Effect of electrode surface properties on enhanced electron transfer activity in microbial fuel cells

The influence of electrode surface chemistry over biofilm growth was evaluated for photo‐bioelectrocatalytic fuel cell. A consortium of photosynthetic bacteria was grown onto different electrodes designed with polyethylenimine (PEI) and multiwall carbon nanotubes as hydrophilic and hydrophobic modifier, respectively. The designed electrodes were loaded with 0.08, 0.17, and 0.33 μg/cm² of PEI to change the hydrophilicity. However, 0.56, 0.72, and 0.83 mg/cm² of multiwall carbon nanotubes were used to alter the hydrophobicity of the electrodes. The surface chemistry of electrode and bio‐interaction was evaluated as a function of contact angle and biofilm formation. The results were compared with those obtained with a carbon paper electrode. The contact angle on the untreated electrode (carbon paper) was 118°, whereas for hydrophobic and hydrophilic electrodes, the maximum and minimum contact angles were 170° and 0°, respectively. Interestingly, the maximum biofilm growth (0.2275 g, wet basis) was observed on highly hydrophobic surface; however, the maximum electrochemical performance (246 mV) was shown by the most hydrophilic electrode surface. PEI‐based electrode with good biofilm formation showed comparatively higher electrogenic activity.     

Rechargeable glucose electrodes for long-term implantation

A unique enzyme electrode was designed using glucose oxidase immobilized on fine graphite powder. The graphite-enzyme is in a fluid state which enables recharging of the system when the enzyme activity decreases, therefore allowing the system to have a long lifetime in a diabetic patient. The sensor was tested using glucose concentration in the range of 20–300 mg d⁻¹. The electrode construction includes a hydrophobic membrane and a platinum electrode for detection of hydrogen peroxide. The signal output is large with minimal noise when tested in buffer. The sensor has been performing at ambient temperature for 4 months when stored overnight at 4° C.     

Urea sensor with single poly(propylene) membrane

Urease was immobilized on the plasma-aminated surface of a hyfrophobic poly(propylene) (PP) membrane. This membrane, with urease matrix on one side while maintaining its original hydrophobic property on the other, was used to construct the urea sensor. The new urea sensors had response sensitivities ranged from 19 mV/decade to 30 mV/decade depending on the conditions of the plasma reaction. The enzyme electrode using single membrane gave a shorter response time as compared to the corresponding conventional electrode employing two seperate PP membranes. The sensitivity of the enzyme electrode increased with increasing buffer pH and reached a maximal level (40 mV/decade) at pH 7.6. The response sensitivity of the electrode was not affected by the change of buffer strength. Deamination of the plasma-modified hydrophobic PP membrane did not occur in aqueous environment judging from the stability of the urea electrode up to 12 days of operation. (c) 1992 John Wiley & Sons. Inc.     

A flexible and wearable glucose sensor based on functional polymers with soft-MEMS techniques

A novel biosensor for glucose measurement using functional polymers was fabricated and tested. The biosensor utilizes the physical and chemical functions of hydrophobic polydimethyl siloxane (PDMS) and hydrophilic 2-methacryloyloxyethyl phosphorylcholine (MPC) copolymerized with dodecyl methacrylate (DMA). The glucose sensor was constructed by immobilizing glucose oxidase (GOD) onto a flexible hydrogen peroxide electrode (Pt working electrode and Ag/AgCl counter/reference electrode). The electrodes were fabricated using microelectromechanical systems (MEMS) techniques onto those functional polymers. The sensor showed novel functions of flexibility and it was stretchable so that the sensor could normally work when it was released after expanding to 120% longer than that of normal length. Also, basic characteristics of the sensor were evaluated. The output current of the hydrogen peroxide electrode was linearly related to the hydrogen peroxide concentration in a range of 0.20-2.50 mmol/l, with a correlation coefficient of 0.998. GOD was then immobilized onto the surface of the sensor using MPC polymer. In this case, the current output of the glucose sensor related to the glucose level over a range of 0.06-2.00 mmol/l, with a correlation coefficient of 0.997. The calibration range includes the reported concentration of tear glucose in normal human subject (0.14 mmol/l).     

