核苷中2'-位取代基对Burkhoideria cepacia脂肪酶选择性酰化核苷的影响

研究了核苷结构中2'-位含有不同取代基时Burkholderia cepacia脂肪酶促3'-位羟基高区域选择性酰化反应.结果表明,该脂肪酶催化核苷类似物酰化的优势产物均为3'-酯,且随着酰基供体链的增长,3'-区域选择性逐渐升高;在酶促核苷癸酰化反应中,当核苷2'-位含有H,F,Cl和Br取代基时,3'-区域选择性最...     

PSIM催化核苷类化合物3'-酯选择性合成中的底物识别

研究了有机溶剂中酶促核苷类化合物3'-酯高区域选择性的合成反应,探讨了核苷结构中R1位取代基变化对酶促酰化反应的影响.结果表明:来源于Burkholderia cepacia的固定化脂肪酶PS IM在有机溶剂THF中的催化效果最佳,且酰化反应的优势位点均为3'-羟基(90%～>99%);随着核苷中R1位取代基由F变化至CF3或酰基供体链长由C6增加至C14,该酰化反应的3'-区域选择性均呈现逐渐增加的趋势,构效关系分析表明,这主要源于PS IM酶特殊的活性中心结构.     

PS IM 催化核苷类化合物3＇-酯选择性合成中的底物识别

研究了有机溶剂中酶促核苷类化合物3＇-酯高区域选择性的合成反应,探讨了核苷结构中R1位取代基变化对酶促酰化反应的影响.结果表明:来源于Burkholderia cepacia的固定化脂肪酶PS IM在有机溶剂THF中的催化效果最佳,且酰化反应的优势位点均为3＇-羟基(90％～＞99％)；随着核苷中R1位取代基由F变化至CF3或酰基供体链长由C6增加至C14,该酰化反应的3＇-区域选择性均呈现逐渐增加的趋势,构效关系分析表明,这主要源于PS IM 酶特殊的活性中心结构.     

溶剂和酰基受体对α-氰基-3-苯氧基苄醇乙酯的酶促对映体选择性转酯化反应的影响

研究了在有机溶剂中固定化Alcaligenes sp.脂肪酶催化α-氰基-3-苯氧基苄醇乙酯的对映体选择性转酯化反应,考察了不同性质的溶剂和酰基受体对酶的催化活性和选择性以及对产物稳定性的影响.结果发现在弱极性溶剂如正己烷中酶具有较高的催化活性但产物e.e.%值低,而且容易分解;在四氢呋喃等溶剂中酶催化活性相对低,但产物e.e.%值高,也较为稳定;但反应时间太长,会导致产物分解及纯度下降;不同酰基受体对酶反应无显著影响,甲醇为最佳酰基受体,太多醇会导致反应速率下降;溶剂水含量大于2.0%时对酶活性和产物稳定性产生明显不利影响.在优化条件下,酶反应可得到(S)-α-氰基-3-苯氧基苄醇产率＞48%,纯度＞99%e.e.     

溶剂和酰基受体对α-氰基-3-苯氧基苄醇乙醇的酶促对映体选择性转酯化反应的影响

研究了在有机溶剂中固定化Alcaligenes sp.脂肪酶催化α-氰基-3-苯氧基苄醇乙酯的对映体选择性转酯化反应，考察了不同性质的溶剂和酰基受体对酶的催化活性和选择性以及对产物稳定性的影响．结果发现在弱极性溶剂如正己烷中酶具有较高的催化活性但产物e．e．％值低，而且容易分解；在四氢呋喃等溶剂中酶催化活性相对低，但产物e．e．％值高，也较为稳定；但反应时间太长，会导致产物分解及纯度下降；不同酰基受体对酶反应无显著影响，甲醇为最佳酰基受体，太多醇会导致反应速率下降；溶剂水含量大于2．0％时对酶活性和产物稳定性产生明显不利影响．在优化条件下，酶反应可得到（S）-α-氰基-3-苯氧基苄醇产率＞48％，纯度＞99％e．e．     

