纳米金-还原氧化石墨烯修饰葡萄糖氧化酶传感器的制备及其电流法检测饮料中的葡萄糖

利用纳米金(Au NPs)与还原氧化石墨烯(rGO)复合纳米材料制备了葡萄糖氧化酶生物传感器并用于饮料中葡萄糖含量的检测。将壳聚糖作为还原剂及稳定剂,通过一步法合成了Au NPs-rGO复合材料,并通过物理吸附固定葡萄糖氧化酶(GOx)来制作GOx生物传感器。该传感器在磷酸盐缓冲溶液(0.1 mol/L,p H6.0)中,-0.45 V(vs.Ag/Ag Cl)电位下电流法检测葡萄糖含量,线性检测范围为0.01~0.88 mmol/L,灵敏度为22.54μA·mmol-1·L·cm-2,检出限为1.01μmol/L,且表观米氏常数为0.497 mmol/L。该传感器用于多种饮料中葡萄糖含量的直接检测,结果满意。     

A Selective Cholesterol Biosensor Based on Composite Film Modified Electrode for Amperometric Detection

An amperometric cholesterol biosensor based on immobilization of cholesterol oxidase in a Prussian blue (PB)/polypyrrole (PPy) composite film on the surface of a glassy carbon electrode was fabricated. Hydrogen peroxide produced by the enzymatic reaction was catalytically reduced on the PB film electrode at 0 V with a sensitivity of 39 μA (mol/L)^?1. Cholesterol in the concentration range of 10^?5 ? 10^?4 mol/L was determined with a detection limit of 6 × 10^?7 mol/L by amperometric method. Normal coexisting compounds in the bio‐samples such as ascorbic acid and uric acid do not interfere with the determination. The excellent properties of the sensor in sensitivity and selectivity are attributed to the PB/PPy layer modified on the sensor.     

Amperometric Glucose Biosensors Based on Integration of Glucose Oxidase onto Prussian Blue/Carbon Nanotubes Nanocomposite Electrodes

Nanosized Prussian blue (PB) particles were synthesized with a chemical reduction method and then the PB nanoparticles were assembled on the surface of multiwall carbon nanotubes modified glassy carbon electrode (PB/MWNTs/GCE). The results showed that the PB/MWNTs nanocomposite exhibits a remarkably improved catalytic activity towards the reduction of hydrogen peroxide. Glucose oxidase (GOD) was immobilized on the PB/MWNTs platform by an electrochemically polymerized o‐phenylenediamine (OPD) film to construct an amperometric glucose biosensor. The biosensor exhibited a wide linear response up to 8 mM with a low detection limit of 12.7 μM (S/N=3). The Michaelis–Menten constant K_m and the maximum current i_max of the biosensor were 18.0 mM and 4.68 μA, respectively. The selectivity and stability of the biosensor were also investigated.     

Preparation of GOD/Sol‐Gel Silica Film on Prussian Blue Modified Electrode for Glucose Biosensor Application

A novel amperometric glucose biosensor was fabricated by in situ incorporating glucose oxidase (GOD) within the sol‐gel silica film on a Prussian blue (PB) modified electrode. The method is simple and controllable, which combined the merits of in situ immobilizing biomolecules in sol‐gel silica film by electrochemical method and the synergic catalysis effects of PB and GOD molecules. Scanning electron microscopy (SEM) showed that the GOD/sol‐gel silica film was homogeneous with a large number of three‐dimensional nanopores, which not only enhanced mass transport, but also maintained the active configuration of the enzyme molecule and prevented the leakage of enzyme, therefore improved the stability and sensitivity of the biosensor. The fabricated biosensor showed fast response time (10 s), high sensitivity (26.6 mA cm^?2 M^?1), long‐term stability, good suppression of interference, and linear range of 0.01 mM–5.8 mM with a low detection limit of 0.94 μM for the detection of glucose. In addition, the biosensor was successfully applied to determine glucose in human serum samples.     

