退浆酶TF-162的酶活力的测试及应用评价

介绍了采用淀粉-碘量分光光度法测试退浆酶活力,通过实验验证,确定测定酶活最佳反应条件和测试条件,同时结合退浆酶在实际应用中的退浆效果,得出用淀粉-碘液显色法检测退浆效果的应用评价方法。     

超声辅助胶内酶切方法的优化及其酶切特点的分析

分别应用20、100和200 W超声功率对牛血清白蛋白以及人尿蛋白组的胶内条带进行胶内酶切,通过比较不同功率鉴定的多肽数和蛋白数,确定最优的超声功率。同时比较超声辅助酶切与传统过夜酶切方法,并分析超声辅助胶内酶切方法的特点。结果表明,超声功率为20 W时对胶粒破坏最小,样品损失最少,在牛血清白蛋白样品中鉴定的多肽数最多为65。在人尿蛋白组2个条带中,20 W方法鉴定的蛋白数分别为168和199,鉴定效果最优。通过对多肽误切率和不完全酶切多肽的定性和定量分析发现,超声酶切方法的误切率高于传统方法,但不完全酶切率较低。通过分析多肽误切位点发现,传统过夜酶切法和超声辅助酶切法的主要误切类型分别为脯氨酸（P）和天冬氨酸/谷氨酸（D/E）,3种超声方法之间各种误切类型所占比例均无明显差别。超声辅助酶切可显著缩短蛋白质胶内酶切时间,其中20 W功率超声酶切的效果最佳。与传统过夜酶切法相比,超声辅助酶切多肽的鉴定结果更好,其多肽的误切率更高,不完全酶切率较低,可用于发现蛋白质组学研究。     

酶解辅助乙醇法提取土茯苓总黄酮条件研究

本文采用植物复合酶酶解辅助乙醇法提取土茯苓总黄酮.以芦丁为标准品,用铝离子显色、紫外-可见分光光度法测定总黄酮含量.采用单因素实验探讨酶添加量、酶解时间和酶解pH值对总黄酮提取率的影响.结果表明:最佳提取条件为酶添加量1.0%,酶解时间3.0h和酶解pH值5.0;最佳条件下,粗提物干粉得率13.9%,干粉中总黄酮含量4.0%,总黄酮提取率为0.53%.     

纳米金标记的重组核糖核酸酶谱分析技术的生物信息学评价

现代科学仪器和分析测试技术业已在众多的领域，特别是在生命科学技术领域获得越来越广泛的应用。常规的测试和分析工作往往限于对样品直接可测试参数的获得和评价，而对更深层次的有效信息的提取和分析评价的应用及研究较少。本文通过对由纳米标记的合成短肽重组的核糖核酸酶RNase S与野生的核糖核酸酶RNase A结构和功能的此较，探讨了在联合使用高效液相色谱（HPLC）分离纯化技术以及紫外-可见光吸收光谱和圆二色性谱（CD）等现代谱仪分析技术在生物信息学方面应用的可行性。研究结果表明。现代科学仪器及分析测试技术在生物信息学等方面有着潜在的发展活力。     

酶电极中酶固定化方法研究进展

酶电极是最常用也是最早开发的生物传感器,而固定化酶作为酶电极的关键也得到了广泛的关注。本文介绍了酶电极及固定化酶的特点,重点讨论了制作酶电极的关键技术即酶固定化的方法,包括吸附法、包埋法、共价键合法和交联法等传统方法和静电纺丝法、纳米技术处理等一些新型的固定化方法,并进一步探讨了各种固定化方法的优缺点。     

碘显色-连续流动法测定雪茄烟草中的淀粉

以磷酸溶液调节反应酸度,基于碘化钾/碘酸钾混合溶液体系在酸性条件下进行氧化还原反应产生碘,在660nm波长下测量碘与淀粉反应溶液的吸光度值,建立了碘显色-连续流动法测定烟草中淀粉的方法。应用于雪茄烟外包烟叶、内包烟叶、芯烟以及烤烟、白肋烟、香料烟叶中淀粉含量的检测,方法操作简便、分析速度快、精密度高;平均回收率为98.5%,相对标准偏差小于2.0%,相对误差不超过±4.0%。研究结果表明,雪茄烟叶中的淀粉含量与白肋烟相当,但显著低于烤烟,也低于香料烟叶中的淀粉量。     

MB—Ⅲ型酶标检测仪的原理及应用

MB-Ⅲ型酶标检测仪是酶联免疫吸附试验(ELISA)的专用仪器,它具有性能稳定、灵敏度高、重复性好、使用安全可靠等特点,广泛应用于临床诊断、微生物学、免疫学、病理学、生物化学、药物学、兽医学及农业科学等领域。拓展了系统软件的应用范围后在检测鸡传染性囊病抗体(IBD)中也取得较好效果。     

