Direct electrochemistry and electrocatalysis of heme-protein based on N,N-dimethylformamide film electrode

The heme-protein including myoglobin (Mb), hemoglobin (Hb) and horseradish peroxidase (HRP) were immobilized on normal graphite electrode by using N,N-dimethylformamide (DMF). The proteins undergo direct electron-transfer reactions. The current is linearly dependent on the scan rate, indicating that the direct electrochemistry of heme-protein in that case is a surface-controlled electrode process. The E°s are linearly dependent on solution pH (redox-Bohr effect), indicating that the electron transfer was proton-coupled. Ultraviolet-visible (UV-vis) and reflection-absorption infrared (RAIR) spectra suggest that the conformation of proteins in the presence of DMF are little different from that proteins alone the conformation changes reversibly in the range of pH 3.0-10.0. The catalytic activity of proteins were examined by hydrogen peroxide and nitrite.     

Direct electrochemistry and electrocatalysis of myoglobin-based nanocomposite membrane electrode

The direct electron transfer of myoglobin (Mb) was achieved based on the immobilization of Mb/Silver nanoparticles (AgNPs) on glassy carbon electrode by multi-wall carbon nanotubes (MWNTs)-chitosan(Chit) film. The immobilized Mb displayed a pair of well-defined and reversible redox peaks with a formal potential (E^θ′) of − 24 mV (vs. Ag/AgCl) in 0.1 M pH 7.0 phosphate buffer solution. The apparent heterogeneous electron transfer rate constants (k_s) of Mb confined to Chit-MWNTs film was evaluated as 5.47 s^{− 1} according to Laviron's equation. The surface concentration (Γ^?) of the electroactive Mb in the Chit-MWNTs film was estimated to be (4.16 ± 0.35) × 10^{− 9} mol cm^{− 2}. Meanwhile, the catalytic ability of Mb toward the reduction of H₂O₂ was studied. Its apparent Michaelis–Menten constant for H₂O₂ was 0.024 mM, showing a good affinity. The linear range for H₂O₂ determination was from 2.5 × 10^{− 5} M to 2.0 × 10^{− 4} M with a detection limit of 1.02 × 10^{− 6} M (S/N = 3). Moreover, the biosensor displays rapid response to H₂O₂ and good stability and reproducibility.     

Direct electrochemistry and electrocatalysis of myoglobin immobilized on L-cysteine self-assembled gold electrode

Myoglobin (Mb) has been successfully immobilized on a self-assembled monolayer (SAM) of L-cysteine (Cys) on a gold electrode, Au/Cys. The presence of a pair of well-defined and nearly reversible waves centered at ca. 0.086 V vs Ag/AgCl (pH 6.5) suggests that the native character of Mb heme Fe(III/II) redox couple has been obtained. The formal potential of Mb on Cys SAM exhibited pH-dependent variation in the pH range of 5-9 with a slope of 55 mV/pH, indicating that the electron transfer is accompanied by a single proton exchange. Thermodynamic and kinetic aspects of Mb adsorption processes on Au/Cys were studied by using voltammetric and quartz-crystal microbalance methods. The Au/Cys electrode with immobilized Mb exhibited electrocatalytic activity toward ascorbic acid (AA) oxidation with an overpotential decrease of over 400 mV and a linear dependence of current on the AA concentration from 0.5 to 5.0 mmol L(-1).     

Direct electrochemistry and electrocatalysis of cytochrome c immobilized on gold nanoparticles-chitosan-carbon nanotubes-modified electrode

A robust and effective nanohybrid film based on gold nanoparticles (GNPs)/chitosan (Chit)/multi-walled carbon nanotubes (MWNTs) was prepared by a layer-by-layer self-assembly technique. Cytochrome c (Cyt c) was successfully immobilized on the nanohybrid film modified glassy carbon (GC) electrode by cyclic voltammetry. The direct electron transfer between Cyt c and the modified electrode was investigated in detail. Cyt c shows a couple of quasi-reversible and well-defined cyclic voltammetry peaks with a formal potential (E^0′) of −0.16 V (versus Ag/AgCl) in pH 7.0 phosphate buffer solution (PBS). The Cyt c/GNPs/Chit/MWNTs modified GC electrode gives an improved electrocatalytic activity towards the reduction of hydrogen peroxide (H₂O₂). The sensitivity is 92.21 μA mM⁻¹ cm⁻² and the calculated apparent Michaelis-Menten constant () is 0.791 mM, indicating a high-catalytic activity of Cyt c. The catalysis currents increase linearly to the H₂O₂ concentration in a wide range of 1.5 × 10⁻⁶ to 5.1 × 10⁻⁴ M with a correlation coefficient 0.999. The detection limit is 9.0 × 10⁻⁷ M (at the ratio of signal to noise, S/N = 3). Moreover, the modified electrode displays rapid response (5 s) to H₂O₂, and possesses good stability and reproducibility.     

