氯化血红素催化反应体系的动力学性质及在葡萄糖分析中的应用

研究了氯化血红素作为过氧化物模拟酶催化显色体系（Ｃ６Ｈ５－ＯＨ－４－ＡＡＰ－Ｈ２Ｏ２）并与葡萄糖氧化反应介联的动力学性质。通过控制模拟酶和各反应物的用量确定了反应成假一级反应的葡萄糖浓度范围，在此反应体系中，葡萄糖浓度可以通过测定染料的吸光度而获得。该法干扰小，回收率达９４％￣９８％。检测限为２．０×１０＾－４ｇ／Ｌ。     

胶束增敏的辣根过氧化物酶催化荧光反应测定葡萄糖的研究

采用葡萄糖氧化酶－辣根过氧化物酶－对羟基苯丙酸－过氧化体系测定葡萄糖，阳离子胶束十六烷基三甲基溴化铵对生荧反应具有催化和增敏作用，可用于１μ１样品中葡萄糖的测定，线性范围０．５－５０μｇ，线性相关系数ｒ＝０．９９６６，相对标准偏差为０．７３％，检测限为０．４６μｇ。     

酶-离子色谱法检测葡萄糖

应用酶反应催化性能高，反应选择性好以及离子色谱法快速，灵敏、准确的特点，率先提出酶、离子色谱法检测葡萄糖的新方法，在含葡萄糖样品溶液中加入一定量的葡萄糖氧化酶（GOD），在葡萄糖氧化酶的催化下，被测葡萄糖被氧化产生H2O2，生成的H2O2与加入的NO2^-进一步反应，生成的NO3^-用离子色谱定性和定量测定，因为被测的葡萄糖与生成的NO3^-存在定量关系，从而可以由NO3^-的浓度计算出葡萄糖的含量，测定葡萄糖的线性范围在5－100mg/L，测定了市售葡萄糖和蜂蜜中葡萄糖的含量，回收率分别为94％和94．5％。     

基于葡萄糖/葡萄糖氧化酶/H2O2/L-酪氨酸/辣根过氧化物酶反应体系的荧光毛细分析法测定血糖

建立了葡萄糖(G)/葡萄糖氧化酶(GOD)/H2O2/L-酪氨酸/辣根过氧化物酶(HRP)新荧光反应新体系,并将此反应体系与荧光毛细分析法(FCA)结合,开发一种能够真正实现血糖测定的新葡萄糖酶荧光毛细分析法(GE-FCA).经优选得到的新反应体系最佳条件为:GOD和HRP浓度均为2000 U/L,L-酪氨酸浓度为1....     

环糊精葡萄糖基转移酶的固定化及其应用：Ⅲ.固定化环糊精葡萄糖基转…

用固定化环糊精葡萄糖基转移酶催化淀粉环化反应，考察了反应时间，乙醇浓度和淀粉液化程度对环糊精产率的影响。环糊精产率最高达５４％，固定化酶经多次重复使用或长时间连连续操作，酶活力降低较少。     

有机溶剂中(R)-醇腈酶催化不对称合成(R)-苯乙醇腈

 研究了来源于杏仁的（R）－醇腈酶在有机溶剂异丙醚中催化苯甲醛与HCN不对称合成（R）－苯乙醇腈，初步探讨了来源于不同杏仁的（R）－醇腈酶的筛选、最适酶量的确定以及底物HCN与苯甲醛的配比、底物浓度、酶的微环境pH和反应温度对不对称合成反应的影响．结果发现，来源于苦杏仁的（R）－醇腈酶优于来源于甜杏仁的（R）－醇腈酶．优化的反应条件为：最适酶量150g／L，HCN与苯甲醛的配比2．5，苯甲醛浓度300mmol／L，酶的微环境pH5．4，反应温度0～5℃．在该优化反应条件下，反应平衡转化率和产物的光学纯度均高达99％以上．     

