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Flow injection and HPLC determination of furosemide using pulsed amperometric detection at microelectrodes.

The flow-injection and HPLC determination of the diuretic drug furosemide using pulsed amperometric detection (PAD) at cylindrical carbon fibre microelectrodes (CFMEs) is reported. Experimental conditions such as pH (6.5) and buffer concentration (0.05 mol l(-1) HPO4(2-)/H2PO4(-)) were optimized using square-wave voltammetry (SWV). Repetitive flow-injection amperometric measurements at +1.25 V for furosemide showed a continuous decrease in the peak current, probably as a consequence of the microelectrode surface fouling. However, a suitable amperometric detection of furosemide was achieved using a PAD program consisting of a two-step potential waveform with alternating anodic and cathodic polarization. The anodic (detection) potential was +1.25 V (time of application 0.1 s), and the cathodic (cleaning) potential was -0.20 V (t=0.2 s). A linear calibration graph was obtained for furosemide in the 5.0 x 10(-7)-1.0 x 10(-4) mol l(-1) concentration range, with a limit of detection of 1.7 x 10(-7) mol l(-1). HPLC-PAD at carbon fibre microelectrodes was used for the determination of furosemide in the presence of several thiouracil drugs and oxytetracycline (OTC). The mobile phase selected was a 25:75 acetonitrile:5.0 x 10(-3) mol l(-1) NaH2PO4 (pH 5.0) mixture. A linear calibration graph was obtained for furosemide in the 1-100 microM range, with a limit of detection of 0.55 microM. The usefulness of this method for the determination of furosemide in real samples was evaluated by performing the analysis of commercial milk samples spiked with furosemide at a concentration level of 4.5 x 10(-7) mol l(-1) (150 ng ml(-1)), as well as with other thiouracil drugs and OTC. A mean recovery of 95+/-5% furosemide was obtained.

Chromatography, High Pressure Liquid↗

A mobile intracellular microelectrode designed to record from neurons in contracting tissue.

Intracellular recording from neurons in moving tissue allows data to be gathered in circumstances that are physiologically more realistic than those requiring pharmacological or mechanical suppression of movement. The construction of a mobile, suspended microelectrode assembly is described. Short glass microelectrodes were attached to a flexible length of 100 micrometer silver wire. A finer wire was inserted in the shank of the microelectrode to carry the electrical signal. Recordings were made from myenteric neurons of the guinea pig ileum, which was moving during the recording session. Intracellular recordings were maintained while the electrodes followed movements of 1 mm or more.

Animals↗

Studies on electrochemical properties and scavenge of superoxide anion in aprotic media by using carbon nanotubes powder microelectrode.

Multiwall carbon nanotubes (MWCNTs) are filled in the cavity at the tip of a microelectrode to form a carbon nanotubes powder microelectrode (CNTs-PME). CNTs-PME was used to study electrochemical properties of superoxide anion in aprotic media. The reversibility of the oxygen/superoxide anion couple (O(2)/O(2)(.-)) at the different powder microelectrode in different aprotic media was compared by cyclic voltammetry (CV). The result indicated that the nearly reversible redox process of the O(2)/O(2)(.-) couple was obtained at a CNTs-PME. The heterogeneous electron transfer rate constant (k(s)) can be measured by steady-state voltammogram and the result is 4.7 x 10(-3) cm s(-1), suggesting that the electrode reaction is a nearly reversible process as expected. The scavenging activities of bilirubin, alpha-tocopherol (vitamin E), are examined, and the experimental results confirm that alpha-tocopherol is the better scavenger toward O(2)(.-) between them.

Bilirubin↗

Amperometric glucose microelectrodes prepared through immobilization of glucose oxidase in redox hydrogels.

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.

Enzymes, Immobilized↗

Dehydrogenase-modified carbon-fiber microelectrodes for the measurement of neurotransmitter dynamics. 1. NADH voltammetry.

