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1.
Glucose microelectrodes have been formed with glucose oxidase immobilized in poly[(vinylpyridine)Os(bipyridine)2Cl] derivative-based redox hydrogels on beveled carbon-fiber microdisk (7 microns diameter) electrodes. In the resulting microelectrode, the steady-state glucose electrooxidation current density is 0.3 mA cm-2 and the sensitivity is 20 mA cm-2 M-1. The current density and sensitivity are 10 times higher than in macroelectrodes made with the same hydrogel. Furthermore, the current is less affected by a change in the partial pressure of oxygen. The higher current density and lower oxygen sensitivity point to the efficient collection of electrons through their diffusion in the redox hydrogel to the electrode surface. These results contrast with those observed for enzyme electrodes based on diffusing mediators, where loss of the enzyme-reduced mediator by radial diffusion to the solution decreases the current densities of microelectrodes relative to similar macroelectrodes.  相似文献   

2.
A new enzymeless glucose sensor has been fabricated via electrospinning technology and subsequent calcination. The morphology and structure of the as-prepared nanofibers have been characterized by scanning electron microscopy (SEM), transmission electron microscopy (TEM), and X-ray diffraction (XRD). The electrocatalytic oxidation of glucose in alkaline medium at nickel oxide modified glassy carbon electrodes has been investigated. The modified electrodes offer excellent electrocatalytic activity toward the glucose oxidation at low positive potential (0.3 V). Glucose has been determined chronoamperometrically at the surface of NiO nanofibers modified electrode in 0.5 mM NaOH. Under the optimized condition, the calibration curve is linear in the concentration range of 2 x 10(-3) mM - 1 mM, and 1 mM - 9.5 mM. The detection limit (signal-to-noise 3) and response time are 3.394 x 10(-6) M and 2 s, respectively. The NiO electrospun nanofibers is easy to prepare and feasible in economy. The modified electrode is steady and can be used repeatedly, so it is reasonable to expect its broad use in non-enzymatic glucose sensor.  相似文献   

3.
The specially deposited Prussian Blue denoted as "artificial peroxidase" was used as a transducer for hydrogen peroxide. The electrocatalyst was stable, highly active, and selective to hydrogen peroxide reduction in the presence of oxygen, which allowed sensing of H2O2 around 0.0 V (Ag/AgCl). Glutamate oxidase was immobilized on the surface of the Prussian Blue-modified electrode in a Nafion layer using a nonaqueous enzymology approach. The calibration range for glutamate in flow injection system was 1 x 10(-7)-1 x 10(-4) M. The lowest concentration of glutamate detected (1 x 10(-7) M) and the highest sensitivity in the linear range of 0.21 A M-1 cm-2 were achieved. The influence of reductants was practically avoided using the low potential of an indicator electrode (0.0 V Ag/AgCl). The attractive performance characteristics of the glutamate biosensor illustrate the advantages of Prussian Blue-based "artificial peroxidase" as transducer for hydrogen peroxide detection.  相似文献   

