葡萄糖氧化酶在氨基功能化三维有序大孔材料上的固定化研究

一类新型功能化三维有序大孔（3DOM）材料被用于固载葡萄糖氧化酶．以聚苯乙烯胶晶为模板,三氨基丙基三乙氧基硅烷和正硅酸乙酯为前驱物,合成了不同氨基含量的3DOM SiO2-NH2材料．并以戊二醛为交联剂,成功地在3DOM SiO2-NH2材料表面嫁接了葡萄糖氧化酶,所得材料具有较高的催化活性．SEM观察表明,3DOM SiO2-NH2材料具有规则整齐相通的大孔孔道结构．FTIR测试表明材料中含有氨基及有机基团,戊二醛及葡萄糖氧化酶以C=N键嫁接在3DOM材料孔壁表面．     

三维规则排列的大孔SiO2材料的制备及表征

 以聚苯乙烯微球离心后形成的三维规则排列的胶晶作模板，以正\r\n硅酸乙酯、水、乙醇和盐酸等配制的溶胶填充微球间的间隙，然后原位\r\n形成凝胶，最后通过焙烧除去微球得到三维规则排列大孔二氧化硅材料\r\n．以苯乙烯为原料，过硫酸钾为引发剂，在70℃下搅拌28h后得到含聚\r\n苯乙烯微球（直径约为600nm）的母液．将母液在900～1000r／min的转\r\n速下离心12～24h得到紧密堆积排列的胶晶．以正硅酸乙酯作为硅源，\r\n按n（Si（OEt）4）∶n（EtOH）∶n（HCl）∶n（H2O）＝1∶3．9∶0．\r\n3∶1．8制成透明的SiO2溶胶．溶胶浓度控制在0．5～1．0mol／L．滴\r\n加在胶晶上的溶胶靠毛细管的作用力填充入微球间隙．重复填充多次（\r\n一般不超过5次）．焙烧在流量为1L／min的空气中进行，升温速度控制\r\n在5℃／min以下．在300℃下恒温5h以除去聚苯乙烯微球，在570℃下焙\r\n烧5h．焙烧后的样品表面可观察到五颜六色的彩光．SEM分析结果表明\r\n，球形孔（孔径约500nm）大小均匀，排列整齐，保持了微球的紧密堆\r\n积排列结构．孔与孔之间由小孔窗相互交连，孔壁比较充实，壁厚约为\r\n130nm．     

三维有序大孔氨基功能化材料的制备及其对Cr(Ⅵ)的吸附性能

以聚苯乙烯胶晶为模板,3-氨基-丙基-三乙氧基硅烷和正硅酸乙酯为前驱物,合成了三维有序大孔(3DOM)氨基功能化SiO2-NH2材料. SEM观察表明,合成的3DOM材料具有规则整齐的大孔通孔结构,平均孔径在535～596 nm之间,孔径收缩率为4.8%～14.5%. FTIR分析表明,材料中含有氨基等有机基团. BET分析表明,材料的比表面积为10.2 m2/g. 合成的3DOM SiO2-NH2材料对Cr(Ⅵ)离子的吸附能力随着材料中氨基含量的增加而增大,最大吸附量为4.31 mmol/g.     

壳聚糖固定化葡萄糖氧化酶生物传感器测定葡萄糖的含量

应用壳聚糖将葡萄糖氧化酶固定于鸡蛋膜上,结合氧电极制得葡萄糖传感器.实验表明,壳聚糖比戊二醛能更好地固定葡萄糖氧化酶,最佳条件为壳聚糖浓度0.3%、固定化酶量0.8 mg、 pH 7.0、缓冲溶液浓度300 mmol/L和温度25 ℃.本葡萄糖传感器的线性范围为0.016～1.10 mmol/L;检出限为8.0 μmol/L(S/N=3), 响应时间<60 s,有很好的稳定性,寿命>3个月.同一个传感器重复使用以及同方法制作的不同传感器之间都有很好的重现性,RSD分别为2.5%(n=10)和4.7%(n=4).实际样品中可能存在的烟酰胺、 VB6、 VB12、 VE、Ca2+、 Mg2+、 K+和Zn2+等对葡萄糖的测定不产生干扰.本传感器已成功地应用于市售饮料中葡萄糖含量的测定.     

