Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:1.10.3.3 (ascorbate oxidase)
778 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

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.
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PMID:On-line biosensors for simultaneous determination of glucose, choline, and glutamate integrated with a microseparation system. 1451 55

Hydrogel-coated microsensors based on carbon fiber electrodes (CFEs) are promising tools for in vivo analysis of endogeneous compounds such as glutamate. However, their construction generally depends on manual fabrication, which often results in poor reproducibility. The aim of this study was to improve the reproducibility and performance of glutamate microsensors. CFEs (10 microm diameter, 300-500 microm long) were coated with a cross-linked redox-polymer hydrogel containing l-glutamate oxidase, horseradish peroxidase and ascorbate oxidase. Various parameters that are likely to influence the reproducibility of the glutamate microsensors were studied. It appeared that the most crucial step in determining the microsensor performance is the manual hydrogel-application procedure. To control this procedure an automated dipcoater was constructed, which allowed mechanical application of the hydrogel on the CFE under standardized conditions. Significant improvements in performance were seen when the CFEs were dipcoated for 10 min at 37 degrees C. Further improvements were obtained when the automated hydrogel application was combined with other cross-link methods, such as electrodeposition and electrostatic complexation. A crucial factor in determining the microsensor performance is the hydrogel thickness. Microscopic observations revealed that, despite the use of an automated dipcoater, the layer thickness was not constant. By combining the automated dipcoat technique with amperometry, the layer thickness could be indirectly monitored and controlled, which resulted in significant improvements of the reproducibility of the sensors.
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PMID:Improving the reproducibility of hydrogel-coated glutamate microsensors by using an automated dipcoater. 1558 41

The functionalized conducting polymer (CP) of 5, 2':5', 2' '-terthiophene-3'-carboxylic acid on a platinum microelectrode was prepared through the electropolymerization process using cyclic voltammetry and was used as a substrate for the immobilization of enzymes. The nanoparticles of the CP were obtained at a high scan rate in the cyclic voltammetric experiment. A needle-type amperometric glutamate microbiosensor based on the covalent immobilization of glutamate oxidase (GlOx) onto the CP layer was fabricated for in vivo measurements. The surfaces of the CP/Pt and GlOx/CP/Pt were characterized by QCM, ESCA, and AFM. The biosensor efficiently detected glutamate through the oxidation of enzymatically generated H2O2 at approximately +0.45 V versus Ag/AgCl. Various experimental parameters, such as pH, temperature, and the applied potential in the detection step were optimized. The interference effects from other biological compounds were examined, and ascorbate and dopamine interferences were observed, which were completely minimized by coimmobilizing ascorbate oxidase and by coating the sensor surface with a cationic polymer, polyethyleneimine. A linear calibration plot for glutamate was obtained between 0.2 and 100 microM with a detection limit of 0.1 +/- 0.03 microM. The proposed glutamate microbiosensor was successfully used for in vivo monitoring of the extracellular glutamate released by cocaine stimulation.
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PMID:Functionalized conducting polymer as an enzyme-immobilizing substrate: an amperometric glutamate microbiosensor for in vivo measurements. 1605 98

Amperometric hydrogel-coated glutamate microsensors form a promising concept to detect glutamate levels directly in brain tissue. These microsensors are constructed by coating a carbon fiber electrode (CFE) (10 microm diameter; 300-500 microm long) with a five-component redox-hydrogel, in which L-glutamate oxidase, horseradish peroxidase, and ascorbate oxidase are wired via poly(ethylene glycol) diglycidyl ether to an osmium-containing redox polymer. Coating with a thin Nafion film completes the construction. Prior to use in vivo, a reliable and reproducible construction of microsensors with a high performance is required. For an optimal microsensor performance, the balance between the five individual hydrogel components is critical. However, due to their small size, hydrogel application to CFE's need to be performed by dip-coating. Dip-coating is a difficult procedure to control and does not allow individual application of hydrogel constituents. To improve the microsensor construction and to better control the dip-coating procedure, we have recently developed an automated device. Throughout this study, automatic dip-coating was performed with premixed solutions, in which the amount of a single component was varied. This allowed us to optimize the hydrogel composition, which resulted in a significant improvement of the microsensor properties in terms of sensitivity, current density, linearity, detection limit, and interference by ascorbic acid.
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PMID:Improving glutamate microsensors by optimizing the composition of the redox hydrogel. 1613 Oct 61