The voltammetry of NADH has been characterized at carbon-fiber microelectrodes at scan rates up to 100 V/s. Electrochemical pretreatment of the electrode dramatically changed the properties of the modified electrode. Anodic pretreatment of the surface resulted in an adsorptive wave for NADH oxidation, while less adsorption was evident under more moderate conditions. The pH of the buffer used for the anodization played a critical role in determining the voltammetric peak shape. Oxidation of NADH at slow scan rates (< 10 V/s) fouled the electrode. In contrast, consistent and reproducible voltammetry of NADH was observed at faster scan rates (100 V/s). This voltammetric measurement was used to monitor NADH generated during the oxidative deamination of glutamate catalyzed by glutamate dehydrogenase. A 150-microns-l.d. microdialysis fiber was used to entrap the enzyme near the microelectrode tip, forming the dehydrogenase-modified carbon-fiber microelectrode.

Biosensing Techniques↗

Subsecond adsorption and desorption of dopamine at carbon-fiber microelectrodes.

High-repetition fast-scan cyclic voltammetry and chronoamperometry were used to quantify and characterize the kinetics of dopamine and dopamine-o-quinone adsorption and desorption at carbon-fiber microelectrodes. A flow injection analysis system was used for the precise introduction and removal of a bolus of electroactive substance on a sub-second time scale to the disk-shaped surface of a microelectrode that was fabricated from a single carbon fiber (Thornel type T650 or P55). Pretreatment of the electrode surfaces consisted of soaking them in purified isopropyl alcohol for a minimum of 10 min, which resulted in S/N increasing by 200-400% for dopamine above that for those that were soaked in reagent grade solvent. Because of adsorption, high scan rates (2,000 V/s) are shown to exhibit equivalent S/N ratios as compared to slower, more traditional scan rates. In addition, the steady-state response to a concentration bolus is shown to occur more rapidly when cyclic voltammetric scans are repeated at short intervals (4 ms). The new methodologies allow for more accurate determinations of the kinetics of neurotransmitter release events (10-500 ms) in biological systems. Brain slice and in vivo experiments using T650 cylinder microelectrodes show that voltammetrically measured uptake kinetics in the caudate are faster using 2,000 V/s and 240 Hz measurements, as compared to 300 V/s and 10 Hz.

Adsorption↗

Self-referencing ceramic-based multisite microelectrodes for the detection and elimination of interferences from the measurement of L-glutamate and other analytes.

A self-referencing technique utilizing two microelectrodes on a ceramic-based multisite array is employed for confirmation and elimination of interferences detected by enzyme-based microelectrodes. The measurement of L-glutamate using glutamate oxidase was the test system; however, other oxidase enzymes such as glucose oxidase can be employed. One recording site was coated with Nafion with L-glutamate oxidase and bovine serum albumin (BSA) cross-linked with glutaraldehyde while the other had Nafion with BSA cross-linked with glutaraldehyde. Differences in the chemistry of the two recording sites allowed for identification and elimination of interfering signals to be removed from the analyte response. The electrode showed low detection limits (LOD = 0.98 +/- 0.09 microM, signal-to-noise ratio of 3), fast response times (T90 approximately 1 s), and excellent linearity (R2 = 0.999 +/- 0.000) over the concentration range of 0-200 microM for calibrations of L-glutamate in vitro. The selectivity and dimensions of the multisite electrode allow in vivo glutamate measurements. This electrode has been applied to in vivo measurements of the clearance of locally applied glutamate and release of glutamate in the prefrontal cortex of anesthetized rats. In addition, a aimilar approach has been applied to the development of a microelectrode for measures of glucose.

Amino Acid Oxidoreductases↗

Intracellular magnesium ion selective microelectrode based on a neutral carrier.