Immobilization of glucose oxidase on thin-film gold electrodes produced by magnetron sputtering and their application in an electrochemical biosensor

An electrochemical biosensor is described consisting of a thin-layer gold film electrode prepared by cathodic sputtering using a poly(vinyl chloride) sheet as substrate, with voltammetric behaviour comparable to that of conventional polycrystalline gold electrodes, coated with the hydrolysed copolymer hydroxyethyl methacrylate-co-methyl methacrylate onto which glucose oxidase was immobilized. The mechanical properties of the plastic foil substrate permit easy construction of circular-shaped electrodes which were employed as working electrodes for batch injection analysis. The electrochemical biosensor fabrication is inexpensive and can be used as disposable enzyme sensor for the detection of hydrogen peroxide. The biosensor was tested for the determination of glucose in serum and a good correlation was obtained with the measurement using the electrochemical and the spectrophotometric methods.     

Extended shelf life of enzyme-based biosensors using a novel stabilization system

The storage stability of amperometric enzyme electrodes has been enhanced by a combination of a soluble, positively charged polymer, diethylaminoethyl (DEAE)-dextran, and a sugar alcohol, lactitol. Two different types of alcohol biosensor have been produced using the enzyme alcohol oxidase, isolated from the methylotrophic yeast Hansenula polymorpha. The first employs enzyme entrapment between two membranes with direct hydrogen peroxide amperometry at +0·65 V. The second was based on the mediated, coupled reaction with horseradish peroxidase and N-methyl phenazimiumtetracyanoquinonedimethane (NMP-TCNQ) on a graphite electrode. In both cases, addition of the stabilizers promoted a considerable increase in the storage stability of the enzyme component, as indicated by an increase in the shelf life of desiccated biosensors under conditions of thermal stress at 37°C. In addition, an L-glutamate biosensor constructed from NMP-TCNQ-modified graphite electrodes and L-glutamate oxidase also exhibited an increase in shelf life when stored, desiccated in the presence of stabilizers.     

Use of a copper-phthalocyanine membrane electrode for rapid preliminary detection of polycyclic mutagens

Preliminary screening of polycyclic mutagens is achieved within 20 min by using a biomimetic electrode composed of an oxygen electrode and a copper-phthalocyanine membrane. When benzo[alpha]pyrene (0.05 mM) was added to the buffer solution in the presence of 0.98 M hydrogen peroxide, the current of the phthalocyanine electrode decreased. A linear relationship was obtained between the current decrease and the benzo[alpha]pyrene concentration over the range 0.19-0.60 mM. The minimum measurable concentration for benzo[alpha]pyrene was 0.01 mM. Such responses were not obtained for other organic compounds such as alcohol, ether, n-hexane and cyclohexane. The copper-phthalocyanine membrane electrode has selectively detected polycyclic mutagens such as amino acid pyrolysis products. The current decrease was 1.18-1.46 microA when 0.05 mM amino acid pyrolysis products were employed.     

A new method for enzyme membrane preparation based on polyurethane technology: Electrode modification for sensor development

The new method developed for enzyme membrane preparation is based on cross-linking poly (vinyl alcohol) (PVAL) with triisocyanate (TIC) in the presence of enzyme. Dimethylsulfoxide (DMSO) was the only solvent found to dissolve PVAL, TIC and enzyme at room temperature, without completely denaturing the latter. The rate of gelation to form the desired network membrane can be controlled by the amount of solvent used. All the enzymes tested (alkaline phosphatase and alcohol, cholesterol and glucose oxidases) dissolved in DMSO and retained sufficient activity for use in electrochemical sensors. Membranes were formed on both graphite and platinized graphite electrodes. The resulting prototype sensors were examined with regard to feasibility of preparation, adhesion of the gels to the electrode surfaces, swelling properties of the gels in DMSO and aqueous buffers, and their electrochemical properties.     

The determination of titratable acid and ammonium ions in picomole amounts

A methodological system mainly designed for the use on intratubular urine samples is described, which permits the determination of titratable acid and ammonium ions in samples of a few nanoliters. The pH measurements were performed by means of antimony micro electrodes, the construction of which are described in detail. The hydroxyl ions were added to the samples from a second antimony electrode system, by an electric current. The amount of hydroxyl ions liberated was equivalent to the amount of current used.The ammonium determinations were based upon the fact that hydrogen ions were liberated from the ammonium ions by formaldehyde. The hydrogen ions were titrated in the same manner as the titratable acid.The use of two electrode systems simultaneously inserted in the droplet permitted recordings of the titration curves. The magnitude of methodological errors of these ultramicro methods are the same as those of corresponding methods using milliliter volumes.     

Comparison of colloidal gold electrode fabrication methods: the preparation of a horseradish peroxidase enzyme electrode.