一种基于壳聚糖固定葡萄糖氧化酶的葡萄糖生物传感器的研究

本文选用生物相容性好的壳聚糖作为基体材料，使其与戊二醛交联成网状结构包埋葡萄糖氧化酶制成电化学传感器。这种壳聚糖膜不仅可以减小葡萄糖氧化酶的流失，而且能为酶提供了适宜的微环境。用红外光谱、紫外光谱及透射电镜对膜的形态和性质进行了表征。实验结果表明该传感器具有很快的响应速度，很好的稳定性和重现性，能选择性地催化葡萄糖并测定其浓度。该传感器的制备方法简单，成本低，于冰箱中放置两周信号保持在90％以上，对葡萄糖测量的线性范围为1×10^-5 - 3.4×10^-3mol•L^-1，当信噪比为3:1时检测限为5×10^-6mol•L^-1。     

有序大孔二氧化硅孔壁聚合物功能化修饰及葡萄糖淀粉酶固载化

介绍了一种简易的制备方法用于制备功能化有序大孔氧化硅@聚合物材料,并将其用作固载酶的新型载体。     

A Glucose Biosensor Based on Immobilization of Glucose Oxidase into 3D Macroporous TiO2

3D macroporous TiO₂ inverse opals have been derived from a sol‐gel procedure using polystyrene colloidal crystals as templates. EDS and SEM showed a face‐centered cubic (FCC) structure TiO₂ inverse opal was obtained. Glucose oxidase (GOx) was successfully immobilized on the surface of indium‐tin oxide (ITO) electrode modified by TiO₂ inverse opal (TiO_2(IO)). Electrochemical properties of GOx/TiO_2(IO)/ITO electrode were characterized by using the three electrodes system. The result of cyclic voltammetry showed that a couple of stable and well‐defined redox peaks for the direct electron transfer of GOx in absence of glucose, and the redox peak height enhanced in presence of 0.1 μM glucose. Compare with the ordinary structured GOx/TiO₂/ITO electrode, inverse opal structured GOx/TiO_2(IO)/ITO electrode has a better respond to the glucose concentration change. Under optimized experimental conditions of solution pH 6.8 and detection potential at 0.30 V versus saturated calomel electrode (SCE), amperometric measurements were performed. The sensitivity and the detection limit of glucose detection was 151 μA cm^?2 mM^?1 and 0.02 μM at a signal‐to‐noise ratio of 3, respectively. The good response was due to the good biocompatibility of TiO₂ and the large effective surface of the three‐dimensionally ordered macroporous structure.     

Immobilization and Characterization of Glucose Oxidase on Single-Walled Carbon Nanotubes and Its Application to Sensing Glucose

The negatively charged （at pH 8.2） glucose oxidase （GOx, pI ca. 4.2） was assembled onto the surface of single-walled carbon nanotubes （SWNT）, which was covered （or wrapped） by a layer of positively charged polyelectrolyte poly（dimethyldiallylammonium chloride） （PDDA）, via the electrostatic interaction forming GOx-PDDASWNT nanocomposites. Fourier transform infrared （FTIR）, UV-Vis and electrochemical impedance spectroscopy （EIS） were used to characterize the growth processes of the nanocomposites. The results indicated that GOx retained its native secondary conformational structure after it was immobilized on the surface of PDDA-SWNT. A biosensor （Nafion-GOx-PDDA-SWNT/GC） was developed by immobilization of GOx-PDDA-SWNT nanocomposites on the surface of glassy carbon （GC） electrode using Nafion （5%） as a binder. The biosensor showed the electrocatalytic activity toward the oxidation of glucose under the presence of ferrocene monocarboxylic acid （FcM） as an electroactive mediator with a good stability, reproducibility and higher biological affinity. Under an optimal condition, the biosensor could be used to detection of glucose, presenting a typical characteristic of Michaelis-Menten kinetics with the apparent Michaelis-Menten constant of KM^app ca. 4.5 mmol/L, with a linear range of the concentration of glucose from 0.5 to 5.5 mmol/L （with correlation coefficient of 0.999） and the detection limit of ca. 83 μmol/L （at a signal-to-noise ratio of 3）. Thus the biosensor was useful in sensing the glucose concentration in serum since the normal glucose concentration in blood serum was around 4.6 mmol/L. The facile procedure of immobilizing GOx used in present work would promote the developments of electrochemical research for enzymes （proteins）, biosensors, biofuel cells and other bioelectrochemical devices.     

三维有序大孔SiO2中的纳米银

A multi-step process was used for preparation of three-dimensionally ordered macroporous (3DOM) SiO2, in which fully accessible Ag nanoparticles are incorporated. The method involves the processes of assembly of polystyrene colloidal crystal, preparation of 3DOM SiO2, and incorporation of Ag nanoparticles within 3DOM SiO2 through in situ Tollens‘ reaction. XRD, SEM and EDXS determination show that the Ag particles deposited on the macroporous walls in nano dimension. The results indicate that lower concentration of silver ammoniate and for-maldehyde in the solution is favorable for forming a very narrow size distribution and uniform shape of nanoparticles. However, the higher the concentration of the solution and the more the loading times, the larger the possibility to form un-uniform particles. Ag nanoparticles can be sintered into larger and spheral particles by calcination at 600℃, but can resist sintering owing to their high dispersivity when loading amount is small. The study provided a simple approach to tailor Ag/3DOM SiO2 composite materials with desired morphology and size of Ag particles within the macropores.     