A magnesium ion selective microelectrode based on a synthetic neutral carrier is presented. The selectivity of Mg2+ over Na+, K+, H+, and Ca2+ is sufficient for assays of intracellular magnesium ion activities. The microelectrodes with an optimized membrane composition have a resistance of about 5 x 10(10) omega and a 90% response time of less than or equal to 3 s for a tip diameter around 1 microns. The lifetime of the microelectrode cell assembly is longer than 1 week and the emf drift after equilibration is less than or equal to 0.3 mV/h.

Animals↗

Polymer microspray with an integrated thick-film microelectrode.

A microfabrication process leading to a sheathless electrospray interface for mass spectrometry analysis is described. Photoablation is performed on a polymer substrate, allowing the integration of a thick-film conductive track in a sealed microchannel. High voltage is supplied close to the outlet, through an embedded microelectrode. The microspray is generated directly from the edge of the substrate without any tip addition. The flexibility of this technology provides a wide range of dimensions for the probe and the microelectrode design, including location, shape, and conductive material used. Thanks to the thick-film microelectrode and the hydrophobicity of the polymer, which avoids solution spreading at the outlet, the device has been found to be an efficient ionization source providing a stable MS signal through time. Moreover, the same device can be used several times without failure. The performance of the microspray has been studied in simple infusion mode for proteins and reserpine MS analyses. The detection limit of reserpine was found to be at the picomolar level in full-scan MS mode. It implies also that approximately 500 zmol was read consumed during 3 min of infusion. A dynamic range from pico- to millimolar level is also underlined.

Microelectrodes↗

Optimization of the geometry and porosity of microelectrode arrays for sensor design.

This paper describes the systematic investigation of a range of microelectrode arrays with varying dimensions fabricated by standard photolithographic and reactive-ion etching techniques. As expected from theory, the electrochemical behavior of microelectrode arrays with a constant individual diameter varied strongly with center-to-center spacing, the larger the spacing the more sigmoidal the recorded voltammogram. Furthermore, the behavior of arrays with a constant relative center-to-center spacing is shown to vary with individual electrode diameter, the arrays with the smallest electrodes producing strongly peaked voltammograms. Peak current densities and signal-to-noise ratios were also obtained for a variety of array geometries, and the use of electrodeposited platinum black electrodes was investigated. To demonstrate one advantage of using a loosely packed microelectrode array in electroanalysis, a ferrocene-mediated enzyme-linked assay involving the biocatalytic reduction of H2O2 was investigated. Results showed an improved temporal response, with current-time transients reaching a steady-state response more quickly using arrays with increased center-center spacings.

Biosensing Techniques↗

Development of an array of ion-selective microelectrodes aimed for the monitoring of extracellular ionic activities.

In this study, we present the development and the characterization of a generic platform for cell culture able to monitor extracellular ionic activities (K+, NH4+) for real-time monitoring of cell-based responses, such as necrosis, apoptosis, or differentiation. The platform for cell culture is equipped with an array of 16 silicon nitride micropipet-based ion-selective microelectrodes with a diameter of either 2 or 6 microm. This array is located at the bottom of a 200-microm-wide and 350-microm-deep microwell where the cells are cultured. The characterization of the ion-selective microelectrode arrays in different standard and physiological solutions is presented. Near-Nernstian slopes were obtained for potassium- (58.6 +/- 0.8 mV/pK, n = 15) and ammonium-selective microelectrodes (59.4 +/- 3.9 mV/pNH4, n = 13). The calibration curves were highly reproducible and showed an average drift of 4.4 +/- 2.3 mV/h (n = 10). Long-term behavior and response after immersion in physiological solutions are also presented. The lifetime of the sensors was found to be extremely long with a high recovery rate.

Ion-Selective Electrodes↗

Reusable platinum nanoparticle modified boron doped diamond microelectrodes for oxidative determination of arsenite.

