Enzymatic alcoholysis reaction of soy phospholipids

Lipase-catalyzed alcoholysis of soy phospholipids was investigated to simultaneously make lysophospholipids and fatty acid esters of individual alcohols. Alcoholysis was carried out by stirring a mixture of soy phospholipids and individual alcohols in equimolar proportions with 10% (by weight of reactants) Mucor miehei lipase at 55°C for 24 h. The products were isolated by column chromatography after removal of the lipase. Lysophospholipids (in 69–78% molar yield) were obtained from soy phospholipids, and the yield of esters of various alcohols also conformed nearly with theoretical yields.     

Chemo-enzymatic synthesis of amino acid-based surfactants

The application of lipases to the synthesis of amino acid-based surfactants was investigated. Low yields (2–9%) were obtained in the acylation of free amino acids, such as l-serine and l-lysine, as well as their ethyl esters and amides with fatty acids, owing in part to low miscibility of the reactants. When the N-carbobenzyloxy (Cbz)-l-amino acids were used in an effort to improve miscibility of the amino acid derivatives with the acyl donor, a dramatic improvement was observed for N-Cbz-l-serine (92% yield) but not for N _α-Cbz- or N _ζ-Cbz-l-lysine (7 and 2% yield, respectively). As an alternative, and efficient synthesis of N _ζ-acyl-l-lysines was developed, based on the regiospecific chemical acylation of copper(II) lysinate. In pursuit of a general route to amino acid-fatty acid surfactants, the utility of a polyol linker was investigated. Thus, the glycerol ester of N _α′ N _ζ-di-Cbz-l-lysine was prepared and evaluated as a substrate for acylation. As expected, this and other glycer-1-yl esters of N-protected amino acids were excellent substrates for lipase-catalyzed acylation. Their reaction with myristic acid in the presence of Novozyme resulted in the regioselective acylation of the primary hydroxyl group of the glycerol moiety to afford the corresponding 1-O-(N-Cbz-l-aminoacyl)-3-O-myris-toylglycerols with conversions of 50–90%. These were readily deprotected to give a range of 1-O-(aminoacyl)-3-O-myristoyl-glycerols with overall yields of 27–71%.     

Two-step enzymatic reaction for the synthesis of pure structured triacylglycerides

Structured triacylglycerides with medium-chain fatty acids (caprylic acid) in sn1- and sn3-positions and a long-chain unsaturated fatty acid (oleic or linoleic acid) in the sn2-position of glycerol (MLM) were synthesized by lipase catalysis in a two-step process. First, pure 2-monoacylglycerides (2-MG) were synthesized by alcoholysis of triacylglycerides (triolein, trilinolein, or peanut oil) in organic solvents with 1,3-regiospecific lipases (from Rhizomucor miehei, Rhizopus delemar, and Rhizopus javanicus). The 2-MG were purified by crystallization and obtained in up to 71.8% yield. These 2-MG were esterified in a second reaction with caprylic acid in n-hexane to form almost pure MLM. For 2-MG obtained from peanut oil, the final product contained more than 90% caprylic acid in the sn1- and sn3-positions, whereas the sn2-position was composed of 98.5% unsaturated long-chain fatty acids. Reaction conditions for both steps were optimized with respect to source and immobilization of lipase, water activity, and solvent.     

Lipase-catalyzed modification of borage oil: Incorporation of capric and eicosapentaenoic acids to form structured lipids

Two immobilized lipases, nonspecific SP435 from Candida antarctica and sn-1,3 specific IM60 from Rhizomucor miehei, were used as biocatalysts for the restructuring of borage oil (Borago officinalis L.) to incorporate capric acid (10:0, medium-chain fatty acid) and eicosapentaenoic acid (20:5n-3) with the free fatty acids as acyl donors. Transesterification (acidolysis) reactions were carried out in hexane, and the products were analyzed by gas-liquid chromatography. The fatty acid profiles of the modified borage oil were different from that of unmodified borage oil. Higher incorporation of 20:5n-3 (10.2%) and 10:0 (26.3%) was obtained with IM60 lipase, compared to 8.8 and 15.5%, respectively, with SP435 lipase. However, SP435 lipase was able to incorporate both 10:0 and 20:5n-3 fatty acids at the sn-2 position, but the IM60 lipase did not. Solvents with log P values between 3.5 and 4.5 supported the acidolysis reaction better than those with log P values between −0.33 and 3.0.     

