Terpene ester synthesis by lipase-catalyzed transesterification

Summary Five lipases were screened for their ability to synthesize terpene esters by transesterification. The nature of terpene alcohol and enzyme, as well as the chain length of the acyl donor used affected the product yields. Lipase AY from Candida rugosa gave the best overall yield (96.2%). Geraniol and tributyrin were also found to be the best reactants.     

Short-chain flavour ester synthesis by immobilized lipase in organic media

Summary Mucor miehei lipase has been adsorbed on Celite and covalently bound to nylon. The obtained derivatives have been studied regarding their ability for synthetize several flavouring esters in biphasic aqueous/organic media. The influence of the immobilization procedure on the synthetic activity of the derivatives was considered. Solvent hydrophobicity and water content in the biphasic system influenced both enzyme stability and equilibrium displacements. In this way, solvents with log P>3.5 and less than 1% water were optimal. It was important to consider pH effects on enzyme microenvironment when using acidic substrates. Optimum temperature and reuse of the catalyst were also checked.     

Engineering of lipase-catalyzed transesterification reaction media using water and diethylamine

The objective of presented study was to maximize yields of 2-methylbutyl esters, derived by transesterification reactions mediated by sn-1,3-specific lipases, through engineering of reaction medium. Effects of water and diethylamine (DEA) concentrations on the efficiency of plant oils 2-methylbutanolysis, catalyzed by either mycelium-bound Mucor circinelloides lipase (powder) or commercial immobilized lipase Lipozyme TL IM, were determined. Water content monitoring in reaction mixtures enabled to optimize the initial water content in terms of preventing the dehydration of enzyme’s microenvironment and increasing 2-methylbutyl esters yields. These yields were found to be increased by addition of either suitable amounts of water (0.5–1.5%) or diethylamine (10–30?mM) to the mixture of substrates. The presented results suggest that at low concentrations, diethylamine molecules contribute to retaining water in the microenvironment of enzyme that gives rise to increased transesterification yields and significantly reduced amounts of residual mono- and 1,2-diacyl-glycerols.     

Selective lipase-catalyzed preparation of diol monobenzoates by transesterification and alcoholysis reactions in organic solvents

Lipases from Mucor miehei (MML) and Candida antarctica (CAL) are able to catalyze the monobenzoylation of the primary hydroxy group of 1,2- 1,4- or 1,5-diols with vinyl benzoate in an organic solvent, the reaction proceeding with high regioselectivity and moderate enantioselectivity. The lipase-catalyzed debenzoylation of 1,2-propanediol dibenzoate by alcoholysis with 1-octanol most satisfactorily occurred with Pseudomonas cepacia lipase absorbed onto celite that allowed also to prepare (R)-1-benzoyloxy-2-methylpropan-3-ol from 2-methyl-1,3-propanediol dibenzoate, a result complementary to MML-catalyzed benzoylation of 2-methyl-1,3-propanediol that affords the (S)-monobenzoate.     

Effect of reaction parameters on synthesis of citronellyl methacrylate by lipase-catalyzed transesterification

The methacrylate ester of citronellol was synthesized using various lipases as catalyst. The effect of different reaction parameters such as amount of lipase, solvent, temperature, and acylating agent on the conversion of citronellol to citronellyl methacrylate was studied. Methyl methacrylate, vinyl methacrylate, and 2,3-butanedione mono-oxime methacrylate were used as acylating agents. Porcine pancreatic lipase (PPL), Candida rugosa lipase (CRL), and Pseudomonas cepacia lipase (Amano-PS) were used as biocatalysts. Diisopropyl ether (DIPE) was found to be the most suitable solvent. The stereoselectivity of CRL in transesterification of (+/-)-citronellol was tested for the optimized reaction parameters.     

Process integration involving lipase-catalyzed ester synthesis reactions

Summary The technical feasibility of integrating bioconversion (using a lipase from Mucor miehei immobilized by ion exchange onto a macroporous resin) and product recovery (via mass transfer towards liquid/vapor equilibrium) was experimentally verified for the production of ethyl acetate from acetic acid and ethanol at 30 and 40 °C. The configuration selected consisted of a packed bed thermostatted reactor inserted between the condenser and the liquid reflux to a distillation column. The concentrations of the aforementioned components and water were measured periodically for a 9 to 10-h period. The extents of conversion of acetic acid ranged from 1 to 32% at 30 °C, and from 13 to 25% at 40 °C, and they were maximum at initial molar fractions of acid of ca. 0.3 and 0.4, respectively. The ratio of the reaction product to the equilibrium reaction product increased from 2 to 32 at 30 °C, and from 5 to 27 at 40 °C when the initial molar fraction of acid in the feed increased from 0.2 to 0.7.     