4.
A novel microscale and surface-based method for the study of the interactions of DNA with other redox-active molecules using DNA-modified electrodes is described. The method is simple, convenient, reliable, reagent-saving, and applicable for DNA studies, especially those involving microsamples. Information such as binding site size (s, in base pairs), binding constant (K), ratio (K0x/KRed) of the binding constants for the oxidized and reduced forms of a bound species, binding free energy (delta Gb), and interaction mode, including changes in the mode of interaction, and "limiting" ratio K0x0/KRed0 at zero ionic strength can be obtained using only 3-15 micrograms of DNA samples. The method was developed using [Co(Phen)3]3+/2+ (Phen = 1,10-phenanthroline)/double-stranded DNA (dsDNA)-modified gold electrodes and [Co(bpy)3]3+/2+ (2,2'-bipyridyl)/dsDNA-modified gold electrodes as model systems. For the [Co(Phen)3]3+/2+/dsDNA-modified gold electrode system, a K2+ of (2.5 +/- 0.3) x 10(5) M-1 and an s of 5 bp were obtained in 5 mM pH 7.1 Tris-HCl buffer solution containing 50 mM NaCl. For [Co(bpy)3]3+/2+/dsDNA-modified gold electrodes, K3+ and s values of (1.3 +/- 0.3) x 10(5) M-1 and 3 bp, respectively, were obtained. While the s values are consistent with those reported in the literature obtained by solution methods, the K values are almost an order of magnitude larger. A transition in the nature of the interaction between dsDNA and [Co(Phen)3]3+/2+, from electrostatic to intercalative with increasing ionic strength, was found in our studies. Negative values of delta E0' for [Co(bpy)3]3+/2+ bound to dsDNA suggest that its interaction with dsDNA is predominantly electrostatic over the ionic strength range of 5-105 mM. The "limiting" ratio K3+0/K2+0 of 22 obtained for [Co(Phen)3]3+/2+ bound to dsDNA at zero ionic strength suggests that electrostatic interactions are predominant over intercalative ones under these limiting conditions. The ratio for [Co(bpy)3]3+/2+ of 16 also indicates that the 3+ form binds to dsDNA more strongly than the 2+ form at zero ionic strength. For [Co(Phen)3]3+/2+/single-stranded DNA (ssDNA)-modified gold electrodes, the nonuniformity of the surface structure of ssDNA-modified gold electrodes greatly complicates the analysis. A system consisting of a dsDNA-modified gold electrode and [Co(tppz)2]3+/2+ (tppz = tetra-2-pyridyl-1,4-pyrazine) was studied by this method, with a K2+ value of (5 +/- 1) x 10(5) M-1 and an 8 value of 7 bp being obtained.  相似文献   

5.
This work presents the design and optimization of amperometric biosensors for the determination of biogenic amines (e.g., histamine, putrescine, cadaverine, tyramine, cystamine, agmatine, spermidine), commonly present in food products, and their application for monitoring of freshness in fish samples. The biosensors were used as the working electrodes of a three-electrode electrochemical cell of wall-jet type, operated at -50 mV vs Ag/AgCl, in a flow injection system. Two different bienzyme electrode designs were considered, one based on the two enzymes [a newly isolated and purified amine oxidase (AO) and horseradish peroxidase (HRP)] simply adsorbed onto graphite electrodes, and one when they were cross-linked to an Os-based redox polymer. The redox hydrogel-based biosensors showed better biosensors characteristics, i.e., sensitivity of 0.194 A M-1 cm-2 for putrescine and 0.073 A M-1 cm-2 for histamine, and detection limits (calculated as three times the signal-to-noise ratio) of 0.17 microM for putrescine and 0.33 microM for histamine. The optimized redox hydrogel-based biosensors were evaluated in terms of stability and selectivity, and were used for the determination of total amine content in fish samples kept for 10 days in different conditions.  相似文献   

6.
Studies of the oxidation of beta-nicotinamide adenine dinucleotide (NADH) at glassy carbon (GCEs) electrode surfaces, modified with nonconventional conducting polymer nanotubules, are reported. In contrast to the situation with conventional carbon electrodes, chemical reversibility of the NADH oxidation reaction was achieved by means of poly(1,2-diaminobenzene) conducting nanotubule coatings. A Delta E(p) of 425 mV (vs Ag/AgCl; pH 7.0) was observed. The NADH amperometric response of the conducting nanotubule modified GCEs was shown to be extremely stable, with 98% of the initial response remaining after 48 h of stirring in the presence of 1 x 10(-4) M NADH solutions (compared to 14% at the poly(1,2-diaminobenzene) modified GCEs). The nonconventional conducting polymer nanotubule-coated electrodes, when tested in amperometric mode for NADH electrochemical oxidation at an applied potential of 450 mV, showed a sensitivity of 99 nA/mM, an operational stability for 2 days, a storage stability of 2 weeks at 4 degrees C, a linearity from 5 x 10(-5) to 1 x 10(-3) M, and good NADH chemical reversibility, all of which make them useful tools for dehydrogenase enzyme probe assembly.  相似文献   

