SEPES/异辛烷反胶团中脂肪酶的特性

<正>反胶团酶反应兼有水相与有机相酶反应的共同优点,但至今尚无工业应用的重要原因是反胶团中酶的活性和稳定性不高.常用的离子型表面活性剂,如二-(2-乙基己基)琥珀酸双酯磺酸钠(AOT)在有机溶剂中形成的反胶团,因相界面与酶分子间的静电作用将造成酶在反胶团中迅速失活.而非离子型表面活性剂形成的反胶团中,酶一般具有较高的活性和稳定性,但这种体系需加入助表面活性剂,给下游加工过程带来麻烦.作者制备了具有不同氧乙烯聚合数(n)的二-(2-乙基己基聚氧乙烯)琥珀酸双酯磺酸钠(简称SEPES),其分子结构与AOT相似,但其极性头与疏水尾间连有聚氧乙烯链非离子性亲水基团.本文报道脂肪酶在SEPES/异辛烷反胶团体系中的特性.     

CTAB混合反胶团萃取工业脂肪酶

研究了CTAB与Tween、含氧有机物形成的混合反胶团对工业脂肪酶进行萃取分离的效果. 实验表明,CTAB-Tween85和CTAB-含氧有机物混合反胶团的萃取率高于单一CTAB反胶团;反萃时CTAB-含氧有机物混合反胶团的反萃率与单一CTAB反胶团的反萃率相似,CTAB-Tween60和Tween40混合反胶团的反萃效果优于单一CTAB反胶团. 通过测定反萃水相的酶活,发现CTAB-TRPO混合反胶团的效果最好,酶活回收率最高,可以达到70%.     

反胶团酶反应-膜分离技术研究进展

将反胶团酶反应与膜分离相结合被认为是使反胶团酶反应技术工业化的最有前途的方法。文中介绍反胶团酶反应－膜分离技术的研究现状,探讨了今后进一步研究开发的方向。     

基于表面活性剂的反胶团萃取技术

基于表面活性剂的反胶团萃取技术是一种新型的有发展前途的生物分离技术。介绍了反胶团萃取技术的驱动力、表面活性剂选择和影响因素,提出了反胶团萃取技术目前存在的问题,探讨了反胶团萃取技术在其酶促反应和纳米材料制备方面的应用及其发展方向。     

反胶团酶反应研究进展

介绍了酶在反胶团中的主要特性,对反胶团酶反应的应用,动力学和过程的最新进展进行了综述。     

反胶团酶催化研究新进展

评述了近年来反胶团酶催化研究的新进展。在AOT/异辛烷反胶团中加入非离子型表面活性剂如Tween 85、小相对分子质量聚乙二醇等可有效降低酶与表面活性剂间的静电和疏水作用 ,显著提高酶的活性。对AOT进行化学修饰及合成结构与磷脂类似的新表面活性剂以用其构建新的反胶团体系 ,酶的活性较常用的AOT/异辛烷反胶团体系有显著提高。在反胶团酶反应动力学研究中考虑水含量或底物在反胶团表面吸附的影响等 ,提出了进一步研究的设想 ,包括开发新型表面活性剂以进一步提高酶的活性和稳定性及有利于产物分离     

聚乙烯醇-壳聚糖复合酶膜的制备及在单甘油酯合成中的应用

引　言脂肪酶有机相酶催化在有机合成及手性药物合成中有重要用途[1],这些反应大都是在油水两相中进行 ,脂肪酶是一种界面酶 ,在两相界面上反应活性最高 .为增大比表面积 ,在反应体系中加入表面活性剂或采用反胶团体系进行脂肪酶催化反应[2 ],但加入表面活性剂后对后续产物的分离纯化带来不利影响 .采用酶膜反应器进行脂肪酶两相反应是近年来的一个发展方向 ,即将脂肪酶固定在膜上 ,油水两相分别走膜两侧 ,并在膜表面接触反应 ,反应生产的产物可及时被两相带走 ,从而避免了两相直接接触乳化的问题[3].目前脂肪酶是通过吸附或交联固定在疏水…     

反胶团技术应用研究进展

反胶团具有独特的优良性能,有着十分广阔的应用前景,近年来正越来越多地受到关注。该文介绍了反胶团理论的最新研究进展,综述了反胶团技术在酶催化反应、萃取分离、纳米粒子的制备等方面的应用研究进展, 提出了反胶团技术目前存在的问题,并对其应用前景作了展望。引用文献22篇。     

