate was carried out using response surface methodology (RSM) based on a four‐factor‐five‐level central composite rotatable design (CCRD). Immobilized lipase from Candida antarctica (Novozym 435) was used as catalyst in this reaction. Various reaction parameters affecting the synthesis of adipate ester, including alcohol/acid molar ratio, amount of enzyme, temperature and reaction time, were investigated. RESULTS: Statistical analysis showed that the amount of enzyme was less significant than the other three factors. The optimal conditions for the enzymatic reaction were obtained at 5.7:1 substrate molar ratio using 0.18 g of enzyme at 53.1 °C for 282.2 min. Under these conditions the esterification percentage was 96.0%. CONCLUSIONS: The results demonstrated that response surface methodology can be applied effectively to optimize the lipase‐catalyzed synthesis of adipate ester. The optimum conditions can obtained be used to scale up the process. Copyright © 2008 Society of Chemical Industry     

Transesterification for biodiesel production catalyzed by combined lipases: Optimization and kinetics

Preparation of biodiesel from waste cooking oil catalyzed by combined lipases in tert‐butanol medium was investigated. Several crucial parameters affecting biodiesel yield were optimized by response surface methodology, such as dosage of combined lipases of Novozym 435 and Lipozyme TLIM, weight ratio of Novozym 435 to Lipozyme TLIM, amount of tert‐butanol, reaction temperature, and molar ratio of oil to methanol. Under the optimized conditions, the highest biodiesel yield was up to 83.5% The proposed model on biodiesel yield had a satisfactory coefficient of R² (= 94.02%), and was experimentally verified. The combined lipases exhibited high‐operational stability. After 30 cycles (300 h) successively, the activity of combined lipases maintained 85% of its original activity. A reaction kinetic model was proposed to describe the system and deduced to be a pseudo‐first‐order reaction, and the calculated activation energy was 51.71 kJ/mol. © 2009 American Institute of Chemical Engineers AIChE J, 2010     

Lipase‐mediated methanolysis of soybean oils for biodiesel production

BACKGROUND: Biodiesel is increasingly perceived as an important component of solutions to the important current issues of fossil fuel shortages and environmental pollution. Biocatalysis of soybean oils using soluble lipase offers an alternative approach to lipase‐catalyzed biodiesel production using immobilized enzyme or whole‐cell catalysis. The central composite design (CCD) of response surface methodology (RSM) was used here to evaluate the effects of enzyme concentration, temperature, molar ratio of methanol to oil and stirring rate on the yield of fatty methyl ester. RESULTS: Lipase NS81006 from a genetically modified Aspergillus oryzae was utilized as the catalyst for the transesterification of soybean oil for biodiesel production. The experimental data showed that enzyme concentration, molar ratio of methanol to oil and stirring rate had the most significant impact on the yield of fatty methyl ester; a quadratic polynomial equation was obtained for methyl ester yield by multiple regression analysis. The predicted biodiesel yield was 0.928 (w/w) under the optimal conditions and the subsequent verification experiments with biodiesel yield of 0.936 ± 0.014 (w/w) confirmed the validity of the predicted model. CONCLUSION: RSM and CCD were suitable techniques to optimize the transesterification of soybean oil for biodiesel production by soluble lipase NS81006. The related lipase NS81006 reuse stability, chemical or genetic modification, and transesterification mechanism should be taken into consideration. Copyright © 2007 Society of Chemical Industry     

A continuous ultrasound‐assisted packed‐bed bioreactor for the lipase‐catalyzed synthesis of caffeic acid phenethyl ester