Boron doped diamond (BDD) macro- and microelectrodes were modified by electrodeposition of platinum nanoparticles using a multipotential step electrodeposition technique and used for the oxidative determination of arsenite, As(III). The formation of Pt nanoparticles was evident from cyclic voltammetry measurement, whereas AFM and SEM revealed the size and size distribution of deposited Pt nanoparticles. Raman spectroscopy illustrated a correlation between the typical BDD signature and the number of platinum deposition cycles. Linear sweep voltammetry performed with the modified BDD microelectrode outperformed its macrocounterpart and resulted in very low detecting currents with enhanced signal-to-noise ratios. With linearity up to 100 ppb and a detection limit of 0.5 ppb, the electrochemical system was applicable for processing tap and river water samples. Over 150 repetitive runs could be performed, and electrochemical etching of platinum allowed the reuse of the BDD microelectrode. The presence of copper and chloride ions, the two most severe interferents at levels commonly found in groundwater, did not interfere with the assay.

Arsenites↗

Modified microelectrodes and multivariate calibration for flow injection amperometric simultaneous determination of ascorbic acid, dopamine, epinephrine and dipyrone.

Flow injection amperometric quantification of ascorbic acid (AA), dopamine (DA), epinephrine (EP) and dipyrone (DI) in mixtures (in the microgram g-1 range) was successfully performed by using an array of microelectrodes with units modified by the electrodeposition of different noble metals, together with multivariate calibration analysis. The four groups of microelectrodes utilized included a pure gold electrode and electrodes modified by electrodeposition of platinum, palladium or a mixture of platinum + palladium. The array of microelectrodes was inserted in a flow cell and the amperometric data acquisition was performed with a four-channel potentiostat. The analysis of the resulting signals was carried out by a multivariate calibration method, using a group of 16 standard mixtures selected by a two-level factorial design. The analysis of synthetic samples and pharmaceutical compounds containing AA and DI led to very similar values to those obtained by the classical iodimetric analysis. The average absolute errors (in microgram g-1) calculated for each analyte were 0.3, 0.2, 0.4 and 0.4 for AA, DA, EP and DI, respectively.

Ascorbic Acid↗

Liquid and solid-state Cl- -sensitive microelectrodes. Characteristics and application to intracellular Cl- activity in Balanus photoreceptor.

When intracellular chloride activity (aiCl) was monitored with chloride-sensitive liquid ion exchanges (CLIX) microelectrodes in Balanus photoreceptors, replacement of extracellular chloride (Cl0) by methanesulfonate or glutamate was followed by a rapid but incomplete loss of aiCl. When propionate was used as the extracellular anion substitute, CLIX electrodes detected an apparent gain in aiCl, while a newly designed Ag-AgCl wire-in glass microelectrode showed a loss of aiCl under the same conditions. This discrepancy in Cl- washout when propionate replaced Cl0 is explained by the differences in selectivity of CLIX and Ag-AgCl electrodes for native intracellular anions and for the extracellular anion substitute which also replaces Cli and interferes in the determination of aiCl. Both electrodes indicate that ECl approximately Em when the cells are bathed in normal barnacle saline, and both electrodes showed the rate of Cl washout (tau approximately 5 min) to be independent of Cli when Cl0 was replaced by glutamate. Details of Ag-AgCl microelectrode construction are presented. These electrodes were tested and found to be insensitive to the organic anion substitutes used in this study. Selectivity data of CLIX electrodes for several anions of biological interest are described.

Animals↗

Halothane mimics oxygen in oxygen microelectrodes.

The effects of halothane and enflurane on the polarographic measurement of oxygen with five platinum and three gold microelectrodes were examined. Oxygen microelectrodes were calibrated in saline solution equilibrated with either nitrogen (N2) or air, then either halothane, 1.0 per cent, or enflurane, 2.0 per cent, was added to the gas mixture. For each electrode, polarographic curves were determined during exposure to five equilibrating gas mixtures: N2, air, N2 plus halothane, air plus halothane, and N2 plus enflurane. Halothane variably increased the current produced (and therefore the estimated oxygen tension) at all polarizing voltages in saline solution equilibrated with either N2 or air. The effect was present in both conical platinum electrodes and recessed-tip gold electrodes and was not prevented by membrane coatings of polystyrol, Rhoplex or collodion. Enflurane did not alter the polarographic measurement of oxygen. It is concluded that tissue oxygen tension measurements, made with these microelectrodes and membranes, may be unreliable in the presence of halothane.

