Fabrication of a Nanobiocomposite Film Containing Heme Proteins and Carbon Nanotubes on a Choline Modified Glassy Carbon Electrode: Direct Electrochemistry and Electrochemical Catalysis

A nanocomposite electrochemical sensing film is assembled on choline (Ch) modified glassy carbon electrode (GCE), which contains multiwalled carbon nanotubes (MWNTs), Nafion cation exchanger, and myoglobin (Mb) or hemoglobin (Hb). The MWNTs provide a 3D porous and conductive network for the enzyme immobilization and Nafion acts as polymeric binder to give cast thin films. Both MWNTs and Nafion provide negative functionalities to bind to the positively charged redox proteins and to attach at the positively charged Ch modified layer, and drive the formation of homogeneous and stable nanocomposite film, the MWNT‐Nafion‐Mb. The nanocomposite film was characterized by field emission scanning electron microscope (FE‐SEM). The Mb in the nanocomposite film showed a pair of well‐defined and nearly reversible cyclic voltammetric peaks at about ?0.32 V vs. SCE at pH 7.0 solution for the heme Fe(III)/Fe(II) redox couple. The immobilized heme proteins can display the features of peroxidase in electrocatalytic reductions of oxygen, hydrogen peroxide, nitric oxide, trichloroacetic acid (TCA), and bromate.     

Direct Electrochemistry of Multi‐Copper Oxidases at Carbon Nanotubes Noncovalently Functionalized with Cellulose Derivatives

This study describes the direct electron transfer of multi‐copper oxidases, i.e., laccase (from Trametes versicolor) and bilirubin oxidase (BOD, from Myrothecium verrucaria) at multiwalled carbon nanotubes (MWNTs) noncovalently functionalized with biopolymers of cellulose derivatives, i.e., hydroxyethyl cellulose (HEC), methyl cellulose (MC), and carboxymethyl cellulose (CMC). The functionalization of the MWNTs with the cellulose derivatives is found to substantially solubilize the MWNTs into aqueous media and to avoid their aggregation on electrode surface. Under anaerobic conditions, the redox properties of laccase and BOD are difficult to be defined with cyclic voltammetry at either laccase/MWNT‐modified or BOD/MWNT‐modified electrodes. The direct electron transfer properties of laccase and BOD are thus studied in terms of the bioelectrocatalytic activities of the laccase/MWNT‐modified and BOD/MWNT‐modified electrodes toward the reduction of oxygen and found to be facilitated at the functionalized MWNTs. The possible application of the laccase‐catalyzed O₂ reduction at the laccase/MWNT‐modified electrode is illustrated by constructing a CNT‐based ascorbate/O₂ biofuel cell with the MWNT‐modified electrode as the anode for the oxidation of ascorbate biofuel.     

Fabrication of Chitosan‐Multiwall Carbon Nanotube Nanocomposite Containing Ferri/Ferrocyanide: Application for Simultaneous Detection of D‐Penicillamine and Tryptophan

We report a simple and effective strategy for fabrication of the nanocomposite containing chitosan (CS) and multiwall carbon nanotube (MWNT) coated on a glassy carbon electrode (GCE). The characterization of the modified electrode (CS‐MWNT/GC) was carried out using scanning electron microscopy (SEM) and UV–vis absorption spectroscopy. The electrochemical behavior of CS‐MWNT/GC electrode was investigated and compared with the electrochemical behavior of chitosan modified GC (CS/GC), multiwalled carbon nanotube modified GC (MWNT/GC) and unmodified GC using cyclic voltammetry (CV) and electron impedance spectroscopy (EIS). The chitosan films are electrochemically inactive; similar background charging currents are observed at bare GC. The chitosan films are permeable to anionic Fe(CN)₆^3?/4? (FC) redox couple. Electrochemical parameters, including apparent diffusion coefficient for the Fe(CN)₆^3?/4? redox probe at FC/CS‐MWNT/GC electrode is comparable to values reported for cast chitosan films. This modified electrode also showed electrocatalytic effect for the simultaneous determination of D‐penicillamine (D‐PA) and tryptophan (Trp). The detection limit of 0.9 μM and 4.0 μM for D‐PA and Trp, respectively, makes this nanocomposite very suitable for determination of them with good sensitivity.     

