Elimination of high-voltage field effects in end column electrochemical detection in capillary electrophoresis by use of on-chip microband electrodes

The influence of the separation voltage on end column electrochemical detection (EC) in capillary electrophoresis (CE) has been investigated using an electrochemical detector chip based on an array of microband electrodes. It is shown, both theoretically and experimentally, that the effect of the CE electric field on the detection can be practically eliminated, without using a decoupler, by positioning the reference electrode sufficiently close to the working electrode. In the present study, this was demonstrated by using an experimental setup in which neighboring microband electrodes on a chip, positioned 30 microns from the end of the CE capillary, were used as working and reference electrodes, respectively. The short distance (i.e., 10 microns) between the working and reference electrode ensured that both of the electrodes were very similarly affected by the presence of the CE electric field. With this experimental setup, no significant influence of the CE voltage on the peak potentials for gold oxide reduction could be seen for CE voltages up to +30 kV. The detector noise level was also found to be reduced.     

Capillary electrophoresis microchip with a carbon nanotube-modified electrochemical detector

Significant improvements in the performance of a capillary electrophoresis (CE) microchip with an electrochemical detector are observed using a carbon nanotube (CNT)-modified working electrode. The CNT-modified electrode allows CE amperometric detection at significantly lower operating potentials and yields substantially enhanced signal-to-noise characteristics. The electrocatalytic detection is coupled to resistance to surface fouling and hence enhanced stability. Such advantages are illustrated in connection with several classes of hydrazine, phenol, purine, and amino acid compounds. Substantial minimization of surface fouling effects has been demonstrated in connection with the monitoring of phenol and tyrosine. Factors affecting the performance of the new CNT detector were assessed and optimized. CNTs from different sources are evaluated, and the effect of an anodic pretreatment is explored. The broad and significant catalytic activity exhibited by CNT-based CE detectors indicates great promise for a wide range of bioanalytical and environmental applications.     

Integrated electrophoresis chips/amperometric detection with sputtered gold working electrodes

An on-chip electrochemical detector for micromachined capillary electrophoresis (CE) systems, based on sputtering a gold working electrode directly onto the capillary outlet, is described. The new on-chip detector preparation requires no microfabrication or alignment procedures nor a decoupling mechanism. The attractive performance of the integrated electrophoresis chips/amperometric detection was demonstrated for the anodic detection of neurotransmitters. The response for dopamine was linear from 20 to 200 μM, with a LOD of 1.0 μM and a sensitivity of 52 pA/μM. Such intimate coupling of capillary electrophoresis chips and electrochemical detection facilitates the realization of complete integrated microanalytical devices.     

Microchip capillary electrophoresis coupled with a boron-doped diamond electrode-based electrochemical detector

The attractive behavior and advantages of a diamond electrode detector for a micromachined capillary electrophoresis (CE) system are discussed. A chemically vapor-deposited boron-doped diamond (BDD) film band (0.3 x 6.0 mm) electrode is used for end-column amperomettic detection. The favorable performance of the diamond electrode microchip detector is indicated from comparison to a commonly used thick-film carbon detector. The diamond electrode offers enhanced sensitivity, lower noise levels, and sharper peaks for several groups of important anaytes (nitroaromatic explosives, organophosphate nerve agents, phenols). The favorable signal-to-background characteristics of the BDD-based CE detector are coupled with a greatly improved resistance to surface fouling and greater isolation from high separation voltages. The enhanced stability is indicated from a RSD of 0.8% for 60 repetitive measurements of 5 ppm 2,4,6-trinitrotoluene (vs RSD of 10.8% at the thick-film carbon electrode). A highly linear response is obtained for the explosives 1,3-dinitrobenzene and 2,4-dinitrotoluene over the 200-1,400 ppb range, with detection limits of 70 and 110 ppb, respectively. Factors influencing the performance of the BDD detector are assessed and optimized. The attractive properties of BDD make it very promising material for electrochemical detection in CE microchip systems and other micromachined flow analyzers.     