BACKGROUND: The focus of this paper is the ultrasound‐assisted synthesis of caffeic acid phenethyl ester (CAPE) from caffeic acid and phenyl ethanol in a continuous packed‐bed bioreactor. Immobilized Novozym^® 435 (from Candida antarctica) is used as the catalyst. A three‐level–three‐factor Box–Behnken design and a response surface methodology (RSM) are employed to evaluate the effects of temperature, flow rate, and ultrasonic power on the percentage molar conversion of CAPE. RESULTS: Based on ridge max analysis, it is concluded that the optimum condition for synthesis is reaction temperature 72.66 °C, flow rate 0.046 mL min^?1, and ultrasonic power 1.64 W cm^?2. The expected molar conversion value is 97.84%. An experiment performed under these optimal conditions resulted in a molar conversion of 92.11 ± 0.75%. The enzyme in the bioreactor was found to be stable for at least 6 days. CONCLUSIONS: The lipase‐catalyzed synthesis of CAPE by an ultrasound‐assisted packed‐bed bioreactor uses mild reaction conditions. Enzymatic synthesis of CAPE is suitable for use in the nutraceutical and food production industries. Copyright © 2011 Society of Chemical Industry     

Lipase‐catalyzed ethanolysis of soybean oil in a solvent‐free system using central composite design and response surface methodology

BACKGROUND: In this work we describe the synthesis of ethyl esters, commonly known as biodiesel, using refined soybean oil and ethanol in a solvent‐free system catalyzed by lipase from Thermomyces lanuginosus. Central composite design and response surface methodology (RSM) were employed to optimize the biodiesel synthesis parameters, which were: reaction time, temperature, substrate molar ratio, enzyme content, and added water, measured as percentage of yield conversion. RESULTS: The optimal conditions obtained were: temperature, 31.5 °C; reaction time, 7 h; substrate molar ratio, 7.5:1 ethanol:soybean oil; enzyme content, 15% (g enzyme g⁻¹ oil); added water, 4% (g water g⁻¹ oil). The experimental yield conversion obtained under these conditions was 96%, which is very close to the maximum predicted value of 94.4%. The reaction time‐course at the optimal values indicated that 5 h was necessary to obtain high yield conversions. CONCLUSION: A high yield conversion was obtained under the optimized conditions, with relative low enzyme content and short time. Comparison of predicted and experimental values showed good correspondence, implying that the empirical model derived from RSM can be used to adequately describe the relationship between the reaction parameters and the response (yield conversion) in lipase‐catalyzed biodiesel synthesis. Copyright © 2008 Society of Chemical Industry     

脂肪酶催化非食用植物油制备生物柴油的过程优化

以麻疯树油、亚麻油、乌柏油为原料油,采用固定化脂肪酶Lipozyme TL IM,在3.0 g油、1 mL正己烷、醇油摩尔比为3.5∶1、固定化酶质量为油质量20%的条件下进行生物柴油的制备,通过脂肪酸甲酯产率和组成分析,以考察生物柴油制备的影响因素,进行反应时间优化.结果表明,酶的催化作用对脂肪酸组分不存在选择性,且...     

复合固定化脂肪酶催化麻疯树油生产生物柴油

对固定化复合脂肪酶催化麻疯树油合成生物柴油进行了研究,利用3因素5水平中心旋转设计的响应曲面法对反应条件进行了优化,研究了复合酶用量、复合酶配比及底物配比对反应的影响。优化结果为复合酶用量为0.27 g,N435占总酶质量的比例为0.15,乙酸甲酯与麻疯树油的摩尔比为10.10,预测生物柴油得率为72.55 %,与实际产率74.34 %吻合较好。并建立了复合酶催化合成生物柴油反应的动力学方程,反应为双底物抑制,符合乒乓机制。     

Kinetic modeling of lipase‐mediated one‐pot chemo‐bio cascade synthesis of R‐1‐phenyl ethyl acetate starting from acetophenone

BACKGROUND: A systematic investigation of mutual interference between a hydrogenation catalyst, Pd/Al₂O₃, and an immobilized lipase in a one‐pot synthesis of R‐1‐phenyl ethyl acetate at 70 °C has been undertaken. This paper reports the kinetic modeling of lipase‐mediated chemo‐bio cascade synthesis of R‐1‐phenyl ethyl acetate starting from acetophenone. RESULTS: The kinetic results revealed that these catalysts were not acting independently but in concert. A mechanism which predicts the experimental observations for this reaction is proposed. CONCLUSION: The parameters of the kinetic model, which are in good agreement with the experimental data, were estimated through numerical data fitting. The reliability of the estimated parameters was analyzed using the Markov Chain Monte Carlo (MCMC) method. Copyright © 2009 Society of Chemical Industry     

