Direct Electrochemistry and Electrocatalytic Properties of Hemoglobin Immobilized on Carbon Nanotubes Ionic Liquid Electrode

Hemoglobin (Hb) was directly immobilized on a multiwalled carbon nanotubes ionic liquid electrode by immersing this electrode in a solution consisting of Hb and 1‐octyl‐pyridinium chloride as an ionic liquid. The immobilized Hb displayed a pair of well‐defined cyclic voltammetric peaks with a formal potential of ?0.187 V in acetate buffer solution, pH 5.0. This modified electrode exhibits fast response, a long linearity range, a low detection limit, high stability and excellent sensitivity through hydrogen peroxide detection with a detection limit (3σ) of 3.18 µM. The electrode was also used for electrocatalysis and voltammetric determination of oxygen and trichloroacetic acid.     

Direct Electrochemistry and Electrocatalysis of Hemoglobin on Aligned Carbon Nanotubes Based Electrodes Modified with Au Nanoparticles and SiO2 Gel

Here we report on the preparation and characterization of new electrodes based on aligned carbon nanotubes (ACNTs) for hemoglobin (Hb) electrochemistry and electrocatalysis. The ACNTs are obtained by a thermal chemical vapor deposition method under normal pressure. Then the electrodes are elaborated by first sputtering a thin Au film (thickness of 200 nm) onto the top of the ACNTs, and then removing the Au layer/ACNTs from the quartz substrate with the aide of hydrofluoric acid (HF) treatment. Field emission scanning electron microscopy (FESEM) demonstrates that after nitric acid (HNO₃) treatment, the nanotubes of the removed Au layer are totally tip‐opened, purified and organized in a perfect vertically aligned architecture. The final ACNTs electrode is obtained by attaching the Au layer of ACNTs onto a glassy carbon electrode. Then the electrode was modified to act as a matrix for hemoglobin (Hb) immobilization and as an electrode for Hb electroanalysis by the assistance of Au nanoparticles (AuNPs) and SiO₂ gel. Due to the individual specific effects of AuNPs, SiO₂ gel and ACNTs, the resulting SiO₂/Hb‐AuNPs/ACNTs electrode showed good direct electrochemistry of Hb with an apparent Michaelis? Menten constant of 0.44 mM. The electrode showed an excellent electrocatalytic activity towards H₂O₂, possessing a linear range from 40 µM to 4 mM and the detection limit was 22 µM based on a signal to noise ratio of 3.     

A Nitrite Biosensor Based on the Direct Electron Transfer of Hemoglobin Immobilized on Carboxyl‐Functionalized Multiwalled Carbon Nanotubes/Polyimide Composite

The high electrically conductive carboxyl‐functionalized multiwalled carbon nanotubes (COOH‐MWCNTs) were used to combine with nonconducting polyimide (PI) to generate a PI/COOH‐MWCNTs membrane. PI served as a matrix to entrap COOH‐MWCNTs and hemoglobin (Hb). COOH‐MWCNTs can improve the conductivity of the composite. The direct electrochemistry measurement indicated that the PI/COOH‐MWCNTs composite enhanced the immobilization of Hb significantly. Besides, the Hb/PI/COOH‐MWCNTs/GCE biosensor possessed excellent electrocatalytic activity for the detection of nitrite. Therefore, PI is a good matrix for Hb immobilization and it has application in sensor construction. This work is promising in the development of sensitive biosensors based on PI/COOH‐MWCNTs composite film.     

血红蛋白在碳纳米管修饰碳糊电极上的直接电化学行为

利用吸附法将血红蛋白(Hb)固定在碳纳米管修饰碳糊电极表面,制成稳定的固载Hb碳纳米管修饰电极,研究了Hb在碳纳米管修饰电极上的直接电化学行为.固载Hb的碳纳米管修饰电极在pH=7.0的PBS（磷酸盐缓冲溶液）中有一对相当可逆的循环伏安氧化还原峰,为Hb血红素辅基Fe(Ⅲ)/Fe(Ⅱ)电对的特征峰.式电位为－0.160 V(vs SCE),随扫描速度变化很小.电子转移数为1.021,近似为一个辅基发生电子转移.Hb在碳纳米管修饰电极表面的电子转移常数为0.0816 s－1,远大于亚甲蓝作媒介体时Hb的电子转移反应速率常数.应用于过氧化氢、三氯乙酸和硝基苯等的电催化还原,固定在碳纳米管修饰碳糊电极的血红蛋白表现出稳定且较高的催化活性.     