Microchip capillary electrophoresis with an integrated indium tin oxide electrode-based electrochemiluminescence detector

This paper describes an indium tin oxide (ITO) electrode-based Ru(bpy)3(2+) electrochemiluminecence (ECL) detector for a microchip capillary electrophoresis (CE). The microchip CE-ECL system described in this article consists of a poly(dimethylsiloxane) (PDMS) layer containing separation and injection channels and an electrode plate with an ITO electrode fabricated by a photolithographic method. The PDMS layer was reversibly bound to the ITO electrode plate, which greatly simplified the alignment of the separation channel with the working electrode and enhanced the photon-capturing efficiency. In our study, the high separation electric field had no significant influence on the ECL detector, and decouplers for isolating the separation electric field were not needed in the microchip CE-ECL system. The ITO electrodes employed in the experiments displayed good durability and stability in the analytical procedures. Proline was selected to perform the microchip device with a limit of detection of 1.2 microM (S/N = 3) and a linear range from 5 to 600 microM.     

Capillary electrophoresis with an integrated on-capillary tubular detector based on a carbon sol-gel-derived platform

An integrated on-capillary tubular electrochemical detector for capillary electrophoresis systems has been fabricated based on sol-gel technique. It consists of a sol-gel carbon composite tubular electrode attached permanently onto the outlet of the separation capillary. The device greatly eases the setting up of capillary electrophoresis with electrochemical detection (CEEC) as it makes possible electrode/capillary alignment without the aid of a micromanipulator since this integrated unit can be simply immersed in the CE separation buffer in an ordinary three-electrode stationary cell. To improve analytical performance of the integrated unit, the external wall of the exit capillary was etched with HF after the polyimide coating of the capillary had been removed. Influences of the working electrode length and the wall thickness at the outlet of capillary on the separation efficiency and amperometric sensitivity were assessed and optimized. The practical applicability of this configuration is demonstrated with the detection of both catecholamines and carbohydrates. The advantages, namely, versatility, convenience, ease of operation, and low-cost, of the new design combined with an excellent performance lead to high stability and low detection limits.     

Application of diamond microelectrodes for end-column electrochemical detection in capillary electrophoresis

Highly boron-doped diamond microelectrodes were employed in an end-column electrochemical detector for capillary electrophoresis (CE). The diamond microline electrodes were fabricated from conducting diamond thin films (exposed surface area, 300 x 50 microm), and their analytical performance as CE detectors was evaluated in a laboratory-made CE installation. The CE-ED system exhibited high separation efficiency for the detection of several catecholamines, including dopamine (DA), norepinephrine (NE), and epinephrine (E), with excellent analytical performance, for example, 155,000 theoretical plates for DA. The diamond-based electrochemical detection system also displayed low detection limits (approximately 20 nM for E at S/N = 3) and a highly reproducible current response with 10 repetitive injections of mixed analytes containing DA, NE, and E (each 50 microM), with relative standard deviations (RSD) of approximately 5%. The performance of the diamond detector in CE was also evaluated in the detection of chlorinated phenols (CP). When compared to the carbon fiber microelectrode, the diamond electrode exhibited lower detection limits in an end-column CE detection resulting from very low noise levels and highly reproducible analyses without electrode polishing due to analyte fouling, which makes it possible to perform easier and more stable CE analysis.     

Dual-channel method for interference-free in-channel amperometric detection in microchip capillary electrophoresis

A novel in-channel amperometric detection method for microchip capillary electrophoresis (CE) has been developed to avoid the interference from applied potential used in the CE separation. Instead of a single separation channel as in conventional CE microchips, we use a dual-channel configuration consisting of two different parallel separation and reference channels. A working electrode (WE) and a reference electrode (RE) are placed equally at a distance 200 microm from its outlet on each channel. Running buffer flows through the reference channel. Our dual-channel CE microchips consist of a poly(dimethylsiloxane) (PDMS) upper plate and a glass lower plate to form a PDMS/glass hybrid chip. Amperometric signals are measured without any potential shift and interference from the applied CE potential, and CE separation maintains its high resolution because this in-channel configuration does not allow additional band broadening that is notorious in end-channel and off-channel configurations. The high performance of this new in-channel electrochemical detection methodology for CE has been demonstrated by analyzing a mixture of electrochemically active biomolecules: dopamine (DA), norepinephrine, and catechol. We have achieved a 0.1 pA detectability from the analysis of DA, which corresponds to a 1.8 nM concentration.     

