键合型手性固定相的研究进展

手性分离在生物医药等领域具有重要意义。高效液相色谱(HPLC)因其经济、快速、高效等特点被广泛应用于手性化合物的分离分析中。手性固定相(CSP)是HPLC实现手性分离的核心,而制备有效CSP的关键在于手性选择剂的筛选。近年来,大量文献报道了新型CSPs的制备,其中键合型CSPs因具有溶剂耐受性和较高稳定性等优点受到了广泛关注。该文对近年来以手性单分子、多糖、环糊精、大环抗生素、冠醚、杯芳烃及生物碱等为手性选择剂制备的新型键合型CSPs进行了归纳整理,并对其发展前景进行了展望。     

Full enzymatic hydrolysis of commercial sucrose laurate by immobilized-stabilized derivatives of lipase from Thermomyces lanuginosa

Sucrose laurate is a detergent that is useful for various biochemical applications because it is a green compound and is easily degradable after hydrolysis with a lipase or esterase. One problem observed in the process of sucrose laurate degradation is that most commercial detergent preparations are impure, necessitating the hydrolysis of all of the sucrose esters present in the preparation, all of them with detergent properties. In this article, a highly active catalyst, which is able to perform the hydrolysis of commercial sucrose laurate, is presented. The use of glyoxyl agarose preparations of a previously aminated Thermomyces lanuginosa lipase (TLL) enabled complete hydrolysis, in less than 30 min, of all of the compounds that comprise the mixture. In addition, this derivative is stable in the presence of 20% ethanol, which is necessary to prevent microbial contamination.     

人神经母细胞瘤的磷酸化膜蛋白质组分析

针对人神经母细胞瘤SH-SY5Y细胞系的磷酸化膜蛋白质组,发展了基于多酶酶解法结合杂化硅胶基质固定化钛离子亲和色谱(Ti4+-IMAC)整体柱富集的分析策略。该方法通过对细胞裂解液进行超速离心,以及1 mol/L NaCl和0.1 mol/L Na2CO3顺序清洗,获得膜蛋白质组分。所提取的蛋白质分别经胰蛋白酶、胰凝乳蛋白酶和胃蛋白酶平行酶解,产生的肽段经Ti4+-IMAC整体柱选择性富集磷酸肽后,采用纳升级反相液相色谱分离和质谱鉴定,成功鉴定到43个磷酸化蛋白质,其中有14个定位于膜上。研究结果表明,采用该策略开展SH-SY5Y细胞系磷酸化膜蛋白质组学分析有望加速对该肿瘤的研究和相关潜在标记物的筛选。     

碳酸钙固定化猪胰脂肪酶的制备及活性研究

本文利用市售普通碳酸钙(general calcium carbonate,GCC)作为猪胰脂肪酶(PPL)固定化的载体材料,研究了不同固定化条件对所制备的固定化酶活性的影响.结果显示PPL固定化于GCC上的最佳制备工艺为:将20 mL酶液与1.50 g GCC放入置于摇床内的三角瓶中,摇床温度设为40℃,摇床转速设为...     

纳豆菌细胞包埋材料的选择

以海藻酸钠（SA）和聚乙烯醇（PVA）为包埋材料，采用固定化细胞技术对纳豆菌生产纳豆激酶进行了研究。研究发现，在PVA中加入SA进行细胞包埋可获得渗透性能好强度高的固定化细胞，通过正交试验进一步确定：当PVA的浓度为11％，SA的浓度为1％，硼酸的浓度为5％，CaCl2的浓度为6％时，固定化细胞的强度最好，可反复使用6批次，活性也很高，产生的纳豆激酶酶活溶纤圈直径积达88mm^2／15μL。     

31P CP/MAS NMR of polycrystalline and immobilized phosphines and catalysts with fast sample spinning