In order to prepare biosensing electrodes which respond to hydrogen peroxide, horseradish peroxidase has been adsorbed to colloidal gold sols and electrodes prepared by deposition of these enzyme-gold sols onto glassy carbon using three methods: evaporation, electrodeposition and electrolyte deposition. In the latter method the enzyme-gold sol is applied to the surface of a glassy carbon disk electrode followed by an equal volume of 2 mM CaCl2. The electrolyte causes the sol to precipitate on the electrode surface, producing an immobilized enzyme electrode. Satisfactory electrodes which gave an electrochemical response to hydrogen peroxide in the presence of the electron transfer mediator ferrocenecarboxylic acid were produced by all three methods. Evaporation of horseradish peroxidase-gold sols produced electrodes with the best reproducibility and the widest linear amperometric response range. These electrodes can also easily be stored in a dry state. Although not as good as evaporation, electrodeposition also produced satisfactory electrodes. Electro-deposition provides the added advantage that it lends itself to the preparation of multi-enzyme/multi-analyte electrodes by the adsorption of different enzymes to separate gold sols, followed by sequential electrodeposition onto discrete areas of a multichannel electrode.     

The biocatalytic effect of Halobacterium halobium on photoelectrochemical hydrogen production

Hydrogen gas can be produced electrochemically by leading a current through two electrodes immersed in a NaCl solution. Bacteriorhodopsin (BR) a protein found in the purple membrane of Halobacterium halobium, is known to pump protons across the membrane upon illumination. In this study, the effect of BR on photoelectrochemical hydrogen production was investigated. A batch type bio-photoelectrochemical reactor was designed and constructed. The photoelectrochemical hydrogen production experiments were performed with free H. halobium packed cells or immobilised H. halobium cells. The cells were either immobilised in polyacrylamide gel (PAG) or on cellulose acetate membrane (CAM). Experiments were also performed with purple membrane fragments of H. halobium immobilised on cellulose acetate membrane. It was found that the presence of bacteriorhodopsin (BR) in the reactor enhances the hydrogen production rate upon illumination. Immobilisation increased the amount of hydrogen produced per mole of BR. Compared to control experiments without BR, the power requirement of the photoelectrochemical reactor per amount of hydrogen produced decreased fourfold when purple membrane fragments immobilised on CAM were used. The presence of BR regulates the pH of the system, increases the hydrogen production rate and causes light-induced proton dissociation, which lowers the electrical power requirement for the electrochemical conversion.     

几种金属单质及其合金修饰电极电解合成L-半胱氨酸的研究

用几种氢过电位较高的金属Ｓｎ、Ｂｉ、ＮｉＺｎ以及Ｃｕ修饰铅电极,用于电解合成Ｌ－半胱氨酸的反应,首先筛选出性能较好的Ｓｎ、Ｎｉ、Ｚｎ电极,然后对该三种金属的合金进行研究,筛选出性能优越的（Ｎｉ－Ｓｎ）／Ｐｂ电极,电极活性大为提高,反应同期转化率明显提高。     

Amperometric biosensor for the detection of hydrogen peroxide using catalase modified electrodes in polyacrylamide

A simple biosensor for the detection of hydrogen peroxide in organic solvents has been developed and coupled to a flow injection analysis (FIA) system. Catalase was entrapped in polyacrylamide gel and placed on the surface of platinum (working electrode) fixed in a Teflon holder with Ag-wire (auxiliary electrode), followed by addition of filter paper soaked in KCl. The entrapped catalase gel was held on the electrode using membranes. The effects of cellulose and polytetrafluroethylene (PTFE) membranes on the electrode response towards hydrogen peroxide have been studied. The modified electrode has been used to study the detection of hydrogen peroxide in solvents like water, dimethyl sulfoxide (DMSO), and 1,4-dioxane using amperometric techniques like cyclic voltammetry (CV) and FIA. The CV of modified catalase electrode showed a broad oxidation peak at -150 mV and a clear reduction peak at -212 mV in the presence of hydrogen peroxide. Comparison of CV with hydrogen peroxide in various solvents has been carried out. The electrode showed an irreversible kinetics with DMSO as the solvent. A flow cell has been designed in order to carry on FIA studies to obtain calibration plots for hydrogen peroxide with the modified electrode. The calibration plots in several solvents such as water, dimethyl sulfoxide, 1,4-dioxane have been obtained. The throughput of the enzyme electrode was 10 injections per hour. Due to the presence of membrane the response time of the electrode is concentration dependent.     