Direct electrochemistry and electrocatalysis of hemoglobin on gold nanoparticle decorated carbon ionic liquid electrode

In this paper a carbon ionic liquid electrode (CILE) was fabricated by using ionic liquid 1-ethyl-3-methylimidazolium ethylsulfate ([EMIM]EtOSO₃) as modifier and further gold nanoparticles were in situ electrodeposited on the surface of CILE. The fabricated Au/CILE was used as a new platform for the immobilization of hemoglobin (Hb) with the help of a Nafion film. Electrochemical experimental results indicated that direct electron transfer of Hb was realized on the surface of Au/CILE with a pair of well-defined quasi-reversible redox peaks appeared. The formal peak potential (E⁰′) was obtained as −0.210 V (vs. SCE) in pH 7.0 phosphate buffer solution (PBS), which was the characteristic of Hb heme Fe(III)/Fe(II) redox couple. The fabricated Nafion/Hb/Au/CILE showed excellent electrocatalytic activity to the reduction of trichloroacetic acid (TCA) and the reduction peak current was in proportional to TCA concentration in the range from 0.2 to 18.0 mmol/L with the detection limit as 0.16 mmol/L (S/N = 3). The proposed electrode showed good stability and reproducibility, and it had the potential application as a new third-generation electrochemical biosensor.     

Direct electrochemistry and electrocatalysis of hemoglobin on undoped nanocrystalline diamond modified glassy carbon electrode

Direct electrochemistry of hemoglobin (Hb) was observed at glassy carbon electrode (GCE) modified with undoped nanocrystalline diamond (UND) and Hb multilayer films via layer-by-layer assembly. UV-VIS absorbance spectroscopy, electrochemical impedance spectroscopy and cyclic voltammograms were employed to characterize the film. The results showed that the UND had the effect of enhancing the electron transfer between Hb and the electrode surface. Hb in the multilayer films maintained its bioactivity and structure. It also exhibited a good catalytic activity towards the reduction of H(2)O(2). The reciprocal of catalytic current showed a linear dependence on the reciprocal of H(2)O(2) concentration ranging from 0.5 microM to 0.25 mM with a detection limit of 0.4 microM. The apparent Michaelis-Menten constant was estimated to be 0.019 mM.     

Direct electrochemistry and electrocatalysis of hemoglobin in gelatine film modified glassy carbon electrode

Direct electrochemical and electrocatalytic behavior of hemoglobin (Hb) immobilized on glass carbon electrode (GCE) containing gelatine (Gel) films was investigated. The characteristics of Hb/Gel film modified GC electrode were performed by using SEM microscopy, UV-vis spectroscopy and electrochemical methods. The immobilized Hb showed a couple of quasi-reversible redox peak with a formal potential of −0.38 V (versus SCE) in 0.1 M pH 7.0 PBS. The formal potential changed linearly from pH 4.03 to 8.41 with a slope value of −52.0 mV pH⁻¹, which suggested that a proton transfer was accompanied with each electron transfer (ET) in the electrochemical reaction. The Hb/gelatine/GCE displayed a rapid amperometric response to the reduction of H₂O₂ and nitrite.     

Direct electrochemistry and electrocatalysis of hemoglobin on a gold ion implantation-modified indium tin oxide electrode

In this paper, a gold nanoparticle-modified indium tin oxide electrode (Au/ITO) was prepared without the use of any cross-linker or stabilizer reagent. The prepared Au/ITO was used as a new platform to achieve the direct electron transfer between Hb and the modified electrode. The proposed electrode exhibited a pair of well-defined redox peaks with a formal potential of ?0.073 V (vs. Ag/AgCl). The immobilized Hb showed excellent electrocatalytic activity toward H₂O₂ and the electrocatalytic current values were linear with the increasing concentration of H₂O₂ ranging from 1.0?×?10^?6?M to 7.0?×?10^?4?M. The detection limit was 2.0?×?10^?7?M (S/N?=?3) and the Michaelis–Menten constant was calculated to be 0.2 mM. The proposed electrode also showed high selectivity, long-term stability, and good reproducibility.     