离子液体中酶促区域选择性合成CFAE

碳水化合物脂肪酸酯(CFAE)作为一类非离子型生物表面活性剂,被广泛用于食品、医药及化妆品工业,一些CFAE还具有抗菌、抗肿瘤等特殊生物活性。非水介质中酶促区域选择性合成CFAE反应的瓶颈在于高极性碳水化合物与酰基供体不易相溶,并具有多个可酰化羟基。传统有机溶剂虽能提高极性底物在体系中溶解度,但常使酶活下降。新型绿色介质离子液体用于CFAE的酶法合成过程具有诸多优点,不仅生物催化剂能维持较好的活性与稳定性,且良好的底物溶解性可改善反应区域选择性及转化速率,反应体系还可重复利用。本文介绍了离子液体中影响酶促区域选择性合成CFAE反应的主要因素,包括酶、底物在离子液体中的溶解性能及底物自身性质等,详述了离子液体中酶促制备CFAE的研究进展,也指出了酶促合成CFAE存在的问题与发展前景。     

非水介质中曲克芦丁乙烯酯的酶促合成

在非水介质中,选择性催化曲克芦丁羟乙基上的伯羟基发生酯交换反应,合成一系列曲克芦丁乙烯酯.同时考察了酶源、酶量、反应介质、酰化试剂链长、溶剂含水量、反应时间等因素对曲克芦丁酶促酯交换反应区域选择性的影响.结果表明:以吡啶为溶剂,其含水量1%时,枯草杆菌碱性蛋白酶催化活性最高,随着酰基供体碳链的减少,产率增大,且反应达平衡的时间缩短.     

高取代度魔芋葡甘聚糖醋酸酯的制备及其热塑性和流变性研究

以魔芋葡甘聚糖(KGM)为原料,浓硫酸为催化剂,乙酸酐为改性剂,制备了KGM醋酸酯.研究了反应条件对KGM醋酸酯取代度(DS)的影响,KGM醋酸酯制备的最佳反应条件是乙醇∶水量比5∶5,反应时间2h,反应温度75℃,催化剂浓硫酸的浓度0.1mol/L,KGM与乙酸酐的量比5∶40(g/mL),取代度高达2.93,其分子量与KGM相比,则明显降低.运用傅立叶红外光谱仪(FTIR)、X射线衍射仪(XRD)、扫描电镜(SEM)、热重分析仪(TGA)、示差扫描量热仪(DSC)和旋转流变仪对KGM和高取代度KGM醋酸酯进行了表征.结果表明,KGM醋酸酯的羰基(CO)特征吸收峰较KGM明显增强,其表观形貌大部分为疏松絮状,KGM及其醋酸酯均为非晶态结构.与KGM相比,KGM醋酸酯的热稳定性下降,分解温度(Td)由KGM的261.10℃降低至KGM醋酸酯的204.56℃,KGM醋酸酯出现了明显的玻璃化转变,其玻璃化转变温度(Tg)为128.49℃.KGM醋酸酯是典型的黏弹性材料,其弹性比率占21.27%,其剪切黏度η对温度变化非常敏感,可通过温度的改变来调节KGM醋酸酯的加工流动性.KGM醋酸酯具有较好的热塑性.     

草酸淀粉酯制备及其性能研究

本文以草酸和玉米淀粉为原料,通过改变原料的摩尔比反应制备了不同取代度(0.1到0.9)的草酸淀粉酯,采用滴定法测定产物的取代度,利用红外光谱和核磁共振表征产物(取代度为0.41)的化学结构,产物中含有羰基的结果表明成功制备了草酸淀粉酯.详细考察了草酸淀粉酯的物理化学性质,利用粘度测定、热重分析、广角X衍射(WXRD)以及湿度吸收等研究不同取代度的草酸淀粉酯的分子量、热稳定性,结晶形态以及吸水性能.结果说明,与玉米淀粉相比,草酸淀粉酯的吸水率随着取代度的提高而增加,其分子量、热稳定性以及结晶性能则呈下降趋势.     