Development of a microfabricated palladium decoupler/electrochemical detector for microchip capillary electrophoresis using a hybrid glass/poly(dimethylsiloxane) device

The fabrication and evaluation of a palladium decoupler and working electrode for microchip capillary electrophoresis (CE) with electrochemical detection is described. The use of the Pd decoupler allows the working electrode to be placed directly in the separation channel and eliminates the band-broadening characteristic of the end-channel configuration. The method used for fabrication of the decoupler and working electrode was based on thin-layer deposition of titanium followed by palladium onto a glass substrate. When employed as the cathode in CE, palladium absorbs the hydrogen gas that is generated by the hydrolysis of water. The effect of the decoupler size on the ability to remove hydrogen was evaluated with regard to reproducibility and longevity. Using boric acid and TES buffer systems, 500 microm was determined to be the optimum decoupler size, with effective voltage isolation lasting for approximately 6 h at a constant field strength of 600 V/cm. The effect of distance between the decoupler and working electrode on noise and resolution for the separation of dopamine and epinephrine was also investigated. It was found that 250 microm was the optimum spacing between the decoupler and working electrode. At this spacing, laser-induced fluorescence detection at various points around the decoupler established that the band broadening due to pressure-induced flow that occurs after the decoupler did not significantly affect the separation efficiency of fluorescein. Limits of detection, sensitivity, and linearity for dopamine (500 nM, 3.5 pA/microM, r(2) = 0.9996) and epinephrine (2.1 microM, 2.6 pA/microM, r(2) = 0.9996) were obtained using the palladium decoupler in combination with a Pd working electrode.     

Cyclic chronopotentiometry as a detection tool for flowing solution systems

Cyclic chronopotentiometry provides a very simple detection method, which may be particularly useful in capillary electrophoresis (CE) and microseparation systems. It has been shown that for disk microelectrodes it is possible to define safe reduction and oxidation currents that would never lead to the formation of H2 or O2 gas bubbles, even if they are applied for an indefinitely long time period. During end-column CE detection, currents passing through the working microelectrode can be completely controlled by the external electronic circuit and they are not affected by the separation current. Consequently, problems created by the offset potential in CE can be completely eliminated. The detection can be accomplished through a variety of different mechanisms; however, generation of the electrode response as a result of analyte adsorption seems to be most common. The method is applicable to many analytes, which do not have to be electroactive. The analytical signal is obtained by monitoring the change in the average electrode potential (calculated for either a cathodic or an anodic half-cycle) caused by an analyte interacting with the electrode. The analytical signal is proportional to the analyte concentration, within a concentration range extending over approximately 2 orders of magnitude.     

Experimental characterization of end-column electrochemical detection in conjunction with nonaqueous capillary electrophoresis

An end-column electrochemical detector arrangement for capillary electrophoresis (CE) based on a 75-microm-i.d. capillary and a 25-microm microdisk electrode is characterized. The investigations were carried out using a nonaqueous (acetonitrile-based) buffer and ferrocene model compounds which offer high reliability for voltammetric measurements. The positioning of the microdisk electrode relative to the capillary outlet is the most important parameter for optimization of detection performance as it determines the characteristics of mass transport toward the electrode and the effect of ohmic potential drop resulting from the electrophoretic current on the actual detection potential. On the basis of spatially resolved studies, it was concluded that for the detection system used the microdisk electrode should be placed in a central position relative to the capillary outlet at a distance within the range of 75-100 microm. The presence of a high-voltage electric field had no negative effect on baseline noise, which was demonstrated by comparison of capillary flow injection based on gravity flow and CE experiments. Even a faster stabilization of the baseline was observed by increasing the separation voltage.     

Three-electrode electrochemical detector and platinum film decoupler integrated with a capillary electrophoresis microchip for amperometric detection

This article demonstrates that a three-electrode electrochemical (EC) detector and an electric decoupler could be fabricated in the same glass chip and integrated with an O2-plasma-treated PDMS layer using microfabrication techniques to form the capillary electrophoresis (CE) microchip. The platinized decoupler could mostly decouple the electrochemical detection circuit from the interference of an separation electric field in 10 mM 2-(N-morpholino)ethanesulfonic acid (MES, pH 6.5) solution. The baseline offset of background current recorded from the working electrode with and without application of a separation electric field was maintained at less than 0.05 pA in 10 mM MES. In addition, the platinized pseudoreference electrode was demonstrated to offer a stable potential in electrochemical detection. As a consequence, the limit of detection of dopamine was 0.125 microM at a S/N = 4. The responses for dopamine to different concentrations were found to be linear between 0.25 and 50 microM with a correlation coefficient of 0.9974 and a sensitivity of 11.76 pA/microM. The totally integrated CE-EC microchip should be able to fulfill the ideal of miniaturization and commercialization.     