Direct electrochemistry and electrocatalysis of myoglobin in dodecyltrimethylammonium bromide film modified carbon ceramic electrode

Direct electrochemistry and electrocatalysis of myoglobin（Mb） were studied with Mb immobilized on dodecyltrimethylammonium bromide（DTAB） film modified carbon ceramic（CC） electrode.Cyclic voltammetry showed a pair of well-defined and nearly reversible redox peaks of Mb（Fe^Ⅱ/Fe^Ⅲ） at about—0.3 V vs.SCE（pH = 6.98）.The currents of the redox peak were linear to scan rate,and rate constant（Ks） was estimated to be 3.03 s^-1.The formal potential（E°’） of Mb in the DTAB/CC electrodes shifted linearly with pH with a slope of -36.44 mV/pH,implying that the electron transfer between DTAB and CC electrodes is accompanied by proton transportation.The immobilized Mb exhibited excellent electrocatalytic response to the reduction of hydrogen peroxide（H₂O₂）.     

Direct electrochemistry and electrocatalysis with hemoglobin in water-soluble quantum dots film on glassy carbon electrode

The direct electrochemistry of hemoglobin can be performed by immobilizing hemoglobin in a water-soluble quantum dots (CdSe-ZnS) film on glassy carbon electrode.     

细胞色素c在Fe2O3修饰电极上的直接电化学和电催化特性

采用全氟磺酸树脂Nafion将金属氧化物Fe2O3颗粒细胞色素c(Cyt c)固定玻碳电极(GCE)表面,制备了Nafion-Cyt c-Fe2O3修饰的玻碳电极,构建了基于直接电子传递的过氧化氢生物传感器。在0.10mol/L pH7.0的磷酸盐缓冲溶液中,修饰电极的循环伏安曲线上显示出一对准可逆的氧化还原峰,式量电位为22mV。Cyt c在修饰电极表面的异相电子转移速率常数为1.21s-1。修饰后的电极对过氧化氢有良好响应,响应时间小于10s,电极的安培响应与过氧化氢浓度在2.0×10-6～3.0×10-3mol/L范围内成线性关系,检出限为1.0×10-6mol/L,米氏常数为1.35mmol/L,显示出较好的亲和力。     

Direct electrochemistry and electrocatalysis of hemoglobin on an indium tin oxide electrode modified with implanted carboxy ions

A flow injection on-line preconcentration system was developed for the determination of lead by hydride generation atomic fluorescence spectrometry (HG-AFS). It is based on a simple micro-column filled with multiwalled carbon nanotubes (MWCNTs). The preconcentration of lead on the MWCNTs was carried out based on the adsorptive retention of analyte via on-line introducing the sample into the micro-column system. A 0.3 mol L^?1 HNO₃ was introduced to elute the retained analyte and merged with KBH₄ solution for HG-AFS detection. Under the optimal experimental conditions, an enhancement factor of 26 was obtained with a sample consumption of 14.4 mL. The limit of detection was 2.8 ng L^?1 and the precision (RSD) of 11 replicate measurements of 0.2?μg L^?1 Pb was 4.4%. The method was validated by analyzing three certified reference materials, and was successfully applied to the determination of trace lead in natural water samples.     

Direct electrochemistry and electrocatalysis of heme proteins on SWCNTs-CTAB modified electrodes

Direct electrochemistry and electrocatalysis of heme proteins including hemoglobin (Hb), myoglobin (Mb) and horseradish peroxidase (HRP) were studied with the protein incorporated single walled carbon nanotubes (SWCNTs)-cetylramethylammonium bromide (CTAB) nanocomposite film modified glassy carbon electrodes (GCEs). The incorporated heme proteins were characterized with Fourier transform infrared spectroscopy (FTIR), ultraviolet visible (UV) spectroscopy, atomic force microscopy (AFM) and electrochemistry, indicating the heme proteins in SWCNTs-CTAB nanocomposite films keep their secondary structure similar to their native states. The direct electron transfer between the heme proteins in SWCNTs-CTAB films and GCE was investigated. The electrochemical parameters such as formal potentials and apparent heterogeneous electrontransfer rate constants (k_s) were estimated by square wave voltammetry with nonlinear regression analysis. The heme protein-SWCNT-CTAB electrodes show excellent electrocatalytic activities for the reduction of H₂O₂ and NO₂⁻, which have been utilized to determine the concentrations of H₂O₂ and NO₂⁻.     