壳聚糖与聚乙二醇交联水凝胶研究

研究对壳聚糖(CS)进行化学修饰得到了不同丙烯酰基取代度(1.03%,3.55%和5.21%)的丙烯酰化羟丙基壳聚糖(AHCS);通过自由基引发反应,AHCS与聚乙二醇二丙烯酸酯(PEGDA)交联得到壳聚糖(CS)与聚乙二醇(PEG)为主体的交联水凝胶。通过SEM观察其为通透性良好的多孔性支架材料。水凝胶的溶胶含量和溶胀度随丙烯酰基取代度的增加而降低,水凝胶中壳聚糖的降解速率也随丙烯酰基取代度的升高而降低。对于同一取代度的交联水凝胶,其在酸性和碱性条件下的溶胀度大于中性环境。细胞试验表明,壳聚糖与聚乙二醇交联水凝胶具有良好的生物相容性。     

Thermolysin catalyses the synthesis of cyclodextrin esters in DMSO

Fatty acid esters of cyclodextrins (CDs) were synthesised in a one-step reaction with native CDs as acyl acceptors and vinyl-activated fatty acid esters as acyl donors. Immobilised preparations of thermolysin, subtilisin, the alkaline protease AL-89 and Candida antarctica lipase B were investigated for their catalytic properties regarding transesterification in solvents of increasing hydrophilicity. The synthesis of cyclodextrin fatty acid esters was proved to be catalysed enzymatically by thermolysin in DMSO. The obtained products were analysed by TLC and their structures characterised by NMR, MS and FTIR spectroscopy. With vinyl decanoate as acyl donor β-CD was esterified at all seven glucose C-2 positions resulting in heptakis(2-O-decanoyl)-β-cyclodextrin as the major product. With vinyl butyrate, substitution occurred at all the C-2 and partially at the C-3 or C-6 positions resulting in an average degree of substitution of nine. Between 20% and 25% (w/w) of the acyl donor was converted to esters in 20 h corresponding to an estimated total conversion of the acyl acceptor in the case of maltosyl-β-CD. In the subtilisin and AL-89 catalysed reactions, product formation was simultaneously catalysed non-enzymatically by inorganic buffer salts in aprotic, hydrophilic solvents and with the lipase no products were formed in any of the solvents investigated.     

Lipase-catalyzed glucose fatty acid ester synthesis in ionic liquids

[reaction: see text] Glucose fatty acid ester synthesis with poly(ethylene glycol)-modified Candida antarctica lipase B (CAL-B) was performed in pure 1-butyl-3-methyl imidazolium tetrafluoroborate [BMIM][BF(4)] (30% conversion) and in pure 1-butyl-3-methyl imidazolium hexafluorophosphate [BMIM][PF(6)] (35% conversion). In a solvent system composed of ionic liquid and 40% t-BuOH conversions up to 90% and isolated yields of up to 89% were achieved using fatty acid vinyl esters as acyl donors and commercial CAL-B.     

Synthesis of Long Chain Fatty Acids Acylated Coumarin Glycoside Esters with Lipase as Catalyst

A series of novel coumarin glycoside esters(1―9) was synthesized through the acylation reaction of 4-methylcoumarin-7-O-β-D-glucoside(11) with different long chain fatty acids catalyzed by lipase in organic medium. The acylation occurred regioselectively at the 6'-OH of glycosyl moiety. The enzymatic synthesis was optimized to achieve 54%―70% yield using immobilized lipase(Novozym 435, 10 mg/mL) as catalyst and acetone and pyridine(9:1, volume ratio, water content<1%) as solvent with an acyl donor/coumarin glycoside molar ratio of 10:1 at a temperature of 40―50 ℃ for 96 h. All the synthesized compounds were confirmed.     

Regioselective enzyme-catalyzed synthesis of sophorolipid esters,amides, and multifunctional monomers