滚筒式填料筛板萃取器反胶团法萃取蛋白质

设计出滚筒式填料筛板萃取器,并应用该萃取器反胶团法萃取蛋白质。萃取溶菌酶和牛血清白蛋白（ＢＳＡ）的实验结果表明,该萃取器萃取效果良好,分相迅速,可澄清分相的水相ＫＣＬ最低浓度为０．０２ｍｏｌ／Ｌ,可有效解决反胶团法萃取蛋白质过程分相困难的问题。报道使用石油醚为溶剂反胶团法萃取蛋白质。石油醚为混合烷烃,价格低廉,易于纯化,萃取效果良好,不会引起酶的失活。     

表面活性剂在生物工程中的应用之一—反胶团萃取

反胶团萃取分离蛋白质是一种新型的、有发展前途的生物产品的分离技术。它是利用表面活性剂在有机溶剂中形成反向胶团,从而实现了对蛋白质的萃取,是表面活性剂在生物工程中的一种成功应用。     

亲和反胶团选择性萃取分离酵母乙醇脱氢酶

The reversed micelles were formed with cationic cetyltrimethylammonium bromide (CTAB) as surfactant and n-hexanol as cosolvent in the CTAB (50mmol L-1)/hexanol (15% by volume)/hexane system. Cibacron Blue 3GA (CB) as an affinity ligand in the aqueous phase was directly introduced to the reversed micelles with electrostatic interaction between anionic CB and cationic surfactant. High molecular weight (Mr) protein, yeast alcohol dehydrogenase (YADH, Mr = 141000) from baker's yeast, has been purified using the affinity reversed micelles by the phase transfer method. Various parameters, such as CB concentration, pH and ionic strength, on YADH forward and backward transfer were studied. YADH can be transferred into and out from the reversed micelles under mild conditions (only by regulation of solution pH and salt concentration) with the successful recovery of most YADH activity. Both forward and backward extractions occurred when the aqueous phase pH>pI with electrostatic attraction between YADH and CTAB. Th     

Non-enzymatic and enzymatic degradation of poly(ethylene glycol)-b-poly(?-caprolactone) diblock copolymer micelles in aqueous solution

The non-enzymatic and enzymatic degradation behaviors of the monomethoxy-poly(ethylene glycol)-b-poly(?-caprolactone) diblock copolymers (MPEG-PCL) micelles in aqueous solution were investigated by DLS, ¹H NMR, SEC and HPLC. It is found that the degradation mechanism of MPEG-PCL micelles in aqueous solution in non-enzymatic case is quite different from that in the presence of enzyme. In non-enzymatic case, the degradation induced by acidic catalysis was not found in low pH aqueous solution but the degradation of the micelles occurred under neutral and basic conditions. The degradation of MPEG-PCL micelles first happens near the interface region of the MPEG shell and PCL core, leading to the part detachment of PEG chains. With increasing degradation time, the degradation inside the PCL core with a random scission on PCL chains occurred. Compared with non-enzymatic degradation, the enzymatic degradation of MPEG-PCL micelles is much fast and the degradation rate of MPEG-PCL micelles is proportional to either the micelles or the enzyme concentration in a certain range. Based on the micelle degradation behaviors that we observed, a possible mechanism for the enzymatic degradation of the MPEG-b-PCL micelles including PCL core erosion which results in cavitization of micellar core and micellar dissociation is proposed.     

Performance of Perstractive Enzyme Reactor for Synthesis of Aspartame Precursor

A perstractive enzyme reactor was used for the synthesis of N-(benzyloxycarbonyl)-L -aspartyl-L -phenylalanine methyl ester (ZAPM), the precursor of the artificial sweetener, aspartame. The synthesis of ZAPM in the reactor proceeded by an enzymatic reaction between N-(benzyloxycarbonyl)-L -aspartic acid (ZA) and L -phenylalanine methyl ester (PM) in the aqueous phase. The synthesized ZAPM in the aqueous phase was mainly extracted into the organic phase, therefore, the concentration of ZAPM in the aqueous phase could be kept low. As a result, high conversion of ZAPM was obtained with this system. The partition coefficients of substances in the aqueous/butyl acetate biphasic system, the mass transfer coefficients of substances through the membrane and the enzymatic kinetics of ZAPM synthesis were determined experimentally. The reaction model which was based on the material mass balance equations was discussed to estimate the performance of the perstractive enzyme reactor system. The calculation values using the model and the experimental data showed good agreement with the concentration changes of the substances in the system. © 1997 SCI     

A comprehensive study of the loss of enzyme activity in a continuous membrane reactor – application to starch hydrolysis

Loss of enzyme activity is a problem associated with enzymatic reactions in continuous recycled membrane reactors (CRMR). It may result from catalyst leakage and also enzyme denaturation due to the effects of pH, temperature, shear effects or adsorption/deposit on membrane. In this study, the relative importance of these various factors has been assessed in order to reduce their adverse effects on starch hydrolysis in a CRMR. The effects of temperature and denaturation by adsorption/deposit on membrane were the most limiting phenomena. Reducing the temperature to overcome thermal denaturation was not a practical solution since this increases viscosity and thereby decreases permeate flux and reactor performance. Insofar that adsorption/deposit of enzymes on the membrane is directly linked to membrane fouling, back‐flushing or regularly purging retentate should reduce this phenomenon by lowering accumulation of high molecular weight products. © 2001 Society of Chemical Industry     