有机介质中脂肪酶催化转酯化反应拆分苯乙氰醇的研究

研究了有机相中脂肪酶催化苯乙氰醇的转酯化反应,拆分苯乙氰醇。考察了酶、溶剂、溶剂水含量、外加苯甲醛和苯甲酸以及底物浓度等因素对反应的影响,结果表明ZJU008号脂肪酶催化活性最高,经实验确定的最佳反应条件为：乙酸乙烯酯为反应物兼溶剂,利用分子筛去除溶剂中微量水分,40 ℃,200 r/min,酶量为10 mg/mL时的最佳底物浓度为200 mmol/L,在上述条件下反应20 h底物转化率为50%,e.e.值大于99%,能将苯乙氰醇有效拆分。外加苯甲醛和苯甲酸不利于反应的进行。     

The kinetic study on lipase-catalyzed transesterification of α-cyano-3-phenoxybenzyl alcohol in organic media

Optically active form of α-cyano-3-phenoxybenzyl alcohol (CPBA), building block of pyrethroid insecticides, was synthesized as its acetate by the combination of anion-exchange resin-catalyzed transcyanation between m-phenoxybenzaldehyde and acetone cyanohydrin, and lipase-catalyzed enantioselective transesterification of the resulting cyanohydrin with vinyl acetate. Through a screening of enzymes, Alcaligenes sp. lipase showed the highest activity and the conversion exceeded 50%. Effects of solvents and temperatures on this reaction were studied. Among four kinds of solvent, diisopropyl ether was a best choice. The optimal temperature was 50 °C. Although the external diffusion limitation could be excluded by raising the rotational speed, internal diffusion could not be ignored, since the enzyme was an immobilized one on the particles with a considerably large diameter. The e.e. values of CPBA ester were measured by polarimeter and NMR.     

Optimization of lipase-catalyzed glucose ester synthesis in ionic liquids

Lipase-catalyzed esterification of glucose with fatty acids in ionic liquids (ILs) mixture was investigated by using supersaturated glucose solution. The effect of ILs mixture ratio, substrate ratio, lipase content, and temperature on the activity and stability of lipase was also studied. The highest yield of sugar ester was obtained in a mixture of 1-butyl-3-methylimidazolium trifluoromethanesulfonate ([Bmim][TfO]) and 1-methyl-3-octylimidazolium bis[(trifluoromethyl)-sulfonyl]amide ([Omim][Tf₂N]) with a volume ratio of 9:1, while Novozym 435 (Candida antarctica type B lipase immobilized on acrylic resin) showed the optimal stability and activity in a mixture of [Bmim][TfO] and [Omim][Tf₂N] with a 1:1 volume ratio. Reuse of lipase and ILs was successfully carried out at the optimized reaction conditions. After 5 times reuse of Novozym 435 and ILs, 78% of initial activity was remained.     

Enzymatic synthesis of derivatives of vitamin A in organic media

The present article provides an enzymatic method of retinol esterification to reduce photodestruction and irritation problems characteristic of retinol. More specifically, it relates to a method of synthesising retinyl adipate, retinyl succinate, retinyl oleate and retinyl lactate greatly appreciated by cosmetic manufacturer. Desired compounds can be synthesised directly using Candida antarctica lipase and Rhizomucor miehei lipase in organic media.     

