Nad Determination By Using A Glucose Dehydrogenase Based Glucose Probe

Abstract

This study describes the determination of NAD by using a glucose based enzyme carbon probe. Linear sweep voltammetric and constant potential studies were carried out in order to choose the best parameters for NAD analysis. The enzyme GDH has been used in solution first and then immobilized on the probe surface. Results indicate NAD can be detected in concentrations of 10^?6 Mol/1 with a good precision.     

基于固定化酶的流动荧光法测定血清中的葡萄糖

本文将固定化酶柱，光纤以及停流技术结合起来，研究了以廉价而易得的硫胺素为荧光底物的葡萄糖的荧光分析，提出了一种快速、简便、精确地测定葡萄糖的新方法。其线性范围为１．０￣６０．０ｍｇ／Ｌ，线性相关系数为０．９９９，检测限为０．１５ｍｇ／Ｌ。以５．０ｍｇ／Ｌ的葡萄糖作精度试验，ＲＳＤ％＝２．１％（ｎ＝１１）。该方法已成功地用于血清中葡萄糖的测定，结果与光度法相符。     

电聚合修饰碳纤维电极及乳酸脱氢酶活性的测定

用电聚合方法将亚甲基绿 (MG)修饰在碳纤维电极 (直径 7μm)上 ,并用该修饰电极测定了乳酸脱氢酶 (LDH)活性。在烟酰胺腺嘌呤二核苷酸 (NAD+ )浓度为 6 .0× 10 - 4mol L ,乳酸浓度为 5 .0× 10 - 3mol/L的pH7.0NaOH KH2 PO4 缓冲介质中 ,电位恒定在 +0 .10V下 ,用电流法测定乳酸脱氢酶活性 ,线性范围为 15～ 2 4 0U/mL ,检测限为 10U/mL ,响应时间为 15s。该修饰微电极稳定性好、灵敏度高、测定干扰小     

Amperometric Glucose Determination by Means of Glucose Oxidase Immobilized on a Cellulose Acetate Film: Dependence on the Immobilization Procedures

Glucose microelectrodes were prepared by immobilizing glucose oxidase onto a cellulose acetate film coating a platinum wire. Hexamethylenediamine (HMDA) and Glutaraldehyde (GA) were employed as spacer and coupling agent, respectively. Sensitivities and linear response ranges were studied as a function of the relative amounts of HMDA and GA. The best sensitivity was found when HMDA and GA were 5% and 2.5% in aqueous solutions, respectively. Taking as a reference the functioning of this biosensor, the roles of HMDA and GA percentages appear to be opposed when the extension of the linear response range is considered. Indeed, an increase of one unit in HMDA percentage (from 5 to 6 %) induces an increase in the extension of the linear response range equal to that obtained with a decrease of one unit of GA percentage (from 2.5 to 1.5%).     

Direct Bioelectrocatalysis of PQQ‐Dependent Glucose Dehydrogenase

The direct bioelectrocatalysis was demonstrated for pyrroloquinoline quinone‐dependent glucose dehydrogenase (PQQ‐dependent GDH) covalently attached to single‐walled carbon nanotubes (SWNTs). The homogeneous ink‐like SWNT suspension was used for both creating the SWNT network on the microelectrode carbon surface and for enzyme immobilization. Functionalization of the SWNT surface by forming active ester groups was found to considerably enhance SWNT solubility in water with a range from 0.1 to 1.0 mg/mL. The PQQ‐dependent GDH immobilized on the surface of the SWNTs exhibited fast heterogeneous electron transfer with a rate constant of 3.6 s^?1. Moreover, the immobilized PQQ‐dependent GDH retained its enzymatic activity for glucose oxidation. A fusion of PQQ‐dependent GDH with SWNTs has a great potential for the development of low‐cost and reagentless glucose sensors and biofuel cells.     

Glucose Dehydrogenase Based Bioelectrode Utilizing a Synergistic Scheme of Substrate Conversion

A Bioelectrode utilizing a synergistic scheme of substrate conversion was built using glucose dehydrogenase from Acinetobacter calcoaceticus immobilized on the surface of a graphite electrode. At saturated glucose concentration the bioelectrode responded to the low reactive substrate hexacyanoferrate(III) with a sensitivity of 0.0035 µA/µM cm². The response of the bioelectrode increased up to the 3.4×10⁴ fold in the presence of high reactive organic electron acceptors (mediators). The increase of the response depended on the concentration of the mediators and their chemical nature. The sensitivity of the bioelectrode to mediators reached 7.3–77 µA/µM cm². The comparison of the bioelectrode sensitivity with kinetic parameters of enzyme action in homogeneous solution revealed good correlation between the sensitivity of the bioelectrode and the predicted value from the kinetic scheme of the reactivity of mediators. This confirms a synergistic scheme of bioelectrode action.     

