Direct Electron Transfer between Glucose Oxidase and Multi-walled Carbon Nanotubes

The direct dedrochemical behavior between the glucose oxfdase ( GOD ) and the multi-wailed carbon nauotubes (MWNTs) has been studied. Two pairs of cyclic voltammetric peaks corresponding to the two different processes, i. e. mass-transportand surface reaction of GOD are observed on this MWNTs. The formal potentials with E^o‘=-0.45Vand E^o‘=-0.55V were obtained respectively.The GOD film was observed on the carbon nanotube by the TEM.     

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。     

Unadulterated Glucose Biosensor Based on Direct Electron Transfer of Glucose Oxidase Encapsulated Chitosan Modified Glassy Carbon Electrode

Glucose oxidase (GOD) was encapsulated in chitosan matrix and immobilized on a glassy carbon electrode, achieving direct electron transfer (DET) reaction between GOD and electrode without any nano‐material. On basis of such DET, a novel glucose biosensor was fabricated for direct bioelectrochemical sensing without any electron‐mediator. GOD incorporated in chitosan films gave a pair of stable, well‐defined, and quasireversible cyclic voltammetric peaks at about ?0.284 (E_pa) and ?0.338 V (E_pc) vs. Ag/AgCl electrode in phosphate buffers. And the peak is located at the potentials characteristic of FAD redox couples of the proteins. The electrochemical parameters, such as midpoint potential (E_1/2) and apparent heterogeneous electron‐transfer rate constants (k_s) were estimated to ?0.311 V and 1.79 s^?1 by voltammetry, respectively. Experimental results indicate that the encapsulated GOD retains its catalytic activity for the oxidation of glucose. Such a GOD encapsulated chitosan based biosensor revealed a relatively rapid response time of less than 2 min, and a sufficient linear detection range for glucose concentration, from 0.60 to 2.80 mmol L^?1 with a detection limit of 0.10 mmol L^?1 and electrode sensitivity of 0.233 μA mmol^?1. The relative standard deviation (RSD) is under 3.2% (n=7) for the determination of practical serum samples. The biologic compounds probably existed in the sample, such as ascorbic acid, uric acid, dopamine, and epinephrine, do not affect the determination of glucose. The proposed method is satisfactory to the determination of human serum samples compared with the routine hexokinase method. Both the unique electrical property and biocompatibility of chitosan enable the construction of a good bio‐sensing platform for achieved DET of GOD and developed the third‐generation glucose biosensors.     

Biphenol as an Electron Transfer Mediator for Glucose Oxidase

Abstract

Biphenol (4,4′-dihydroxy-biphenyl) was found to be an electron transfer mediator for glucose oxidase (GOD) of Aspergillus niger. At a glassy carbon electrode, a 1.44×10^-4 M solution of biphenol in phosphate-buffered saline (PBS) at pH 7.4 gives an quasi-reversible, one-electron, pH-sensitive couple at 255mV (relative to the standard calomel electrode). The apparent second-order rate constant for electron transfer from reduced GOD to oxidized biphenol was determined to be 3×10⁵ M^-1 s^-1. When biphenol and GOD are cophysiadsorbed on a graphite electrode immersed in PBS and held at 400mV, a glucose-dependent current response is noted. In addition to the predominant quasi-reversible biphenol redox couple, repetitive cyclic voltammetry at a graphite electrode gave rise to a polybiphenol polymer; this is most marked at a pH above the biphenol pK_a of about 9.5. At pH 7.4, the polymerization is less significant. Polybiphenol formed either side of the pK_a remains electrochemically active (E°_app. = 245mV) but no longer mediates GOD.     