Novel enzyme-mediated synthetic routes were developed to provide a new family of sophorolipid derivatives and glycolopid-based amphiphilic monomers. These compounds are of great interest for their potential use in immunoregulation, as well as for other biological properties. In the present work, an efficient lipase-catalyzed conversion of sophorolipid ethyl ester to (a) the 6'-monoacylated derivatives using Novozym 435, (b) 6' '-monoacylated derivatives using Lipase PS-C, (c) secondary amide derivatives using Novozym 435, and (d) 6',6' '-diacylated amide derivatives using Novozym 435 in an one-pot reaction and (e) the regioselective monoacylation of an amide derivative at the 6'- and 6' '-positions using Novozym 435 and Lipase PS-C, respectively, are described. The ethyl ester produced by esterification of the sophorolipid mixture with sodium ethoxide was subjected to acylation catalyzed by Novozym 435 in dry tetrahedrofuran (THF) with vinyl acetate and vinyl methacrylate to produce 6'-monoacylated derivatives. In contrast, Lipase PS-C catalyzed acylations of sophorolipid ethyl ester in dry THF with vinyl acetate and vinyl methacrylate to give the corresponding 6'-monoacylated derivatives. Novozym 435 mediated amidation of sophorolipid ethyl ester in dry THF with phenethylamine, tyramine, p-methoxyphenethylamine, 2-(p-tolyl)ethylamine, and p-fluorophenethylamine generated the corresponding secondary amides but not tertiary amides. The formation of diacyl derivatives of amides was achieved by their treatment with vinyl acetate and vinyl methacrylate in dry THF using Novozym 435 as catalyst. The conversion of sophorolipid ethyl ester to the same diacyl derivatives of amide (i.e., both amidation and acylation) in high yield was also demonstrated in dry THF by a one-pot reaction using Novozym 435. Furthermore, regioselective monoacylation of a sophorolipid amide at 6' and 6' ' in dry THF with vinyl acetate and vinyl methacrylate using Novozym 435 and Lipase PS-C was also demonstrated.     

Kinetic resolution of fluorinated propargyl alcohols by lipase-catalyzed enantioselective transesterification

In order to obtain optically active fluorinated propargyl alcohols, a lipase-catalyzed kinetic resolution has been carried out. The effect of lipase types, organic solvents, reaction temperature, and acyl donors was examined in the lipase-catalyzed transesterification of 1,1,1-trifluoro-4-phenyl-3-butyn-2-ol. Various enantiomerically pure fluorinated propargyl alcohols have been successfully prepared in good enantiomeric excess (>84%) by Novozym 435-catalyzed transesterification with vinyl butanoate at 60 °C in n-hexane. In some cases, the enantiomeric purities were excellent (>99% ee).     

Enzymatic synthesis of sorbitan methacrylate according to acyl donors

Recently, sugar polymers have been considered for use as biomaterials in medical applications. These biomaterials are already used extensively in burn dressings, artificial membranes, and contact lenses. In this study, we investigated the optimum conditions under which the enzymatic synthesis of sorbitan methacrylate can be affected using Novozym 435 in t-butanol from sorbitan and several acyl donors (ethyl methacrylate, methyl methacrylate, and vinyl methacrylate). The enzymatic synthesis of sorbitan methacrylate, catalyzed by Novozym 435 in t-butanol, reached an approx 68% conversion yield at 50 g/L of 1,4-sorbitan, 5% (w/v) of enzyme content, and a 1∶5 molar ratio of sorbitan to ethyl methacrylate, with a reaction time of 36 h. Using methyl methacrylate as the acyl donor, we achieved a conversion yield of approx 78% at 50 g/L of 1,4-sorbitan, 7% (w/v) of enzyme content, at a 1∶5 molar ratio, with a reaction time of 36 h. Sorbitan methacrylate synthesis using vinyl methacrylate as the acyl donor was expected to result in a superior conversion yield at 3% (w/v) of enzyme content, and at a molar ratio greater than 1∶2.5. Higher molar ratios of acyl donor resulted in more rapid conversion rates. Vinyl methacrylate can be applied to obtain higher yields than are realized when using ethyl methacrylate or methyl methacrylate as acyl donors in esterification reactions catalyzed by Novozym 435 in organic solvents. Enzyme recycling resulted in a drastic reduction in conversion yields.     