Movable contactless-conductivity detector for microchip capillary electrophoresis

A new movable contactless-conductivity detection system for microchip capillary electrophoresis is introduced. Such a versatile system relies on positioning the detector at different points along the separation channel via "sliding" the electrode holder. The new movable microchip detection system offers distinct improvements compared to common fixed-location conductivity detectors. For example, placing the detector at different locations along the microchannel offers useful insights into the separation process. Three-dimensional plots of resolution/channel length/separation voltage can be used for optimizing the separation process and selecting the analysis time. The system enables rapid switching between "total" (unresolved) and "individual" (resolved/fingerprint) signals on the basis of placing the detector at the beginning and end of the separation channel, respectively. By moving the detector to a shorter effective separation length, after eluting fast-migrating ions, shorter analysis times can be achieved (through faster detection of late-eluting analytes). These and other improvements in the analytical performance and insights into the separation process are illustrated in connection with the detection of low-energy ionic explosives and nerve agent degradation products.     

Development of rotating electrochemical detectors for capillary electrophoresis

A rotating disk electrode (RDE) has been evaluated and optimized for the detection of electroactive species separated by capillary electrophoresis (CE). With catechol as a working model, the limit of detection was estimated to be 0.3 microM, i.e., approximately 2.5-fold better than that of the stationary disk electrode (0.7 microM). Separation efficiency was significantly improved as exemplified by an increase of theoretical plates from 26,000 plates/m at 0 rpm to 67,000 plates/m at 500 rpm. Of particular importance was the capability of RDE to alleviate electrode passivation and electrical interference associated with high separation potential fields. Therefore, rotation amperometry was especially useful for analytes such as phenolic compounds that tended to rapidly foul the electrode surface. The RDE/ CE system was capable of separation and determination of pentachlorophenol in contaminated soils, and the result obtained agreed well with conventional liquid chromatography, an EPA recommended procedure.     

In-channel electrochemical detection for microchip capillary electrophoresis using an electrically isolated potentiostat

A new electrode configuration for microchip capillary electrophoresis (CE) with electrochemical (EC) detection is described. This approach makes it possible to place the working electrode directly in the separation channel. The "in-channel" EC detection was accomplished without the use of a decoupler through the utilization of a specially designed, electrically isolated potentiostat. The effect of the working electrode position on the separation performance (in terms of plate height and peak skew) of poly(dimethylsiloxane)-based microchip CEEC devices was evaluated by comparing the more commonly used end-channel configuration with this new in-channel approach. Using catechol as the test analyte, it was found that in-channel EC detection decreased the total plate height by a factor of 4.6 and lowered the peak skew by a factor of 1.3. A similar trend was observed for the small, inorganic ion nitrite. Furthermore, a fluorescent and electrochemically active amino acid derivative was used to directly compare the separation performance of in-channel EC detection to that of a widely used laser-induced fluorescence (LIF) detection scheme. In this case, it was found that the plate height and peak skew for both detection schemes were essentially equal, and the separation performance of in-channel EC detection is comparable to LIF detection.     

Microchip capillary electrophoresis with electrochemical detection

A novel microchip capillary electrophoresis system with electrochemical detection, using the replaceable microelectrode, is first reported. This kind of electrode can be fabricated in general laboratories and can be replaced quickly with electrodes of different materials according to the requirements of experiments. The end-column electrochemical detection on microchip CE was achieved by fixing the working electrode (such as carbon fiber, Pt, or Au, etc.) through a guide tube on the end of the separation channel. The experiment results indicate that the alignment of the electrode with the channel outlet can be carried out accurately and reproducibly, and therefore, the detection device has low noise and good reproducibility. The detection limit of dopamine is 2.4 x 10(-7) M, which is the lowest result reported so far. The separation and detection of dopamine, 5-hydroxytryptamine and epinephrine using carbon fiber and Pt microdisk electrodes within 50 s was successfully performed.     

