Direct electrodeposition of a biocomposite consisting of reduced graphene oxide, chitosan and glucose oxidase on a glassy carbon electrode for direct sensing of glucose

We have electrodeposited a composite film consisting of graphene oxide, chitosan and glucose oxidase directly on a glassy carbon electrode (GCE) through electrochemical reduction of a solution of the 3 components under controlled direct electrical potential. The procedure takes only several minutes, and the thickness of the resulting film is uniform and controllable. The GOx has uncompromised bioactivity and exhibits reversible 2-proton and 2-electron transfer in presence of glucose. It therefore can be used amperometric sensing of glucose. The biosensor has a fast response (<3 s), a detection limit of 0.4 μM (which is 50-fold lower compared to the biosensor prepared by drop-casting solutions of the same materials onto an GCE), and a linear response in the 0.4 μM to 2 mM concentration range (which again is much better than that of the biosensor prepared by the drop-casting method). Other features include high reproducibility, long-time storage stability, and satisfactory selectivity. We presume that the direct single-step electrodeposition of this nanocomposite offers a promising approach towards novel types of highly sensitive and stable electrochemical biosensors.

Amperometric determination of glucose with a ferrocene-mediated glucose oxidase/polyacrylamide gel electrode

Enzyme electrodes were constructed by immobilization of glucose oxidase and ferrocene into cross-linked polyacrylamide gels. Electrogenerated ferrocinium ion acts as a direct electron mediator between glucose oxidase and a reticulated vitreous carbon (RVC)/graphite support bed. The electrode is easily constructed, gives a current response proportional to glucose concentrations up to 30 mM, and has good chemical stability in water and air.     

Nonenzymatic glucose detection by using a three-dimensionally ordered, macroporous platinum template

A three-dimensionally ordered, macroporous, inverse-opal platinum film was synthesized electrochemically by the inverted colloidal-crystal template technique. The inverse-opal film that contains platinum nanoparticles showed improved electrocatalytic activity toward glucose oxidation with respect to the directly deposited platinum; this improvement is due to the interconnected porous structure and the greatly enhanced effective surface area. In addition, the inverse-opal Pt-film electrode responds more sensitively to glucose than to common interfering species of ascorbic acid, uric acid, and p-acetamidophenol due to their different electrochemical reaction mechanisms. Results showed that the ordered macroporous materials with enhanced selectivity and sensitivity are promising for fabrication of nonenzymatic glucose biosensors.     

Oxidation of glucose to gluconic acid by glucose oxidase in a membrane bioreactor

Glucose oxidase (GO) (EC 1.1.3.4) was used as catalyst for oxidizing glucose into gluconic acid utilizing a 10-mL Bioengineering Enzyme Membrane Reactor® or a 400-mL Millipore Stirred Ultrafiltration Cell (MSUC) coupled with a Millipore UF membrane (cutoff of 100 kDa) and operated for 12 h under an agitation of 100 rpm, pH 5.5, and 30°C. The effect of feeding rate (0.10, 0.15, or 0.20 min^?1), glucose (2.5 or 5.0 mM), and GO (1.0 or 2.0 mg/mL) concentrations on the catalysis were studied. A yield of about 75% was attained when the MSUC filled with 1.0 mg/mL of GO was fed with 2.5 mM glucose solution at a rate of 0.15 min^?1.     

葡萄糖氧化酶-辣根过氧化物酶-葡萄糖-愈创木酚-荧光反应体系测定血糖

利用愈创木酚的荧光特性,将其导入葡萄糖-葡萄糖氧化酶(GOD)-辣根过氧化物酶(HRP)的荧光猝灭反应体系,并与荧光毛细分析技术结合,开发了一种能测定微量葡萄糖的新方法.本方法的的最佳测定条件为:反应时间15 min,反应温度30C,GOD和HRP的浓度分别为4500U/L、2500U/L,愈创木酚浓度为150μmol...     

On-line biosensors for simultaneous determination of glucose, choline, and glutamate integrated with a microseparation system

An effective microseparation system integrated with ring-disc electrodes and two microfluidic devices was fabricated for in vivo determination using a microdialysis pump. The major interference of ascorbic acid (AA) was excluded by direct oxidation with ascorbate oxidase. Glucose, glutamate, and choline were successfully determined simultaneously through the biosensors modified with a bilayer of osmium-poly(4-vinylpyridine)gel-horseradish peroxidase (Os-gel-HRP)/glucose oxidase (GOD), glutamate oxidase (GlutaOD) or choline oxidase (ChOD). To stabilize the biosensors, 0.2% polyethylenimine (PEI) was mixed with the oxidases. The cathodic currents of glucose, glutamate, and choline biosensors started to increase after the standard solutions were injected into the microseparation system. The on-line biosensors show a wide calibration range (10(-7)-10(-5) mol/L) with a detection limit of 10(-8) mol/L at the working potential of -50 mV. The variations of glucose, glutamate, and choline were determined simultaneously in a free moving rat when we perfused the medial frontal cortex with 100 micro mol/L N-methyl-D-aspartate (NMDA) solution, which is the agonist of the NMDA receptor.     