固定化假丝酵母99-125脂肪酶催化酯化脂肪酸低碳醇酯反应条件的研究

以硅藻土和纺织品为载体 ,采用吸附法制备固定化脂肪酶 ,研究了固定化假丝酵母 99 1 2 5脂肪酶在有机溶剂中催化脂肪酸低碳醇酯酯化合成过程中 ,有机溶剂性质、脂肪酸与低碳醇的结构、pH值、反应温度和体系含水量、低碳醇的抑制作用等因素对酯化过程的影响。试验结果表明 :底物低碳醇需要采用流加方式加入体系 ,石油醚是最适宜的有机溶剂 ,脂肪酸与醇的碳链越长 ,越易于酯化 ;固定化脂肪酶对直链醇的选择性优于支链醇。以石油醚为有机溶剂 ,在反应温度为 40℃、pH值为 7时 ,硬脂酸与甲醇的酯化率达 95 % ;反应后期应除去体系中的水以避免酶失活。固定化酶间歇催化油酸与甲醇的酯化时 ,重复使用 1 5次 (每次 2 4h) ,其操作半衰期约为 360h。     

脂肪酶不对称立体选择性能改善的研究进展

脂肪酶已广泛用于制备光学纯手性化合物的不对称反应中，但大多数酶催化拆分外消旋化合物的立体选择性不很理想. 目前，改善脂肪酶立体选择性的研究主要从改造脂肪酶酶蛋白结构、优化体系的反应条件、改善反应过程以及对映选择性抑制等方面进行. 通过微波照射也能在一定程度上改善脂肪酶催化反应的立体选择性. 本文主要介绍了几种改善脂肪酶催化不对称反应的立体选择性的方法.     

Borago officinalis oil: Fatty acid fractionation by immobilized Candida rugosa lipase

γ-Linolenic acid (Z,Z,Z-6,9,12-octadecatrienoic acid), a very important polyunsaturated fatty acid is found in the free fatty acid fraction prepared by the hydrolysis of borage oil. Our aim was to enrich this fraction in γ-linolenic acid using selective esterification. Candida rugosa lipase was used as catalyst after immobilization on the following ion-exchange resins: Amberlite IRC50, IRA35, IRA93, and Duolite A7, A368, A568. In every case, immobilization modified the lipae’s specificity: palmitic, stearic, oleic, and linoleic acids were preferentially esterified compared to γ-linolenic acid, thus allowing a γ-linolenic acid enrichment of 3.0.     

Immobilization of lipase on poly(N-vinyl-2-pyrrolidone-co-2-hydroxyethyl methacrylate) hydrogel for the synthesis of butyl oleate

Lipase from Candida rugosa was immobilized by entrapment on poly(N-vinyl-2-pyrrolidone-co-2-hydroxyethyl methacrylate) [poly(VP-co-HEMA)] hydrogel, cross-linked with ethylene glycol dimethacrylate (EDMA). The immobilized enzyme was used in the esterification of oleic acid with butanol in hexane. The activities of the immobilized enzyme preparations and the leaching of the enzyme from the hydrogel supports with respect to composition were investigated. The thermal, solvent, and storage stability of the immobilized preparations also were determined. Increasing the percentage VP from 0 to 90, which corresponds to the increase in the hydrophilicity of the hydrogels, increased the activity of the immobilized enzyme. Lipase immobilized onto VP(%):HEMA(%), 90:10 hydrogel had the highest activity. Increasing the hydrophobicity of the hydrogel (increasing the percentage HEMA) seemed to decrease leaching of the enzyme from the support. Immobilized lipase on 100% HEMA hydrogel indicated highest entrapment and lowest leaching by hexane washing. The lipase immobilized on VP(%):HEMA(%), 50:50 hydrogel showed highest thermal, solvent, and storage stability compared to lipase immobilized on other hydrogel compositions as well as the native lipase.     

Utilization of Catalytic Properties of the Encapsulated Lipase with Calix[4]arene-Adorned Sporopollenin

The dihydrazide calix[4]arene was immobilized onto sporopollenin in order to encapsulate Candida rugosa lipase (CRL) via sol-gel entrapment. The kinetic resolution of the new encapsulated lipase was investigated for enantioselective hydrolysis of racemic naproxen methyl ester and 2-phenoxypropionic acid methyl ester. The results demonstrated that the activity and enantioselectivity of the lipase were improved when the lipase was encapsulated in the presence of calix[4]arene-immobilized sporopollenin. The encapsulated lipase showed an excellent rate of enantioselectivity against the (R/S)-naproxen methyl and (R/S)-2-phenoxypropionic acid methyl esters, with E = 350 and 295, respectively, compared to the free enzyme.     

脂肪酶和醇腈酶在有机合成中的应用

有机相酶催化反应常用于高附加值产品的合成。脂肪酶是有机合成中一种重要的工具酶，可以催化酯交换反应。脂肪酶催化的酯的氨解反应已经用于手性酰胺的合成，烷氧羰基化也广泛应用于一些天然化合物的化学选择性修饰。以前在有机合成中很少实用的裂合酶如(R)-醇腈酶在合成具有旋光性的氰醇中的应用也开始引起人们的关注。