Immobilization of d-glucose oxidase onto a hydrogen peroxide permselective membrane and application for an enzyme electrode

A hydrogen peroxide permselective membrane with asymmetric structure was prepared and d-glucose oxidase (EC 1.1.3.4) was immobilized onto the porous layer. The activity of the immobilized d-glucose oxidase membrane was 0.34 units cm^?2 and the activity yield was 6.8% of that of the native enzyme. Optimum pH, optimum temperature, pH stability and temperature stability were found to be pH 5.0, 30–40°C, pH 4.0–7.0 and below 55°C, respectively. The apparent Michaelis constant of the immobilized d-glucose oxidase membrane was 1.6 × 10^?3 mol l^?1 and that of free enzyme was 4.8 × 10^?2 mol l^?1. An enzyme electrode was constructed by combination of a hydrogen peroxide electrode with the immobilized d-glucose oxidase membrane. The enzyme electrode responded linearly to d-glucose over the concentration 0–1000 mg dl^?1 within 10 s. When the enzyme electrode was applied to the determination of d-glucose in human serum, within day precision (CV) was 1.29% for d-glucose concentration with a mean value of 106.8 mg dl^?1. The correlation coefficient between the enzyme electrode method and the conventional colorimetric method using a free enzyme was 0.984. The immobilized d-glucose oxidase membrane was sufficiently stable to perform 1000 assays (2 to 4 weeks operation) for the determination of d-glucose in human whole blood. The dried membrane retained 77% of its initial activity after storage at 4°C for 16 months.     

Electrochemical modulation of antigen-antibody binding

The label-free amperometric detection of a rabbit IgG antigen by an anti-rabbit IgG antibody is achieved by observing the electrochemistry at a glassy carbon electrode modified with antibody entrapped in an electrodeposited polypyrrole membrane. In a flow injection apparatus the electrode is pulsed between -0.2 and +0.4 V versus Ag/AgCl. The pulsing of the electrode switches the polypyrrole membrane between the oxidised and reduced states. When antigen is injected into the flow stream a change in current is observed at the electrode despite the antigen or antibody being redox inactive at the potentials employed. It is proposed that this current is due to a change in the flux of ions into and out of the polypyrrole matrix during a pulse when the poly-anionic antigen is present. The immunoreaction was reversible because the 200 ms pulse at each potential was too short to allow secondary bonding forces (hydrogen bonding and hydrophobic forces) which are responsible for the strength of the antibody-antigen complex to be established. The consequence of the reversibility of the antigen-antibody binding is a low apparent affinity constant but an easily regenerated recognition interface.     

Electricity production by Geobacter sulfurreducens attached to electrodes

Previous studies have suggested that members of the Geobacteraceae can use electrodes as electron acceptors for anaerobic respiration. In order to better understand this electron transfer process for energy production, Geobacter sulfurreducens was inoculated into chambers in which a graphite electrode served as the sole electron acceptor and acetate or hydrogen was the electron donor. The electron-accepting electrodes were maintained at oxidizing potentials by connecting them to similar electrodes in oxygenated medium (fuel cells) or to potentiostats that poised electrodes at +0.2 V versus an Ag/AgCl reference electrode (poised potential). When a small inoculum of G. sulfurreducens was introduced into electrode-containing chambers, electrical current production was dependent upon oxidation of acetate to carbon dioxide and increased exponentially, indicating for the first time that electrode reduction supported the growth of this organism. When the medium was replaced with an anaerobic buffer lacking nutrients required for growth, acetate-dependent electrical current production was unaffected and cells attached to these electrodes continued to generate electrical current for weeks. This represents the first report of microbial electricity production solely by cells attached to an electrode. Electrode-attached cells completely oxidized acetate to levels below detection (<10 micro M), and hydrogen was metabolized to a threshold of 3 Pa. The rates of electron transfer to electrodes (0.21 to 1.2 micro mol of electrons/mg of protein/min) were similar to those observed for respiration with Fe(III) citrate as the electron acceptor (E(o)' =+0.37 V). The production of current in microbial fuel cell (65 mA/m(2) of electrode surface) or poised-potential (163 to 1,143 mA/m(2)) mode was greater than what has been reported for other microbial systems, even those that employed higher cell densities and electron-shuttling compounds. Since acetate was completely oxidized, the efficiency of conversion of organic electron donor to electricity was significantly higher than in previously described microbial fuel cells. These results suggest that the effectiveness of microbial fuel cells can be increased with organisms such as G. sulfurreducens that can attach to electrodes and remain viable for long periods of time while completely oxidizing organic substrates with quantitative transfer of electrons to an electrode.     