The effect of substrate concentrations on the production of biodiesel by lipase‐catalysed transesterification of vegetable oils

All the kinetic studies, found in the literature, on the production of biodiesel (fatty acids methyl esters) considered the esterifications of free fatty acids rather than the transesterification of the vegetable oil itself. The main industrial interest, however, is for the production of biodiesel with the triglyceride (oil) being the substrate. A mathematical model taking into account the mechanism of the methanolysis reaction starting from the vegetable oil as substrate, rather than the free fatty acids, has been developed. From the proposed model equation, the regions where the effect of alcohol inhibition fades, at different substrate concentrations, were identified. The proposed model equation can be used to predict the rate of methanolysis of vegetable oils in a batch or a continuous reactor and to determine the optimal conditions for biodiesel production. Copyright © 2005 Society of Chemical Industry     

Optimized Production of Biodiesel from Waste Cooking Oil by Lipase Immobilized on Magnetic Nanoparticles

Biodiesel, a non-toxic and biodegradable fuel, has recently become a major source of renewable alternative fuels. Utilization of lipase as a biocatalyst to produce biodiesel has advantages over common alkaline catalysts such as mild reaction conditions, easy product separation, and use of waste cooking oil as raw material. In this study, Pseudomonas cepacia lipase immobilized onto magnetic nanoparticles (MNP) was used for biodiesel production from waste cooking oil. The optimal dosage of lipase-bound MNP was 40% (w/w of oil) and there was little difference between stepwise addition of methanol at 12 h- and 24 h-intervals. Reaction temperature, substrate molar ratio (methanol/oil), and water content (w/w of oil) were optimized using response surface methodology (RSM). The optimal reaction conditions were 44.2 °C, substrate molar ratio of 5.2, and water content of 12.5%. The predicted and experimental molar conversions of fatty acid methyl esters (FAME) were 80% and 79%, respectively.     

生物柴油合成中脂肪酶的应用研究进展

生物柴油是以动植物油脂为原料制造的可再生能源,可作为石油柴油的替代或部分替代燃料.生物柴油的发展不仅有利于解决能源问题,而且可以减少温室气体的排放量.本文对脂肪酶的催化活性和选择性,脂肪酶的固定化、不同反应体系中生物柴油的脂肪酶催化合成及其操作条件,不同酰基受体对脂肪酶催化制备生物柴油的影响等问题的研究进展进行了综述.     

Lipase‐catalyzed alcoholysis of vegetable oils

Fatty acid alkyl esters were produced from various vegetable oils by transesterification with different alcohols using immobilized lipases. Using n‐hexane as organic solvent, all immobilized lipases tested were found to be active during methanolysis. Highest conversion (97%) was observed with Thermomyces lanuginosa lipase after 24 h. In contrast, this lipase was almost inactive in a solvent‐free reaction medium using methanol or 2‐propanol as alcohol substrates. This could be overcome by a three‐step addition of methanol, which works efficiently for a range of vegetable oils (e.g. cottonseed, peanut, sunflower, palm olein, coconut and palm kernel) using immobilized lipases from Pseudomonas fluorescens (AK lipase) and Rhizomucor miehei (RM lipase). Repeated batch reactions showed that Rhizomucor miehei lipase was very stable over 120 h. AK and RM lipases also showed acceptable conversion levels for cottonseed oil with ethanol, 1‐propanol, 1‐butanol and isobutanol (50‐65% conversion after 24 h) in solvent‐free conditions. Methyl and isopropyl fatty acid esters obtained by enzymatic alcoholysis of natural vegetable oils can find application in biodiesel fuels and cosmetics industry, respectively.     