芽孢杆菌所产果胶裂解酶检测条件的研究

果胶酶(Pectinase)一般是指分解植物组织中果胶质的一类含多种酶的复合酶,是一类具有广泛用途的生物酶.酸性果胶酶广泛应用于果胶和白酒的提取和澄清;碱性果胶酶可用于香料油和类胡萝卜素等医用原料的提取等过程.近年来又发现了碱性果胶酶新的用途,如植物病毒的纯化、纸浆漂白和纺织品的生物精练等,碱性果胶酶已经成为棉织物处理过程中重要的生物制剂.酶活力是衡量酶生物催化功能的重要指标,但酶活力的测定通常会受到测定温度、PH、底物浓度等多种因素的影响,因此,选择适宜的酶活力测定条件,提高测定结果的准确性,对于更好的开发和利用微生物果胶酶十分必要.本研究通过对芽孢杆菌所产果胶裂解酶酶学性质的研究,在此基础上确定了芽孢杆菌所产果胶裂解酶适宜的作用条件温度为50℃,底物聚半乳糖醛酸的浓度为0.2%,酶反应的PH值为9.4.     

固定化酶在仪器分析中的应用

对固定化酶的特点和制备方法，固定化酶反应器在高效液相色谱，流动注射分析和生物传感器中的应用作了介绍。引用文献４３篇。     

基质辅助激光解吸电离飞行时间质谱法分析核糖核酸酶

应用基质辅助激光解吸电离飞行时间质谱法(MALD I-TOF-MS)成功地对核糖核酸酶相对分子量进行测定。探讨三种不同基质对其分析结果的影响,实验中发现α-氰基-4-羟基肉桂酸是适宜的基质,该方法是传统生物方法无法比拟的。     

固定化酶及其在食品和生物领域的研究进展

固定化酶技术是酶工程的核心,渗透在现代各种工艺之中,成为不可或缺的工具。本文对其发展、优缺点和固定技术做了介绍,并简述了其在生物和食品方面的一些研究和应用。     

基于微流控和酶比色的微创血糖连续检测仪

连续血糖检测对糖尿病的诊断与治疗具有十分重要的意义。本文设计了一种集成化、自动化的微创血糖连续检测仪器,该仪器通过微流控芯片透皮抽取组织液,利用单片机精确测量透皮抽取组织液的体积,并采用酶比色法检测组织液的葡萄糖浓度,利用组织液与血液的葡萄糖浓度相关性实现连续血糖检测。由于透皮抽取的组织液体积很小且分散在皮肤表面,为了便于收集,利用生理盐水对抽取出的组织液进行稀释,稀释后的组织液中葡萄糖浓度在3～50mg/dL。为了测量低浓度葡萄糖,实验选取了1～50mg/dL中的10个浓度的葡萄糖溶液进行吸光度测量,根据光谱数据与葡萄糖浓度建立吸光度模型,结果表明该酶比色检测方法在1～50mg/dL葡萄糖浓度内具有良好的线性度,测量相对标准偏差小于0.65%。该仪器能够实现自动化控制,为糖尿病的诊断提供依据,在微创血糖连续检测领域具有良好的应用前景。     

Mass spectrometry for enzyme assays and inhibitor screening: an emerging application in pharmaceutical research

Robust methods that monitor enzyme activity and inhibitor potency are crucial to drug discovery and development. Over the past 20 years, mass spectrometric methods have increasingly been used to measure enzyme activity and kinetics. However, for rapid screening of inhibitory compounds, various forms of fluorescence and chemiluminscence readout have continued to dominate the market. As the sensitivity, speed, and miniaturization of mass spectrometry methods continue to advance, opportunities to couple mass spectrometry with screening will continue to come to the forefront. To appreciate the tremendous potential for MS‐based screening assays, it becomes necessary to understand the current state of capabilities in this arena. Thus, this review is intended to capture how mass spectrometry for studying enzymes activity has progressed from simple qualitative questions (i.e., is the product detected?) to quantitative measures of enzyme activity and kinetics and then as a tool for rapidly screening inhibitory compounds as an alternative to current methods of high throughput drug screening. © 2007 Wiley Periodicals, Inc., Mass Spec Rev     

Imaging enzyme activity with polarization-sensitive confocal fluorescence microscopy

We describe a technique for imaging enzyme activity through steady‐state fluorescence anisotropy measurements on a per‐pixel basis with a confocal microscope. With this method, enzyme activity is reported by changes in the fluorescence anisotropy of a fluorescently labelled substrate. Enzymatic cleavage of the substrate yields smaller labelled fragments that tumble more readily than the intact substrate and therefore yield a lower anisotropy. Anisotropy is recovered to an accuracy of 7% or better on and off the optical axis to depths of 210 µm using objective numerical apertures as high as 0.75. Enzyme imaging experiments were performed with Bodipy‐FL‐labelled bovine serum albumin (BSA) attached to sepharose beads as a substrate for trypsin and proteinase K. Anisotropy images acquired up to 1 h after enzyme addition revealed more rapid digestion of BSA with proteinase K than with trypsin, but in both cases anisotropy decreased by at least five‐fold. Fluorescence lifetime and time‐resolved anisotropy decay measurements were made on the construct in fluid solution to reveal the effects of enzyme activity. The Bodipy‐FL lifetime increased from 1.34 ns for the construct without enzyme to 5.98 ns after 1 h in the presence of proteinase K. Anisotropy decays yielded average rotational correlation times of 1.13 ns before enzymatic action and 0.27 ns after enzymatic action, consistent with the presence of smaller Bodipy‐containing protein fragments. These results suggest wide applicability of the technique in biological systems when used in conjunction with appropriately designed constructs.     