Activated platinum electrodes as transducer for a glucose sensor using glucose oxidase in a photopolymer membrane

A planar platinum electrode was covered by a photopolymer membrane containing glucose oxidase (GOD) to construct an amperometric glucose sensor. The application of a photopolymer system in membrane formation gives the opportunity to manufacture cheap biosensors with good reproducibility by means of automated techniques and to miniaturise sensors using photolithography. The electrodes were pretreated mechanically and chemically resulting in a half-wave potential (E_1/2) of the H₂O₂ oxidation shifted towards more negative potentials. This shift allows the determination of glucose at a low working potential (300 mV vs. SCE) without addition of mediators. The important advantage of such applied potential decreasing lies in minimising the interference of oxidisable substances such as uric acid, bilirubin and paracetamol. The selectivity to ascorbic acid could also be proved without the application of additional protection layers. The glucose sensor developed has a high life-time, selectivity and sensitivity and a linear working concentration range from 0.05 up to 10 mmol/l of glucose. The sensor was used for the glucose determination in human serum samples with a very good correlation to a common photometric reference method. Received: 13 July 1996 / Revised: 11 September 1996 / Accepted: 14 September 1996     

Electrodeposition of a biocompatible hydroxyapatite matrix to immobilize glucose oxidase for sensitive glucose biosensing

Microchimica Acta - The electrochemical quartz crystal microbalance was used to quantitatively examine the electrodeposition of hydroxyapatite (HA) at a gold electrode. Potentiostatic...     

Investigation of the origin of the glucose response in a glucose oxidase|polyaniline system

The origin of the signal seen in response to glucose in a polyaniline|glucose oxidase system is explored by immittance spectroscopy, by comparing data from an equivalent circuit model and the parameters obtained from a solution of the faradaic branch of the frequency dispersion for a coupled chemical—electrochemical reaction mechanism. It was shown that an RC subcircuit in the equivalent circuit model was sensitive to peroxide concentration, and the interaction of peroxide with polyaniline at potentials where it either oxidised or reduced the polyaniline was discussed. This information was used to compare the data obtained in a bulk and entrapped glucose oxidaselglucose system, and it was seen that the origin of the response could not be fully attributed to peroxide interaction in the latter case. Under anaerobic conditions with entrapped enzyme, it was proposed that a complex between the gluconolactone product of the enzyme reaction and the polymer leads to a more conducting polymer, with inherent charge compensation, and this results in the observed enhanced current signal.     

Fiber-optic glucose biosensor based on glucose oxidase immobilised in a silica gel matrix

A monolithic silica gel matrix with entrapped glucose oxidase was constructed as a bioactive element in an optical biosensor for glucose determination. Physicochemical and biochemical characterizations of the catalytic matrix were performed, and the intrinsic fluorescence of immobilised glucose oxidase (GOD) was investigated in the UV and visible range by performing steady state and time course measurements. In all cases, the silica gel matrix proved to be a suitable support for optical biosensing owing to its superior optical properties (e.g., high transmittance and reliable fluorescence and GOD absorption spectra after immobilisation). From steady state measurements, calibration curves were obtained as a function of glucose concentration. When time course measurements were performed, the silica gel support displayed a larger linear calibration range and higher sensitivity than other immobilisation systems. In addition, a glucose optical biosensor was developed and characterised using as catalytic element GOD immobilised on a gel disk bound to a bundle of optical fibres.     

Investigation of a novel solid-phase chemiluminescent analytical system, incorporating photographic detection, for the measurement of glucose

A method has been developed for the rapid determination of substances by use of solid-phase reagents and a luminescence indicator reaction coupled with photographic detection of the light. The viability of the assay has been demonstrated for glucose estimations. The method uses small sample sizes (5-20 mul) and shows good sensitivity, e.g., detection of glucose down to 28 nmole.     