Direct Electrochemistry and Electrocatalysis Behaviors of Glucose Oxidase Based on Hyaluronic Acid-Carbon Nanotubes-Ionic Liquid Composite Film

Multi‐walled carbon nanotubes (MWNTs) were dispersed in the ionic liquid [BMIM][BF₄] to form a uniform black suspension. Based on it, a novel glucose oxidase (GOx)‐hyaluronic (HA)‐[BMIM][BF₄]‐MWNTs/GCE modified electrode was fabricated. UV‐vis spectroscopy confirmed that GOx immobilized in the composite film retained its native structure. The experimental results of EIS indicated MWNTs, [BMIM][BF₄] and HA were successfully immobilized on the surface of GCE and [BMIM][BF₄]‐MWNTs could obviously improve the diffusion of ferricyanide toward the electrode surface. The experimental results of CV showed that a pair of well‐defined and quasi‐reversible peaks of GOx at the modified electrode was exhibited, and the redox reaction of GOx at the modified electrode was surface‐confined and quasi‐reversible electrochemical process. The average surface coverage of GOx and the apparent Michaelis‐Menten constant were 8.5×10⁻⁹ mol/cm² and 9.8 mmol/L, respectively. The cathodic peak current of GOx and the glucose concentration showed linear relationship in the range from 0.1 to 2.0 mmol/L with a detection limit of 0.03 mmol/L (S/N=3). As a result, the method presented here could be easily extended to immobilize and obtain the direct electrochemistry of other redox enzymes or proteins.     

Prussian Blue Modified Carbon Ionic Liquid Electrode: Electrochemical Characterization and Its Application for Hydrogen Peroxide and Glucose Measurements

Prussian blue modified carbon ionic liquid electrodes (PB‐CILEs) were fabricated using chemical and electrochemical procedures. Chemically fabricated PB‐CILE exhibited an excellent sensitivity (0.0866 μA μM^?1), low detection limit (0.01 μM) and two linear ranges (0.01–1 and 1–600 μM) toward hydrogen peroxide. Then, glucose oxidase (GOx) was immobilized on the surface of PB‐CILE to fabricate glucose biosensor using three different procedures involving cross linking with glutaraldehyde (GLU) and bovine serum albumin (BSA), entrapment into the Nafion matrix and covering with a sol‐gel layer. Glucose biosensor fabricated using cross linking procedure showed the best sensitivity (0.0019 μA μM^?1) and operational stability for glucose.     

Composite film of carbon nanotubes and chitosan for preparation of amperometric hydrogen peroxide biosensor

A new amperometric biosensor for hydrogen peroxide was developed based on cross-linking horseradish peroxidase (HRP) by glutaraldehyde with multiwall carbon nanotubes/chitosan (MWNTs/chitosan) composite film coated on a glassy carbon electrode. MWNTs were firstly dissolved in a chitosan solution. Then the morphology of MWNTs/chitosan composite film was characterized by field-emission scanning electron microscopy. The results showed that MWNTs were well soluble in chitosan and robust films could be formed on the surface. HRP was cross-linked by glutaraldehyde with MWNTs/chitosan film to prepare a hydrogen peroxide biosensor. The enzyme electrode exhibited excellent electrocatalytic activity and rapid response for H₂O₂ in the absence of a mediator. The linear range of detection towards H₂O₂ (applied potential: −0.2 V) was from 1.67 × 10⁻⁵ to 7.40 × 10⁻⁴ M with correction coefficient of 0.998. The biosensor had good repeatability and stability for the determination of H₂O₂. There were no interferences from ascorbic acid, glucose, citrate acid and lactic acid.     

A Hydrogen Peroxide Biosensor Based on Room Temperature Ionic Liquid Functionalized Graphene Modified Carbon Ceramic Electrode

A room temperature ionic liquid (IL) 1‐butyl‐3‐methylimidazolium hexafluorophosphate functionalized graphene (GE) was prepared and a hydrogen peroxide (H₂O₂) biosensor was fabricated by immobilizing hemoglobin (Hb) into the IL‐GE composite film. UV‐visible and Fourier transform infrared spectra of the composite film indicated that Hb retained its native structure in the film. Electrochemical investigation of the biosensor showed a pair of well‐defined, quasi‐reversible redox peaks with E_pa=?0.209 V and E_pc= ?0.302 V (vs. SCE) in pH 7.0 phosphate buffer solution at the scan rate of 100 mV/s. To the reduction of H₂O₂, the biosensor had a good linear range from 8.0×10^?7 to 1.8×10^?4 mol/L with a detection limit of 3.0×10^?7 mol/L. The apparent Michaelis‐Menten constant K^app_M was estimated to be 3.4×10^?5 mol/L.     