聚苯乙烯胶晶膜及三维有序大孔SiO2膜的制备及表征

采用垂直沉积法组装了三维聚苯乙烯胶晶膜,并用其为模板制备了三维有序大孔（3DOM）SiO2膜．SEM观察表明,制备的胶晶膜和3DOMSiO2膜具有fcc结构,有序性很好．考察乳液浓度对胶晶膜结构的影响表明,浓度越高,胶晶膜越厚,有序性也越高,膜在30层内都能很好的粘附在载玻片上．通过调整前驱物溶液的浓度和滴加方式,可得到表面为球形或孔状的3DOM SiO2膜．     

氨基化树枝状介孔二氧化硅固定葡萄糖氧化酶用于检测葡萄糖

以三乙胺为碱源合成了树枝状介孔二氧化硅纳米粒子(DMSNs),并用3-氨基丙基三乙氧基硅烷(APTES)进行氨基修饰合成了氨基化树枝状介孔二氧化硅纳米粒子(DMSNs-NH₂),将其用于葡萄糖氧化酶(GOD)的固定化研究.采用扫描电子显微镜、透射电子显微镜、红外光谱仪、X射线衍射仪、氮气吸附仪及热重分析仪对固定化GOD(DMSNs-NH₂-GOD)进行了表征,测定了其活性及蛋白载量.结果表明,固定化GOD的直径约为200 nm,形状均一,呈分散的球形微粒;在最佳固定条件下,蛋白载量达225 mg/g,酶活性达215 U/mg;固定化GOD检测葡萄糖的最低检测限为0.0014 mg/mL.利用固定化GOD检测了血清和饮料中的葡萄糖,重复使用36次以上其相对酶活性仍剩余80%.该方法操作方便、准确度高,提高了酶的pH稳定性、热稳定性及重复使用性,降低了检测成本.     

HEC/SiO2凝胶复合物包埋固定化葡萄糖氧化酶的研究

溶胶-凝胶技术;HEC/SiO2凝胶复合物包埋固定化葡萄糖氧化酶的研究     

Effect of Immobilization Supports on the Amperometric Response of Silver Nanoparticles Enhanced Glucose Oxidase Electrodes

Silver nanoparticles enhanced glucose oxidase electrodes, which are based on polyvinyl butyral and chitosan, were fabricated and investigated. The sensitivity and maximum current i _max of the chitosan‐based electrode are 1.06 mA · M^?1 and 25.64 µA, respectively. However, the sensitivity and i _max of the polyvinyl butyral‐based electrode are 0.34 mA · M^?1 and 7.22 µA. The chitosan‐based electrode has a sensitivity 3.1 times higher and a maximum current 3.6 times larger than those of the polyvinyl butyral‐based electrode. Therefore, it is more sensitive and stable than the polyvinyl butyral‐based electrode.     

蜘蛛丝素和聚乙烯醇固定葡萄糖氧化酶 制备葡萄糖传感器

用蜘蛛丝素和聚乙烯醇的混合材料把葡萄糖氧化酶固定在氧电极表面,制成葡萄糖氧化酶电极。传感器对葡萄糖有灵敏的响应,平均响应时间为20s,电位变化值与葡萄糖浓度在3.0×10     

炭气凝胶纳米颗粒固定葡萄糖氧化酶的直接电化学

利用间苯二酚和甲醛在碱性环境下制备炭气凝胶(CA), 通过扫描电镜(SEM)、透射电镜(TEM)、比表面积测试Brunauer-Emmett-Teller (BET)等方法分析载体的形貌结构; 以CA为载体通过吸附法固定葡萄糖氧化酶(GOD)并修饰玻碳(GC)电极, 得到GOD/CA/GC电极. 在0.1 mol·L^-1磷酸盐缓冲溶液中, 利用循环伏安法研究了GOD/CA/GC 电极的直接电化学行为和对葡萄糖的催化性能. 结果表明, 以CA为载体可以很好地固定GOD并保持其生物活性, 在无任何电子媒介体存在时, GOD在电极上实现了直接电子转移, GOD/CA/GC电极对葡萄糖具有很好的电催化性能.     

Preparation of Three‐Dimensional Ordered Macroporous Prussian Blue Film Electrode for Glucose Biosensor Application

Ordered 3D interconnected macroporous Prussian blue (PB) films were electrochemically fabricated by using colloidal crystals of polystyrene beads as sacrificial templates. The prepared PB film electrodes have excellent catalytic activity towards the reduction of hydrogen peroxide. The PB structure was further used as functional interface for fabricating an enzyme‐based glucose sensor by using surface modification technique based on the electrostatic interactions. The resulted sensor has higher functional density, and larger surface area. The interconnected macroporous structure allows enhanced mass transport. These characteristics of the sensor enable us to detect glucose with high sensitivity. Therefore, the present 3D ordered macroporous film sensor exhibits wide linear detection range towards glucose, acceptable reproducibility and operational and storage stability. The present approach is promising for the generation of high‐enzyme‐content thin films with tailored bioactivity.     

