| Capillary electrophoresis(CE),also known as high-performance capillary electrophoresis (HPCE)or capillary separation(CESM),is a kind of New Liquid Separation Technology with high efficiency and selectivity,which carries out separation by taking capillary tubes for separation passage and high voltage electricity for motive force and in accordance with the difference of mobility and distribution behaviors of all the components of samples.This method has some advantages of high efficiency,fast analysis,high automaticity,little sample consumption and little contamination.Being given great concerns by separation and analysis scientists,it has been one of the most-watched and most advanced technologies in biochemistry and analytical chemistry in last ten years,especially in recent years.Carbohydrate is a kind of substance which widely exists in nature and in organism.Over a long period of time,it has been considered that the main function of carbohydrate is to provide energy.But the newly emerging research on carbohydrate biochemistry shows that it has many other biological activities and functions besides energy-providing.Being the basic unit of protein and having close relationship with vital activity,amino acid has special physiological function in antibody and is one of the indispensable nutrition component in organism.Carbohydrate and amino acid are both the important conponents of life organism,so that it has great value in biology,medicine,food,health and other fields to do research on them by Capillary Eleetrophoresis.Chemical derivation has been developed in the separation of amino acids and carbohydrates, as these biological molecules have no or weak ultraviolet absorption and the low contents cannot be detected directly by UV detection.At present,the widely used derivatizations of carbohydrate are acylhydrazine, amino-pyridine,and aniline ect.And the widely used derivatizations of amino acids are o-phthalaldehyde(OPA),9-fluorenylmethyl chloroformate(FMOC-C1)and Dansyl Chloride (DNS-C1),but these reagents have also been reported some various shortcomings in their application,such as short detection wavelengths,poor stability of reagents and serious interference in the determination of biological samples.In this thesis,new labeling reagents have been synthesized and applied to the separation of amino acids and carbohydrates by capillary electrophoresis,the results are satisfactory.The thesis consists of three chapters.Chapter OneThe developments of capillary electrophoresis(CE)and the its applications in pharmaceutical analysis,biomolecular separation,genetic engineering,clinical medicine and many other fields have been simply introduced.And the application of capillary electrophoresis on carbohydrate seperation and amino acid seperation are introduced in detail.At the same time, the separation strategy of analyte by capillary electrophoresis and the related basic knowledge are reviewed,too.Chapter TwoThe applications of 1-naphthyl-3-methyl-5-pyrazolone(NMP)for the separation of monosaccharides by CE have been investigated.The principal contents of this chapter consist of four parts.2.1 A simple and rapid method for the separation of derivatized monosaccharides using 1-naphthyl-3-methyl-5-pyrazolone(NMP)as derivatization reagent by capillary zone electrophoresis(CZE)has been developed.Under the CZE mode,the separation conditions have been studied and optimized.In optimal conditions,9 monosaccharide derivatives are separated within 18 min.The proposed method has been applied to the determination of monosaccharides in Nitraria and bee-pollen samples with satisfactory results.2.2 Under the capillary zone electrophoresis(CZE)the separation of derivatized monosaccharides using 1-phenyl-3-methyl-5-pyrazolone(PMP)as derivatization reagent has been studied.The separation conditions have been optimized,and under the optimal conditions,9 monosaccharide derivatives are separated within 16 min.The separation is efficient and rapid,the results are satisfactory.2.3 The separation differences between NMP-derivatized monosaccharides and PMP-derivatized monosaccharides by capillary zone electrophoresis(CZE)has been compared and summarized.The results indicate that the NMP-derivatized monosaccharides have higher detective sensitivity.2.4 Under the micellar electrokinetic chromatography(MEKC)mode the separation of derivatized monosaccharides using 1-naphthyl-3-methyl-5-pyrazolone(NMP)as derivatization reagent has been studied.8 NMP-derivatized monosaccharides achieve baseline resolution within 10 min under the proposed conditions.Chapter ThreeThe applications of 2-(11H-benzo[a]carbazol- 11 -yl)ethyl carbonochloridate(BCEC-C1)for the separation of amino acids have been investigated.The principal contents of this chapter consist of two parts.3.1 A simple and rapid method for the separation of derivatized amino acids using 2-(11H-benzo[a]carbazol-11-yl)ethyl carbonochloridate(BCEC-Cl)as derivatization reagent by non-aqueous capillary electrophoresis has been proposed.The separation conditions have been studied and optimized,the optimum conditions are as follows: 0.7mol/L acetic acid,60 mmol/L ammonium acetate buffer,voltage 30kV,column temperature at 18℃and DAD detection at 279 nm.The samples are introduced atmospherically with an injection time of 12 s.The results indicate that 16 derivatized amino acids achieve baseline resolution under the proposed conditions.3.2 Under the micellar electrokinetic chromatography(MEKC)mode,the chiral separation of BCEC-Cl-derivatized amino acids has been studied.To optimize the conditions,the background electrolyte concentration,buffer pH,column temperature,voltage and other factors have been studied.It has been found that the buffer concentration and pH value have a greater impact on the separation,but the influences of temperature,voltage and additives on resolution are not obvious.On the basis of the optimization,15 pairs of amino acid enantiomers are all separated within 20 min with the resolution between 1.41(Phe)～12.04(Asp).This method is simple and rapid,the rusult is satisfactory. |
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