7.
A series of glucose oxidase (GOx) hybrids (GOx-phe-nothiazine-labeled poly(ethylene oxide) (PT-PEO)) capable of direct electrical communication with electrodes is synthesized by covalently modifying PT-PEO to lysine residues on the enzyme surface. The length of the PEO chain and the number of PT groups are systematically altered. After the PT-PEO modification, all the hybrids maintain more than 50% of enzyme activity relative to that of native GOx, although loss of the activity becomes greater with increasing PEO chain length. The catalytic current, i(cat), is observed at a potential more positive than 0.55 V after the addition of glucose, due to the intramolecular electron transfer (El) from reduced forms of flavin adenine dinucletide (FADH2/FADH) to PT+ that are electrogenerated at the electrode. The i(cat) value increases with the number of PT groups, indicating that most of the modified PT groups act as mediators. The magnitude of the i(cat) increase depends on the PEO chain length and reveals a maximum for PT-PEO with the molecular weight of 3,000. In contrast, the i(cat) is almost constant for GOx-2-(10-phenothiazyl)propionic acid (PT-PA) hybrids with more than two PT groups synthesized by covalently modifying PT-PA to surface lysines, indicating that only a few key PT groups function as mediators. The maximum rate constant (130 s(-1)) for the ET from FADH2/FADH to PT+ is obtained for the GOx hybrid modified with five PT-PEO groups with the molecular weight of 3,000.  相似文献   

8.
Supramolecular organized multilayers composed of glucose oxidase (GOx) and osmium-derivatized poly(allylamine) redox polymer have been self-assembled electrostatically from Os-polyelectrolyte solutions of variable pH (5.5-8.8) leading to a decrease of the linear charge density in the PAH-Os with increasing pH. The layer-by-layer enzyme multilayers were studied by ellipsometry, quartz crystal microbalance, AFM, cyclic voltammetry, and electrocatalytic oxidation of beta-D-glucose. At higher adsorption solution pH, an increase in the film thickness, enzyme loading, and redox charge was observed. While the electrocatalytic response increases with the increase of the adsorption solution pH (decrease of the polyelectrolyte linear charge), the FADH2 oxidation bimolecular rate constant has a maximum in the pH range 7.0-7.5 where a change in the film growth mechanism is observed.  相似文献   

9.
A triethylammonium-sensitive electrode was constructed using sodium tetrakis[3,5-bis(2-methoxyhexafluoro-2-propyl)phenyl]borate as an ion-exchanger and benzyl 2-nitrophenyl ether as a solvent mediator in a poly(vinylchloride) membrane matrix and was used to determine the pH difference across a cell membrane. The method is based on monitoring of the pH gradient-induced uptake of triethylammonium in situ. The triethylammonium electrode exhibited a near-Nernstian response to triethylammonium in the concentration range of 5 x 10(-6)-1 x 10(-2) M with a slope of 58.5 mV per concentration decade in a buffer solution composed of 150 mM NaCl and 10 mM NaH2PO4/Na2HPO4 (pH 7.5). The limit of detection was 1 microM. In experiments using liposomes, the uptake of triethylammonium into liposomes was quantitatively induced according to the pH difference across the liposomal membrane. The transmembrane pH differences in Escherichia coli cells and the light-induced pH differences across the envelope vesicles of Halobacterium halobium were successfully determined by the present method.  相似文献   

10.
Cyclic voltammetric and electrogenerated chemiluminescent data were used to study the binding of tris(2,2'-bipyridine)-osmium(II), Os(bpy)3(2+), an electrostatic binder, to calf thymus DNA. The oxidized form of the osmium complex, Os(bpy)3(3+), has a stronger association to DNA than the reduced form, Os(bpy)3(2+), as indicated by the negative shift of E0' of the CV waves (K3+/K2+ = 3.35). The calculated binding constant, K2+, and binding site size, s, for the Os(byp)3(2+)-DNA system depended slightly on whether a mobile or a static equilibrium was assumed. In 10 mM NaCl, 10 mM Tris pH 7, K2+ = (7.3 +/- 0.4) x 10(3) M-1 and s = 3 base pairs (mobile) and K2+ = (5.0 +/- 0.2) x 10(3) M-1 and s = 3 base pairs (static). Electrogenerated chemiluminescence (ECL) was produced upon oxidation of Os(bpy)3(2+) at a Pt electrode in a solution containing 10 mM C2O4(2-) and 10 mM phosphate at pH 5. Addition of DNA caused a decrease in the emission intensity (I); a plot of I vs relative DNA concentration yielded K2+ = (6.5 +/- 0.5) x 10(3) M-1 and s = 3 base pairs. The osmium complex produced ECL when bound to the DNA molecule with an efficiency of 30% that of the unbound chelate.  相似文献   