A wireless electrochemiluminescence detector applied to direct and indirect detection for electrophoresis on a microfabricated glass device

A novel electrochemiluminescence (ECL) detector is presented in this article. The detector is applied for micellar electrokinetic chromatographic separation of dichlorotris(2,2'-bipyridyl)ruthenium(II) hydrate [Ru-(bpy)] and dichlorotris(1,10-phenanthroline)ruthenium-(II) hydrate [Ru(phen)] on a microfabricated glass device. It consists of a microfabricated "U"-shape floating platinum electrode placed across the separation channel. The legs of the U function respectively as working and counter electrode. The required potential difference for the ECL reaction is generated at the Pt electrode by the electric field available in the separation channel during electrophoretic separation. Initial experiments demonstrate a micellar electrokinetic separation and direct ECL detection of 10(-16) mol of Ru(phen) (10(-6) M) and 4.5 x 10(-16) mol of Ru(bpy) (5 x 10(-6) M). Also, preliminary results show the indirect detection of three amino acids. The high voltage at the location of detection does not interfere with the electrochemistry.     

Dual microcoil NMR probe coupled to cyclic ce for continuous separation and analyte isolation

Capillary electrophoresis (CE)-nuclear magnetic resonance (NMR) spectroscopy combines the separation efficiency of CE and the information-rich detection capabilities of NMR. However, the temporally narrow CE peaks reduce NMR sensitivity and prevent on-line multidimensional NMR acquisitions. In this work, cyclic CE with multicoil NMR instrumentation is developed to perform CE in multiple closed loops. As a proof of concept, a two-loop five-junction capillary configuration creates two connected yet independently operable fluidic loops. With appropriate voltage switching, analytes can be directed as desired around or between the loops, and a particular analyte band can be parked in one NMR detector coil while CE continues in the second loop and monitored with a second NMR detector coil. The separation of a mixture of amino acids (Ala, Val, Thr) is achieved in two cycles. After one CE cycle, Ala is separated and COSY data are recorded in one loop while Val and Thr are separated in the second loop. At the end of the second cycle, both Val and Thr are separated and multidimensional NMR spectra acquired. With this instrumentation and appropriate protocols, two-dimensional NMR data acquisition and CE separation are achieved simultaneously.     

Capillary electrophoresis coupled with electrochemiluminescence detection using porous etched joint

A new setup to couple capillary electrophoresis (CE) with electrochemiluminescence (ECL) detection is described in which the electrical connection of CE is achieved through a porous section at a distance of 7 mm from the CE capillary outlet. Because the porous capillary wall allowed the CE current to pass through and there was no electric field gradient beyond that section, the influence of CE high-voltage field on the ECL procedure was eliminated. The porous section formed by etching the capillary with hydrofluoric acid after only one side of the circumference of 2-3 mm of polyimide coating of the capillary was removed, while keeping the polyimide coating on the other part to protect the capillary from HF etching makes the capillary joint much more robust since only a part of the circumference of it is etched. A standard three-electrode configuration was used in experiments with Pt wire as a counter electrode, Ag/AgCl as a reference electrode, and a 300-microm diameter Pt disk as a working electrode. Compared with CE-ECL conventional decoupler designs, the present setup with a porous joint has no added dead volume created. Moreover, the dead volume can be increasingly decreased by shortening the distance ( approximately 100 microm) between the working electrode and the end of the separation capillary. The versatility in choice of capillaries and separation buffers within this design is the main advantage over the use of small i.d. capillary and low conductivity buffer in some CE-ECL systems. The performance of this setup is illustrated by the analyses of tripropylamine and proline.     

Electrochemical detection method for nonelectroactive and electroactive analytes in microchip electrophoresis

In this work, we establish an indirect amperometric detection method via mounting a single carbon fiber disk working electrode in the end part of a microchannel. This in-channel configuration for microchip capillary electrophoresis brings about that the potential of the working electrode in the case of electrochemical reduction reaction is coupled by the separation electric field, while the potential of the working electrode in the case of electrochemical oxidation reaction is not coupled by the separation electric field. Such a special performance provides a convenient and sensitive approach for indirectly detecting nonelectroactive analytes that relies on amperometric response of dissolved oxygen in solution and directly detecting electroactive analytes based on their own amperometric response on the carbon fiber electrode. This method has shown its essential importance in the analysis of inorganic cations, biomolecules, and electroosmotic flow rates. Based on preliminary results, a detection limit of 1.0 microM for K(+) and Na(+) have been achieved.