Prussian-Blue-based amperometric biosensors in flow-injection analysis

Optimisation of the electrodeposition of Prussian Blue onto mirrored glassy carbon electrodes yielded a modified electrode practically insensitive to oxygen reduction. At the same time the electrode activity towards hydrogen peroxide reduction was extremely high. This allowed the detection of hydrogen peroxide by electroreduction over a wide potential range. Flow-injection investigations of this electrode inserted into a flowthrough electrochemical cell of the confined wall-jet type showed that the response for hydrogen peroxide is limited by diffusion. Glucose and alcohol biosensors were made by immobilisation of glucose oxidase and alcohol oxidase respectively, within a Nafion layer, onto the top of the Prussian-Blue-modified electrodes. By increasing the density of Nafion and decreasing the measuring potential the glucose biosensor was made completely insensitive to both ascorbate and acetominophes.     

Which Parameters Affect the Response of the Channel Biosensor?

The important parameters in defining the response of the portable channel biosensor described previously are explored by connecting the portable flow cell to a gravity feed flow system and using a highly defined enzyme immobilization protocol which ensures the enzyme reaction is a surface reaction. The enzyme glucose oxidase (GOD) was immobilized by covalent attachment to a self‐assembled monolayer modified gold surface. As a glucose solution flowed down the rectangular duct defined by the flow cell, it passed over the enzyme layer where the enzyme reaction produced hydrogen peroxide. The hydrogen peroxide was swept further downstream to the detector electrode. The response of such an enzyme electrode was shown to be limited by mass transport of the cosubstrate oxygen to the enzyme layer. Increasing the amount of oxygen in the sample meant the response of the biosensor became limited by the enzyme kinetics. The influence of parameters such as flow rate, height of the channel, enzyme layer length and the gap between the enzyme layer and the detector electrode were explored.     

Fe修饰多壁碳纳米管电极高效产H2O2

为提高电芬顿(Electro-Fenton)体系H2O2的产率, 制备了多壁碳纳米管(MWNT)电极, 并与石墨/气体扩散(GDC)电极进行了比较. 结果表明, MWNT电极H2O2产率高于GDC电极. 采用电沉积方法, 制备了Fe修饰MWNT(Fe-MWNT)电极, 发现Fe对MWNT电极的修饰不仅可以提高体系的H2O2产率, 而且电流效率可以提高8%左右, 与GDC电极的电流效率接近. Fe-MWNT电极有望成为一种新型的阴极材料应用于Electro-Fenton体系中.     

A dual-electrode approach for highly selective detection of glucose based on diffusion layer theory: experiments and simulation

A dual-electrode configuration for the highly selective detection of glucose in the diffusion layer of the substrate electrode is presented. In this approach, a glassy carbon electrode (GCE, substrate) modified with a conductive layer of glucose oxidase/Nafion/graphite (GNG) was used to create an interference-free region in its diffusion layer by electrochemical depletion of interfering electroactive species. A Pt microelectrode (tip, 5 microm in radius) was located in the diffusion layer of the GNG-modified GCE (GNG-G) with the help of scanning electrochemical microscopy. Consequently, the tip of the electrode could sense glucose selectively by detecting the amount of hydrogen peroxide (H2O2) formed from the oxidization of glucose on the glucose oxidase layer. The influences of parameters, including tip-substrate distance, substrate potential, and electrolyzing time, on the interference-removing efficiency of this dual-electrode approach have been investigated systematically. When the electrolyzing time was 30 s, the tip-substrate distance was 1.8 a (9.0 microm) (where a is the radius of the tip electrode), the potentials of the tip and substrate electrodes were 0.7 V and 0.4 V, respectively, and a mixture of ascorbic acid (0.3 mM), uric acid (0.3 mM), and 4-acetaminophen (0.3 mM) had no influence on the glucose detection. In addition, the current-time responses of the tip electrode at different tip-substrate distances in a solution containing interfering species were numerically simulated. The results from the simulation are in good agreement with the experimental data. This research provides a concept of detection in the diffusion layer of a substrate electrode, as an interference-free region, for developing novel microelectrochemical devices.     