Direct Electrochemistry of Hemoglobin and its Electrocatalysis Based on a Carbon Nanotube Paste Electrode

A new hemoglobin (Hb) and carbon nanotube (CNT) modified carbon paste electrode was fabricated by simply mixing the Hb, CNT with carbon powder and liquid paraffin homogeneously. To prevent the leakage of Hb from the electrode surface, a Nafion film was further applied on the surface of the Hb‐CNT composite paste electrode. The modified electrode was characterized by scanning electron microscopy (SEM) and electrochemical impedance spectroscopy (EIS). Direct electrochemistry of hemoglobin in this paste electrode was easily achieved and a pair of well‐defined quasi‐reversible redox peaks of a heme Fe(III)/Fe(II) couple appeared with a formal potential (E^0′) of ?0.441 V (vs. SCE) in pH 7.0 phosphate buffer solution (PBS). The electrochemical behaviors of Hb in the composite electrode were carefully studied. The fabricated modified bioelectrode showed good electrocatalytic ability for reduction of H₂O₂ and trichloroacetic acid (TCA), which shows potential applications in third generation biosensors.     

Direct Electrochemistry and Electrocatalysis of Hemoglobin Immobilized in a Polymeric Ionic Liquid Film

A polymer film based on polymeric ionic liquid, which was poly(1‐vinyl‐3‐butylimidazolium chloride) (poly(ViBuIm⁺Cl^?)for short), was firstly used as matrix to immobilize hemoglobin (Hb). FTIR and UV‐vis spectra demonstrated that the native structure of Hb was well preserved after entrapped into the polymer film. The Hb immobilized in the poly(ViBuIm⁺Cl^?) film showed a fast direct electron transfer for the Hb‐Fe^III/Fe^II redox couple. Based on the direct electron transfer of the immobilized Hb, polyvinyl alcohol (PVA)/Hb/poly(ViBuIm⁺Cl^?)/GC electrode displayed good sensitivity and wide linear range for the detection of H₂O₂. The linear range of the PVA/Hb/poly(ViBuIm⁺Cl^?)/GC electrode to H₂O₂ is from 3.5 to 224 μM with a limit of detection of 1.17 μM. Such an avenue, which integrated polymeric ionic liquid and redox protein via a simple method, may provide a novel and efficient platform for the fabrication of biosensors, biofuel cells and other bioelectrochemical devices.     

Direct Electrochemistry of Hemoglobin on Single‐Walled Carbon Nanotubes Modified Carbon Ionic Liquid Electrode and Its Electrocatalysis

In this article we report on the fabrication of a carbon ionic liquid electrode (CILE) by using a room temperature ionic liquid of 1‐butyl‐3‐methylimidazolium hexafluorophosphate (BMIMPF₆) as binder. It was further modified by single‐walled carbon nanotubes (SWCNTs) to get a SWCNTs modified CILE denoted as SWCNTs/CILE. The redox protein of hemoglobin (Hb) was further immobilized on the surface of SWCNTs/CILE with the help of Nafion film. UV‐vis and FT‐IR spectra indicated that the immobilized Hb retained its native conformation in the composite film. The direct electrochemistry of Hb on the SWCNTs/CILE was carefully studied in pH 7.0 phosphate buffer solution (PBS). Cyclic voltammetric results indicated that a pair of well‐defined and quasireversible voltammetric peaks of Hb heme Fe(III)/Fe(II) was obtained with the formal potential (E°') at ?0.306 V (vs. SCE). The electrochemical parameters such as the electron transfer coefficient (α), the electron transfer number (n) and the apparent heterogeneous electron transfer rate constant (k_s) were calculated as 0.34, 0.989 and 0.538 s^?1, respectively. The fabricated Hb modified electrode showed good electrocatalytic ability to the reduction of trichloroacetic acid (TCA) in the concentration range from 20.0 to 150.0 mmol/L with the detection limit of 10.0 mmol/L (3σ).     

Direct Electrochemistry and Electrocatalysis of Hemoglobin at PAMAM Dendrimer-MWNTs-Au Nanoparticles Composite Film Modified Glassy Carbon Electrode

The direct electron transfer of hemoglobin at the PAMAM-MWNTs-AuNPs composite film modified glassy carbon electrode was studied. In a phosphate buffer solution(PBS, pH=7.0), the formal potential(E 0' ) of Hb was –0.105 V versus SCE, the electron transfer rate constant was 4.66 s –1 . E 0′ of Hb at the modified electrode was linearly varied in a pH range of 5.0—8.0 with a slope of –49.2 mV/pH. The Hb/PAMAM-MWNTs-AuNPs/GCE gave an ex-cellent electrocatalytic response to the reduction of hydrogen peroxide. The catalytic current increased linearly with H 2 O 2 concentration in a range of 1.0×10 ?6 to 2.2×10 ?3 mol/L. The detection limit was 2.0×10 ?7 mol/L at a signal to noise ratio of 3. The Michaelis-Menten constant(K ma pp ) was 2.95 mmol/L.     