Fabrication of Tyrosinase Biosensor Based on Multiwalled Carbon Nanotubes‐Chitosan Composite and Its Application to Rapid Determination of Coliforms

A tyrosinase (Tyr) biosensor was fabricated by immobilizing Tyr on the surface of multiwalled carbon nanotubes (MWNTs)‐chitosan (Chit) composite modified glassy carbon electrode (GCE). The MWNTs‐Chit composite film provided a biocompatible platform for the Tyr to retain the bioactivity and the MWNTs possessed excellent inherent conductivity to enhance the electron transfer rate. The Tyr/MWNTs‐Chit/GCE biosensor showed high sensitivity (412 mA/M), broad linear response (1.0×10^?8–2.8×10^?5 M), low detection limit (5.0 nM) and good stability (remained 93% after 10 days) for determination of phenol. The biosensor was further applied to rapid detection of the coliforms, represented by Escherichia coli (E. coli) in this work. The current responses were proportional to the quantity of coliforms in the range of 10⁴–10⁶ cfu/mL. After 5.0 h of incubation, E. coli could be detected as low as 10 cfu/mL.     

Enzyme biosensor based on the immobilization of HRP on SiO<Subscript>2</Subscript>/BSA/Au composite nanoparticles

In this work, an enzyme biosensor based on the immobilization of horseradish peroxidase (HRP) on SiO₂/BSA/Au/thionine/nafion-modified gold electrode was fabricated successfully. Firstly, nafion was dropped on the surface of the gold electrode to form a nafion film followed by chemisorption of thionine (Thi) as an electron mediator via the ion-exchange interaction between the Thi and nafion. Subsequently, the SiO₂/BSA/Au composite nanoparticles were assembled onto Thi film through the covalent bounding with the amino groups of Thi. Finally, HRP was immobilized on the SiO₂/BSA/Au composite nanoparticles due to the covalent conjugation to construct an enzyme biosensor. The surface topographies of the SiO₂/BSA/Au composite nanoparticles were investigated by using scanning electronic microscopy. The stepwise self-assemble procedure of the biosensor was further characterized by means of cyclic voltammetry and chronoamperometry. The enzyme biosensor showed high sensitivity, good stability and selectivity, a wide linear response to hydrogen peroxide (H₂O₂) in the range of 8.0 × 10^-6 ∼ 3.72 × 10^-3 mol/L, with a detection limit of 2.0 × 10^-6 mol/L. The Michaelies-Menten constant K_M^app K_M^{app} value was estimated to be 2.3 mM.     

New Nanocomposite Based on Prussian Blue Nanoparticles/Carbon Nanotubes/Chitosan and Its Application for Assembling of Amperometric Glucose Biosensor

Abstract

A new nanocomposite was developed by combination of prussian blue (PB) nanoparticles and multiwalled carbon nanotubes (MWNTs) in the matrix of biopolymer chitosan (CHIT). The PB and MWNTs had a synergistic electrocatalytic effect toward the reduction of hydrogen peroxide. The CHIT/MWNTs/PB nanocomposite‐modified glassy carbon (GC) electrode could amplify the reduction current of hydrogen peroxide by ～35 times compared with that of CHIT/MWNTs/GC electrode and reduce the response time from ～60 s for CHIT/PB/GC to 3 s. Besides, the CHIT/MWNTs/PB nanocomposite‐modified GC electrode could reduce hydrogen peroxide at a much lower applied potential and inhibit the responses of interferents such as ascorbic acid (AA) uric acid (UA) and acetaminophen (AC). With glucose oxidase (GOx) as an enzyme model, a new glucose biosensor was fabricated. The biosensor exhibited excellent sensitivity (the detection limit is down to 2.5 µM), fast response time (less than 5 s), wide linear range (from 4 µM to 2 mM), and good selection.     