11.
Oxidoreductases, such as glucose oxidase, can be electrically "wired" to electrodes by electrostatic complexing or by covalent binding of redox polymers so that the electrons flow from the enzyme, through the polymer, to the electrode. We describe two materials for amperometric biosensors based on a cross-linkable poly(vinylpyridine) complex of [Os-(bpy)2Cl]+2+ that communicates electrically with flavin adenine dinucleotide redox centers of enzymes such as glucose oxidase. The uncomplexed pyridines of the poly(vinylpyridine) are quaternized with two types of groups, one promoting hydrophilicity (2-bromoethanol or 3-bromopropionic acid), the other containing an active ester (N-hydroxysuccinimide) that forms amide bonds with both lysines on the enzyme surface and with an added polyamine cross-linking agent (triethylenetetraamine, trien). In the presence of glucose oxidase and trien this polymer forms rugged, cross-linked, electroactive films on the surface of electrodes, thereby eliminating the requirement for a membrane for containing the enzyme and redox couple. The glucose response time of the resulting electrodes is less than 10 s. The glucose response under N2 shows an apparent Michaelis constant, Km' = 7.3 mM, and limiting current densities, jmax, between 100 and 800 microA/cm2. Currents are decreased by 30-50% in air-saturated solutions because of competition between O2 and the Os(III) complex for electrons from the reduced enzyme. Rotating ring desk experiments in air-saturated solutions containing 10 mM glucose show that about 20% of the active enzyme is electrooxidized via the Os(III) complex, while the rest is oxidized by O2. These results suggest that only part of the active enzyme is in electrical contact with the electrode.  相似文献   

12.
A new extracellular flavin adenine dinucleotide (FAD)-dependent glucose dehydrogenase from Glomerella cingulata (GcGDH) was electrochemically studied as a recognition element in glucose biosensors. The redox enzyme was recombinantly produced in Pichia pastoris and homogeneously purified, and its glucose-oxidizing properties on spectrographic graphite electrodes were investigated. Six different Os polymers, the redox potentials of which ranged in a broad potential window between +15 and +489 mV versus the normal hydrogen electrode (NHE), were used to immobilize and "wire" GcGDH to the spectrographic graphite electrode's surface. The GcGDH/Os polymer modified electrodes were evaluated by chronoamperometry using flow injection analysis. The current response was investigated using a stepwisely increased applied potential. It was observed that the ratio of GcGDH/Os polymer and the overall loading of the enzyme electrode significantly affect the performance of the enzyme electrode for glucose oxidation. The best-suited Os polymer [Os(4,4'-dimethyl-2,2'-bipyridine)(2)(PVI)Cl](+) had a potential of +309 mV versus NHE, and the optimum GcGDH/Os polymer ratio was 1:2 yielding a maximum current density of 493 μA·cm(-2) at a 30 mM glucose concentration.  相似文献   