Direct electrochemistry and electrocatalysis of hemoglobin in sodium alginate film on a BMIMPF6 modified carbon paste electrode

A room temperature ionic liquid (RTIL) modified carbon paste electrode was constructed based on the substitute of paraffin with 1-butyl-3-methyl-imidazolium hexafluorophosphate (BMIMPF₆) as binder for carbon paste. Direct electrochemistry and electrocatalytic behaviors of hemoglobin (Hb) entrapped in the sodium alginate (SA) hydrogel film on the surface of this carbon ionic liquid electrode (CILE) were investigated. The presence of IL in the CILE increased the electron transfer rate and provided a biocompatible interface. Hb remained its bioactivity on the surface of CILE and the SA/Hb modified electrode showed a pair of well-defined, quasi-reversible cyclic voltammetric peaks with the apparent standard potential (E^0′) at about −0.344 V (vs. SCE) in pH 7.0 Britton–Robinson (B–R) buffer solution, which was attributed to the Hb Fe(III)/Fe(II) redox couple. UV–Vis absorption spectra indicated that heme microenvironment of Hb in SA film was similar to its native status. Hb showed a thin-layer electrochemical behavior in the SA film with the direct electron transfer achieved on CILE without the help of electron mediator. Electrochemical investigation indicated that Hb took place one proton with one electron electrode process and the average surface coverage of Hb in the SA film was 3.2 × 10⁻¹⁰ mol/cm². The immobilized Hb showed excellent electrocatalytic responses to the reduction of H₂O₂ and nitrite.     

Direct electrochemistry and electrocatalysis of reduced glutathione on CNFs-PDDA/PB nanocomposite film modified ITO electrode for biosensors

The electrochemical oxidation of reduced glutathione (GSH) catalyzed by electro generated Berlin green at carbon nanofibers-poly(diallyldimethylammonium chloride)/Prussian blue (CNFs-PDDA/PB) nanocomposite film modified ITO electrode has been studied. The CNFs-PDDA/PB nanocomposite film were fabricated by casting the composite CNFs enfolded PDDA on ITO electrode followed by electrochemical deposition of PB on the CNFs-PDDA matrix using cyclic voltammetry (CV). Electron microscopy (TEM, AFM), and Fourier transform infrared spectroscopy (FT-IR) studies were used to characterize the morphology and structure of the nanocomposite. The fabricated CNFs-PDDA/PB/ITO nanocomposite film electrode shows significant improvement of redox activity of PB due to the excellent electron transfer ability of CNFs. It was also found to possess prominent electrocatalytic activity toward the oxidation of glutathione with high sensitivity as high as 2.07 μA dm(3) mol(-1) cm(-2). A nontoxic, stable and convenient method for the detection of GSH in the concentration range of 6.0×10(-6) to 1.74×10(-5) M has been developed and it showed improved sensor performance compared to the unmodified PB electrode. The high sensitivity, wider linear range, good reproducibility, and the minimal surface fouling make this CNFs/PDDA/PB nanocomposite film a promising candidate for GSH sensors.     

Direct electrochemistry and electrocatalysis of hemoglobin immobilized by methacrylic acid

The direct electrochemistry of hemoglobin (Hb) incorporated in methacrylic acid (MAA) film on a paraffin-impregnated graphite electrode (PIGE) was described. A pair of well-defined and quasi-reversible cyclic voltametric peaks are obtained. The formal potentials (E ^0′) linearly depend on the pH of solution, indicating that the electron transfer was proton-coupled. Ultraviolet-visible (UV-Vis) spectra showed that the secondary structure of Hb in the MAA film was similar to individual Hb. The immobilized Hb retained its biological activity well and exhibited a nice response to the reduction of both NO₂⁻, and H₂O₂, on the basis of which a new biosensor has been developed. Published in Russian in Elektrokhimiya, 2008, Vol. 44, No. 9, pp. 1079–1086. The text was submitted by the authors in English.     