Carbon Nanotubes‐Based Amperometric Cholesterol Biosensor Fabricated Through Layer‐by‐Layer Technique

A carbon nanotubes‐based amperometric cholesterol biosensor has been fabricated through layer‐by‐layer (LBL) deposition of a cationic polyelectrolyte (PDDA, poly(diallyldimethylammonium chloride)) and cholesterol oxidase (ChOx) on multi‐walled carbon nanotubes (MWNTs)‐modified gold electrode, followed by electrochemical generation of a nonconducting poly(o‐phenylenediamine) (PPD) film as the protective coating. Electrochemical impedance measurements have shown that PDDA/ChOx multilayer film could be formed uniformly on MWNTs‐modified gold electrode. Due to the strong electrocatalytic properties of MWNTs toward H₂O₂ and the low permeability of PPD film for electroacitve species, such as ascorbic acid, uric acid and acetaminophen, the biosensor has shown high sensitivity and good anti‐interferent ability in the detection of cholesterol. The effect of the pH value of the detection solution on the response of the biosensor was also investigated. A linear range up to 6.0 mM has been observed for the biosensor with a detection limit of 0.2 mM. The apparent Michaelis‐Menten constant and the maximum response current density were calculated to be 7.17 mM and 7.32 μA cm^?2, respectively.     

基于碳纳米管和铁氰酸镍纳米颗粒协同作用的葡萄糖生物传感器

将制备的铁氰酸镍纳米颗粒(NiNP)与多壁碳纳米管(CNT)混合, 分散于壳聚糖溶液中, 形成一种新的纳米复合成分(NiNP-CNT-CHIT), 将其修饰在玻碳电极表面. 新复合膜体现了NiNP和CNT之间的协同作用, 由于CNT的良好的传递电子性能, 促使NiNP催化氧化还原能力有了较大的提高. 此NiNP-CNT-CHIT复合膜修饰的玻碳电极在较低电位下对过氧化氢具有良好的电催化性能, 与NiNP-CHIT膜比较, 测定H₂O₂的灵敏度增大了50倍. 通过戊二醛在电极表面固定葡萄糖氧化酶制备了一种新的葡萄糖传感器. 该传感器在－0.2 V下对葡萄糖的线性范围为0.05～10 mmol/L, 检测下限为10 μmol/L.     

葡萄糖氧化酶在羟基磷灰石／Nation复合膜修饰电极上的直接电化学及其对葡萄糖的生物传感

将葡萄糖氧化酶固定于羟基磷灰石（HAp）-Nation复合膜,构建了高灵敏、高选择性的葡萄糖传感器．羟基磷灰石和Nation良好的协同作用,可以有效地提高传感器的稳定性与灵敏度．实验结果表明：固定在复合膜修饰电极上的葡萄糖氧化酶呈现出一对较好的近乎可逆的氧化还原峰,并且对葡萄糖的氧化有良好的催化作用,同时消耗溶解氧,从而导致溶解氧还原峰的降低．在-0.8V处,随葡萄糖浓度的增加,葡萄糖氧化酶催化葡萄糖氧化时消耗溶解氧的量增加,溶解氧还原电流逐渐降低,因此该修饰电极可以作为葡萄糖传感器实现对葡萄糖的高灵敏检测．在0．12～2．16mmol·L^-1浓度范围内,溶解氧还原电流的降低与葡萄糖的浓度成正比,据此可以测定出溶液中葡萄糖的浓度,该传感器的检出限和灵敏度分别为0．02mmol·L^-1（SIN=3）和6．75mA·mol·L^-1．因此,HAp-Nation复合膜为酶的固定和直接电化学研究提供了一个新的有效平台,在构建新型无试剂葡萄糖传感器方面具有较大的应用前景．     

Fabrication of Glucose Biosensor Based on Encapsulation of Glucose‐Oxidase on Sol‐Gel Composite at the Surface of Glassy Carbon Electrode Modified with Carbon Nanotubes and Celestine Blue