Electrochemical Behavior of K3[Fe(CN)6] on the Electrode Coated with Modified Multi‐Walled Carbon Nanotubes

The electrochemical behavior of K₃[Fe(CN)₆] was studied on an ITO electrode that was coated with β‐cyclodextrin (CD) modified multi‐walled carbon nanotubes (MWNTs) and with carboxyl modified multi‐walled carbon nanotubes (MWNT‐COOHs). MWNT‐COOHs showed an excellent electrocatalytic effect on the redox of K₃[Fe(CN)₆] while MWNT‐CDs had a subdued effect on the electrochemical response of K₃[Fe(CN)₆]. It is probably due to mismatching between K₃[Fe(CN)₆] and cyclodextrin, which hampers the contact of K₃[Fe(CN)₆] with carbon nanotubes. Moreover, the electrochemical behavior of K₃[Fe(CN)₆] on the MWNT‐COOHs coated ITO electrode at various scan rates also was measured. The results indicated that both potential difference between redox peaks and peak current of K₃[Fe(CN)₆] increased with increasing scan rate. A good linearity of peak current versus scan rate was observed.     

多璧碳管/聚中性红修饰电极的制备及对鸟嘌呤和腺嘌呤的应用研究

结合纳米材料的电催化特性和中性红聚合物薄膜的分子识别能力, 以玻碳电极为基体制备了多壁碳管/聚中性红(MWNT/PNR)修饰电极, 并用表面扫描电镜和循环伏安法进行了表征. 实验表明, 该修饰电极对腺嘌呤(A)和鸟嘌呤(G)都表现出了良好的电催化性能. 在最佳条件下, 用示差脉冲伏安法对A和G进行了测定, 其氧化峰电流于A和G的浓度分别在0.01～4 μmol/L和0.01～8 μmol/L范围内呈良好的线性关系, 检测限均为5×10-9 mol/L (S/N=3). 该修饰电极可以用来同时测定DNA中的A和G.     

A Nitric Oxide Biosensor Based on Myoglobin Adsorbed on Multi‐Walled Carbon Nanotubes

Myoglobin (Myb) of horse heart is incorporated on multi‐walled carbon nanotubes (MWNTs) and immobilized at a glassy carbon (GC) electrode surface. Its electrochemical behavior and enzyme activity are characterized by employing electrochemical methods. The results indicate that MWNTs can obviously promote the direct electron transfer between Myb and electrode, and that the Myb on MWNTs behaves as an enzyme‐like activity towards the electrochemical reduction of nitric oxide (NO). Accordingly, an unmediated NO biosensor is constructed. Experimental results reveal that the peak current related to NO is linearly proportional to its concentration in the range of 2.0×10^?7–4.0×10^?5 mol/L. The detection limit is estimated to be 8.0×10^?8 mol/L. Considering a relative standard deviation of 2.1% in seven independent determinations of 1.0×10^?5 mol/L NO, this biosensor shows a good reproducibility. The biosensor based on Myb/MWNTs modified electrode can be used for the rapid determination of trace NO in aqueous solution with a good stability, nice selectivity and easy construction.     

Disposable electrochemical biosensor with multiwalled carbon nanotubes-chitosan composite layer for the detection of deep DNA damage.

A novel electrochemical DNA-based biosensor for the detection of deep DNA damage was designed employing the bionanocomposite layer of multiwalled carbon nanotubes (MWNT) in chitosan (CHIT) deposited on a screen printed carbon electrode (SPCE). The biocomponent represented by double-stranded (ds) herring sperm DNA was immobilized on this composite using layer-by-layer coverage to form a robust film. Individual and complex electrode modifiers are characterized by a differential pulse voltammetry (DPV) with the DNA redox marker [Co(phen)(3)](3+), cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) with [Fe(CN)(6)](3-) as a redox probe in a phosphate buffer solution (PBS). A good correlation between the CV and EIS parameters has been found, thus confirming a strong effect of MWNT on the enhancement of the electroconductivity of the electrode surface and that of CHIT on the MWNT distribution at the electrode surface. Differences between the CV and EIS signals of the electrodes without and with DNA are used to detect deep damage to DNA, advantageously using simple working procedures in the same experiment.     