新型气体扩散电极体系高效产H2O2的研究

以自制新型石墨/聚四氟乙烯(PTFE)气体扩散电极在无隔膜体系中进行双氧水发生工艺的优化研究, 主要探讨了不同石墨和PTFE质量比、阴极电位、pH值和氧气流速对H2O2产率的影响. 结果表明, 以石墨和PTFE质量比为2:1的气体扩散电极为阴极, 在pH=3, Na2SO4浓度为0.1 mol•L−1, 氧气流速为0.4 L•min−1, 阴极电位为−0.55 V (vs SCE)时, 2 h后H2O2可以达到60 mg•L−1. 该新型体系有较高的H2O2产率和电流效率(可达60%以上), 且pH值适用范围较广, 可望应用于水中污染物的处理.     

Electric detection of DNA with PDMS microgap electrodes and silver nanoparticles

In this work a novel microdevice sensor has been developed by plating gold on the PDMS surface to generate a sandwich-type gap electrode for DNA detection. The microdevice utilizes a gold band electrode-PDMS-gold band electrode configuration and the minimum detectable volume could be as low as 5 μL. The 20 μm PDMS-based gap was chemically modified with DNA capture probes and DNA sandwich hybrids were formed with the addition of DNA target and silver nanoparticle probes. To increase detection sensitivity, parallel detection zones have been developed in which the relevant resistances decrease substantially upon hybridyzation. By measuring the change in electrical conductivity, the DNA target in the concentration range of 1000-0.1 nM can be assayed and the limit of lowest detectable concentration was achieved at 0.01 nM.     

Bio-sensing using recessed gold-filled capillary amperometric electrodes

A novel recessed electrode is reported for amperometric detection of hydrogen peroxide and via glucose oxidase for the detection of glucose. The electrode utilised electrodeposited platinum over a gold wire surface, which proved to be an effective peroxide-detecting surface. Compared with a traditional exposed electrode surface, the recessed tip facilitated an extended linear range for glucose from 4 to over 14 mM. Bio-fouling, as assessed by exposure to bovine serum albumin, was also significantly reduced. Though response time at the recess was increased, it was within an acceptable range for physiological monitoring. Moreover, the recess enabled precise measurement of the hydrogen peroxide diffusion coefficient; this was based on a bipartite expression for the transient amperometric current at the recessed structure following a step change in ambient hydrogen peroxide concentration. An important aspect of the diffusion measurement was the curve fitting routine used to map on to the theoretical response curve.     

Development of a diamine biosensor

An amperometric diamine sensor is developed for clinical applications in diagnosis of bacterial vaginosis (BV). The sensor is based on crosslinked putrescine oxidase (PUO) which catalyzes the conversion of diamines (mainly putrescine and cadaverine) to products including hydrogen peroxide. The hydrogen peroxide is detected anodically at platinum electrode polarized at 0.5 V versus Ag/AgCl. Platinum-plated gold electrodes used as a substrate for the sensor construction, are batch-fabricated on a flexible polyimide foil (Kapton(R), DuPont). A three-electrode cell configuration is used in all amperometric measurements. The sensor construction is based on three layers: an inner layer to reject the interference effect of oxidizable molecules, an outer diffusion controlling layer, and in addition, an enzyme middle layer. The enzyme layer was immobilized by crosslinking PUO with bovine serum albumin (BSA) using glutaraldehyde (GA). An optimization study of the enzyme solution composition was carried out. With the optimized enzyme layer, the biosensor showed a very high sensitivity and fast response time of ca. 20 s. The sensor has a linear dynamic range from (0.5-300 muM) for putrescine that covers the expected biological levels of the analyte. Details on sensor fabrication and characterization are given in the present work.     