Real-time monitoring of L-glutamate release from various neuronal regions of mouse hippocampal slices under ischemia (a glucose-free hypoxia condition) is described. A glass capillary microelectrode with a tip size of approximately 10 microm containing a very small volume ( approximately 2 microL) of a solution of glutamate oxidase (GluOx) and ascorbate oxidase was used. First, the amperometric response behavior of the electrode at 0 V versus Ag/AgCl was characterized with a standard glutamate solution in terms of continuous measurements, effect of oxygen, viscosity of solution and concentration dependence. The electrode was applied to the real-time monitoring of L-glutamate released from different neuronal regions of acute hippocampal slices submerged in a hypoxia solution. The time-resolved amounts of L-glutamate released at various neuronal regions (CA1, CA3 and DG) of mouse hippocampal slices were quantified and compared with the reported L-glutamate fluxes using difference-image analysis during ischemia.
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PMID:Real-time monitoring of L-glutamate release from mouse brain slices under ischemia with a glass capillary-based enzyme electrode. 1615 99

Enzyme-based biosensors have the potential to directly detect extracellular concentrations of glutamate in brain tissue with a high spatial and temporal resolution. To optimize their analytical performance, much attention has been paid to the architectural construction of these biosensors. In particular, the coupling of enzymes to the electrode surface has received much interest, which has resulted in many (derivatives of) first-, second-, and third-generation type of biosensors. However, it is remarkable that in the literature little attention, if any, has been paid to the influence of the quality of the enzyme itself on the analytical performance of a biosensor. Previously we have reported that different batches of ascorbate oxidase significantly altered the performance of our glutamate microsensor.(1) In this note, it is shown that a simple enzyme purification procedure as buffer exchange leads to a more uniform enzyme quality and also significantly improves the reproducibility and performance of the microsensor. In our opinion, this is an important observation and of general interest for the construction of enzyme-based biosensors.
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PMID:Improving the performance of glutamate microsensors by purification of ascorbate oxidase. 1657 35

Glutamate microsensors form a promising analytical tool for monitoring neuronally derived glutamate directly in the brain. However, when a microsensor is implanted in brain tissue, many factors can diminish its performance. Consequently, a thorough characterization and evaluation of a microsensor is required concerning all factors that may possibly be encountered in vivo. The present report deals with the validation of a hydrogel-coated glutamate microsensor. This microsensor is constructed by coating a carbon fiber electrode (10-microm diameter; 300-500 microm long) with a five-component redox hydrogel, in which L-glutamate oxidase, horseradish peroxidase, and ascorbate oxidase are wired via poly(ethylene glycol) diglycidyl ether to an osmium-containing redox polymer. A thin Nafion coating completes the construction. Although this microsensor was previously used in vivo, information concerning its validation is limited. In the present study, attention was given to its selectivity, specificity, calibration, oxygen dependency, biofouling, operating potential dependency, and linear range. In addition, successful microsensor experiments in microdialysate, in vitro (in organotypic hippocampal slice cultures), and in vivo (in anesthesized rats) are shown.
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PMID:Evaluation of hydrogel-coated glutamate microsensors. 1668 39