13.
The electrochemical oxidation of the adenine moiety in NAD+ and other adenine nucleotides at carbon paste electrodes gives rise to redox-active products which strongly adsorb on the electrode surface. Carbon paste electrodes modified with the oxidation products of NAD+ show excellent electrocatalytic activity toward NADH oxidation, reducing its overpotential by about 400 mV. The rate constant for the catalytic oxidation of NADH, determined by rotating disk electrode measurements and extrapolation to zero concentration of NADH, was found to be 2.5 x 10(5) M-1 s-1. The catalytic oxidation current allows the amperometric detection of NADH at an applied potential of +50 mV (Ag/AgCl) with a detection limit of 4.0 x 10(-7) M and linear response up to 1.0 x 10(-5) M NADH. These modified electrodes can be used as amperometric transducers in the design of biosensors based on coupled dehydrogenase enzymes and, in fact, we have designed an amperometric biosensor for glycerol based on the glycerol dehydrogenase (GlDH) system. The enzyme GlDH and its cofactor NAD+ were co-immobilized in a carbon paste electrode using an electropolymerized layer of nonconducting poly(o-phenylenediamine) (PPD). After partial oxidation of the immobilized NAD+, the modified electrode allows the amperometric detection of the NADH enzymatically obtained at applied potential above 0 V (Ag/AgCl). The resulting biosensor shows a fast and linear response to glycerol within the concentration range of 1.0 x 10(-6)-1.0 x 10(-4) M with a detection limit of 4.3 x 10(-7) M. The amperometric response remains stable for at least 3 days. The biosensor was applied to the determination of glycerol in a plant-extract syrup, with results in good agreement with those for the standard spectrophotometric method.  相似文献   

14.
Graphene oxide(GO) has received considerable attention for glucose detection because of high surface area, abundant functional groups, and good biocompatibility. Defects and functional groups of the GO are beneficial to immobilization of glucose oxidase(GOD), but sacrificing electron-transfer capability for highly-sensitive detection. In order to obtain high GOD loading and highly-sensitive detection of biosensors, we first designed and fabricated a graphene-laminated electrode by combining GO and edgefunctionalized graphene(FG) layers together onto glassy-carbon electrode. The graphene-laminated electrodes exhibited faster electron transfer rate, higher GOD loading of 3.80 × 10-9 mol·cm-2, and higher detection sensitivity of 46.71 μA·mM-1·cm-2 than other graphene-based biosensors reported in literature. Such high performance is mainly attributed to the abundant functional groups of GO, high electrical conductivity of FG, and strong interactions between components in the graphene-laminated electrodes.By virtue of their high enzyme loading and highly-sensitive detection, the graphene-laminated electrodes show great potential to be widely used as high-performance biosensors in the field of medical diagnosis.  相似文献   

15.
Different redox polymers based on poly(allylamine) with covalently attached ferrocene and pyridine groups that coordinate iron and ruthenium complexes were prepared, and hydrogels were obtained by cross-linking them with epichlorohydrin. Charge propagation from the underlying electrode, through the redox polymer and electrical communication with the enzyme FADH(2) of glucose oxidase, was studied by cyclic voltammetry and electrochemical impedance spectroscopy. The effects of electrolyte composition, concentration of enzyme and substrate, and electrode potential are reported. The role of different redox mediators covalently attached to the polymer backbone is discussed in terms of driving force and electrostatic barriers.  相似文献   

16.
New salicylate-selective electrodes based on aluminum-(III) and tin(IV) salophens are described. The electrodes were prepared by incorporating the ionophores into plasticized poly(vinyl chloride) (PVC) membranes, which were directly coated on the surface of graphite electrodes. These novel electrodes display high selectivity for salicylate with respect to many common inorganic and organic anions. The influence of membrane composition and pH and the effect of lipophilic cationic and anionic additives on the response properties of the electrodes were investigated. The electrode based on aluminum salophen, with 32% PVC, 65.8% plasticizer, and 2.2% ionophore, shows the best potentiometric response characteristics and displays a linear log [Sal-] vs EMF response over the concentration range 1 x 10(-6) - 1 x 10(-1) M in phosphate buffer solutions of pH 7.0, with a Nernstian slope of -59.2 mV/decade of salicylate concentration. Highest selectivity was observed for the membrane incorporating 38.8% PVC, 57.3% plasticizer, 2.6% Sn(salophen), and 1.3% sodium tetraphenylborate. The electrodes exhibit fast response times and micromolar detection limits (approximately 1 x 10(-6) M salicylate) and could be used over a wide pH range of 3-8. Applications of the electrodes for determination of salicylate in pharmaceutical preparations and biological samples are reported.  相似文献   