利用卵壳膜固定化葡萄糖氧化酶制备酶电极

报道了以卵壳膜为载体，用戊二醛将葡萄糖氧化酶固定于膜上，并探讨了酶膜制备过程的各种影响因素。在固定过程中，当戊二醛浓度为０．１％，酶用量为２０ＵＩ时，酶回收率为１７％。渗透性是影响酶膜重现的主因。以此酶膜制备的葡萄糖电极，线性范围为１．０×１０５～１．６×１０－３ｍｏｌ／Ｌ检测限为５×１０－６ｍｏ１／Ｌ，寿命两个月。     

有机溶剂—水以液相体系脂肪酶不对称水解合成S—（＋）—萘普生

采用有机溶剂－水双液相体系作为反应介质，并用键合有一Ｃ６Ｈ５的无定型多孔硅胶作为分散在两相间的接触面。用圆柱状假丝酵母脂肪酶对萘普生甲酯进行不对称水解，考察了酶浓度、体系ＰＨ值、温度、分散剂对反应的影响，当转化率为２４．３％时，产品萘普生的对映体守量值为９４．９％，剩余底物萘普生甲酯的对映体过量值为３０．５％，当以异辛烷作为有机相时，该体系酶催化水解的对映体比率约为５０。     

测定麦芽糖转糖苷反应体系组成的高效液相色谱法

建立了分析低聚异麦芽糖组成的高效液相色谱法，采用Spherisorb-NH2色谱柱，示差折光检测器，乙腈－水（体积比70：30）为流动相，外标法定量测定；结果显示各糖质量浓度在0．1－10g/L范围内与峰面积呈良好线性关系，相关系数为0．999 0－0．999 7；应用法跟踪了pH5．0的柠檬酸－柠檬酸钠缓冲溶液中以α－葡萄糖转苷酶为催化剂在58℃温度下的麦芽糖转糖苷反应，分析了反应体系组成随时间的变化，得到了上述反应条件下麦芽糖最大限度地转化为低聚异麦芽糖的最佳反应时间为24h；该法快捷、简单、准确，可用于低聚异麦芽糖生产的质量控制。     

α—葡萄糖基甜菊糖的酶促合成反应（Ⅱ）

研究了以球形纤维素弱阴离子交换剂化载体，固定化葡萄糖基转移酶ＧＴ－１转化甜菊糖为α－葡萄糖基甜菊糖反应的各种影响因素。结果表明，甜菊糖的酶促转经到达一定程度后，继续增大酶用量及淀粉用量，对转化结果没有太多改善。     

Simultaneous use of multiple fluorescent reporter dyes for glucose sensing in aqueous solution

The simultaneous use of several fluorescent reporter dyes in a multicomponent boronic acid-based glucose sensing system is reported. In one application, two dyes with widely different emission wavelengths are used to report changes in glucose concentration. A third glucose-insensitive dye was then added to act as a reference dye and provide for a ratiometric correction to the two reporter dye signals. The inclusion of such a reference dye reduces errors arising from sources such as fluctuations in lamp intensity and sample dilution. The simultaneous use of multiple fluorescent reporter dyes     

High-performance size exclusion chromatography as a rapid method for the separation of steroid hormone receptors

Triazine dyes, bound to polyethylene glycol, have been used to influence the partition of some enzymes within a dextran-polyethylene glycol-water two-phase system. The enzymes, present in a protein extract from baker's yeast, included glucose 6-phosphate dehydrogenase, glyceraldehyde phosphate dehydrogenase, 3-phospho-glycerate kinase and alcohol dehydrogenase. The partition coefficients of the enzymes could be changed by a factor of 10-500 in favour of the polyethylene glycol-rich phase, while the partition of bulk protein was much less affected. The influence of the concentration of polymer-bound dye and phase-forming polymers, temperature, pH, kind and concentration of salt and the presence of nucleotides on this affinity partitioning effect was studied. The extraction was effective even at high concentrations of dye and protein (40 g/l). A partial purification (32-fold) of glucose 6-phosphate dehydrogenase was carried out by an extraction in five steps.     