Phospholipase A2-catalyzed hydrolysis of lecithin in a continuous reversed-micellar membrane bioreactor

Lysophospholipids and free fatty acids produced by lecithin hydrolysis are important natural compounds with high potential for application in the food, chemical, and pharmaceutical industries. In this work, the enzymatic hydrolysis of lecithin (essentially phosphatidylcholine) catalyzed by porcine pancreatic phospholipase A₂ (phosphatide 2-acyl-hydrolase, EC 3.1.1.4), encapsulated in mixed reversed micelles of lecithin andbis(2-ethylhexyl) sodium sulfosuccinate (AOT) in isooctane, was carried out in a continuous reversed-micellar membrane bioreactor. A tubular ceramic membrane with a 10,000 molecular weight (MW) cutoff was installed in an ultrafiltration module to retain the phospholipase A₂ (MW 14,000) and to continuously separate the products from the reaction media. Water and co-factor (Ca⁺⁺)-containing reversed micelles of lecithin/AOT in isooctane were supplemented to the reactor to compensate for the permeation of reversed micelles and to continuously supply the substrate. The influence of relevant parameters, such as substrate, AOT and enzyme concentrations, water content and fluid hydrodynamics, on the performance of the ultrafiltration membrane bioreactor was investigated. Fluid axial velocity and substrate concentration were the major factors that affected the transport processes through the membrane. Permeate flow rate increased significantly with fluid axial velocity and decreased with substrate concentration; on the other hand, water and enzyme concentrations were identified as critical parameters for the final conversion of lecithin. The relationship between productivity and normalized residence time was analyzed for each set of experimental parameters tested. Operational stability of the bioreactor was tested in a long-term operation to confirm the high stability of this catalytic system.     

Enzyme Separation Using Supported Liquid Membrane Filled with Reversed Micelles

Abstract

A phenomenological model describing the transfer of lysozyme between a bulk aqueous phase and a reversed micellar phase in a stirred membrane cell has been confirmed. Transport of the enzyme at the interface at low surfactant concentrations is dominant, while that through the membrane is the rate-determining step at high surfactant concentrations. Complete separation of α-chymotrypsin from lysozyme using a supported liquid membrane filled with reversed micelles demonstrates the feasibility of the present process for enzyme separation.     

Intensification of enzymatic hydrolysis of penicillin G: Part 2. model for enzymatic reaction with reactive extraction

During the enzymatic hydrolysis of the potassium salt of Penicillin-G (PenGK) into Phenylacetic acid (PAA) and potassium salt of 6-Aminopenicillanic acid (6-APA), the pH of the reaction mixture falls on account of accumulation of PAA. This lowers the stability and activity of the enzyme used, viz., Penicillin-G acylase (PGA). A new approach of extracting the PAA by a long-chain tertiary amine, which is in the dispersed phase, as a liquid ion exchanger (LIX), is presented. A mathematical model has been developed for this slurry phase reactor with PenGK in the continuous aqueous phase, the amine alongwith a diluent in the dispersed organic phase and the immobilized PGA enzyme as the solid catalyst. Effects of various parameters affecting the conversion of PenG have been discussed. The model has been solved for batch and semi-batch modes. It has been shown that the semi-batch mode yields a higher productivity. This approach can also be advantageously used for other intermediates like 7-ADCA for cephalosporins.     

Stability and proteolytic activity of papain in reverse micellar and aqueous media: A kinetic and spectroscopic study

The stability and proteolytic activity of papain were studied in reverse micellar systems, and in aqueous media. In reverse micelles the maximum activity obtained was 80% of the enzyme activity in aqueous solution. Higher papain stability was found in reverse micellar systems compared with that in aqueous solution with half-lives of 24 and 10 days respectively. Electron spin resonance (ESR) spectroscopy studies of aqueous and reverse micellar systems were performed in an attempt to explain the observed enzyme stability and activity profiles. For this purpose a spin label—TEMPOacetamide—was covalently linked to the Cys-25 residue of the papain active center. ESR spectra of labeled papain indicated that catalytic activity of papain could be related to the conformational rigidity near the reaction center. The lower activities obtained in reverse micelles could be a result of the greater degree of mobility and polarity observed in these systems, which can be attributed to papain unfolding. The greater stability found for papain in reverse micelles could be the result of the limited extent of this denaturing process owing to the organized surfactant molecules around the enzyme.