| For the obvious advantages, such as minimal reagent consumption, rapid and efficient separations and high resolving power, the broad impact of capillary electrophoresis (CE) has been the subject of extensive study in the fields of analysis of biological samples. With the development of column coating technology and additives, the selectivity and sensitivity of CE methods was improved and it made CE more suitable for the separations and detections of trace analytes in complex biological samples.This dissertation was focused on a new column technology firstly. A series of significant coating capillary based on different carboxyphenylboronic acid functionalized fused-silica capillary was devised and the revelant mechanism was discussed. The coating capillaries have been applied to separate and determine the neurotransmitters in urine samples. In addition, the separation of normal lymphocytes and RAJI cells in this coating capillary was also investigated.Considering for the advantages of additives and the characteristics of graphene oxide (GO), we studied on the study of the feasibility and the effects on the CE performances of GO additive. A parameter, retention factor K’was used to describe the variation of CE performance in theory. Based on the theory of K’, two groups of neutral pollutants were separated by CE using GO as additive and satisfied results were obtained. At the same time, the effects of GO additives in the separations and repeatability of proteins were also investigated.Furthermore, aim to the further appliacaion of CE in the field of biological analysis, four organic acids constituents of Plateau alfalfa roots have been identified and detected by a novel CE strategy which combined chitosan (CS) trapping and CTAB assisted sample stacking.This dissertation consists of five chapters:Chapter 1:In this chapter, the developments and applications of CE were introduced. The improvements and progresses of column coating technology and additives in CE were reviewed intensively. And the impacts of CE in the fields of analysis of biological samples were summarized. The further developments over the research direction and applications of CE were discussed.Chapter 2:A significant coating capillary based on carboxyphenylboronic acid functionalized fused-silica capillary was devised in this work. The separations of neurotransmitters in the different buffer solutions, such as phosphate buffer solution (PBS) and borax buffer solution, were investigated by these coating capillaries. The assumed mechanism of this coating capillary was summarized as follow:The groups of boronic acid in the coating capillaries could bind with diols in the neurotransmitters. The different affinity for the neurotransmitters would finally contribute to the improved concentration and separation of analytes. The regenerations of coating capillaries (simply named reactive capillary) would also be realized based on that pH could typically reverse covalent cyclic ester formation, while the ester dissociated when surrounding pH was changed to acidic. Compared with the other coating capillary in this work, the meta-carboxyphenylboronic acid coated capillary (m-coated capillary) showed the better performances. And then the m-coated capillary was applied to separate and determine the dopamine, epinephrine, isoprenaline hydrochloride and norepinephrine bitartrate in urine samples. Under the optimum conditions, the linearity was obtained over the magnitude range of 5.0～100.0 ng/mL for above neurotransmitters. The limits of detection (LODs) were 1.34～ 2.42 ng/mL and the recoveries were calculated at 92.6～108.1%. The feasibility of this m-coated capillary in cells separation was also investigated. The CE methods based on this m-coated capillary could achieve the separations of RAJI cells and lymphocytes.Chapter 3:Combined with the theory of chromatograph and CE, the effects of GO additive on the CE separations were discussed in this section. Retention factor K’was introduced to describe the variation of CE conditions when GO was used as an additive,where tB+GO and tB were the migration time of analyte (the EOF marker) in the buffer with GO and without GO, respectively. Moreover, the influences of GO additive under different CE conditions were investigated and the expression of K’was corrected as:where C was a constant related to the other CE conditions. In this expression, it meaned that the tendency of K’would be similar if the amount of GO additive was constant. It’s suggested that K could describe the effects of GO additive in CE performances.Chapter 4:Base on the theory of K’, two groups of the neutral pollutants were separated by GO additive within 4 min. The effects of voltage, pH and the added amount of GO were studied. Furthermore, the effects of GO additives on the repeatability and resolution of protein separations were also investigated.Chapter 5:In this section, four organic acids constituents of Plateau alfalfa roots have been identified and detected by a novel capillary electrophoresis (CE) strategy which combined chitosan (CS) trapping and cetyltrimethyl ammonium bromide (CTAB) assisted sample stacking. Under the optimized condition, organic acids, i.e., aconitic acid, gallic acid, citric acid and L-malic acid were concentrated and separated within 3 min. Validation parameters of this method (such as detection limits, linearity and precision) were also investigated and the limit of detection (LOD) was 2.41-53.9 ng ml-1. Linearity was obtained over the magnitude range of 5-4000 ng mL-1 approximately for different organic acids and 3×102～1.5×104 folds enrichment was achieved. The method was applied to the determination of organic acids in roots of normal grown Plateau alfalfa and stressing affected Plateau alfalfa. Satisfactory results and recoveries were obtained. |
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