A Cheap Amperometric and Optical Sensor for Glucose Determination

A cheap amperometric and optical sensor for glucose, based on an ITO electrode coated with electrodeposited Co/Al hydrotalcite (HT) is described. Cobalt based HT shows a reversible electrochromic behavior which can be exploited for the development of an optical sensor. Working in the optical mode, the linearity range of the sensor is between 0.008 and 0.13 mM with a sensitivity of 1.14 mM^?1?cm^?2, whereas when working in the amperometric mode, the linearity ranges from 0.002 to 1.5 mM with a sensitivity of 4.24×10^?4 A?mM^?1?cm^?2. The sensor has been successfully employed for the determination of glucose in a serum sample.     

葡萄糖脱氢酶微型生物传感器的研制及应用

以甲苯胺兰（ＴＢ）修饰碳糊微电极为基体，将葡萄糖脱氢酶（ＧＤＨ）用丝素蛋白膜固定于修饰微电极表面制成了生物传感器，在ｐＨ７．０的ＮａＯＨ－ＮａＨ２ＰＯ４缓冲溶液中，烟酰胺腺嘌呤二核苷酸（ＮＡＤ）的浓度为１．０４×１０＾－３ｍｏｌ／Ｌ的条件下，其响应电流与葡萄糖浓度在１．０×１０＾－４～３．２×１０＾－３ｍｏｌ／Ｌ范围内有良好线性关系，响应时间为２０ｓ；检测限为４．０×１０＾－５ｍｏｌ／Ｌ。该传感器     

酶法测定葡萄糖

利用已糖激酶、６－磷酸葡萄糖脱氢酶偶联而催化葡萄糖反应的原理,通过还原型烟酰胺腺嘌呤二核苷酸磷酸的吸光度变化率得出 酶促反应速度,采用标准曲线法了葡萄糖的含量。检出限为７×１０＾－７ｍｏｌ／Ｌ。     

基于表面等离子体激元共振的葡萄糖光化学传感器的研究

采用自行设计组装的表面等离子体激元共振光化学传感器实验装置，在不加其它化学试剂的条件下，研究了传感器对葡萄糖溶液的响应特性。实验结果表明，此传感器测定葡萄糖溶液的稳定性，可逆性良好，响应迅速，使用寿命长。     

反胶束固定化醇脱氢酶的制备及应用研究

研究十六烷基三甲基溴化铵(CTAB)-辛烷-己醇反胶束体系固定化醇脱氢酶(ADH)的制备及应用。考察了含水量、CTAB和己醇用量对于ADH固定化的影响。对游离酶和固定化酶的催化动力学性质研究表明:酶促反应的最适pH值分别为8.2和8.8,最适温度分别为31℃和20℃,米氏常数分别为12mmol/L和7mmol/L。30℃时,游离酶存放150min后失活90%,固定化酶失活50%,表明反胶束固定化ADH有较好的热稳定性。应用此体系测定了试样中乙醇的含量。     

Continuous Amperometric Determination of Glucose Using an Immobilized Enzyme Reactor in Combination with an Immersible Dialysis Probe

Abstract

Glucose was continuously determined by reaction in a packed-bed enzyme reactor containing glucose oxidase and catalase. Oxygen consumption was measured amperometrically with a polarographic Clark electrode. Glucose was sampled through a dialysis probe immersed in the solution to be measured. An extension of the normal range for the enzyme was achieved by modulating the flow rate through the dialysis probe and a linear response was obtained in the range of 1.0-60 mM glucose. The correlation between the glucose transfer and the membrane area of the dialysis probe was also studied. Six different membranes were used, all showing variations in the adhesion of yeast cells.     

A Chemiluminescence Optical Fiber Glucose Biosensor Based on Co-immobilizing Glucose Oxidase and Horseradish Peroxidase in a Sol-gel Film

IntroductionIn recent years chemiluminescence (CL)biosensor prepared by immobilization of a sensitivereagent such as peroxidase or oxidase onto a solidmatrix has attracted much attention due to the highsensitivity of the chemiluminescent reaction of thesensitive reagent even with a simple instrument.Generally,CL biosensors can be divided into twocategories.One consists of hydrogen peroxide sen-sors prepared by immobilizing a kind of peroxidaseonto a suitable solid support[1,2 ] ,and the immo…     

基于共价固定化苯嵌蒽酮荧光猝灭的药根碱光化学传感器

将3-烯丙氧基苯嵌蒽酮与丙烯酸胺和N,N’-亚甲基双丙烯酰胺一起共聚在硅烷化含乙烯基团的石英玻片上,制成了可传感药根碱的光极膜。药根碱能快速可逆挥灭光极膜的荧光。在1．00×10－5~4．00×10~(-4)mol/L范围内,膜对药根碱响应呈线性关系。由于共价固定化作用阻止了敏感试剂的流失,传感器显示出好的稳定性。常见无机阴、阳离子及一些常见药物对测定无显著干扰。传感器用于尿样中药根碱测定的回收率在102％~105％之间。     