The fabrication of a colloidal gold-carbon nanotubes composite film on a gold electrode and its application for the determination of cytochrome c

Colloid Au (Au_nano) with a diameter of about 20 nm was prepared and used in combination with the multi-wall carbon nanotubes (MWNTs) to modify a gold electrode. Dihexadecylphosphate (DHP) dispersed in Au_nano aqueous solution was used to solubilize MWNTs. Deposition of Au_nano on MWNTs was realized as illustrated by TEM micrographs. The DHP formed a network that connected Au_nano and MWNTs to the gold electrode surface. The Au_nano–MWNTs–DHP composite film on the gold electrode surface was characterized by electrochemical impedance spectroscopy (EIS) and cyclic voltammmetry (CV). The composite film modified gold electrode was used to detect cytochrome c and a pair of well-defined redox waves was obtained. It was found that the composite film promoted the redox of horse heart cytochrome c and its effect was developed for the determination of cytochrome c.     

Self‐Assembly of Platinum Nanoparticle/Multiwalled Carbon Nanotube Multilayer Film on Au Substrate Electrode

A novel approach to assemble multilayer films of Pt nanoparticle/multiwalled carbon nanotube (MWNTs) composites on Au substrate has been developed for the purpose of improving the methanol oxidation efficiency by providing high catalytic surface area. MWNTs were firstly functionalized with 4‐mercaptobenzene and then assembled on an Au substrate electrode. Pt nanoparticles were fabricated and attached to the surface of the functionalized MWNTs subsequently. Thus a layer of Pt/MWNT composites were assembled on the Au substrate electrode. Repeating above process can assemble different layers of film of Pt/MWNTs composites on the Au electrode. Cyclic voltammetry shows that the Au electrode modified with two layers of film of Pt/MWNT composites exhibits high catalytic ability and long‐term stability for methanol oxidation. The layer‐by‐layer self‐assembly technique provides an efficient strategy to construct complex nanostructure for improving the methanol oxidation efficiency by providing high catalytic surface area.     

Self-assembly of bilayer lipid membrane at multiwalled carbon nanotubes towards the development of photo-switched functional device

Bilayer lipid membrane (BLM) was self-assembled on a uniquely fabricated hydrophilic surface, containing N atoms from the carbon source of ethylene amine, of the multi-walled carbon nanotubes (MWNTs) to form the BLM/MWNTs nanocomposites. Cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS), and photoelectric experiments were taken to study the properties of the BLM/MWNTs nanocomposites. The thickness of the BLM, which was calculated from the CV data obtained at BLM/MWNTs electrode, turned out to be 4.38 nm, suggesting that the lipid self-assembled at the nanotubes surface was consistent with a bilayer structure. C₆₀-incorporated BLM could also be self-assembled at the nanotubes surface (C₆₀-BLM/MWNTs). The formation of BLM on the MWNTs surface blocked the diffusion of [Fe(CN)₆]^3/4− redox ions across BLM to the MWNTs electrode as no redox current was observed by CV measurement, whereas the incorporation of the electron mediator, C_60, resumed a pair of redox peaks at C₆₀-BLMs/MWNTs electrode. Moreover, the incorporation of C₆₀ led to a four order of magnitude reduction of the resistance of C₆₀-BLM/MWNTs (369.3 Ω) than that of BLM/MWNTs (3.238 × 10⁶ Ω). MWNTs electrode exhibited an intrinsic cathodic photocurrent (166 μA cm⁻²) while BLM/MWNTs electrode blocked photocurrent response of the MWNTs. Interestingly, C₆₀-BLM/MWNTs electrode resumed partial photoelectric properties (photo current: 65 μA cm⁻²) due to the electron mediation effect of C₆₀ incorporated into the lipid membrane. As a result, the novel self-assembled BLM/MWNTs nanocomposites provided a simple yet useful model to study the C₆₀-mediated photoelectric properties of the BLM/MWNTs which may be applicable to develop new biosensors and molecular devices.     

Voltammetric Characterization of Grafted Polymer Modified with ZnO Nanoparticles on Glassy Carbon Electrode