Experimental design of the kinetic resolution of a key precursor of high‐value bioactive myo‐inositols by an immobilized lipase

BACKGROUND: The response surface methodology was successfully applied to the optimization of the reaction variables for the kinetic resolution of a precursor of high‐value myo‐inositols, ( ± )‐1,2‐O‐isopropylidene‐3,6‐di‐O‐benzyl‐myo‐inositol (( ± )‐1), by Novozym 435. The resolutions were run separately, with two acylating agents, ethyl acetate and vinyl acetate, in a solvent‐free system. The variables analyzed were reaction temperature, substrate concentration, water concentration and enzyme activity. A statistical model was employed for the evaluation of the influence of the variables on conversion and enantiomeric excess (ee). RESULTS: The optimal conditions for this resolution using vinyl acetate as acylating agent were 45 °C, 5 mg mL^?1 of substrate, 71 U of enzyme activity and 0%w/w of water concentration. The high conversion (49.2 %) and ee (>99%) reached in the chemoenzymatic synthesis of acylated product, L‐(?)‐5‐O‐Acetyl‐3,6‐di‐O‐benzyl‐1,2‐O‐isopropylidene‐myo‐inositol, secure the efficient synthesis of the D enantiomorph present in the original racemic mixture (( ± )‐1) as well. CONCLUSIONS: The use of the experimental design strategy was productive, leading to a 14‐fold increase in the productivity of the reaction compared with the non‐optimized conditions. Both derivative L‐(?)‐2 and remaining substrate D‐(+)‐1 were obtained at high ee. © 2012 Society of Chemical Industry     

固定化脂肪酶催化制备香叶树籽生物柴油研究

研究了Novozym 435和Lipozyme TLIM混合脂肪酶催化香叶树籽油制备生物柴油,2种酶按1:3质量比混合使用时,既可提高反应转化率,又可降低酶的使用成本.应用响应面优化法确定了固定化酶催化香叶树籽生物柴油的最优工艺参数,采用叔丁醇作为反应体系的溶剂,最优反应条件为反应温度38.5℃、甲醇与油摩尔比4:1、叔丁醇与油体积比1:1.5、酶用量为油质量的4%,此时反应转化率达90.09%.分析表明香叶树籽油的甘油三酯主要由短链脂肪酸甘油酯组成,生物柴油中原油的甘油三酯已完全转变成脂肪酸甲酯.     

微波辅助固体酸催化废弃油脂合成生物柴油工艺的优化

以废弃油脂为原料,采用微波辐射辅助固体酸催化合成生物柴油。考察反应温度、微波功率、微波加热时间、醇油摩尔比及催化剂的用量对生物柴油产率的影响,并采用响应面法建立二次回归模型对合成工艺进行了优化。分析结果显示,在反应温度为82℃、反应时间为93 min、醇油摩尔比7∶1、催化剂的用量为3.2%时,生物柴油的产率可达到94.58%,与在相同工艺条件下未经微波处理的产率提高了32.47%。     

响应面法优化固定化洋葱伯克霍尔德菌脂肪酶回收工艺的研究

酶法制备生物柴油工艺引起了越来越多的关注。本文以脂肪酶回收前后脂肪酸甲酯得率之比作为检测指标,采用响应面法对自行制备的固定化洋葱伯克霍尔德菌脂肪酶的回收工艺参数进行了优化,并利用软件SAS 9.0对实验数据进行分析。确定的最佳回收条件:回收溶剂用量20.4 mL,回收温度40.2℃,回收时间20 m in。此条件下,验证试验脂肪酶回收率最大可达98.26%,与回归模型预测最优条件下脂肪酶回收率99.59%非常接近,且使用30次后,其回收效率依然接近100%。R2=96.63%显示该回归模型在分析脂肪酶回收率方面具有极高的准确性和可信度。     

Production of specific‐structured triacylglycerols by lipase‐catalyzed reactions: a review

Regiospecificity is one of the major advantages of using lipase technology for the modification of oils and fats to produce high‐value added products, such as cocoa butter equivalents, human milk fat substitutes, and other specific‐structured lipids. Due to the high cost of biocatalysts, the mainstream applications of lipases for normal oils and fats are still limited. Therefore, positional specificity of lipases has the priority and will be the target property to be exploited for commercial and industrial developments, because no chemical method has such a specificity and is promising or possible for this task. In this paper, encouraging products resulting from this regiospecificity are reviewed together with the critical evaluation of their reaction schemes, side reactions and by‐products, sources of substrate oils and acyl donors, and production processes.