酶促制备丝裂霉素类似物反应动力学模型研究

基于固定化T. laibacchii脂肪酶催化2-甲基-1,4-苯醌与正丁胺的Michael加成反应建立了酶促制备丝裂霉素类似物2-甲基-3-正丁胺酰-1-氢-4-醌的反应动力学模型.该反应在柠檬酸缓冲溶液(pH值为7.0)中进行,最终产率可达98%.该文提出了修正的有序双双和随机双双机理,采用King-Altman方法得到相关微分方程组以表示即时反应速率.通过联合解微分方程和最优化方法确定动力学模型参数,使用ode45程序解微分方程组,并运用Fmincon软件计算动力学常数.研究结果表明:模型拟合值与实验值的平均相对偏差为11.25%,且偏差服从关于y=0的轴对称分布.当固定化酶粒径为0.5 mm、搅拌转速为200 rpm时可以忽略内外扩散限制.该文建立的动力学模型为固定化酶固有动力学模型.     

Microcalorimetric Determination of Serum Triglycerides and Cholesterol

Abstract

Reaction heats measured in a microcalorimeter between varying amounts of human serum and 700 IU of lipase (EC 3.1.1.3) and those between serum and 8 U of cholesterol oxidase (EC 1.1.3.6) were significantly linear with the contents of triglycerides (r=1.00) and cholesterol (r=0.99), respectively. However, when sera from 13 human volunteers were subject to comparative study between the micro-calorimetric and spectrophotometric methods, the correlation became less appreciable (r=0.70 and r=0.71) presumably due to the individual variations in free glycerol and cholesterol ester-constituents that were deliberately omitted in the microcalorimetric method but were routinely included in the standard (multi-enzymatic) spectro-photometric clinical method. The contribution of interfering substances, if any, is not totally ruled out.     

水含量对C.cylindracea脂肪酶催化有机硅化合物生物转化的影响

研究了水含量对有机溶剂中脂肪酶催化有机硅化合物的生物转化的影响。水含量可影响酶生物转化反应的反应速度，化学反应平衡及化学反应最适温度，在正己烷有机溶剂中酶催化有机硅醇酯化反应的最适水含量为０．０６（ｗ／ｗ）％。     

脂肪酶催化改性乳脂肪的应用研究

乳脂肪具有令人愉悦的风味但成分复杂，应用于食品领域时存在饱和脂肪酸含量较高，不利于健康，且在低温下涂抹性能较差等问题。利用脂肪酶的专一性和特殊性，可催化乳脂肪生成特殊结构的甘油三酿，得到风味饱满、营养价值高、利于应用的产物。现概述目前国内外脂肪酶酶解乳脂肪的研究进展及改性乳脂肪的应用，并介绍了用脂肪酶催化改性后乳脂肪的优点及应用，主要包括：生成更高含量的短链脂肪酸酯，提高低熔点甘油三酯的含量和降低饱和脂肪酸的含量三个方面。     

脂肪酶催化棕榈油甘油解反应合成单甘酯

研究了以猪胰脂肪酶为催化剂在无有机溶剂体系中由棕榈油通过甘油解瓜在合成单甘油酯，结果表明，当反应温度为２０℃，甘油与油脂摩尔比２：１，甘油中含水质量分数ｗ为４％，反应时间２０ｈ左右时获得的单甘酯ｗ达２４．１７％，降低体系的反应温度到１２～１３℃，单甘酯含量进一步提高，４ｄ后达到平衡，单甘酯ｗ为３０．９３％。     

固定化脂肪酶活性的X射线微区分析

利用X射线微区分析的方法，对吸附交联法得到的固定化脂肪酶的微观活性进行了分析。结果表明：以合成出的大孔吸附树脂为固定化酶栽体，酶活较高，活性脂肪酶分布较均匀。并得到了固定化脂肪酶的活性定位的最佳条件。     