Graphite electrode as a sole electron donor for reductive dechlorination of tetrachlorethene by Geobacter lovleyi

The possibility that graphite electrodes can serve as the direct electron donor for microbially catalyzed reductive dechlorination was investigated with Geobacter lovleyi. In an initial evaluation of whether G. lovleyi could interact electronically with graphite electrodes, cells were provided with acetate as the electron donor and an electrode as the sole electron acceptor. Current was produced at levels that were ca. 10-fold lower than those previously reported for Geobacter sulfurreducens under similar conditions, and G. lovleyi anode biofilms were correspondingly thinner. When an electrode poised at -300 mV (versus a standard hydrogen electrode) was provided as the electron donor, G. lovleyi effectively reduced fumarate to succinate. The stoichiometry of electrons consumed to succinate produced was 2:1, the ratio expected if the electrode served as the sole electron donor for fumarate reduction. G. lovleyi effectively reduced tetrachloroethene (PCE) to cis-dichloroethene with a poised electrode as the sole electron donor at rates comparable to those obtained when acetate serves as the electron donor. Cells were less abundant on the electrodes when the electrodes served as an electron donor than when they served as an electron acceptor. PCE was not reduced in controls without cells or when the current supply to cells was interrupted. These results demonstrate that G. lovleyi can use a poised electrode as a direct electron donor for reductive dechlorination of PCE. The ability to colocalize dechlorinating microorganisms with electrodes has several potential advantages for bioremediation of subsurface chlorinated contaminants, especially in source zones where electron donor delivery is challenging and often limits dechlorination.     

Application of L-(+)-Lactate electrode for clinical analysis and monitoring of tissue culture medium

L-(+)-Lactate oxidase (EC 1.1.3.2) was immobilized onto the porous side of a cellulose acetate membrane with asymmetric structure which has selective permeability to hydrogen peroxide. The lactate electrode was constructed by combination of a hydrogen peroxide electrode with the immobilized enzyme membrane. Properties of the enzyme membrane and characteristics of the lactate electrode were clarified for the determination of L-(+)-lactic acid. The lactate electrode responded linearly to L-(+)-lactic acid over the final concentration 0-0.25 mmol/L within 30 s. When the enzyme electrode was applied to the determination of L-(+)-lactic acid in control serum, within-day precision (CV), analytical recovery, and correlation coefficient between the electrode method and the colorimetric method were 1.4% with a mean value of 4.54 mmol/L, 98.0%, and 0.986, respectively. The lactate electrode was sufficiently stable to perform 1040 assays over 13 days operation for the determination of L-(+)-lactic acid. The dried immobilized enzyme membrane retained 84% of its initial activity after storage at 4 degrees C for 12 months. Moreover, the enzyme electrode was applied to the monitoring of culture medium for human melanoma cells. L-(+)-Lactate production and D-glucose consumption were closely related to cell numbers.     

Development of a biosensor for assaying postmortem nucleotide degradation in fish tissues

An enzyme sensor system has been developed to assess the freshness level in fish tissue. The system was designed to measure the K value, the concentration ratio of [Hx + HxR] and [Hx + HxR + IMP], where Hx, HxR, and IMP are hypoxanthine, inosine and inosine-5'-monophosphate, respectively. The [Hx + HxR] concentration in tissue extract was measured by nucleoside phosphorylase and xanthine oxidase immobilized on a preactivated nylon membrane and attached to the tip of a polarographic electrode. The electrode amperometrically detected the products of degradation, hydrogen peroxide and uric acid. For determination of [IMP + HxR + Hx], IMP was first converted to HxR by nucleotidase immobilized on the wall of a polystyrene tube. The enzyme electrode consisting of nucleoside phosphorylase and xanthine oxidase provided excellent reproducible results for at least 40 repeated assays and immobilized nucleotidase was good for at least 40 assays as well. The K value for each sample could be determined in ca. 10 min. When applied to K value measurements in several fish meats, the results obtained agreed well with those obtained by the conventional enzymatic method.     

Electrode System for the Determination of Microbial Populations

Determinations of microbial populations were carried out by using a new electrode system composed of two electrodes. Each electrode was constructed from a platinum anode and a silver peroxide cathode. The anode of the reference electrode was covered with cellulose dialysis membrane. The response time of the electrode system was 15 min in culture broth, and current differences between the two electrodes were proportional to populations of microbial cells in cultures of Saccharomyces cerevisiae and Lactobacillus fermentum. Current differences were reproducible; the average relative error was 5%. Furthermore, cell populations of S. cerevisiae in a fermentor could be continuously estimated by using this electrochemical method.