Biosensor for determination of carboxylic acids in wines based on the inhibition of sarcosine oxidase

The flow-through amperometric biosensor is presented for determination of carboxylic acids. It is based on two sensor layers that are deposited on a platinum electrode. The inner layer serves to eliminate interferences by limiting diffusion of electrochemically active substances such as ascorbic acid. This layer is electro-polymerized using an equimolar mixture of o-phenylenediamine and resorcinol. The outer layer is prepared by cross-linking the enzyme sarcosine oxidase and bovine serum albumin using glutaraldehyde. The formation of enzymatically produced hydrogen peroxide is monitored at 600 mV vs. an Ag/AgCl reference electrode. The addition of carboxylic acids causes competitive inhibition of the enzyme and a decrease in signal. The assay was optimized for determination of carboxylic acids in wine samples. Following 10-fold dilution, most samples contain 1–10 mM individual carboxylic acids and thus a 5 mM concentration of sarcosine was chosen as being optimal for competition. In case of real samples, the biosensor measures the sum of all carboxylic acids, which serves as a parameter describing the quality of wines. Results from testing several wine samples are reported.     

Amperometric Determination of Hydrogen Peroxide and its Mathematical Simulation for Horseradish Peroxidase Immobilized on a Sonogel Carbon Electrode

A mathematical model of a horseradish peroxidase biosensor was applied to simulate the amperometric response for the detection of hydrogen peroxide. The development of the mathematical model was based on the Michaelis–Menten equation and Fick’s Second Law. The theoretical study is based on the determination of physico-chemical and geometric parameters of a horseradish peroxidase biosensor as well as the kinetic parameters of reaction mechanism such as diffusion coefficients of hydrogen peroxide, the thickness of enzymatic layer, and the Michaelis–Menten kinetic constant. The theoretical analysis provides an accurate estimate of parameters affecting the biosensor performance such as the diffusion coefficient of hydrogen peroxide in the biomembrane that was estimated to be 56?×?10^?12 m²/s. The thickness of diffusion layer was estimated to be 80–100?µm and the biomembrane 7.5?µm. The experimental and numerical values of kinetic parameters were 0.92 and 0.98?µM for the Michaelis–Menten constants and 0.010 and 0.012?µM/s for the catalytic activity rates. The model was validated for hydrogen peroxide detection and exhibited a good agreement with the experimental measurements.     

Mass transfer to horizontal gas-generating electrodes

Mass transfer to a horizontal electrode during electrolytic evolution of oxygen and hydrogen at current densities of 100 to 10 000 A/m² is studied. The mass transfer intensity is evaluated from the diffusion layer thickness, which varies from 60 to 5 μm at such current densities. Calculations show that the decrease in the diffusion layer thickness is due to bubbles with a stationary interphase surface crossing the diffusion layer. During the hydrogen evolution, the diffusion layer thickness is nearly the same for vertical and horizontal electrodes. During the oxygen evolution, the diffusion layer is much thinner for a horizontal electrode. Additional decrease of the diffusion layer thickness during the evolution of oxygen is associated with the lesser solution density in the near-electrode layer and with its transport away by means of natural convection.     

Electrocatalytic Sensor for Hydrogen Peroxide Based on Immobilized Benzoquinone

An amperometric chemosensor for the detection of hydrogen peroxide is reported. The sensor is based on 1,4-benzoquinone immobilized on the gold electrode using self-assembled monolayer of short chain symmetrical dithiol as an anchor layer. Sensor analysis was performed by cyclic voltammetry at the potential range from −0.6 V till +0.9 V as well as in the anodic or cathodic potential ranges only. The results indicate oxidative electrochemical decomposition of hydrogen peroxide at the potential of ∼+0.4 V leading to the formation of oxygen while at cathodic potentials a reduction of the formed oxygen as well as of the hydrogen peroxide occur. A decrease in the oxidation potential of hydrogen peroxide on the gold electrode coated by self-assembled monolayer with 1,4-benzoquinone in comparison with that measured on the electrodes coated by the same self-assembled monolayer without 1,4-benzoquinone, indicates electrocatalytic effect of this moiety on oxidative decomposition of hydrogen peroxide. Analytical evaluation of the sensor performance was done in the voltammetric as well as in the chronoamperometric mode. The sensor exhibited linear response over the concentration range till 2.5 mM with a limit of detection ∼4 μM.     