Myoglobin on multi-walled carbon nanotubes modified electrode: direct electrochemistry and electrocatalysis

Electrochemical behavior of myoglobin (Myb) incorporated on multi-walled carbon nanotubes (MWNTs) modified GC electrode is investigated by electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV). The results indicate that Myb can be strongly adsorbed on the surface of the MWNT-modified electrode to form an approximate monolayer film. MWNTs can greatly promote the redox of horse heart Myb, on which a pair of well-defined and nearly reversible CV peaks for Myb Fe(III)/Fe(II) redox couple were obtained in pH 7.0 buffers. The formal potential of Myb on MWNT modified electrode shifts linearly with pH with a slope of −56.4 mV/pH, showing that the electron transfer is accompanied by single proton transportation. The high efficiency of the Myb/MWNT modified electrode towards the catalytic electro-reduction of nitric oxide has been observed. Potential application of Myb/MWNT modified electrode as biosensors to monitor NO is proposed.     

Direct electrochemistry and electrocatalysis of hemoglobin on chitosan-room temperature ionic liquid-TiO(2)-graphene nanocomposite film modified electrode

TiO₂-graphene nanocomposite was prepared by hydrolysis of titanium isopropoxide in colloidal suspension of graphene oxide and in situ hydrothermal treatment. The direct electrochemistry and electrocatalysis of hemoglobin in room temperature ionic liquid 1-Butyl-3-methylimidazolium hexafluorophosphate, chitosan and TiO₂-graphene nanocomposite modified glassy carbon electrode were investigated. The biosensor was examined by using UV-vis spectroscopy, scanning electron microscopy and electrochemical methods. The results indicated that hemoglobin remained its bioactivity on the modified electrode, showing a couple of well-defined and quasi-reversible redox peaks, corresponding to hemoglobin Fe^III/Fe^II couple. The kinetic parameters for the electrode reaction, such as the formal potential (E^o'), the electron transfer rate constant (k_s), the apparent coverage (Γ), and Michaelis–Menten constant (K_m) were evaluated. The biosensor showed good electrochemical responses to the reduction of H₂O₂ in the ranges of 1–1170 μM. The detection limit was 0.3 μM (S/N = 3). The properties of this composite film, together with the bioelectrochemical catalytic activity, could make them useful in the development of bioelectronic devices, and investigation of electrochemistry of other heme proteins at functional interface.     

Direct electrochemistry and electrocatalysis of horseradish peroxidase immobilized on L-glutathione self-assembled monolayers

A novel hydrogen peroxide biosensor has been fabricated based on covalently linked horseradish peroxidase （HRP） onto L- glutathione self-assembled monolayers （SAMs）. The SAMs-based electrode was characterized by electrochemical methods, and direct electrochemistry of HRP can be achieved with formal potential of-0.242 V （vs. saturated Ag/AgCl） in pH 7 phosphate buffer solution （PBS）, the redox peak current is linear to scan rate and rate constant can be calculated to be 0.042 s^-1. The HRP-SAMs- based biosensors show its better electrocatalysis to hydrogen peroxide in the concentration range of 1 × 10^-6 mol/L to 1.2 × 10^-3 mol/L with a detection limit of 4 × 10^-7 mol/L. The apparent Michealis-Menten constant is 3.12 mmol/L. The biosensor can effectively eliminate the interferences of dopamine, ascorbic acid, uric acid, catechol and p-acetaminophen.     

Direct electrochemistry and electrocatalysis of hemoglobin entrapped in composite matrix based on chitosan and CaCO3 nanoparticles

The direct electron transfer between hemoglobin (Hb) and the underlying glassy carbon electrode (GCE) can be readily achieved via a high biocompatible composite system based on biopolymer chitosan (CHT) and inorganic CaCO₃ nanoparticles (nano-CaCO₃). Cyclic voltammetry of Hb-CHT/nano-CaCO₃/GCE showed a pair of stable and quasi-reversible peaks for HbFe(III)/Fe(II) redox couple in pH 7.0 buffer. The electrochemical reaction of Hb immobilized in CHT/nano-CaCO₃ composite matrix exhibited a surface-controlled process accompanied by electron and proton transfer. The electron transfer rate constant was estimated to be 1.8 s⁻¹. This modified electrode showed a high thermal stability up to 60 °C. The apparent Michaelis–Menten constant was calculated to be 7.5 × 10⁻⁴ M, indicating a high catalytic activity of the immobilized Hb toward H₂O₂. The interaction between Hb and this nano-hybrid material was also investigated using FT-IR and UV–vis spectroscopy, indicating that Hb retained its native structure in this hybrid matrix.