The patterns of development of the vestibular nuclei (VN) and their main connections involving glutamate neurotransmission offer a good model for studying the function of the glial-derived neuromodulator D-serine in synaptic plasticity. In this study we show that D-serine is present in the VN and we analyzed its distribution and the levels of expression of serine racemase and D-amino acid oxidase (D-AAO) at different stages of postnatal (P) development. From birth to P21, high levels of D-serine were detected in glial cells and processes in all parts of the VN. This period corresponded to high expression of serine racemase and low expression of D-AAO. On the other hand, in the mature VN D-serine displayed very low levels and was mainly localized in neuronal cell bodies and dendrites. This drop of D-serine in adult stages corresponded to an increasing expression of D-AAO at mature stages. High levels of glial D-serine during the first 3 weeks of postnatal development correspond to an intense period of plasticity and synaptogenesis and maturation of VN afferents, suggesting that D-serine could be involved in these phenomena. These results demonstrate for the first time that changes in D-serine levels and distribution occur during postnatal development in the central nervous system. The strong decrease of D-serine levels and the glial-to-neuronal switch suggests that D-serine may have distinct functional roles depending on the developmental stage of the vestibular network.
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PMID:Changes in D-serine levels and localization during postnatal development of the rat vestibular nuclei. 1673 85

An alternative approach to production of amperometric microbiosensors, which combines electrochemical electrometallization and electropolymerisation of phenylene diamine film with covalent binding enzymes, is presented. In this respect, for a sensitive detection of hydrogen peroxide (HP) at +0.4V versus Ag/AgCl (detection limit of 0.5 microM, s/n=3), carbon fiber microelectrodes (30 microm in diameter and 500 microm long) were covered with ruthenium. To obtain a highly selective detection of HP, in the presence of different interfering compounds (ascorbic acid, uric acid, etc.), an additive semi-permeable polymer film was formed on the top of the ruthenium layer by electropolymerisation of m-phenylene diamine (m-PD). The enzymatic selective layers were formed by covalent cross-linking the enzymes (glucose oxidase, lactate oxidase or glutamate oxidase) with BSA by glutaraldehyde in the presence of ascorbate oxidase. An additional polymeric layer based on polyurethane and Nafion was deposited on the top of the enzymatic membrane (glucose oxidase, lactate oxidase, or glutamate oxidase) in order to extend the dynamic range of biosensors up to 4mM for glucose (R=0.997; Y[nA]=-0.22+9.68x[glucose, mM]), 1.75mM for lactate (R=0.991; Y[nA]=0.43+15.36x[lactate, mM]) and 0.25 mM for glutamate (R=0.999; Y[nA]=0.02+29.14x[glutamate, mM]). The developed microbiosensors exhibited also negligible influences from interfering compounds at their physiological concentrations. Microbiosensors remained stable during 10h in a flow injection system at 36 degrees C and pH 7.4. The microbiosensors developed are now used in vivo and, as an example, we report here the data obtained with the glucose biosensor.
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PMID:Highly selective microbiosensors for in vivo measurement of glucose, lactate and glutamate. 1772 13

Poly(gamma-benzyl-L-glutamate) (PBLG) has been a popular model polypeptide for a range of physicochemical studies, and its modifiable ester side chains make it an attractive platform for various potential applications. Thin films of Poly(gamma-benzyl-L-glutamate) PBLG were surface grafted within nanoporous anodic alumina (AAO) by surface-initiated polymerization of the N-carboxy anhydride of benzyl-L-glutamate (BLG-NCA). The grafting process was characterized by optical waveguide spectroscopy (OWS), infrared spectroscopy (FT-IR), and scanning electron microscopy (SEM). OWS was able to track the PBLG layer thickness increase in situ, and ex situ FT-IR gave complementary information on the PBLG chain's secondary structure. Transitions in the PBLG growth rate could be correlated with transitions in the polypeptide secondary structure. The emergence of a three-dimensional, anisotropic PBLG morphology within the cylindrical pores of the AAO membrane was also identified as the grafted PBLG average layer thickness increased. Comparison of the PBLG/AAO results with those on a planar silicon dioxide surface indicated that both the conformational transitions and the PBLG nanostructure development could be attributed to the confining geometry within the pores of the nanoporous AAO matrix. The use of a nanoporous AAO matrix, combined with the surface grafting of a thin film of PBLG chains with multiple modifiable side chains, could potentially offer a nanoporous platform with a very high density of functional sites.
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PMID:In situ characterization of N-carboxy anhydride polymerization in nanoporous anodic alumina. 1922 3


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