Studies On The Fabrication Of High Performance Capillary Electrophoresis Microchips With Integrated End-channel Amperometric Detection

Recently, microfluidic analytical systems aiming at miniaturization, integration, portability of analytical instruments have been growing rapidly. The bulky and complex equipments required by laser-induced fluorescence (LIF) and mass-spectrum (MS) impede the integration of LIF or MS with the microchips. Moreover, the cost of LIF and MS is so expensive that they are usually not available in ordinary analytical laboratory. Electrochemical detection in amperometric mode (AD) is an attractive alternative to utilize for microchip analysis owing to its such impressive advantages as high sensitivity, no requirement of derivatization for many compounds, ideal compatibility with integration and miniaturization, and less expensive in instrumentation.Microchip capillary electrophoresis-amperometric detection (μCE-AD) faces a major challenge, that is, the high-voltage field interference with the amperometric detection. To eliminate or reduce the interference, some workers used off-channel detection mode in-cooperation of a field decoupler or in-channel detection mode in combination with an electrically isolated potentiostat. Most workers, however, employed end-channel detection mode, because it is much simpler in chip fabrication and/or instrumentation in comparison to the off-channel and in-channel detection modes. Nevertheless it can not eliminate the interference of separation voltage thoroughly.The present work is aimed to fabricate high performance capillary electrophoresis microchips with integrated end-channel amperometric detection, and investigate their separation and detection performance. A major thrust is to diminish the separation voltage interference by optimizing the geometric parameters of the separation channels and the detection electrodes. The thesis is composed of four parts:In chapter 1, the recent progress in microchip capillary electrophoresis with amperometric detection was reviewed. It includes microchip design, fabrication of integrated electrodes, elimination of high-voltage interference with the detection, separation field-non-isolating detection, electrode and channel arrays, and applications of microchip capillary electrophoresis with amperometric detection in analysis.In chapter 2, a integrated glass capillary electrophoresis microchips with a replaceable working electrode for amperometric detection was fabricated. With dopamine (DA) and catechol (CA) being used as model analytes, based on the glass chips, the influences of channel cross-sectional area and channel-to-electrode distance on the separation voltage interference with end-channel amperometric detection, accordingly on the separation and detection performances of theμCE system were investigated. By using a chip with the smallest channel cross-section (312μm² with top width of 37.3μm and depth of 8.9μm), the residual separation voltage field in the detection cell was significantly reduced, so that detection was conducted at a channel-to-electrode distance of 20μm to achieve better separation and detection performances. Detection limits of 0.46 and 0.94μmol/L were achieved for DA and CA, respectively, and a theoretical plate number of 9.1×10³ m^-1 was obtained for DA. Relative standard deviations in peak heights observed for seven runs of a standard solution containing the two analytes (0.1 mmol/L for each) were 1.3 % and 3.0 % for DA and CA, respectively.In chapter 3, a fully integrated high performance polycarbonate (PC) microchip for CE with end-channel amperometric detection has been developed. The on-chip integrated three-electrode system consisting of a gold working electrode, an Ag/AgCl reference electrode and a platinum counter electrode was fabricated by photo-directed electroless plating combined with electroplating. The working electrode was positioned against the separation channel exit to reduce post-channel band broadening. The electrophoresis high voltage interference with the amperometric detection was assessed with respect to detection noise and potential shifts at various working-to-reference electrode spacing. It was observed that the electrophoresis HV interference caused by positioning the working electrode against the channel exit could be diminished by using an on-chip integrated reference electrode that was positioned in close proximity (100μm) to the working electrode. TheμCE-AD microchip was demonstrated for the separation of model analytes, including DA and CA. Detection limits of 132 and 164 nmol/L were achieved for DA and CA, respectively, and a theoretical plate number of 2.5×10⁴ m^-1 was obtained for DA. Relative standard deviations in peak heights observed for five runs of a standard solution containing the two analytes (0.1 mmol/L for each) were 1.2 % and 3.1 % for DA and CA, respectively. The chip could be continuously used for more than 8 h without significant deterioration in analytical performance.In chapter 4, a dual-channel PC electrophoresis microchip with dual end-channel amperometric detectors was prepared, and the possibility of simultaneous analysis of two samples was investigated. Significant difference in the separation and detection behavior between two channels was observed. The minor differences in the channel geometry and two working electrodes aligning against two independent channels were the most possible reasons for the observed nonidentical analytical performance. Thereupon, it is a key point to prepare highly identical channels, electrodes and the spacing between the channels and the working electrodes. The main novelty of the present work is summarized as:1. Revealing the close relationship between the channel cross-sectional area and the interference of separation voltage on detection. Put forth an idea that the working electrode could be positioned closed to the channel outlet using the chip with less cross-sectional area, which could be helpful to improving the separation and detection performance of CE-AD system.2. Developing of a novel technique of photo-directed electroless plating combined with electroplating to fabricate the integrated three-electrode amperometric detection system on PC sheets. Established a convenient technique by hot embossing to channel fabrication and chip bonding, which can be proceeded in ordinary analytical laboratory.3. Establishing a high performance microchip capillary electrophoresis with end-channel amperometric detection system, where post-channel band broadening was reduced by aligning the working electrode against the separation channel exit, and separation voltage interference caused by positioning the working electrode against the channel exit was diminished by using an on-chip integrated reference electrode that was positioned in close proximity to the working electrode.