Electrochemistry and Electrocatalysis of Hemoglobin on 1‐Pyrenebutanoic Acid Succinimidyl Ester/Multiwalled Carbon Nanotube and Au Nanoparticle Modified Electrode

A biocompatible nanocomposite film was fabricated for hemoglobin (Hb) molecules immobilization. This film consists of multiwalled carbon nanotubes (MWNTs), 1‐pyrenebutanoic acid, succinimidyl ester (PASE), hemoglobin (Hb) and Au nanoparticles (AuNPs). Herein, PASE molecules physically adsorbed onto MWNTs, and its groups then covalently bond with Hb. AuNPs were then linked with Hb/PASE/MWNTs via electrostatic adsorption force. UV‐visible adsorption spectra, Fourier transform infrared spectra, scanning electron microscope and electrochemical impedance spectroscopy have characterized the film. Cyclic voltammetry (CV) scans showed that in the film Hb has well‐defined redox reaction, with the formal potential (E°) at about ?0.27 V (vs. Ag/AgCl). Herein, differential pulse voltammetry (DPV) was employed to electrochemically detect the Hb in the film with a detection limit of 9.3×10^?8 M. The proposed method was also succeeded for Hb detection in clinical blood samples. Furthermore, the Hb in the film showed good electrocatalytic activities to the reduction of H₂O₂, TCA, NaNO₂ and O₂.     

铂纳米颗粒修饰直立碳纳米管电极的葡萄糖生物传感器

基于Pt纳米颗粒修饰直立的碳纳米管电极制备了葡萄糖生物传感器.铂纳米颗粒是利用电位脉冲沉积法修饰到直立碳纳米管上的,可以增强电极对酶反应过程当中产生的过氧化氢的催化行为.用扫描电镜和透射电镜观察了直立碳纳米管在修饰Pt纳米颗粒前后的形态.该酶电极对葡萄糖的氧化表现出很好的响应,线性范围为1×10-5～7×10-3mol/L,响应时间小于5s,并且有很好的重现性.     

Direct Electrochemistry Transfer and Electrocatalysis of Hemoglobin Immobilized in Porous Mn_2O_3

Novel porous Mn2O3 with good crystallinity was synthesized via hard-template method. Hb-Mn2O3 na-nocomposite was prepared and used for biosensor construction. The Hb-Mn2O3-Nafion modified electrode shows fast direct electron transfer and displays good electrocatalytic response to the reduction of H2O2. The response time is less than 5 s, the sensitivity is as high as 493 μA·L·mmol-1·cm-2 in a linear range of 1-100 μmol/L, and the detection limit is 0.16 μmol/L. This modified electrode also shows good stabil...     

Direct Electrochemistry and Electrocatalysis of Hemoglobin–TiO2 Whisker Film Modified Glassy Carbon Electrode

In this study, Hb TiO₂ whisker nanocomposites are prepared by incorporating TiO₂ whisker with hemoglobin (Hb). Our studies illustrate that the self‐assembled Hb TiO₂ whisker films could efficiently facilitate the direct electron transfer of Hb on glassy carbon electrode (GCE). Moreover, our results demonstrate that the catalytic activity to the reduction of H₂O₂ could be observed on the Hb TiO₂ whisker modified GCE and the photovoltaic effect of TiO₂ whisker can greatly enhance the detection sensitivity of electrocatalytic reduction.     

碳糊电极上无机膜固载血红蛋白的直接电化学

报道了用硅溶胶-凝胶(Sol-gel)膜将血红蛋白(Hb)固载于碳糊电极上的直接电化学行为.研究结果表明,Hb-Sol-gel修饰的碳糊电极在pH=7.0的缓冲溶液中于-0.275V(vs.Ag/AgCl)处有一对可逆的循环伏安氧化-还原峰,为Hb血红素辅基Fe(Ⅲ)/Fe(Ⅱ)电对的特征峰.HbFe(Ⅲ)/Fe(Ⅱ)电对的式量电位在pH5.0～11.0范围内与溶液pH值呈线性关系,表明Hb的电化学还原很可能是一个质子伴随着一个电子的电极过程.FTIR光谱证实,Sol-gel膜对Hb的固载没有破坏其天然结构.Hb-Sol-gel修饰的碳糊电极能够催化还原H₂O₂,可望将其用于制作第三代生物传感器.     