脂肪酶催化牛油甘油解反应合成单甘酯

以猪胰脂肪酶为催化剂在无有机溶剂体系中由牛油通过甘油解反应可合成单甘油酯．研究表明，当反应温度为４２℃，甘油与油脂摩尔比为４∶１，甘油中水含量为４％时获得的单甘酯含量最高，达１９．５１％，反应在２４ｈ左右达到平衡．将反应混合物置于常温下储存，单甘酯含量继续提高，６ｄ后达到平衡，为２８．７２％．     

Phenylmethanesulfonyl fluoride pretreatment stabilizes plasma lipidome in lipidomic and metabolomic analysis

Though it is standard practice to test the stability of analytes in the matrix for routine bioanalytical method, stability evaluation is always impractical and skipped in untargeted lipidomic and metabolomic analysis because analytes in these studies are enormous, diverse and sometimes unknown. Lipidome represents a major class of plasma metabolome and shows great potential to be diagnostic and prognostic biomarkers. However, lipidome also faces stability problems because plasma contains kinds of lipid degradation enzyme. Here, using liquid chromatography time of flight mass spectrometry based lipidomic methodology, plasma levels of various lipids including triglyceride (TG), diglyceride (DG), free fatty acid (FFA), phosphatidylethanolamine (PE) phosphatidylcholine (PC), lyso-phosphatidylcholine (LPC), lyso-phosphatidylethanolamine (LPE), and sphingomyelin (SM) were dynamically determined within 4 h at ambient temperature. In mouse and rat plasma, the levels of most TG, DG, PC and PE species significantly decreased with respect to time, whereas those of LPC, LPE and FFA significantly increased with respect to time. However, such changes did not occur in human plasma, thus indicating hepatic lipase and esterase might involve in the species-specified degradation of lipid classes in plasma. Phenylmethanesulfonyl fluoride (PMSF) pretreatment prevented such lipidome instability in mouse plasma. The results suggested the instability of plasma lipidome should be highly concerned, and the enhancement of ex vivo stability of plasma lipidome could enable more reliable clinical translation of lipidomic data for biomarker discovery.     

A microconductometric biosensor based on lipase extracted from Candida rugosa for direct and rapid detection of organophosphate pesticides

Organophosphate pesticides (OPs) have been intensively used as insecticides in agriculture; after entering the aquatic environment, they may affect a wide range of organisms. A conductometric enzymatic biosensor based on lipase extracted from Candida rugosa (CRL) has therefore been developed for the direct and rapid quantitative detection of organophosphate pesticides: diazinon, methyl parathion and methyl paraoxon in water. The biosensor signal and response time were obtained under optimum conditions, the enzyme being immobilised in the presence of gold nanoparticles. Under these conditions, the enzymatic biosensor was able to measure concentrations as low as 60 µg/L of diazinon, 26 µg/L of methyl parathion and 25 µg/L of methyl paraoxon very rapidly (response time: 3 min). Moreover, this CRL biosensor was not sensitive to interferences such as carbamates. It presented good storage stability for 21 days when kept at 4°C and it was successfully applied to real samples.     

Ring‐Closing and Cross‐Metathesis with Artificial Metalloenzymes Created by Covalent Active Site‐Directed Hybridization of a Lipase

A series of Grubbs‐type catalysts that contain lipase‐inhibiting phosphoester functionalities have been synthesized and reacted with the lipase cutinase, which leads to artificial metalloenzymes for olefin metathesis. The resulting hybrids comprise the organometallic fragment that is covalently bound to the active amino acid residue of the enzyme host in an orthogonal orientation. Differences in reactivity as well as accessibility of the active site by the functionalized inhibitor became evident through variation of the anchoring motif and substituents on the N‐heterocyclic carbene ligand. Such observations led to the design of a hybrid that is active in the ring‐closing metathesis and the cross‐metathesis of N,N‐diallyl‐p‐toluenesulfonamide and allylbenzene, respectively, the latter being the first example of its kind in the field of artificial metalloenzymes.