Cross‐polarization (CP) at fast magic angle spinning (MAS) frequencies leads to a splitting of the Hartmann–Hahn (HH) matching profile into a centerband and additional bands of higher orders. The matching profiles differ with the substance categories. Therefore, signal intensity is usually lost, when e.g. the routine standard NH₄H₂PO₄ is used for optimizing the ¹H–³¹P HH match prior to measuring phosphines and their metal complexes in polycrystalline or immobilized form. Here, a variety of model compounds, such as Ph₂PCH₂CH₂PPh₂ and (CO)₂Ni(PPh₃)₂, which can be used as ³¹P CP standards for analogous substances or materials are presented. Investigating the influences of MAS frequency, contact time, ¹H pulse power and sample volume on the matching profiles of the model compounds leads to general trends. Thereby, a new strategy for measuring difficult samples with CP at high MAS rates has been developed: their optimum CP parameters are derived from the most intense maxima in the HH matching profiles of the corresponding model compounds. This new strategy is compared with variations of a conventional ramp sequence. Although the latter generally provide smaller signal half‐widths, the new strategy leads to higher signal intensities. The new method was successfully applied to polycrystalline and immobilized phosphines and catalysts. Copyright © 2003 John Wiley & Sons, Ltd.     

Wood cellulignin as an alternative matrix for enzyme immobilization

The objective of this work was to select an efficient methodology for preparing active samples of Candida rugosa lipase immobilized in wood cellulignin, to be applied in hydrolysis and ester reactions. For this purpose, lipase was immobilized in the matrix by physical adsorption (pure cellulignin) and covalent binding (activated cellulignin with glutaraldeyde or carbonyldiimidazole [CDI]) in the presence or absence of polyethylene glycol (PEG) (Molecular mass of 1500 Daltons) as stabilizing agent. The activating agent and the presence of PEG-1500 in the immobilization procedure showed a strong influence on enzyme retention in the support. The values for enzyme retention ranged from 20 to 68%, and the highest yield was obtained when the enzyme was immobilized in cellulignin activated with CDI in the presence of PEG-1500. This immobilized derivative presented high hydrolytic (193.27 μM/[mg·min]) and synthetic (522.92 μM/[g·min]) activities when compared with those obtained by other techniques. The superiority of this immobilized system was confirmed by additional analyses, such as infrared spectroscopy and elemental analysis, which demonstrated an appropriate enzyme fixation and the highest level of protein incorporation in the support. Further information on the immobilized derivative was obtained by assessing the recycle potential in both aqueous and nonaqueous media.     

Enzymatic ring-opening polymerization of 2,2-dimethyltrimethylene carbonate catalyzed by PPL immobilized on silica nanoparticles

Silica nanoparticles were first used as the carrier for the porcine pancreas lipase (PPL) immobilization. The result of transmission electron microscopy (TEM) showed that the immobilized lipase was still in nanosize after enzyme immobilization. The ring-opening polymerization of 2,2-dimethyltrimethylene carbonate (DTC) catalyzed by this immobilized PPL (IMPPL) was explored. ¹H NMR spectra suggested no evidence of decarboxylation during propagation. Influences of IMPPL concentration and reaction temperature on the molecular weight and yield of poly(DTC) were studied. The recovery and reuse of IMPPL for the ring-opening polymerization of DTC was also investigated. The recycling IMPPL showed even higher catalytic activity and a higher molecular weight of polycarbonate could be achieved.     

Diffusion and reaction in an immobilized enzyme starch saccharification catalyst

Effectiveness factors were predicted from measurements of basic parameters made on single oligosaccharides, and the prediction was compared to experimental effec tiveness factors for the reaction of each oligosaccharide in the immobilized enzyme catalyst. Kinetic parameters were obtained for the hydrolysis of each oligosaccharide catalyzed by soluble glucoamylase, and were fit with a subsite model equation capable of generalization to all sizes of oligosaccharide. Diffusion coefficients in free solution were determined from movement out of a capillary tube. Spatial characteristics of the immobilized enzyme bed were obtained from pulse response experiments, allowing the calculation of effective diffusivities. Experimental effectiveness factors plotted against modulus were in reasonable agreement with the predictions.