The equilibrium and kinetics of N-acetyl-tryptophan phenylethyl ester synthesis by agarose-chymotrypsin in organic media

The synthesis of N-acetyl tryptophan phenylethyl ester in organic media is catalyzed by suspended agarose beads with multipoint covalently attached chymotrypsin. A dilute aqueous phase is trapped within the gel beads and may be manipulated separately from the organic phase. The equilibrium position becomes more favorable as the solvent polarity decreases, with K(eq) increasing 38 times between 2-butanone and 1,1,1-trichloroethane. The more apolar solvents also give faster kinetics. Addition of cosolvents (up to 10% dimethylformamide or 20% acetonitrile) does not affect the rate but does substantially reduce the equilibrium yield, presumably also by making the organic phase more polar. With trichloroethane as solvent the enzyme appears to be kinetically saturated with 1M phenylethanol. Doubling this concentration also does not cause the expected increase in equilibrium conversion, probably again because K(eq) is reduced in the more polar organic phase. Increased temperature raises the reaction rate as expected but has little effect on the equilibrium. (c) 1992 John Wiley & Sons, Inc.     

Optimized lipase-catalyzed synthesis of adipate ester in a solvent-free system

Immobilized Candida antarctica lipase-catalyzed esterification of adipic acid and oleyl alcohol was investigated in a solvent-free system (SFS). Optimum conditions for adipate ester synthesis in a stirred-tank reactor were determined by the response surface methodology (RSM) approach with respect to important reaction parameters including time, temperature, agitation speed, and amount of enzyme. A high conversion yield was achieved using low enzyme amounts of 2.5% w/w at 60°C, reaction time of 438 min, and agitation speed of 500 rpm. The good correlation between predicted value (96.0%) and actual value (95.5%) implies that the model derived from RSM allows better understanding of the effect of important reaction parameters on the lipase-catalyzed synthesis of adipate ester in an organic solvent-free system. Higher volumetric productivity compared to a solvent-based system was also offered by SFS. The results demonstrate that the solvent-free system is efficient for enzymatic synthesis of adipate ester.     

Markedly enhancing lipase-catalyzed synthesis of nucleoside drugs' ester by using a mixture system containing organic solvents and ionic liquid

Eightfold higher yields and three times faster reaction rates were achieved by means of using a mixture solvent system composed of 90% acetone and 10% [BMIM]BF4 in the lipase-catalyzed regioselective synthesis of polymerizable ester of nucleoside drugs.     

Improving lipase-catalyzed enantioselective ammonolysis of phenylglycine methyl ester in organic solvent by in situ racemization

Lipase-catalyzed enantioselective ammonolysis of phenylglycine methyl ester was processed by in situ racemization with ammonium carbamate as the acyl acceptor. Using 1 mM benzaldehyde or 0.6 mM chloropyridoxal as the racemizing catalyst, 80% substrate conversion with an enantiomeric excess of the product of 95% were achieved at 20 °C after 7 h reaction.     

Improvement of the enantioselectivity of lipase-catalyzed naproxen ester hydrolysis in organic solvent

A method is presented to improve the enantioselectivity of lipase-catalyzed hydrolysis of naproxen methyl ester in water-saturated isooctane. It is shown that coupling of the enantioselective hydrolysis of Naproxen methyl ester with the photo-dissociation methanol leads to the photocatalytic conversion of methanol into water, by which the equilibrium constant (K) of the lipase-catalyzed hydrolysis was changed. The equilibrium yield and enantiomeric excess are increased. Because the lipase would not dissolve in the organic solvent, it was adsorbed on photocatalyst particles, which may facilitate the isolation of enzyme from reaction system.     

Optimization of carbohydrate fatty acid ester synthesis in organic media by a lipase from Candida antarctica

The effect of solvents and solvent mixtures on the synthesis of myristic acid esters of different carbohydrates with an immobilized lipase from C. antarctica was investigated. The rate of myristyl glucose synthesized by the enzyme was increased from 3.7 to 20.2 micromol min(-1) g(-1) by changing the solvent from pure tert-butanol to a mixture of tert-butanol:pyridine (55:45 v/v), by increasing the temperature from 45 degrees C to 60 degrees C, and by optimizing the relative amounts of glucose, myristic acid, and the enzyme preparation. Addition of more than 2% DMSO to the tert-butanol:pyridine system resulted in a reduction of enzyme activity. Lowering the water content of the enzyme preparation below 0.85% (w/w) resulted in significant decreases in enzyme activity, while increasing the water content up to 2.17% (w/w) did not significantly affect the enzyme activity. The highest yields of myristyl glucose were obtained when an excess of unsolubilized glucose was present in the reaction system. In this case, all of the initially solubilized and a significant amount of the initially unsolubilized glucose was converted to the ester within 24 h of incubation, resulting in a myristyl glucose concentration of 34 mg/mL(-1). Myristic acid esters of fructose (22.3 micromol min(-1) g(-1)), alpha-D-methyl-glucopyranoside (26.9 micromol min(-1) g(-1)) and maltose (1.9 micromol min(-1) g(-1)) could also be prepared using the tert-butanol:pyridine solvent system. No synthesis activity was observed with maltotriose, cellobiose, sucrose, and lactose as substrate.     