Direct Electrochemistry and Electrocatalysis of Myoglobin Immobilized on Gold Nanoparticles/Carbon Nanotubes Nanohybrid Film

A novel nanohybrid material, constructed by gold nanoparticles (GNPs) and multiwalled carbon nanotubes (MWNTs), was designed for immobilization and biosensing of myoglobin (Mb). Morphology of the nanohybrid film was characterized by SEM. UV‐vis spectroscopy demonstrated that Mb on the composite film could retain its native structure. Direct electrochemistry of Mb immobilized on the GNPs/MWNTs film was investigated. The immobilized Mb showed a couple of quasireversible and well‐defined cyclic voltammetry peaks with a formal potential of about ?0.35 V (vs. Ag/AgCl) in pH 6.0 phosphate buffer solution (PBS) solution. Furthermore, the modified electrode also displayed good sensitivity, wide linear range and long‐term stability to the detection of hydrogen peroxide. The experiment results demonstrated that the hybrid matrix provided a biocompatible microenvironment for protein and supplied a necessary pathway for its direct electron transfer.     

Direct Electrochemistry and Electrocatalysis of Hemoglobin/ZnO‐Chitosan/nano‐Au Modified Glassy Carbon Electrode

The highly efficient H₂O₂ biosensor was fabricated on the basis of the complex films of hemoglobin (Hb), nano ZnO, chitosan (CHIT) dispersed solution and nano Au immobilized on glassy carbon electrode (GCE). Biocompatible ZnO‐CHIT composition provided a suitable microenvironment to keep Hb bioactivity (Michaelis‐Menten constant of 0.075 mmol L^?1). The presence of nano Au in matrix could effectively enhance electron transfer between Hb and electrode. The electrochemical behaviors and effects of solution pH values were carefully examined in this paper. The (ZnO‐CHIT)‐Au‐Hb/GCE demonstrated excellently electrocatalytical ability for H₂O₂. This biosensor had a fast response to H₂O₂ less than 4 s and excellent linear relationships were obtained in the concentration range from1.94×10^?7 to 1.73×10^?3 mol L^?1 with the detection limit of 9.7×10^?8 mol L^?1 (S/N=3) under the optimum conditions. Moreover, the stability and reproducibility of this biosensor were evaluated with satisfactory results.     

Amperometric Glucose Biosensor Based on Platinum Nanoparticles Combined Aligned Carbon Nanotubes Electrode

A sensitive amperometric glucose biosensor based on platinum nanoparticles (PtNPs) combined aligned carbon nanotubes (ACNTs) electrode was investigated. PtNPs which can enhance the electrocatalytic activity of the electrode for electrooxidating hydrogen peroxide by enzymatic reaction were electrocrystallized on 4‐aminobenzene monolayer‐grafted ACNTs electrode by potential‐step method. These PtNPs combined ACNTs' (PtNPs/ACNTs) surfaces were characterized by scanning electron microscopy (SEM) and transmission electron microscopy (TEM). The highly dispersed PtNPs on ACNTs can be obtained. The enzyme electrode exhibits excellent response performance to glucose with linear range from 1×10^?5–7×10^?3 mol L^?1 and fast response time within 5 s. Furthermore, this glucose biosensor also has good reproducibility. It is demonstrated that the PtNPs/ACNTs electrode with high electrocatalytic activity is a suitable basic electrode for preparing enzyme electrodes.     

Electrocatalysis via Direct Electrochemistry of Myoglobin Immobilized on Colloidal Gold Nanoparticles

The direct electron transfer between immobilized myoglobin (Mb) and colloidal gold modified carbon paste electrode was studied. The Mb immobilized on the colloidal gold nanoparticles displayed a pair of redox peaks in 0.1 M pH 7.0 PBS with a formal potential of –(0.108 ± 0.002) V (vs. NHE). The response showed a surface‐controlled electrode process with an electron transfer rate constant of (26.7 ± 3.7) s ^?1 at scan rates from 10 to 100 mV s^?1 and a diffusion‐controlled process involving the diffusion of proton at scan rates more than 100 mV s^?1. The immobilized Mb maintained its activity and could electrocatalyze the reduction of both hydrogen peroxide and nitrite. Thus, the novel renewable reagentless sensors for hydrogen peroxide and nitrite were developed, respectively. The activity of Mb with respect to the pseudo peroxidase with a K_M^app value of 0.65 mM could respond linearly to hydrogen peroxide concentration from 4.6 to 28 μM. The sensor exhibited a fast amperometric response to NO₂^? reduction and reached 93% of steady‐state current within 5 s. The linear range for NO₂^? determination was from 8.0 to 112 μM with a detection limit of 0.7 μM at 3σ.     