The interaction of boronic acid-substituted viologens with pyranine: the effects of quencher charge on fluorescence quenching and glucose response

The fluorescence sensing of several monosaccharides using boronic acid-substituted viologen quenchers in combination with the fluorescent dye pyranine (HPTS) is reported. In this two-component sensing system, fluorescence quenching by the viologen is modulated by monosaccharides to provide a fluorescence signal. A series of viologen quenchers with different charges were prepared and tested for their ability both to quench the fluorescence of HPTS and to sense changes in glucose concentration in aqueous solution at pH 7.4. Both quenching efficiency and sugar sensing were found to be strongly dependent upon viologen charge. The molar ratio between HPTS and each of the viologen quenchers was varied in order to obtain an optimal ratio that provided a fairly linear fluorescence signal across a physiological glucose concentration range. Both the quenching and sugar sensing results are explained by electrostatic interaction between dye and quencher.     

Optical glucose detection across the visible spectrum using anionic fluorescent dyes and a viologen quencher in a two-component saccharide sensing system

A very general system is described in which anionic fluorescent dyes possessing a wide range of absorbance and emission wavelengths are used in combination with a boronic acid-modified viologen quencher to sense glucose at pH 7.4 in buffered aqueous solution. The present study demonstrates this capability with the use of eleven anionic fluorescent dyes of various structural types. Signal modulation occurs as the monosaccharide binds to the viologen quencher and alters its efficiency in quenching the fluorescence of the anionic dyes. The degree of quenching and the magnitude of the glucose signal were found to correlate roughly with the number of anionic groups on the dye. Optimal quencher : dye ratios were determined for each dye to provide a fairly linear signal in response to changes in glucose concentration across the physiological range.     

An improved enzymatic assay for glucose determination in blood serum using a l,1′-dimethylferricinium dye

l,l′-dimethylferricinium (DMFe⁺),a stable and pH-insensitive blue dye, was prepared via enzymatic oxidation of a 1,1′-dimethylferrocene (DMFe):2-hydroxypropyl-β-cyclodextrin (HPCD) watersoluble inclusion complex, using bilirubin oxidase immobilized onto porous aminopropyl glass beads via glutaraldehyde activation. In the presence of glucose, DMFe⁺ was reduced to DMFe by reacting with the reduced glucose oxidase (FADH₂), and the absorbance decrease was followed at 650 nm. In acetate pH 5.2 buffer, the response to glucose in blood serum was nonlinear, especially in the low concentration range, because of a competition for the reduced glucose oxidase between the DMFe⁺ dye and oxygen. At this pH, endogenous ceruloplasmin was also observed to oxidize residual DMFe (16%) in the dye preparation, causing an increase in absorbance at 650 nm. An assay protocol was then developed using maleate buffer, pH 6.5, to overcome these interferences as well as mutarotation of α-D-glucose. The results obtained for glucose in the blood serum samples agreed well with those of the reference hexokinase/glucose-6-phosphate dehydrogenase method.     

Continuous glucose detection using boronic acid-substituted viologens in fluorescent hydrogels: linker effects and extension to fiber optics

A fluorescent anionic dye and a viologen appended with boronic acids, which serve as glucose receptors, have been synthesized and immobilized into a poly(2-hydroxyethyl methacrylate) hydrogel for use as a continuous glucose monitor. The fluorescence of the dye is modulated by the quenching efficiency of the viologen-based receptor, which in turn is dependent on the glucose concentration. Two monomeric versions of the quencher/receptor unit were prepared and their performance within the hydrogel evaluated. By tethering the quencher/receptor to the hydrogel matrix using a single-point attachment, slightly improved glucose sensing was observed. The hydrogels were tested for their ability to continuously and reversibly detect glucose over the course of several hours. The tests were carried out using a cuvette-based system, as well as a fiber-optic-based configuration. Under physiological conditions (0.1 M phosphate buffer, pH 7.4, 37 degrees C), the fluorescent hydrogels display an excellent dynamic response to glucose concentrations within the biologically significant range (2.5-20 mM).     

A 3,5‐DistyrylBODIPY Dye Functionalized with Boronic Acid Groups for Direct Electrochemical Glucose Sensing

The synthesis and characterization of a novel BODIPY dye functionalized with bis‐boronic acid groups to enable direct glucose sensing through selective recognition of carbohydrates is reported. Styrylation with boronic acid groups at the 3,5‐positions of the BODIPY core results in an extension of the π‐conjugation system of the dye and in a red‐shift of the main absorption band from 500 to 637 nm. The functionalized BODIPY dye was adsorbed on a glassy carbon electrode using the drop and dry method. Modified and bare electrodes were characterized using cyclic voltammetry and scanning electrochemical microscopy, while glucose detection was carried out by using differential pulse voltammetry and chronoamperometry. The detection limit was determined to be 1.42 μM. The dye was found to be selective and sensitive towards glucose, since likely interferences have only minor effects on the glucose detection.     