葡萄糖氧化酶共价交联于蛋膜上的葡萄糖传感器

以牛血清白蛋白－戊二醛为交联剂，将葡萄糖氧化酶固定地鸡蛋膜上，氧电极作电化学敏感元件，制成葡萄糖氧化酶电极。传感器的响应范围为４．０×１０＾－６－２．４×１０＾－３ｍｏｌ／Ｌ；检测限为１．２１０＾－６ｍｏｌ／Ｌ。该传感器具有线性范围宽，灵敏度高，使用寿命长等优点。     

Glucose Biosensor Based on the Fabrication of Glucose Oxidase in the Bio-Inspired Polydopamine-Gold Nanoparticle Composite Film

A highly efficient enzyme immobilization method has been developed for electrochemical biosensors using polydopamine films with gold nanoparticles (AuNPs) embedded. This simple enzyme fabrication method can be performed in very mild conditions and stored in a long time with high bioactivity. The fabricated amperometric glucose biosensor exhibited a high and reproducible sensitivity, wide linear dynamic range and low limit of detection (LOD) (0.1 μmol·L^?1). A low value of 1.5 mmol·L^?1 for the apparent Michaelis‐Menten constant K^app_M was obtained. The high sensitivity, wide linear range, good reproducibility and stability make this biosensor a promising candidate for portable amperometric glucose biosensor.     

Spatial Architecture of Modified Carbon Nanotubes/Electrochemically Reduced Graphene Oxide Nanomaterial for Fast Electron Transfer. Application in Glucose Biosensor

Electron transfer (ET) reactions in bioelectrocatalysis of enzymes at electrode surfaces require not only the efficient immobilization, but also highly conductive nanostructured platform, which allows for retaining its bioactivity and structural conformation. The novel architecture of spatially separated electrochemically reduced graphene oxide (ERGO) by multi‐walled carbon nanotubes functionalized with 4‐(pyrrole‐1‐yl) benzoic acid (MWCNT/PyBA) with the accurate porous structure could be an alternative for earlier approaches to the construction of bioelectrocatalytic systems with rapid diffusion of reagents from the solution to the enzyme molecule. The formation of ERGO/MWCNT/PyBA system was confirmed by electrochemical, spectroscopic and microscopic methods. The cyclic voltammetry experiments revealed that the presence of ERGO in the conductive material affects the electronic communication between the enzyme molecule and modified electrode surface greatly improving its ET properties resulting in a double increase of the heterogeneous ET rate constant value (k_s=6.5 s^?1). The fabricated glucose oxidase based biosensor sensitively detects glucose, therefore, ERGO/MWCNT/PyBA architecture could provide a novel and efficient platform for immobilization of redox enzymes.     

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.     

Biosensing Membrane Base on Ferulic Acid and Glucose Oxidase for an Amperometric Glucose Biosensor

A biosensing membrane base on ferulic acid and glucose oxidase is synthesized onto a carbon paste electrode by electropolymerization via cyclic voltammetry in aqueous media at neutral pH at a single step. The developed biosensors exhibit a linear response from 0.082 to 34 mM glucose concentration, with a coefficient of determination R² equal to 0.997. The biosensors display a sensitivity of 1.1 μAmM⁻¹ cm⁻², a detection limit of 0.025 mM, and 0.082 mM as glucose quantification limit. The studies reveal stable, repeatable, and reproducible biosensors response. The results indicate that the novel poly-ferulic acid membrane synthesized by electropolymerization is a promising method for glucose oxidase immobilization towards the development of glucose biosensors. The developed glucose biosensors exhibit a broader linear glucose response than other polymer-based glucose biosensors.     

Amperometric Glucose Biosensor Based on Ultrafine Platinum Nanoparticles

Abstract

In the present paper the ultrafine and highly dispersed platinum nanoparticles (average size 3 nm) were used for the construction of a glucose biosensor in a simple method. An excellent response to glucose has been obtained with a high sensitivity (137.7 µA mM^?1 cm^?2) and fast response time (5 s). The biosensor showed a detection limit of 5 µM (at the ratio of signal to noise, S/N=3) and a linear range form 0.2 to 3.2 mM with a correlation coefficient r=0.999. The apparent Michaelis–Menten constant (k _m) and the maximum current were estimated to be 9.36 and 1.507 mA mM^?1 cm^?2, respectively. In addition, effects of pH value, applied potential and the interferents on the amperometric response of the sensor were investigated and discussed.     