Construction and Characterization of Direct Electron Transfer-Type Continuous Glucose Monitoring System Employing Thermostable Glucose Dehydrogenase Complex

Abstract

We demonstrated a direct electron transfer-type enzyme electrode using thermostable FAD-glucose dehydrogenase (FADGDH) consisting of three distinct subunits (an FAD-containing catalytic subunit, a cytochrome subunit, and a chaperone-like subunit) and its application in developing a continuous glucose monitoring (CGM) system without a synthetic electron mediator. An FADGDH-immobilized electrode showed current signals according to glucose concentration in the absence of a synthetic electron mediator. The sensor containing the FADGDH complex showed a stable response for 72 h at 37°C. Furthermore, the CGM response was well fitted to the gradient change in the glucose concentration obtained from system calibration.     

Immobilized Fullerene C60‐Enzyme‐Based Electrochemical Glucose Sensor

A mixed‐valence cluster of cobalt(II) hexacyanoferrate and fullerene C60‐enzyme‐based electrochemical glucose sensor was developed. A water insoluble fullerene C60‐glucose oxidase (C60‐GOD) was prepared and applied as an immobilized enzyme on a glassy carbon electrode with cobalt(II) hexacyanoferrate for analysis of glucose. The glucose in 0.1 M KCl/phosphate buffer solution at pH = 6 was measured with an applied electrode potential at 0.0 mV (vs Ag/AgCl reference electrode). The C60‐GOD‐based electrochemical glucose sensor exhibited efficient electro‐catalytic activity toward the liberated hydrogen peroxide and allowed cathodic detection of glucose. The C60‐GOD electrochemical glucose sensor also showed quite good selectivity to glucose with no interference from easily oxidizable biospecies, e.g. uric acid, ascorbic acid, cysteine, tyrosine, acetaminophen and galactose. The current of H₂O₂ reduced by cobalt(II) hexacyanoferrate was found to be proportional to the concentration of glucose in aqueous solutions. The immobilized C60‐GOD enzyme‐based glucose sensor exhibited a good linear response up to 8 mM glucose with a sensitivity of 5.60 × 10² nA/mM and a quite short response time of 5 sec. The C60‐GOD‐based glucose sensor also showed a good sensitivity with a detection limit of 1.6 × 10^‐6 M and a high reproducibility with a relative standard deviation (RSD) of 4.26%. Effects of pH and temperature on the responses of the immobilized C60‐GOD/cobalt(II) hexacyanoferrate‐based electrochemical glucose sensor were also studied and discussed.     

葡萄糖氧化酶催化荧光测定的非酶体系研究

多数氧化酶及其底物都是重要的临床检验指标[1]，目前普遍采用酶荧光法或酶显色法，通过氧化酶催化氧化底物生成H刃。，过氧化物酶（HRP）催化H刃2氧化生成荧光／生色底物，实现间接测定［’j．酶法测定的缺点是体系不稳定、反应条件苛刻、费用较高．最近Mitani等［’j报道了非酶法化学发光测定葡萄糖氧化酶，灵敏度高，但需要特殊试剂，且测定时间较长．我们用Fenton反应［‘j将H。O。转化为羟基自由基（“OH），进而氧化对苯二甲酸生成荧光物质，建立了荧光测定葡萄糖氧化酶的非酶法，详细研究了荧光指示剂的选择、Fenton试剂的改进…     

固定化酶法测定微量组分

以壳聚糖为载体对醇脱氢酶(ADH)和脲酶(UASE)进行固定化。固定化的最适条件:ADH和UASE的偶联时间分别为2.5h和1.5h,交联剂戊二醛浓度为0.6%和0.5%,偶联pH值为6.8和5.0。对游离酶及固定化酶的性质研究表明,ADH酶促反应的最适宜pH为8.0和8.4,最适宜温度为33℃和35℃,米氏常数为48mmol/L和13mmol/L;UASE酶促反应的最适宜pH均为7.0,最适宜温度为60℃和72℃,米氏常数为0.4mmol/L和2mmol/L。固定化酶与游离酶相比在复用性上具有优势,用固定化酶测定了试样中尿素含量以及模拟样中银离子含量。     

Enzymes Immobilized on a Tubular Reactor for Fast Amperometric Determination of Glucose in Brazilian Honey

Abstract

Glucose present in honey was rapidly determined by the differential amperometric method using two tubular reactors containing glucose oxidase and peroxidase. The linear dynamic range extends from 5 × 10^?5 to 2 × 10^?4mol L^?1, at pH 7.0. At flow rate of 1.5 mL min^?1 and injecting 150-µL sample volumes, a sampling frequency of the 33 determinations per hour is afforded. The reproducibility of the methods showed a relative standard deviation (RSD) < 4%. The detection limit of this method is 1.7 × 10^?5 mol L^?1. The samples analyses were compared with the parallel spectrophotometric determination.