Flow injection analysis of hydrogen peroxide using glass-beads modified with manganese(III)-tetra(4-carboxyphenyl)porphine derivative and its analytical application to the determination of serum glucose.

A flow injection analysis system of hydrogen peroxide was developed. The present system is based on measuring of the absorbance of a quinoid dye formed by the following reaction catalyzed by peroxidase: Phenol + 4-Aminoantipyrine + 2H2O2 --> Peroxidase --> Quinoid dye + 4H2O. A column packed with aminopropyl-glass beads modified with amanganese(III)-tetra(4-carboxyphenyl)porphine derivative (Mn-TCPP(G) column), which has peroxidase-like activity, was used in place of an immobilized peroxidase column in the above reaction. The linear range of the calibration curve was 0.4-80 microg/ml hydrogen peroxide. The relative standard deviation of this system was 2.97% (n = 100, 10 microg/ml hydrogen peroxide 20 microl injection). The Mn-TCPP(G) column has sufficiently stability for the continuous injection of hydrogen peroxide untill 100 times. The advantageous feature of the Mn-TCPP(G) column was a less-electrostatic interaction between the mother glass beads and the anionic chromogen or quinoid dye formed and the stability in terms of the storage, temperature and moisture. The determination of serum glucose was achieved by attaching an immobilized glucose oxidase column to this system without deproteinization.     

基于芬顿反应和硫磺素T构建免标记荧光传感器用于葡萄糖的检测

基于芬顿反应和硫磺素T(ThT)构建新奇的免标记荧光传感器用于葡萄糖的检测。当无葡萄糖存在时,ThT诱导富G-DNA探针形成G-四链体/ThT复合物,ThT的荧光强度显著增强;当葡萄糖存在时,葡萄糖氧化酶催化葡萄糖产生H2 O2,在Fe^2+催化的芬顿反应作用下,H2 O2转化为羟基自由基(·OH),·OH引发DNA的氧化损伤导致富G-DNA探针裂解为短寡核苷酸片段而丧失形成G-四链体/ThT的能力,ThT的荧光强度显著降低,从而实现对葡萄糖的检测。在优化的检测条件下,G-四链体/ThT荧光强度变化和葡萄糖浓度在0.5~45μmol/L的范围内呈现较好的线性关系(R^2=0.99268),检出限为0.1μmol/L。利用本法对葡萄糖加标的血液样品进行分析,葡萄糖的回收率为90.7%~118.3%,相对标准偏差为1.7%~5.8%,方法可用于血糖检测。     

Optimization of Culture Conditions for Enhanced Decolorization of Cibacron Red FN-2BL by <Emphasis Type="Italic">Schizophyllum</Emphasis><Emphasis Type="Italic">commune</Emphasis> IBL-6

The objective of this study was to exploit the decolorization potential of a newly isolated white-rot fungus Schizophyllum commune IBL-6 for the biodegradation of reactive textile dye Cibacron Red FN-2BL. In the initial decolorization study of 10 days, it was observed that S. commune IBL-6 was a better decolorizer of Cibacron Red FN-2BL. Various process parameters like composition of basal nutrient medium, pH, temperature, additional carbon and nitrogen sources, and initial dyestuff concentration were optimized to develop an economic decolorization process. The optimum dye decolorization was achieved in basal nutrient medium II containing 0.1% Cibacron Red FN-2BL and supplemented with 1% glucose after 3 days incubation at pH 4.5 and 30 degrees C. All the additional carbon sources were found to enhance decolorization process, whereas most of the nitrogen supplements caused fungal-growth inhibition. The pattern of enzymes involved in the biodegradation of this dye was studied, and manganese peroxidase was found to be the major peroxidase with minor lignin peroxidase and laccase activities.