Subtilisin-catalysed peptide synthesis and transesterification in organic solvents

Summary Subtilisin from Bacillus subtilis was modified with polyethylene glycol (PEG), or adsorbed either on celite or porous glass, or directly used as a suspended powder to catalyse peptide synthesis and transesterification reactions in organic solvents. The rather low yield of peptide synthesis probably resulted from the enzyme tendency to catalyse hydrolysis and transesterification side reactions. The kinetics of transesterification catalysed by PEG-subtilisin was consistent with a ping-pong mechanism modified by a hydrolytic branch. Initial rates of transesterification were found to be dependent on alcohol and organic base concentrations in the reaction mixture. The high affinity of benzyloxycarbonyl-l-serine-methyl ester for the enzyme indicated that a change in substrate specificity of subtilisin occurred in organic phase. The 50-fold increase in the rate of synthesis of benzyloxycarbonyl-l-serine-l-phenylalanine amide which was observed when PEG-subtilisin was used instead of immobilized or powdered enzyme, suggested that a higher flexibility of the polypeptide chain modified by the covalent attachment of a number of soluble PEG moieties occurred in organic solvents. This also resulted in a lower stability of PEG-subtilisin at high temperature.Offprint requests to: A. Puigserver     

Polyvinyl alcohol--trypsin as a catalyst for amino acid ester synthesis in organic media

The bovine trypsin-catalyzed synthesis of N-alpha-benzoyl-DL-arginine esters from N-benzoyl-DL-arginine were studied in various organic solvents. Trypsin was immobilized to polyvinyl alcohol (PVA) by adsorption from its aqueous solutions. Immobilized enzyme showed higher catalytic activities than free enzyme for amino acid esterification in ethanol. The yield of ester is strongly dependent upon the PVA/trypsin ratio and water content in the reaction medium. The rate and equilibrium constant of the ester formation reaction are also dependent on water content.     

Synthesis of a novel macrolactone by lipase-catalyzed intra-esterification of hydroxy-fatty acid in organic media.

The unsaturations and groups bound to the ring and to the lateral chain of lactones give a large diversity in this class of molecules. In this work we produced enzymatically a macrolactone in organic media. The substrate used was a hydroxy-fatty acid: (+)-coriolic acid and the enzymes tested were free or immobilized microbial lipases. The immobilized lipase from Candida antarctica seems to be the most adequate catalyst offering a high reaction yield. The intra-esterification was studied as a function of temperature and type of solvent. Higher yields were obtained when using diisopropyl-ether at 35 degrees C. This reaction, involving an alcohol group on an internal position on the carbon chain of the substrate hydroxy-acid, produces an original lactone: 13S-octadeca-(9Z,11E)-dienolide. The product was purified and characterized using (1)H nuclear magnetic resonance spectroscopy, mass spectrometry and infrared spectroscopy.     

Study on response surface methodology (RSM) of lipase-catalyzed synthesis of palm-based wax ester

The synthesis of wax ester using refined, bleached and deodorized (RBD) palm oil and oleyl alcohol catalyzed by lipozyme IM was carried out. Response surface methodology (RSM) based on a five-level, four-variable central composite rotatable design (CCRD) was used to evaluate the interactive effects of synthesis, of reaction time (2.5–10 h), temperature (30–70 °C), amount of enzyme (0.1–0.2 g) and substrate molar ratio (palm oil to oleyl alcohol, 1:1–1:5) on the percentage yield of wax esters. The optimum conditions derived via RSM were: reaction time 7.38 h, temperature 53.9 °C, amount of enzyme 0.149 g, and substrate molar ratio 1:3.41. The actual experimental yield was 84.6% under optimum condition, which compared well to the maximum predicted value of 85.4%.