17.
Ten nitrate-selective electrodes based upon rubbery membranes containing various betaine salts as sensors covalently bound to a cross-linked polystyrene-block-polybutadiene-block-polystyrene (SBS) polymer have been produced. The membranes were robust, highly selective, and effective over a pH range of 2-8. The best nitrate-selective electrode fabricated contained 10% m/m dicumyl peroxide, 40% 2-nitrophenyloctyl ether, 6.5% triallyl leucine betaine chloride and 43.5% SBS. The characteristics of this electrode were a linear Nernstian range of 1 x 10(-1) to 5 x 10(-6) mol dm(-3) NO3-, a limit of detection of 3.4 x 10(-7) mol dm(-3) NO3-, and a selectivity coefficient for nitrate against chloride, K(NO3-,Cl-)pot, of 3.4 x 10(-3). These figures represent a significant improvement on current commercial nitrate sensors. Response times were 1 min or less; stability of response and electrode lifetime in continuous use was also very satisfactory.  相似文献   

18.
A new technique suitable for automated, large-scale fabrication of enzyme electrodes by air-spraying enzymes in organic inks is presented. Model oxidoreductases, tyrosinase (Tyr) and glucose oxidase (GOx), were adapted to octane-based ink by entrapment in a system of reverse micelles (RM) of surfactant AOT in octane to separate and stabilize the catalytically active forms of the enzymes in nonpolar organic media. Nonpolar caoutchouk polymer was also used to create a kind of "dry micelles" at the electrode/solution interface. Enzyme/RM/polymer-containing organic inks were air-brushed onto conductive supports and were subsequently covered by sprayed Nafion membranes. The air-brushed enzyme electrodes exhibited relevant bioelectrocatalytic activity toward catechol and glucose, with a linear detection range of 0.1-100 microM catechol and 0.5-7 mM glucose; the sensitivities were 2.41 A M(-1) cm(-2) and 2.98 mA M(-1) cm(-2) for Tyr and GOx electrodes, respectively. The proposed technique of air-brushing enzymes in organic inks enables automated construction of disposable enzyme electrodes of various designs on a mass-production scale.  相似文献   

19.
A new approach to construct a multilayered enzyme film on the Au surface for use as a biosensing interface is described. The film was prepared by alternate layer-by-layer depositions of G4 poly(amidoamine) dendrimers and periodate-oxidized glucose oxidase (GOx). The cyclic voltammograms obtained from the Au electrodes modified with the GOx/dendrimer multilayers revealed that bioelectrocatalytic response is directly correlated to the number of deposited bilayers, that is, to the amount of active enzyme immobilized on the Au electrode surface. From the analysis of voltammetric signals, the coverage of active enzyme per GOx/dendrimer bilayer during the multilayer-forming steps was estimated, which demonstrates that the multilayer is constructed in a spatially ordered manner. Also, with the ellipsometric measurements, a linear increment of the film thickness was registered, supporting the formation of the proposed multilayered structure. The E5D5 electrode showed the sensitivity of 14.7 microA x mM(-1) glucose x cm(-2) and remained stable over 20 days under day-by-day calibrations. The proposed method is simple and would be applicable to the constructions of thickness- and sensitivity-controllable biosensing interfaces composed of multienzymes as well as a single enzyme.  相似文献   

20.
Novel polymeric membrane (PME) and coated graphite (CGE) sulfate-selective electrodes based on a recently synthesized Schiff base complex of Zn(II) were prepared. The electrodes reveal a Nernstian behavior over wide SO4(2-) ion concentration ranges (5.0 x 10(-5)-1.0 x 10(-1) M for PME and 1.0 x 10(-7)-1.0 x 10(-1) M for CGE) and very low detection limits (2.8 x 10(-5) M for PME and 8.5 x 10(-8) M for CGE). The potentiometric response is independent of the pH of the solution in the pH range 3.0-7.0. The electrodes manifest advantages of low resistance, very fast response, and, most importantly, good selectivities relative to a wide variety of other anions. In fact, the selectivity behavior of the proposed SO4(2) ion-selective electrodes shows a great improvement compared to the previously reported electrodes for sulfate ion. The electrodes can be used for at least 3 months without any appreciable divergence in potentials. The electrodes were used as an indicator electrode in the potentiometric titration of sulfate and barium ions and in the determination of iron in ferrous sulfate tablets.  相似文献   

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