Research And Development On Quantitative Capillary Electrophoresis And Quantitative Micellar Electrokinetic Capillary Chromatography

Capillary electrophoresis (CE) has been developed into a major analytical tool since 1980s, and has been widely used in various fields such as pharmaceutical, environmental, biotechnology and chemical analyses. However, there are still some limitations in the technique, especially those associated with sample introduction. The lack of a reliable mode of sample introduction hampers the application of CE in quantitative analysis, thereby limiting its scope of utilization. We have been committed to the research and development on a high-precision quantitative capillary electrophoresis system and a high-precision quantitative micellar capillary electrophoresis system. In this dissertation, a high-precision quantitative capillary electrophoresis system with rotary type of injector was developed and tested for the separation and analysis of real samples. For the first time, we constructed a high-precision quantitative micellar electokinetic capillary chromatography (qMEKC) system and applied it to actual sample analysis.This dissertation is divided into four chapters as follows:In Chapter 1, we briefly introduced the background and significance of the project, mainly focused on capillary electrophoresis and micellar electrokinetic capillary chromatography (MEKC). The development and practical application of CE and MEKC in many fields were reviewed.The overview of injection technology of traditional capillary electrophoresis was provided and the problems and difficulties of current injection technology in capillary electrophoresis were summarized.In Chapter 2, a high-precision quantitative capillary electrophoresis system was constructed and optimized.High-precision quantitative capillary electrophoresis system set up: a 20 nL injection rotary type of injector was introduced. And we isolated the injection valve from the electric field by using an electric decoupler, allowing consecutive injections without interrupting the electric voltage and current, and therefore make the system more stable.High-precision quantitative capillary electrophoresis system optimization: We divided the whole capillary into 4 sections and studied the effect of the capillary length of every section on the separation performance. Considering the geometry restriction of the connection, the experimentally adaptable capillary length was chosen to be 15 cm for section from capillary end in the inlet reservoir to the nano-valve injector, 7 cm for section from nano-valve injector to the electrical decoupler and 20 cm for section from detective window to the capillary end in the outlet reservoir.Chapter 3 focused on the quantitative capillary electrophoresis system and its application in analysis of actual samples of caffeine, theobromine and theophylline.The performance of quantitative capillary electrophoresis system was tested. Based on this system, consecutive injections and separations were performed without voltage interruption. Reproducibilities in terms of relative standard deviation (RSD) lower than 0.8% for retention time and 1.7% for peak area were achieved. The effectiveness of the system was demonstrated by the quantitative analysis of caffeine, theobromine and theophylline in real samples, such as tea leaf, roasted coffee, Coca Cola and Theophylline Tablets.In Chapter 4, a high-precision quantitative micellar capillary electrochromatography system was established for the first time, and its performance was evaluatedby the analysis of cephalosporins.The effect of EOF and micelle concentration on the column efficiency was studied. Relative standard deviations (RSDs) in terms of migration time and peak area of benzoic acid, benzene, toluene and DMSO as measured by using this quantitative method, electrokinetic method and hydrodynamic method were calculated. The results showed obviously that sampling accuracy and precision of the quantitative nano-valve method were superior to those of traditional electrokinetic method and hydrodynamic methods. The relative standard deviation of DMSO for peak area was 1.89% by nano-valve injection, better than 5.48% by hydrodynamic injection and 2.32% by electrokinetic injection.Establishment of chromatography conditions for cephalosporins: the type of buffer, SDS concentration, buffer concentration, buffer pH value and the electrophoresis voltage were studied. And the best conditions for chromatography were 10 mM of borax, 20mM SDS (pH8.5) buffer as a background, 18 kV operating voltage. Under the condition, baseline separation of the four cephalosporin drugs was achieved.A nano valve injector and an electric decoupler were introduced to improve the accuracy and precision of quantitative injector technology. On the basis, the quantitative capillary electrophoresis system was optimized and applied in the analysis of real samples. And for the first time, a high-precision quantitative micellar capillary electrochromatography system was constructed and tested, greatly expanding the application scope of the electrokinetic separation technologies.