Highly Efficient Regioselective Synthesis of 5′-<Emphasis Type="Italic">O</Emphasis>-lauroyl-5-azacytidine Catalyzed by <Emphasis Type="Italic">Candida antarctica</Emphasis> Lipase B

Enzymatic regioselective acylation of 5-azacytidine with vinyl laurate was successfully conducted with an immobilized lipase from Candida antarctica type B (i.e., Novozym 435) for the first time. The acylation of 5-azacytidine took place at its primary hydroxyl group and the desired product 5′-O -lauroyl-5-azacytidine could be prepared with high reaction rate, high conversion, and excellent regioselectivity. The influences of several key variables on the enzymatic acylation were also systematically examined. Pyridine was found to be the best reaction medium. The optimum initial water activity, the molar ratio of vinyl laurate to 5-azacytidine and reaction temperature were 0.07, 30:1, and 50 °C, respectively. Under the optimized conditions described above, the initial reaction rate, the substrate conversion, and the regioselectivity were as high as 0.58 mM/min, 95.5%, and >99%, respectively, after a reaction time of around 5 h.     

Kinetics and modeling of (R,S)‐1‐phenylethanol acylation over lipase

The kinetics of the acylation of (R,S)‐1‐phenylethanol was investigated using lipase as a catalyst. The main parameters were temperature, reaction atmosphere, different acyl donors, and different amounts of acyl donor as well as the presence of some additives in the reaction mixture. The initial reaction rate increased with increasing temperature and with a decreasing amount of an acyl donor. The activated esters, such as isopropenyl‐ and vinyl acetate, exhibited very high acylation rates for R‐1‐phenylethanol, whereas low rates were obtained with ethyl acetate and 2‐methoxyethyl acetate. The addition of water and acetophenone decreased the acylation rate. A kinetic model was developed based on a sequential step mechanism, in which enzyme was reacting in the first step with an acyl donor followed by the reaction of a modified enzyme complex with the reactant, R‐1‐phenylethanol. Comparison with experimental data obtained at different temperatures allowed simplification of this model, leading to a kinetic equation with just one apparent parameter. The influence of the amount of acyl donor, ethyl acetate, could be quantitatively described by taking into account the competitive inhibition of the ethanol produced. The rate constants and apparent activation energy for experiments performed under different temperatures and the amounts of acylation agent were determined. The apparent activation energy was 24.5 kJ/mol. © 2010 Wiley Periodicals, Inc. Int J Chem Kinet 42: 629–639, 2010     

Synthesis of Geraniol Esters in a Continuous-Flow Packed-Bed Reactor of Immobilized Lipase: Optimization of Process Parameters and Kinetic Modeling

With increasing demand for perfumes, flavors, beverages, and pharmaceuticals, the various associated industries are resorting to different approaches to enhance yields of desired compounds. The use of fixed-bed biocatalytic reactors in some of the processes for making fine chemicals will be of great value because the reaction times could be reduced substantially as well as high conversion and yields obtained. In the current study, a continuous-flow packed-bed reactor of immobilized Candida antarctica lipase B (Novozym 435) was employed for synthesis of various geraniol esters. Optimization of process parameters such as biocatalyst screening, effect of solvent, mole ratio, temperature and acyl donors was studied in a continuous-flow packed-bed reactor. Maximum conversion of ~ 87% of geranyl propionate was achieved in 15 min residence time at 70 °C using geraniol and propionic acid with a 1:1 mol ratio. Novozym 435 was found to be the most active and stable biocatalyst among all tested. Ternary complex mechanism with propionic acid inhibition was found to fit the data.     

Enzyme catalyzed production of biodiesel from olive oil

Biodiesel (fatty acid methyl esters) was produced by transesterification of triglycerides (triolein) present in olive oil with methanol and Novozym435. The effect of the molar ratio of methanol to triolein, semibatch (stepwise addition of methanol) vs batch operation, enzyme activity, and reaction temperature on overall conversion was determined. Stepwise methanolysis with a 3:1 methanol to triolein molar ratio and an overall ratio of 8:1 gave the best results. The final conversion and yield of biodiesel were unaffected by initial enzyme concentrations greater than 500 U/mL olive oil. The optimum reaction temperature was 60 degrees C. Comparison of conversion data between a test-tube scale reactor and a 2-L batch reactor revealed that the difference in conversion was within 10%. Experiments were also carried out with used cooking oil; the conversion with used cooking oil was slightly lower but no major differences were observed. The efficacy of Novozym435 was determined by reusing the enzyme; although the enzyme's relative activity decreased with reuse, it still retained 95% of its activity after five batches and more than 70% after as many as eight batches.