Author Hua, Lin
Title Development of electrochemical detection techniques for capillary electrophoresis.
Institute Thesis (Ph.D.) National Institute of Education, Nanyang Technological University
Year 2002
Supervisor Tan, Swee Ngin
Call no. QP519.9.C36 Hua
This thesis is concerned with the development of new electrochemical detection (ECD) techniques in capillary electrophoresis (CE) for the analysis of biomolecules. Electrochemical detection for CE is simple in its design and construction with both high sensitivity and selectivity. In this research project, a new silica sol-gel derived carbon composite amperometric detector is developed for improving the sensitivity and stability of the CE detection system.

Chapter 1 is an introduction to CE which is a relatively new technique for the separation and analysis of chemical compounds from small molecules to biomacromolecules. A compilation of articles and reviews published on CE over the last decade illustrates the exponential growth in number, which reflects that this technique has been widely applied. An overview of a modern CE system and the various modes of CE are included in this chapter. The aims of the thesis are also discussed.

Chapter 2 covers the theoretical aspects of separation in CE. The various aspects are as follows: electroosmotic flow (EOF), electrophoretic mobility, and the influences of electrophoretic parameters on separation. Charged solutes are separated due to differences in their electrophoretic mobility, and will tend to migrate toward the detector following the EOF. The important parameters of a CE separation, including efficiency, resolution, and time, have also been discussed.

In Chapter 3, recent advances in the design of ECD systems in CE are presented. In particular, instrumental design to aid in decoupling the CE and EC systems electrically and in aligning them physically are described in detail. Electrode materials used in CE for ECD are reviewed as widely as possible. The electrochemical techniques, including cyclic voltammetry (CV) and amperometry, have been discussed as they have been used in this work.

Chapter 4 describes the development of a new electrode based on carbon sol-gel composite material and its application for amperometric detection in CE. The performance of the sol-gel carbon composite electrode (CCE) is first evaluated in a typical CE application for the detection of purine-based compounds. Application of the CCE is also demonstrated for the detection of phenolic compounds in a micellar system. Separation resolution for non-ionic phenolic compounds can significantly be enhanced by introducing sodium dodecyl sulfate (SDS) at a concentration above its critical micelle concentration (cmc) to the buffer. Another design of the CCE incorporating the electrocatalyst cuprous oxide (Cu2O) is employed for the analysis of sugars and organic acids based on dynamic modification with cetyltrimethylammonium bromide (CTAB). It has been found that the presence of surfactant in the separation buffer does not adversely influence the electrochemical detection using a sol-gel derived carbon electrode.

A sol-gel CCE has been employed (described in Chapter 5) as an electrochemical detector in a wall-jet arrangement for the determination of neurotransmitters. The use of simple anodic electrochemical pre-treatment has shown to produce an improved and reasonably stable electrode response compared to that of the untreated electrode. Following the electrochemical pre-treatment, the sample mixture containing dopamine, catechol, epinephrine and norepinephrine has been separated under optimized conditions in CE system.

As reported in Chapter 6, chemically modified sol-gel CCEs containing Cu2O exhibit a sensitive electrocatalytic response for the oxidation of carbohydrates. The electrochemical pre-treatment results in a marked enhancement of the current responses for all sugars examined, compared to the untreated electrode. The stability of an electrochemically preanodized Cu2O-CCE has also been investigated in CE experiments.

Chapter 7 describes the application of CE-ECD for the analysis of kaempferol and its derivatives in balsam flowers. The influence of buffer pH on separation has been investigated and optimized. The detection potential has also been evaluated and optimized. The application of this method for the separation and detection of these compounds present in balsam flowers (Impatiens balsamina) is reported.

An integrated on-capillary tubular electrochemical detector for CE systems based on sol-gel technique has been reported in Chapter 8. It consists of a sol-gel carbon composite tubular electrode attached permanently onto the outlet of the separation capillary. To improve analytical performance of the integrated unit, the external wall of the exit capillary has been etched with HF after the polyimide coating of the capillary has been removed. Influences of the working electrode length and the wall thickness at the outlet of capillary on the separation efficiency and amperometric sensitivity have been assessed and optimized. The practical applicability of this configuration has been demonstrated with the detection of both catecholamines and carbohydrates.