In this study, a grafted polymer (GP) with ZnO nanoparticles (GP/ZnO NPs) was attached on the surface of glassy carbon electrode (GCE), in order to produce a new modified electrode (GP/ZnO NPs-GCE). The gamma irradiation method was used to grafted polystyrene (polymer) with acrylonitrile (monomer), while slow evaporation process was used to prepare the new modified electrode. The cyclic voltammetry (CV) of K₄[Fe(CN)₆] was used to study the electrochemical properties GP/ZnO NPs-GCE. The peak separation (ΔE_pa-c) was 500 mV between the redox peaks of Fe(II)/Fe(III) in an aqueous solution of 1 M KCl and the current ratio of redox current peaks (I_pa/I_pc) was ≈ 1 for the modified electrode. This indicated that the modified electrode has s good reversibility and conductivity, wherefore; it was applied in the voltammetric filed. It was found that the modified electrode GP/ZnO NPs-GCE have a reasonable solubility and stability at various pH medium. Additionally, the sensitivity of the electrochemical analysis by cyclic voltammetric (CV) method is extensively subjected to the pH medium and the scan rate (SR). A couple of redox current peaks of K₄[Fe(CN)₆] in KCl solution was observed with a reversible process: Fe³⁺/Fe²⁺. Finally a good diffusion coefficient of electroactive species (D) for the new modified electrode was found in this study by chronoamperometry method using Cottrell equation.     

Electrocrystallized Ag nanoparticle on functional multi-walled carbon nanotube surfaces for hydrazine oxidation

4-Aminobenzoic acid was covalently grafted on multi-walled carbon nanotubes (MWNTs) by amine cation radical formation in the electrooxidation process of the amino-containing compound. Then, silver (Ag) nanoparticles were electrocrystallized on 4-aminobenzoic acid monolayer-grafted MWNTs by a potential-step method. The structure and nature of the resulting Ag/MWNT composites were characterized by transmission electron microscopy and X-ray diffraction. The electrocatalytic properties of the Ag/MWNT electrode for hydrazine oxidation have been investigated by cyclic voltammetry, high electrocatalytic activity of the Ag/MWNT electrode can be observed. This may be attributed to the small particle size of the silver particles. The results imply that the Ag/MWNT composites have a good application potential in fuel cells.     

Direct Electrochemistry of Myoglobin on a Room Temperature Ionic Liquid Modified Electrode and Its Application to Nitric Oxide Biosensing

Myoglobin (Myb) was successfully immobilized on a room temperature ionic liquid (RTIL), 1‐ethyl‐3‐methyl imidazolium tetrafluoroborate ([EMIM][BF₄]) modified basal plane graphite (BPG) electrode. The electrochemical behavior of Myb on RTIL modified BPG electrode was explored and the results from cyclic voltammetry (CV) showed a well‐defined and quasi‐reversible CV peaks with a formal potential of ?0.379 V (versus Ag/AgCl) in a phosphate buffer solution (pH 7.0). RTIL shows an obvious promotion for the direct electron‐transfer between Myb and BPG electrode. Myb adsorbed on electrode surface exhibits an obvious electrocatalytic activity for the reduction of nitric oxide (NO). The catalytic current is corresponding linearly to the NO concentration in the range of 7.0×10^?7 to 7.0×10^?6 M with a limit of detection of 2.0×10^?7 M (three times the ratio of signal to noise, S/N=3).     

功能化多壁碳纳米管的光电性质

表面光电压谱;循环伏安;金刚石薄膜电极;功能化多壁碳纳米管的光电性质     

碳纳米管尺寸对电化学反应活性的影响

制备不同尺寸的多壁碳纳米管(MWNT)修饰电极,应用循环伏安法研究了相同管径、不同管长和相同管长、不同管径的多壁碳纳米管修饰电极在K3Fe(CN)6溶液中的电化学行为及其对尿酸、多巴胺等生物分子的电催化作用,以及尺寸效应对碳纳米管修饰电极电化学活性的影响规律.结果显示,在同一条件下,短管的MWNT比长管的更能有效促进K3Fe(CN)6的电子传递,更有利于对生物分子的电催化;管径对它的电化学行为及生物电催化活性影响较小,无明显规律.主要原因在于碳纳米管管端、管壁的不同电化学活性.     

Amperometric Biosensors for Glucose Based on Layer‐by‐Layer Assembled Functionalized Carbon Nanotube and Poly (Neutral Red) Multilayer Film