4-巯基苯甲酸功能化纳米金粒子固定葡萄糖氧化酶/漆酶基燃料电池性能

以4-巯基苯甲酸修饰纳米金粒子作为固酶载体和导电基体构建了新型纳米结构固酶葡萄糖/O₂燃料电池,其制备简单,长期使用性能稳定。利用纳米金粒子通过表面修饰基团和酶分子活性中心附近疏水结合位之间的相互作用固定葡萄糖氧化酶(GO_x)和漆酶(Lac)分子,分别制备了固酶阳极-4-巯基苯甲酸功能化纳米金粒子固定葡萄糖氧化酶修饰金盘电极GO_x/4-MBA@GNP/Au和固酶阴极-4-巯基苯甲酸功能化纳米金粒子固定漆酶修饰金盘电极Lac/4-MBA@GNP/Au。电化学实验结果表明,两种电极在不引入任何外加电子中介的条件下,均可以实现酶活性中心-纳米金粒子之间的直接电子迁移,而且具有较快的催化反应能力(固酶阳极和阴极的转化速率分别为1.3和0.5 s^-1;催化葡萄糖氧化和氧气还原的起始电位分别为-0.23和0.76 V)。评估了固酶阳极和阴极组装成的纳米结构固酶葡萄糖/O₂燃料电池的能量输出性能。该燃料电池在没有Nafion薄膜和阳极无N₂气保护下,开路电压和最大输出能量密度分别可达0.56 V和760.0 μW/cm²,使用一周后输出能量密度仍然可以达到最初值的~88%。进一步测试结果显示,该燃料电池呈现出与游离漆酶类似的pH依赖关系和热稳定性,这些实验结果均暗示:影响整个酶燃料电池性能的关键在于漆酶基阴极催化氧还原的过程。此外,这种燃料电池的性能虽然受到共存干扰物抗坏血酸的影响,但在人类血清中测试结果显示其仍然具有较高的输出能量密度(132.0 μW/cm²,开路电压0.40 V)。本文研究结果给出了设计高性能葡萄糖/O₂燃料电池的新思路,同时也为研究固酶燃料电池的构效关系提供了实验依据和有价值的启示。     

Poly(acrylic acid)-silicon Hybrids Prepared via a RAFT-mediated Process and Covalent Immobilization of Glucose Oxidase

A surface modification technique was proposed for the modification of silicon surface with glucose oxidase (GOD). The silicon surface was first graft copolymerized with acrylic acid (AAc) via surface-initiated reversible addition-fragmentation chain-transfer (RAFT)-mediated process. With the aid of a water-soluble carbodiimide, GOD was then covalently immobilized on the silicon surface through the amide linkage between the amino group of GOD and the carboxyl group of the grafted AAc polymer. The changes in the surface composition after polymer grafting and enzyme immobilization on the silicon surface were investigated using X-ray photoelectron spectroscopy (XPS). The amount of GOD immobilized could be varied by changing the thickness of the polymer layer and the immobilization time. The GOD-functionalized silicon hybrids are potential useful in the application of the silicon-based biosensors.     

双模板法制备具有介孔孔壁的三维有序大孔二氧化铈及其改善的低温还原性能

以聚甲基丙烯酸甲酯(PMMA)为硬模板,三嵌段共聚物F127、十六烷基三甲基溴化铵(CTAB)或聚乙二醇(PEG)为软模板剂(表面活性剂),柠檬酸为络合剂,硝酸铈为金属前驱体,采用双模板法成功地合成出具有介孔孔壁的三维有序大孔(3DOM)结构的立方相CeO2样品CeO2-F127,CeO2-CTAB和CeO2-PEG,...     

葡萄糖氧化酶在炭黑上的固定及直接电化学

经红外光谱和电化学测量证明, 用简单的吸附法能将葡萄糖氧化酶(GOx)固定在炭黑(CB)表面. 电化学测量表明, 固定在CB上的GOx能进行准可逆的直接电化学反应, 其式量电位(E0’)为-0.436 V, 在40-150 mV·s-1范围内, 不随扫描速率而变化. 电化学反应速率常数(ks)为0.800 s-1, 比文献报道的大30多倍. 而且, 固定在CB上的GOx能保持其对葡萄糖氧化的生物电催化活性. 即使在保存两周后, 其电催化活性仅下降了5%, 表明固定在CB上的GOx有良好的稳定性.