Synthesis of Hydrogen Peroxide Using Dielectric Barrier Discharge Associated with Fibrous Materials

A new synthesis pathway toward hydrogen peroxide has been investigated using non-thermal plasma. This work is aimed at studying the activation of oxygen/hydrogen mixtures by a cylindrical dielectric barrier discharge. An experimental device has been especially developed for this application, it mainly differs from other cylindrical discharges in that the liquid ground electrode, and subsequently the reactor, can be regulated in temperature. The formation of hydrogen peroxide is reported (1) in a gas phase discharge and (2) in surface discharge. The gas phase discharge, characterized by an empty discharge gap, lead to a low activation of O₂ into O₂/H₂ mixtures and poor selectivity toward H₂O₂. The modification of the discharge into a surface discharge, by introducing in the gap fibrous materials, considerably improves the efficiency of the process. The influence of the temperature on H₂O₂ formation is discussed and correlated to the formation of a water layer on fibre surface. This layer appears to be a crucial point into H₂O₂ plasma synthesis. The presence of TiO₂ on the fibre surface is reported as improving the stabilisation of hydrogen peroxide. The formation of a complex between H₂O₂ and TiO₂ is suggested and discussed. The formation of H₂O₂ in the gas phase or in the aqueous condensed phase is finally discussed. The investigation of the influence of the reactant gas composition and the presence or not of water, lead to the conclusion that (1) both H₂ and O₂ are required to achieve the synthesis reaction; (2) H₂O₂ is formed in the gas phase and then solubilised and/or stabilised in the water layer. A global reaction pathway is finally proposed to summarize the synthesis reaction.     

The role of anions in oxygen reduction in neutral and basic media on gold single-crystal electrodes

Oxygen reduction on well-defined single-crystal electrodes in different basic and neutral media has been studied using the impinging jet system. The results obtained with this system in 0.1 M NaOH are comparable to those reported in the literature for rotating disk electrodes, indicating that the impinging jet system behaves as a system in which the thickness of the diffusion layer is stationary. The activity of the Au(100) electrode is considerably higher than the rest of the basal planes in all media and yields water when E>–0.2 V and hydrogen peroxide for E<–0.2 V on the SHE scale. For Au(111) and Au(110) the activity of the electrode is much smaller and the final product is always hydrogen peroxide. The transition between both mechanisms for the Au(100) is, essentially, independent of the solution pH. In acid media the final product is always hydrogen peroxide for all the electrodes studied. The differences between the activities of the electrodes have been explained in the light of the different adsorption properties of the surfaces in relation to HO₂ ^–. In the case of the Au(100) electrode, the existence of a negative charge density on the metal inhibits further reduction of HO₂ ^–.Dedicated to Prof. Dr. Wolf Vielstich on the occasion of his 80th birthday in recognition of his numerous contributions to interfacial electrochemistry     

Fabrication of Translucid Gold-Nanocellulose Electrodes and their Potential Application as Hydrogen Peroxide Sensor

In this work, we describe the fabrication of a transparent gold electrode based on nanocellulose. The electrode was prepared via electron beam evaporation of gold onto nanocellulose films previously spread over a glass slip. Electrodes with different thickness of Au was fabricated, and the material's optical, morphological and electrical properties were assessed. Finally, as a proof of concept, a possible application of this electrode in hydrogen peroxide sensing was performed. The results show that a thin layer of gold on a nanocellulose translucid film allows obtaining a conductive transparent surface that could be used to design a transparent electrode.     

Hydrogen transport through Pd-Ni alloy electrodeposited on Pd substrate

Hydrogen transport through a Pd-Ni alloy electrodeposited on a Pd substrate (Pd-Ni/Pd bilayer symmetric electrode) has been investigated using cyclic voltammetry and a.c. impedance spectroscopy combined with the electrochemical hydrogen permeation method. The permeation build-up current transients and the measured impedance spectra were analyzed using the time-lag method for the bilayer electrode and a complex non-linear least squares data-fitting method based upon the derived Faradaic admittance for the hydrogen absorption into and diffusion through the bilayer electrode under the permeable boundary condition, respectively. The value of the hydrogen diffusivity in the Pd-Ni layer was lower than that in the Pd layer. Furthermore, the values of the charge transfer resistance and equilibrium absorption constant for the Pd-Ni/Pd bilayer electrode were higher than those for the Pd single layer electrode. From the experimental results, the role of the thin Ni(OH)₂ film formed on the Pd-Ni layer surface in the hydrogen transport through the Pd-Ni/Pd bilayer electrode is discussed in terms of its passivating effect and extremely large hydrogen solubility. Received: 22 January 1997 / Accepted: 15 April 1997     