Calibration↗

Microelectrode-guided thalamotomy for Parkinson's disease.

OBJECTIVE: To describe the outcomes in our first 40 microelectrode-guided thalamotomies for parkinsonian tremor. METHODS: Twenty-four left-sided and 16 right-sided thalamotomies were performed between October 1984 and January 1996; the mean follow-up period was 35.8 months (range, 1-152 mo). The results were evaluated retrospectively and semiquantitatively by a disinterested observer (MNL) and correlated with the quality of the microelectrode recording and the number and size of radiofrequency lesions made. The first 20 and second 20 procedures were evaluated separately. RESULTS: At the last follow-up, the Unified Parkinson's Disease Rating Scale showed no or virtually no tremor in the upper limb in 75% of patients or in the lower limb in 73% of patients. No significant persistent complications were found. These results were achieved at the expense of having to repeat the procedure on 11 sides (in 5 because of technical problems and in 6 for no obvious reason). Total or nearly total abolition of tremor occurred after the first procedure in 40% of the first 20 operations and in 65% of the second 20. Eight of the first 20 procedures and 2 of the second 20 failed for technical reasons. Lesions were made larger in the second 20 procedures than in the first 20. With the use of an electrode with a 1.1 x 3-mm bare tip for 60 seconds, it seems that lesions had to be created at 60 degrees C or more to produce a successful result. CONCLUSION: Thalamotomy with microelectrode recording is an effective procedure with which to treat tremor in patients with Parkinson's disease and may involve fewer complications than conventional techniques. The procedure appears to involve a learning curve.

Adult↗

Magnetic resonance compatibility of multichannel silicon microelectrode systems for neural recording and stimulation: design criteria, tests, and recommendations.

Magnetic resonance (MR) compatibility of biomedical implants and devices represents a challenge for designers and potential risks for users. This paper addresses these problems and presents the first MR-compatible multichannel silicon chronic microelectrode system, used for recording and electrical stimulation of the central nervous system for animal models. A standard chronic assembly, from the Center for Neural Communication Technology at the University of Michigan, was tested on a 2 Tesla magnet to detect forces, heating, and image distortions, and modified to minimize or eliminate susceptibility artifacts, tissue damage, and electrode displacement, maintaining good image quality and safety to the animals. Multiple commercial connectors were tested for MR compatibility and several options for the reference electrode were also tested to minimize image artifacts and provide a stable biocompatible reference for shortand long-term neural recordings. Different holding screws were tested to anchor the microelectrode assembly on the top of the skull. The final selection of this part was based on MR-compatibility, biocompatibility, durability, and mechanical and chemical stability. The required adaptor to interconnect the MR-compatible microelectrode with standard data acquisition systems was also designed and fabricated. The final design is fully MR-compatible and has been successfully tested on guinea pigs.

Animals↗

Silicon-substrate intracortical microelectrode arrays for long-term recording of neuronal spike activity in cerebral cortex.

This study investigated the use of planar, silicon-substrate microelectrodes for chronic unit recording in the cerebral cortex. The 16-channel microelectrodes consisted of four penetrating shanks with four recording sites on each shank. The chronic electrode assembly included an integrated silicon ribbon cable and percutaneous connector. In a consecutive series of six rats, 5/6 (83%) of the implanted microelectrodes recorded neuronal spike activity for more than six weeks, with four of the implants (66%) remaining functional for more than 28 weeks. In each animal, more than 80% of the electrode sites recorded spike activity over sequential recording sessions during the postoperative time period. These results provide a performance baseline to support further electrode system development for intracortical neural implant systems for medical applications.

Action Potentials↗