A glucose sensor based on glucose dehydrogenase adsorbed on a modified carbon electrode

A glucose sensor is prepared by adsorption of the mediator Meldola blue (N,N-dimethyl-7-amino-1,2-benzophenoxazinium ion, as well as glucose dehydrogenase, on the surface of a carbon electrode. The nicotinamide coenzyme, whhich is present in the solution, is reduced in the enzymatic reaction and is re-oxidized amperometrically at 0 mV vs. Ag/AgCl. The properties of such electrodes depend on whether the mediator or the enzyme is adsorbed first; possible models for the molecular arrangements at the surface are discussed. The modified electrode is mounted in a flow-through cell in a flow-injection system and tested with 50-μl injections of β-d-glucose. The calibration graphs were linear in the range 5 × 10^?6—2 × 10^?3 M βd glucose with the highest sensitivity at pH 6.0. The membrane-free enzyme electrode has a fast response; peak widths are 12 s at half height (flow rate 0.7 ml min^?1, making it possible to process 100 samples h^?1.     

Direct electrochemistry of glucose oxidase in a colloid Au-dihexadecylphosphate composite film and its application to develop a glucose biosensor

Colloid Au (Au(nano)) with a diameter of about 10 nm was prepared and used in combination with dihexadecylphosphate (DHP) to immobilize glucose oxidase (GOD) onto the surface of a graphite electrode (GE). The direct electrochemistry of GOD confined in the composite film was investigated. The immobilized GOD displayed a pair of redox peaks with a formal potential of -0.475 mV in pH 7.0 O(2)-free phosphate buffers at scan rate of 150 mV s(-1). The GOD in the composite film retained its bioactivity and could catalyze the reduction of dissolved oxygen. Upon the addition of glucose, the reduction peak current of dissolved oxygen decreased, which could be developed for glucose determination. A calibration linear range of glucose was 0.5-9.3 mM with a detection limit of 0.1 mM and a sensitivity of 1.14 microA mM(-1). The glucose biosensor showed good reproducibility and stability. The general interferences that coexisted in human serum sample such as ascorbic acid and uric acid did not affect glucose determination.     

Integration of a highly ordered gold nanowires array with glucose oxidase for ultra-sensitive glucose detection

A highly sensitive amperometric nanobiosensor has been developed by integration of glucose oxidase (GO_x) with a gold nanowires array (AuNWA) by cross-linking with a mixture of glutaraldehyde (GLA) and bovine serum albumin (BSA). An initial investigation of the morphology of the synthesized AuNWA by field emission scanning electron microscopy (FESEM) and field emission transmission electron microscopy (FETEM) revealed that the nanowires array was highly ordered with rough surface, and the electrochemical features of the AuNWA with/without modification were also investigated. The integrated AuNWA–BSA–GLA–GO_x nanobiosensor with Nafion membrane gave a very high sensitivity of 298.2 μA cm⁻² mM⁻¹ for amperometric detection of glucose, while also achieving a low detection limit of 0.1 μM, and a wide linear range of 5–6000 μM. Furthermore, the nanobiosensor exhibited excellent anti-interference ability towards uric acid (UA) and ascorbic acid (AA) with the aid of Nafion membrane, and the results obtained for the analysis of human blood serum indicated that the device is capable of glucose detection in real samples.     

Immobilization of glucose oxidase on nylon membranes and its application in a flow-through glucose reactor

The immobilization of glucose oxidase on hydrolyzed nylon-6,6 was studied. Various spacers were introduced on the support before the coupling of the enzyme. Best results were obtained when the membrane was covered with denatured bovine serum albumin (BSA) before spacer coupling and immobilization of glucose oxidase (GOD). The influence of various factors (pH, ionic strength, etc.) on the activity of the free and immobilized enzyme was investigated. It was found that the behavior of the fixed glucose oxidase and the free enzyme is very similar. The covalently immobilized enzyme had a lifetime of around 2 months (50% of initial activity).     

Flow-injection analysis of glucose without enzyme based on electrocatalytic oxidation of glucose at a nickel electrode

This paper reports a flow-injection analysis (FIA) of glucose not using enzyme based on the electrocatalytic oxidation of glucose at a nickel electrode. The electrocatalytic mechanism and quantificational method of glucose have been investigated. The current intensity of the electrocatalytic oxidation to glucose at the potential of 550 mV is proportional to the concentration of glucose over the range of 0.10-2.50 mmol l⁻¹, with a 0.04 mmol l⁻¹ detection limit (S/N = 3) and a correlation coefficient of 0.9991. The relative standard deviation (R.S.D.) is less than 4.3% (n = 5) for the determination of practical serum samples. The biologic compounds probably existed in the sample, such as ascorbic acid, uric acid, dopamine and epinephrine, do not disturb the determination of glucose. The result is satisfactory for the determination of glucose in human serum sample as comparison to that from the routine hexokinase method.     