A carbon electrode sputtered with palladium and gold for the amperometric detection of hydrogen peroxide

A modified carbon electrode for the amperometric determination of hydrogen peroxide is described. By deposition of a 15-nm thick layer of a 40:60 mixture of palladium and gold on the surface of the electrode the overvoltages for both the oxidation and the reduction can be decreased by at least 800 mV. When applied as an electrochemical sensor in a flow-injection system, linear calibration graphs were obtained between 10^?7 and 5 × 10^?3 M hydrogen peroxide. The modified electrodes were stable for months.     

Pulse Voltammetric Biosensing System for the Rapid Determination of Glucose With Micro-Enzyme Sensor

Abstract

An electrochemically prepared micro-enzyme electrode whose diameter is 50 jim is combined with an Pt auxiliary electrode and a reference electrode to assemble a three electrode device for the rapid determination of glucose. Since the device is very small, glucose sample whose volume is only 2 μ1 can be successfully determined. Pulse voltammetry is shown to be an effective approach for making the sensing device work without any attachments such as magnetic stirrer and pump. The transient sensor output, oxidizing current for the hydrogen peroxide generated by the immobilized glucose oxidase, shows a good linearity in the glucose concentration range from 1 mM to 20 mM.     

Pathways of diffusion mixed with subsequent reactions with examples of hydrogen extraction from hydride-forming electrode and oxygen reduction at gas diffusion electrode

This paper covers the role of the rate-determining step (RDS) in anodic hydrogen extraction from hydride-forming electrode. In general, hydrogen extraction from the electrode proceeds through the following steps: (1) hydrogen diffusion within the electrode, (2) hydrogen transfer from absorbed state to adsorbed state, (3) electrochemical oxidation of hydrogen to hydrogen ion involving charge transfer, and (4) hydrogen ion conduction through the electrolyte. In most theoretical and experimental investigations, it has been assumed that the RDS of anodic hydrogen extraction is hydrogen diffusion through the electrode. In real situation, however, the overall rate of hydrogen extraction is simultaneously determined by the rates of two or more reaction steps including hydrogen diffusion. The present work provides the overview of anodic hydrogen extraction in case that diffusion is coupled with interfacial charge transfer, interfacial hydrogen transfer, and hydrogen ion conduction through the electrolyte as well as the purely diffusion-controlled hydrogen extraction. In addition, the mixed controlled diffusion model was also exemplified with oxygen reduction at gas diffusion electrode of fuel cell system.     

Flow-through PCR on a 3D qiandu-shaped polydimethylsiloxane (PDMS) microdevice employing a single heater: toward microscale multiplex PCR

Consistent temperature control in an on-chip flow-through polymerase chain reaction (PCR) employing two or more heaters is one of the main obstacles for device miniaturization and integration when realizing micro total analysis systems (μTAS), and also leads to operational complexity. In this study, we propose a qiandu (right triangular prism)-shaped polydimethylsiloxane (PDMS) microdevice with serpentine microchannels fabricated on its slanted plane, and apply the device for an on-chip flow-through PCR employing a single heater. The inclined nature of the qiandu-shaped microdevice enables the formation of a surface temperature gradient along the slanted plane of the microdevice in a height-dependent manner by the use of a single heater, and enables liquid to traverse over wide ranges of temperatures, including the three temperature zones--denaturation, annealing, and extension temperatures--required in a typical PCR. The feasibility of the qiandu-shaped PDMS microdevice as a versatile platform for performing a flow-through PCR was examined by employing multiple templates and varying the inclination angle of the device. In addition, the potential of performing a multiplex PCR using a single qiandu-shaped PDMS microdevice was explored. A 409 bp long gene fragment effective as a marker for diagnosing lung cancer and a 230 bp long gene fragment from a plasmid vector were simultaneously amplified in less than 25 min on a single microdevice, paving the way for a microscale, multiplex PCR on a single device employing a single heater.