掺杂纳米普鲁士蓝溶胶-凝胶修饰葡萄糖生物传感器

采用溶胶-凝胶法制备了纳米普鲁士蓝微粒,将含纳米普鲁士蓝微粒的TiO2溶胶-凝胶固定在玻碳电极表面得到纳米普鲁士蓝修饰电极,该电极对H2O2产生灵敏的响应,线性范围为0．5～400μmoL／L,较常规普鲁士蓝修饰电极(线性范围为25～500μmol／L)灵敏。电极表面再用溶胶．凝胶法固定葡萄糖氧化酶后构建了葡萄糖生物传感器,响应范围0～20mmoL／L,葡萄糖氧化酶表观米氏常数为8．04mmoL／L。实验表明,该法适合于批量制作高灵敏和高重现性的生物传感器。     

Glucose Biosensor Using Glucose Oxidase Immobilized in Polyaniline

A biosensor for glucose utilizing kinetics of glucose oxidase (EC 1.1.3.4.) was developed. The enzyme was immobilized on polyaniline by covalent bonding, using glutaraldehyde as a bifunctional agent. The system showed a linear response up to 2.2 mM of glucose with a response time of 2.5–4.0 min. In addition, the immobilized enzyme had a higher activity between pH 6.5 and 7.5. The system retained 50% of its activity after 30 d of daily use. The optical absorption spectra of the polyaniline/glucose oxidase electrode after glucose had been added to the buffer solution showed that the absorption band around 800 nm had changed considerably when glucose was allowed to react with the electrode. This optical variation makes polyaniline a very promising polymer for use as a support in optical sensor for clinical application.     

A Novel Potentiometric Glucose Biosensor Based on Polyaniline Semiconductor Films

Abstract

Application of polyaniline semiconductor films to potentiometric biosensor development provides certain advantages comparing with the known systems. Using self-doped polyaniline instead of common polymer as pH transducer the stable potentiometric response of 70 mV/pH was obtained. Taking as an example glucose biosensor we showed that polyaniline based electrode possessed three-four fold increased potential shift than glucose-sensitive field-effect transistor did. One can increase the sensitivity of potentiometric biosensor using thick ion-exchange membranes (in our case Nafion) in order to concentrate product near electrode surface. Such sensor possessed higher response time.     

基于金纳米棒-壳聚糖复合膜的葡萄糖生物传感器

本文采用金纳米棒-壳聚糖复合膜固定葡萄糖氧化酶构建电流型葡萄糖生物传感器.通过电化学交流阻抗法和循环伏安法对酶膜状态进行了表征,得到了相应的等效电路和动力学参数.实验结果表明,金纳米棒-壳聚糖复合膜可以辅助电子传递,提高电极的电流响应,并使生物传感器的使用温度范围有很大的扩展.此传感器表现出对葡萄糖溶液浓度的优良响应,线性范围在2.78×10^-5mol/L—2.22×10^-3mol/L,响应灵敏度约为7.819μA·cm^-2(mmol/L)^-1,表观米氏常数为10mmol/L.本工作还研究了温度和溶液pH值对电极电流响应的影响.     

基于WS2量子点材料固定葡萄糖氧化酶的直接电化学及其生物传感研究

采用水热法制备水溶性WS₂量子点（WS₂ QDs）材料,并将该材料进一步用于葡萄糖氧化酶（GOx）的有效固定,构建GOx/W₂ QDs/GCE传感界面. 采用透射电镜、紫外-可见光谱和电化学等方法对材料的形貌、GOx的固定化过程,以及传感器的直接电化学和电催化性能进行了表征. 结果表明,WS₂ QDs材料能够有效促进GOx与电极之间的直接电子转移. 并且,基于该传感器对葡萄糖良好的电催化作用,该方法有效实现了对葡萄糖的高灵敏检测,其线性范围为25 ~ 100 μmol·L^-1和100 ~ 600 μmol·L^-1,检测限为5.0 μmol·L^-1（S/N=3）. 该传感器具有良好的选择性、重现性和稳定性,可用于实际样品血糖的分析测定.     