Chemical polymerization of m-phenylenediamine, in the presence of glucose oxidase, produces an enzyme-retaining electrooxidisable polymer used to produce a biosensor for amperometric detection of glucose from brain dialysate

A new procedure involving chemical polymerization of a monomer of m-phenylenediamine (m-ppd) containing glucose oxidase (GOx) and subsequent electro-synthesis of the functional GOx containing polymer onto platinum needle electrodes (PTNE) was used for the amperometric analysis of glucose concentration in brain dialysates. Monomer solutions of o-phenylenediamine (o-ppd) and m-ppd were polymerized by low concentrations of glutaraldehyde (GA) and precipitated from solution. The 1,3 position of the amines on the benzene was amenable to stable polymerization by GA but polymerization of o-ppd (1,2 position) by GA was unstable and degraded. Polymerization of m-ppd appears to proceed by dehydration synthesis. GA induced polymerization of m-ppd polymer in the presence of GOx produced a polymer with strongly bound, functional GOx. This GOx-m-ppd polymer formed a stable matrix that was effectively employed in flow injection analysis (FIA) of glucose. If maintained under O(2) free atmosphere after chemical polymerization, the GOx-m-ppd polymer retained the ability to be electropolymerized. PTNE coated with GOx-m-ppd polymer by repeated dip/amperometry produced stable, sensitive amperometric glucose sensors with good interference exclusion properties and long shelf life. Scanning EM demonstrated that amperometry modified the structure of the GOx-m-ppd on the PTNE surface. GOx-m-ppd PTNE glucose sensors and bare PTNE were placed in a radial flow cell and FIA was employed for the simultaneous measurement of glucose and ascorbic acid, respectively, from dialysates of brain tissue.     

Cast thin film biosensor design based on a Nafion backbone, a multiwalled carbon nanotube conduit, and a glucose oxidase function

Novel electroanalytical sensing nanobiocomposite materials are reported. These materials are prepared by mixing multiwalled carbon nanotubes (MWNTs), a Nafion cation exchanger, and glucose oxidase (GOD) in appropriate amounts. The MWNTs are cylindrical with a diameter in the range 40-60 nm and with a length of up to several micrometers, and they provide electrical conductivity. Nafion acts as a polymer backbone to give stable and homogeneous cast thin films. Both MWNTs and Nafion provide negative functionalities to bind to positively charged redox enzymes such as glucose oxidase. The resulting biosensing composite material is inexpensive, reliable, and easy to use. The homogeneity of the MWNT-Nafion-GOD nanobiocomposite films was characterized by atomic force microscopy (AFM). Amperometric transducers fabricated with these materials were characterized electrochemically using cyclic voltammetry and amperometry in the presence of hydrogen peroxide and in the presence of glucose. Their linear response to hydrogen peroxide was demonstrated. The glucose biosensor sensitivity was strongly influenced by the glucose oxidase concentration within the nanobiocomposite film. The optimized glucose biosensor (2.5 mg/mL GOD) displayed a sensitivity of 330 nA/mM, a linear range of up to 2 mM, a detection limit of 4 microM, and a response time of <3 s.     

Synthesis of a new isoxazolidine from diacetone glucose

The synthesis of a new furo-pyran-based isoxazolidine, making use of an intramolecular oxime–olefin cycloaddition (IOOC) reaction, is reported here, starting from diacetone glucose.     

An optical glucose biosensor based on glucose oxidase immobilized on a swim bladder membrane

An optical glucose biosensor using a swim bladder membrane as an enzyme immobilization platform and an oxygen-sensitive membrane as an optical oxygen transducer has been developed. During the enzymatic reaction, glucose is oxidized by glucose oxidase with a concomitant consumption of dissolved oxygen resulting in an increase in the fluorescence intensity of the optical oxygen transducer. The fluorescence intensity is directly related to the glucose concentration. The effects of pH, temperature, buffer concentration, and selectivity have been studied in detail. The immobilized enzyme retained 80% of its initial activity after being kept for more than 10 months at 4°C. The glucose biosensor has been successfully applied to the determination of glucose content in human blood serum and urine samples. Martin M.F. Choi was on sabbatical leave at The University of North Carolina at Chapel Hill from July 2004 to July 2005.