实现微米级灭菌范围控制的微细等离子体射流装置

为了实现微细等离子体的精准灭菌,设计了新型微细等离子体射流装置,并对该装置产生的含氧活性粒子(Reac-tive Oxygen Species,ROS)和含氮活性粒子(Reactive Nitrogen Species,RNS)分布范围及其灭菌范围进行研究.淀粉碘化钾混合溶液里的碘离子可以被微细等离子体射流产生的ROS...     

Field-emission scanning electron microscopy analysis of morphology and enzyme distribution within an industrial biocatalytic particle

Field-emission scanning electron microscopy (FESEM) was used in a technical feasibility study to obtain insight into the internal morphology and the intraparticle enzyme distribution of Assemblase, an industrial biocatalytic particle containing immobilized penicillin-G acylase. The results were compared with previous studies based on light and transmission electron microscopic techniques. The integrated FESEM approach yielded the same quantitative results as the microscopic techniques used previously. Given this technical equivalence, the integrated approach offers several advantages. First, the single preparation method and detection system avoids interpretation discrepancies between corresponding areas that were examined for different properties with different detection techniques in different samples. Second, the specimen size suitable for whole particle study is virtually unlimited, which simplifies sectioning and puts less stringent demands on the embedding technique. Furthermore, the sensitivity toward enzyme presence and distribution increases because the epitopes inside thick sections become available for labeling. Quick and unambiguous analysis of the relation between particle morphology and enzyme distribution is important because this information may be used in the future for the design of enzyme distributions in which the particle morphology can be used as a control parameter.     

肉制品中淀粉含量测定方法的比较及关键点

本文论述了测定肉制品中淀粉含量的两种方法,一种是食品中淀粉含量的测定方法-淀粉酶水解法,另一种是肉制品中淀粉含量的测定方法-碘量法,详细阐述了这两种方法的原理、试剂设备、步骤,并对这两种方法的稳定性、适用性、优劣、关键点,以及测定结果进行了分析比较.结果表明两种方法均适用于肉制品中淀粉含量检测,但方法一更加简便准确.     

生乳中次氯酸钠的检测研究

建立了生乳中次氯酸钠的检测方法。试样中次氯酸根在弱酸性条件下与碘化钾生成游离碘,以淀粉溶液为指示剂,硫代硫酸钠标准滴定溶液滴定,计算氯的含量。方法处理简单,易操作,回收率高,试验成本低,分析周期短,因此该方法可作为生乳中次氯酸钠的快速检测方法。     

Morphologic changes of starch granules in the apple cv. Mutsu during ripening and storage

Unripe (A), semiripe (B), and ripe (C) apples (Malus domestica cv. Mutsu) stored (2–4 °C, 90–95% RH) for 1 week were studied. The decrease in the starch content was as a function of storage time and ripeness. The size (area, perimeter and circularity) of starch granules was determined by image analyzer (Semper6, Synoptics) interfaced with a standard light microscope (Olympus, BH²) using a bright field illumination. Area (Ar) (larger than 100 pixel), perimeter (Pe), and circularity (Ci) of the granules were measured. The distribution of Ar, Pe, and Ci was calculated by the Distribution Fitting Statistical program; normality of frequency distribution of data (Chi-square test), average ± standard deviation, t-values were presented. All data were analyzed by principal component analysis. Area and perimeter decreased, circularity increased with the storage time and ripeness. Critical differences were observed in the semiripe apple cortex (CoB0) and the ripe apple skin (SC0) compared with the other samples (unripe skin and cortex) at the harvest. In both samples, most starch granules swelled (19–24%), and the starch contents were significantly less than in others. The stage of ripeness could be more exactly followed by image analysis than by the starch iodine test, or the color of apple skin. The ultrastructure (SEM) of starch did show typical changes as a function of storage time. More smaller starch granules could be seen in the stored apples than in the fresh ones. Damaged (pits, cavities) starch granules were found in all samples. Carbon-13 NMR spectra of isolated apple cortex starch were obtained with cross polarization (CP), magic angle spinning (MAS), and high power proton irradiation (HD). There were shoulders on the C-1 resonance and on the C-2, 3, 4, 5 cluster. These shoulders indicate the presence of amorphous starch.