Direct Electrochemistry and Application in Electrocatalysis of Hemoglobin in a Polyacrylic Resin‐Gold Colloid Nanocomposite Film

A novel nanocomposite integrating the good biocompatibility of polyacrylic resin nanoparticles (PAR) and the good conductivity of colloidal gold nanoparticles was proposed to construct the matrix for the immobilization of hemoglobin (Hb) on the surface of a glassy carbon electrode (GCE). UV‐vis spectra demonstrated that Hb preserved its native structure after being entrapped into the composite film. The direct electrochemistry of hemoglobin (Hb) in this nanocomposite films showed a pair of well‐defined and quasi‐reversible cyclic voltammetric peaks with a formal potential of ?0.307 mV and a constant electron transfer rate of 2.51±0.2 s^?1. The resultant amperometric biosensor showed fast responses to the analytes with excellent detection limits of 0.2 µM for H₂O₂ and 0.89 µM for TCA (S/N=3), and high sensitivity of 1108.6 for H₂O₂ and 77.14 mA cm^?2 M^?1 for TCA, respectively. The linear current response was found in the range from 0.59 to 7.3 µM (R²=0.9996) for H₂O₂ and from 5 to 85 µM (R²=0.9996) for TCA, while the superior apparent Michaelis–Menten constant was 0.012 mM for H₂O₂ and 0.536 mM for TCA, respectively. Therefore, the PAR‐Au‐Hb nanocomposite as a novel matrix opens up a possibility for further study on the direct electrochemistry of other proteins.     

Direct Electrochemistry of Hemoglobin on Vertically Aligned Carbon Hybrid TiO2 Nanotubes and Its Highly Sensitive Biosensor Performance

The present work is focused on developing a novel biomaterial platform to achieve enhanced direct electron transfer (DET) of hemoprotein and higher biosensor performance on vertically aligned carbon hybrid TiO₂ nanotubes (C‐TiO₂ NTs). Using a simple surfactant‐assisted method, controllable hybridization of TiO₂ NTs with conductive amorphous carbon species is realized. The obtained C‐TiO₂ NTs is ingeniously chosen to serve as an ideal "vessel" for protein immobilization and biosensor applications. Results show that the appropriate hybridization of C into TiO₂ NTs leads to a much better conductivity of TiO₂ NTs without destroying their preponderant tubular structures or damaging their excellent biocompatibility and hydrophilicity. When used in loading proteins, the C‐TiO₂ NTs can be used as a super vessel for rapid and substantive immobilization of hemoglobin (Hb), with a large surface electroactive Hb coverage ( Γ ??) of 3.3×10^?9 mol·cm^?2. Enhanced DET of Hb is commendably observed on the constructed Hb/C‐TiO₂ NTs biosensor with a couple of well‐defined redox peaks in a fast electron transfer process. The biosensor further exhibits fast response, high sensitivity and stability for the amperometric biosensing of H₂O₂ with the detection limit as low as 3.1×10^?8 mol/L.     

Direct Electrochemistry of Hemoglobin Entrapped in Composite Electrodeposited Chitosan‐Multiwall Carbon Nanotubes and Nanogold Particles Membrane and Its Electrocatalytic Application

Hemoglobin was entrapped in composite electrodeposited chitosan‐multiwall carbon nanotubes (MCNTs) film by assembling gold nanoparticles and hemoglobin step by step. In phosphate buffer solution (pH 7), a pair of well‐defined and quasireversible redox peaks appeared with formal potential at ?0.289 V and peak separation of 100 mV. The redox peaks respected for the direct electrochemistry of hemoglobin at the surface of chitosan‐MCNTs‐gold nanoparticles modified electrode. The parameters of experiments have also been optimized. The composite electrode showed excellent electrocatalysis to peroxide hydrogen and oxygen, the peak current was linearly proportional to H₂O₂ concentration in the range from 1×10^?6 mol/L to 4.7×10^?4 mol/L with a detection limit of 5.0×10^?7 mol/L, and this biosensor exhibited high stability, good reproducibility and better selectivity. The biosensor showed a Michaelis–Menten kinetic response as H₂O₂ concentration is larger than 5.0×10^?4 mol/L, the apparent Michaelis–Menten constant for hydrogen peroxide was calculated to be 1.61 μmol/L.