A simple procedure was developed to prepare a glassy carbon electrode modified with multi walled carbon nanotubes (MWCNTs) and Celestin blue. Cyclic voltammograms of the modified electrode show stable and a well defined redox couple with surface confined characteristic at wide pH range (2–12). The formal potential of redox couple (E′) shifts linearly toward the negative direction with increasing solution pH. The surface coverage of Celestine blue immobilized on CNTs glassy carbon electrode was approximately 1.95×10^?10 mol cm^?2. The charge transfer coefficient (α) and heterogeneous electron transfer rate constants (k_s) for GC/MWCNTs/Celestine blue were 0.43 and 1.26 s^?1, respectively. The modified electrode show strong catalytic effect for reduction of hydrogen peroxide and oxygen at reduced overpotential. The glucose biosensor was fabricated by covering a thin film of sol‐gel composite containing glucose oxides (GOx) on the surface of Celestine blue /MWCNTs modified GC electrode. The biosensor can be used successfully for selective detection of glucose based on the decreasing of cathodic peak current of oxygen. The detection limit, sensitivity and liner calibration rang were 0.3 μM, 18.3 μA/mM and 10 μM–6.0 mM, respectively. The accuracy of the biosensor for glucose detection was evaluated by detection of glucose in a serum sample, using standard addition protocol. In addition biosensor can reach 90% of steady currents in about 3.0 sec and interference effect of the electroactive existing species (ascorbic acid–uric acid and acetaminophen) was eliminated. Furthermore, the apparent Michaelis–Menten constant 2.4 mM, of GOx on the nano composite exhibits excellent bioelectrocatalytic activity of immobilized enzyme toward glucose oxidation. Excellent electrochemical reversibility of redox couple, high stability, technically simple and possibility of preparation at short period of time are of great advantages of this procedure for modification of glucose biosensor.     

Prussian Blue electrodeposited on MWNTs-PANI hybrid composites for H2O2 detection

A Prussian Blue (PB)/polyaniline (PANI)/multi-walled carbon nanotubes (MWNTs) composite film was fabricated by step-by-step electrodeposition on glassy carbon electrode (GCE). The electrode prepared exhibits enhanced electrocatalytic behavior and good stability for detection of H₂O₂ at an applied potential of 0.0 V. The effects of MWNTs thickness, electrodeposition time of PANI and rotating rate on the current response of the composite modified electrode toward H₂O₂ were optimized to obtain the maximal sensitivity. A linear range from 8 × 10⁻⁹ to 5 × 10⁻⁶ M for H₂O₂ detection has been observed at the PB/PANI/MWNTs modified GCE with a correlation coefficient of 0.997. The detection limit is 5 × 10⁻⁹ M on signal-to-noise ratio of 3. To the best of our knowledge, this is the lowest detection limit for H₂O₂ detection. The electrode also shows high sensitivity (526.43 μA μM⁻¹ cm⁻²) for H₂O₂ detection which is more than three orders of magnitude higher than the reported.     

Electrocatalytic Oxidation of Glucose at Carbon Nanotubes Supported PtRu Nanoparticles and Its Detection

PtRu nanoparticles were supported on multiwall carbon nanotubes (MWNTs), which were further fabricated as an electrode for nonenzymatic glucose sensing. Transmission electron microscope and X‐ray diffraction patterns were used for characterization of the PtRu nanoparticles on MWNTs. Cyclic voltammetry and chronopotentiometry were applied to investigate the performance of the PtRu/MWNTs nanocomposite electrode for nonenzymatic oxidation of glucose. The PtRu/MWNTs electrode shows high electrocatalytic activity towards the oxidation of glucose in 0.1 M NaOH solution and thus can be used to selectively detect glucose. Under the optimal potential (+0.55 V vs. Ag/AgCl), the biosensor effectively performs a selective electrochemical analysis of glucose in the presence of common interferents, such as ascorbic acid (AA), dopamine (DP) and uric acid (UA). Wide linear calibration ranging from 1 mM to 15 mM, high sensitivity of 28.26 μA cm^?2 mM^?1, low detection limit of 2.5×10^?5 M, and fast response time of 10 s were achieved for the detection of glucose at the PtRu/MWNTs electrode.     