Abstract

Multiwalled carbon nanotubes (MWNTs) were treated with a mixture of concentrated sulfuric and nitric acid to introduce carboxylic acid groups to the nanotubes. Conducting polymer film was prepared by electrochemical polymerization of neutral red (NR). By using a layer‐by‐layer method, homogeneous and stable MWNTs and poly (neutral red) (PNR) multilayer films were alternately assembled on glassy carbon (GC) electrodes. With the introduction of PNR, the MWNTs/PNR multilayer film system showed synergy between the MWNTs and PNR, with a significant improvement of redox activity due to the excellent electron‐transfer ability of carbon nanotubes (CNTs) and PNR. The electropolymerization is advantageous, providing both prolonged long‐term stability and improved catalytic activity of the resulting modified electrodes. The MWNTs/PNR multilayer film modified glassy carbon electrode allows low potential detection of hydrogen peroxide with high sensitivity and fast response time. As compared to MWNTs and PNR‐modified GC electrodes, the magnitude of the amperometric response of the MWNTs/PNR composite‐modified GC electrode is more than three‐fold greater than that of the MWNTs modified GC electrode, and nine‐fold greater than that of the PNR‐modified GC electrode. With the immobilization of glucose oxidase onto the electrode surface using glutaric dialdehyde, a biosensor that responds sensitively to glucose has been constructed. In pH 6.98 phosphate buffer, nearly interference‐free determination of glucose has been realized at ?0.2 V vs. SCE with a linear range from 50 µM to 10 mM and response time <10s. The detection limit was 10 µM glucose (S/N=3).     

功能化碳纳米管的电化学性质及在6-巯基嘌呤分析检测中的应用

采用修饰单壁碳纳米管(SWNT、SWNT-COOH或SWNT-OH)及多壁碳纳米管(MWNT、MWNT-COOH或MWNT-OH)的石墨电极研究配位阴离子[Fe(CN)6]3-和配位阳离子[Co(phen)3]3+的电化学行为与吸附性能,借助[Co(phen)3]3+在碳纳米管(CNT)的强吸附特性制备[Co(phen)3]3+/CNT/C修饰电极,以其应用于6-MP的分析检测.结果表明:1)在CNT修饰电极上[Fe(CN)6]3-/4-呈现很好的氧化还原可逆性,而[Co(phen)3]3+则显示明显的吸附控制特征.2)[Co(phen)3]3+在多壁碳纳米管修饰电极上的吸附量较单壁碳纳米管大,但经羧基化或羟基化后,吸附量减小,而且在羧基化表面的吸附量较羟基化的大.3)[Co(phen)3]3+与6-MP间存在明显的相互作用,其配位产物的还原峰电流与6-MP浓度呈线性关系.     

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.     

Electro-deposition of platinum nanoparticles on 4-mercaptobenzene-functionalized multi-walled carbon nanotubes

A new method to electro-deposit platinum nanoparticles on the surface of multi-walled carbon nanotubes (MWNTs) functionalized with 4-mercaptobenzene has been described. X-ray photoelectron spectroscopy results reveal that 4-mercaptobenzene was attached to the surface of MWNTs. Transmission electron microscope and X-ray diffraction analysis confirm that platinum nanoparticles were highly dispersed on the surface of MWNTs, and the average size of the platinum particle is 4.2 nm. The electrocatalytic properties of the Pt/MWNT composite electrode for methanol oxidation were investigated by cyclic voltammetry, and the results show that the fabricated composites exhibit high catalytic activity and good long-term stability. The study provides a feasible approach to fabricate Pt/MWNT composite electrode for direct methanol fuel cell.     

Fabrication of multilayer-film-modified gold electrode composed of myoglobin,chitosan, and polyelectrolyte-wrapped multi-wall carbon nanotubes by layer-by-layer assembled technique and electrochemical catalysis for hydrogen peroxide and trichloroacetic acid

Electroactive multilayer film of myoglobin (Mb)-, chitosan (CS)-, and poly(dimethyldiallylammonium chloride) (PDDA)-wrapped multi-wall carbon nanotubes (MWNTs) is fabricated on a gold electrode via layer-by-layer (LBL) technique. The assembled multilayer films is characterized by scanning electron microscopy (SEM), UV-vis spectroscopy, cyclic voltammetry (CV), and electrochemical impedance spectroscopy (EIS). UV-vis spectroscopy showed that Mb in the films retained its near-native structure. The stable multilayerfilm-modified gold electrodes showed good electroactivity in protein-free buffer solution, which is originated from protein heme Fe(III)/Fe(II) redox couple. The modified electrode exhibited good electrocatalytic property toward reduction of H₂O₂ and trichloroacetic acid, indicating the potential application as amperometric biosensor. Published in Russian in Elektrokhimiya, 2008, Vol. 44, No. 11, pp. 1366–1376. The text was submitted by the authors in English.