A Fast Made Disposable Glucose Biosensor Incorporating the Oxidase and a Ferrocene in an Alginate Gel

Abstract

A simple procedure is described for co-immobilization of glucose oxidase and dimethylaminomethylferrocene in a sodium alginate gel on the surface of pyrolytic graphite electrode. The film is electrochemically active and the peak current is a function of D-glucose concentration with the apparent Michaelis-Menten constant K_m = (1.2±0.3) × 10^?2 M. The optimal concentration range for biosensoric applications in 0.001–0.010 M.

     

基于多壁碳纳米管和聚丙烯胺层层自组装的葡萄糖生物传感器

经混酸处理后的多壁碳纳米管(MWCNTs)末端及侧壁带有含氧基团,能与阳离子聚电解质通过静电作用结合,也能与酶蛋白非特异性结合。利用层层自组装法在铂(Pt)电极表面构建了聚丙烯胺(PAA)-MWC-NTs-葡萄糖氧化酶(GOx)膜,研究了自组装薄膜的表面微观形貌和电化学性质。组装层数为6层时最优,对葡萄糖响应线性范围为5.0×10-4~2.10×10-2mol/L;检出限为1.0×10-4mol/L(S/N=3);灵敏度为4.95μA/(mmolcm2),响应时间3.80s;GOx表观米氏常数为17.79mmol。对抗坏血酸等具有较强的抗干扰能力,10天后电极响应电流保持最初的91.79%。3次平行实验的RSD为4.85%。     

纳米金胶增强葡萄糖生物传感器的构建

将纳米金胶(AuNPs)和羟基磷灰石(HAp)按一定比例混合制备了新型复合膜用于葡萄糖氧化酶(GOD)的固定,构建了高灵敏的葡萄糖传感器。由于纳米金胶的存在,葡萄糖氧化酶的直接电化学性质得以增强,在去除氧气的PBS(pH 7.0)介质中,固定在复合膜内的GOD表现出一对良好的氧化还原峰。在饱和氧气条件下,当加入一定量的葡萄糖时,由于GOD催化葡萄糖氧化消耗溶液中的溶解氧,-0.8 V处溶解氧的还原峰电流降低,且峰电流降低的量与葡萄糖浓度在0.02～1.62 mmol/L范围内呈线性相关,检出限为5.0μmol/L,检测灵敏度达9.91 mA.mol-1.L,可实现对葡萄糖的快速检测。     

Study on the United Glucose Oxidase Electrode Constructed by Composite Electrode

A novel type of united glucose oxidase (GOD) electrode was designed. Glucose oxidase and ferrocene (Fc), which was a mediator, were added into the composite electrode that was constructed by graphite powder, acetylene black, and epoxy resin. These three materials in composite electrode kept constant proportion in weight. And the optimum amounts of GOD and Fc among united enzyme electrode were 5% and 2%, respectively. The glucose was detected linearly in the concentration range 0.01–9.0 mM with a 20-s steady-state response time and 36 nA/mM of the sensitivity at 0.15 V applied potential. And electrode fouling problem and the response current from the interferents were avoided. The response current of the united GOD electrode had no obvious deterioration within 80 days when stored at 4°C in a refrigerator. The detecting results of human serum by the united GOD electrode had good consistency with that by standard enzyme method. The maximum deviation between these two detecting values was 5%. It might be used for detecting the blood sugar in clinical assay.     

超细银－金复合颗粒增强酶生物传感器的研究

用琥珀酸二异辛酯磺酸钠／环已烷反胶束体系合成憎水纳米银－金复合颗粒， 并用此纳米银－金颗粒与聚乙烯醇缩丁醛构成复合固酶模基质，用溶胶－凝胶法固 定葡萄糖氧化酶，构建葡萄糖生物传感器。实验表明，纳米憎水银－金颗粒可以大 幅度提高固定化酶的催化活性，响应电流从相应浓度的几十纳安增强几万纳安。探 讨了纳米颗粒效应在固定化酶中所起的作用，为纳米颗粒在生物传感器领域中的应 用提供了可参考的实验和理论依据。