Electrosynthesis and characterization of poly(hydroxy-methylated-3,4-ethylenedioxythiophene) film in aqueous micellar solution and its biosensing application

A new and efficient synthetic route to hydroxymethylated-3,4-ethylenedioxylthiophene（EDOT-MeOH） was developed by a simple four-step sequence,and its global yield was approximately 41.06%.The poly（hydroxymethylated-3, 4-ethylenedioxylthiophene）（PEDOT-MeOH） film was electrosynthesized in aqueous sodium dodecylsulfate micellar solutions and characterized by different methods.The EDOT-MeOH possessed better water solubility,and lower onset oxidation potential than EDOT.The as-obtained PEDOT-MeOH film displayed good reversible redox activity,stability and capacitance properties in a monomer-free electrolyte,especially the good solubility of PEDOT-MeOH film in strong polar organic solvents such as dimethyl sulfoxide and tetrahydrofuran created a potential application in many different fields. Fluorescent spectra indicated that PEDOT-MeOH was a yellow-green-light-emitter with maximum emission at 568 nm.The as-formed PEDOT-MeOH film had good biocompatibility and was used for fabricating the electrochemical vitamin C biosensor.The proposed biosensor showed a linear range of 3×10^-6 mol/L to 1.2×10^-2 mol/L with the detection limit of 1μmol/L,a sensitivity of 95.6μA（mmol/L）^-1 cm^-2,and a current response time less than 10 s and a fairly good stability (The relative standard deviation was 0.43%for 20 successive assays,the proposed biosensor still retained 93.5%of bioactivity after 15 days storage.This result indicated that the prepared PEDOT-MeOH film as immobilization matrix of biologically-active species could be a promising candidate for the design and application of biosensor.     

Synergistic Effect of MWNTs/CeO2/CHIT Film for Detection of CdSe Nanoparticle Labeled Sequence‐Specific of 35S Promoter of Cauliflower Mosaic Virus Gene

A porous composite film was fabricated combining the advantages of multiwalled carbon nanotubes, CeO₂ and chitosan. The synergistic effect of the film improved the immobilization of probe ssDNA. The loaded probe ssDNA was used for detection of CdSe quantum dots labeled target DNA. The DNA hybridization reaction was detected by differential pulse anodic stripping voltammetry of Cd²⁺ after the oxidative release of labeled CdSe quantum dots. The established DNA biosensor can discriminate different target sequences associated with 35S promoter of cauliflower mosaic virus gene with relatively wide linear range and low detection limit (2.4×10^?13 mol/L).     

Cast thin film biosensor design based on a Nafion backbone, a multiwalled carbon nanotube conduit, and a glucose oxidase function

Novel electroanalytical sensing nanobiocomposite materials are reported. These materials are prepared by mixing multiwalled carbon nanotubes (MWNTs), a Nafion cation exchanger, and glucose oxidase (GOD) in appropriate amounts. The MWNTs are cylindrical with a diameter in the range 40-60 nm and with a length of up to several micrometers, and they provide electrical conductivity. Nafion acts as a polymer backbone to give stable and homogeneous cast thin films. Both MWNTs and Nafion provide negative functionalities to bind to positively charged redox enzymes such as glucose oxidase. The resulting biosensing composite material is inexpensive, reliable, and easy to use. The homogeneity of the MWNT-Nafion-GOD nanobiocomposite films was characterized by atomic force microscopy (AFM). Amperometric transducers fabricated with these materials were characterized electrochemically using cyclic voltammetry and amperometry in the presence of hydrogen peroxide and in the presence of glucose. Their linear response to hydrogen peroxide was demonstrated. The glucose biosensor sensitivity was strongly influenced by the glucose oxidase concentration within the nanobiocomposite film. The optimized glucose biosensor (2.5 mg/mL GOD) displayed a sensitivity of 330 nA/mM, a linear range of up to 2 mM, a detection limit of 4 microM, and a response time of <3 s.