| High-performance capillary electrophoresis (HPCE) emerged in the mid-late 1980s in the 20th century, which is a kind of new liquid separation technology with high voltage electricity as motive force and capillary column as separation passage. It has some advantages including high efficiency, quick analysis, little sample consumption, high automaticity and various separation modes. As an important branch of HPCE, nonaqueous capillary electrophoresis (NACE) not only has the advantages of capillary electrophoresis, but also possesses its own unique superiority comparing with aqueous phase system. In recent years the rapid development of NACE has been widely used in life science, medical drugs, food, health care, environmental science and other fields. High performance liquid chromatography (HPLC), which developed from the early 1970s, is one of the fastest developing and the most widely used methods in analytical chemistry. The separation is based on different distribution coefficients, adsorption capacities, molecular sizes, ion-exchange effects of samples displayed in the mobile phase and stationary phase. The responsible mechanisms for the separation in liquid chromatography are different from those in capillary electrophoresis. In view of this, two methods can each other offer complementary analyses.Amino acids are basic units of proteins and play an important role in protein synthesis and life activity of organisms. Peptides are material bases of life, affecting many physiological functions in organisms, such as growth, development, immune defence, cell differentiation, resistance to aging, etc. Therefore, the research of their separation by NACE is of great significance on the research in biology, medicine, food and other areas. Fatty acids are important substances wildly distributed in food, living organisms, and biological fluids. Many kinds of fatty acids play significant physiologically roles at trace levels in the regulation of a variety of physiological and biological functions. Hence, isolation and analysis of these compounds in liquid chromatography has great significance.As most amino acids, small peptides and fatty acids show neither natural absorption in the visible or ultra-violet (UV) regions nor fluorescence, chemical derivation has been developed. In this paper, new labeling reagents were synthesized and applied to the separation of amino acids and small peptides by nonaqueous capillary electrophoresis, to fatty acids by high performance liquid chromatography. The results are satisfactory.The thesis consists of four chapters.Chapter OneThe developments of nonaqueous capillary electrophoresis and its applications in chiral separation, inorganic ions, small organic molecules, biological molecules and many other fields were simply introduced. At the same time, the separation strategy of analytes by NACE and the related basic knowledge were reviewed.Chapter TwoA simple and rapid method for the separation of derivatized amino acids using 2-[2-(7H-dibenzo[a,g]carbazole-ethoxy)]-ethyl chloroformate (DBCEC-C1) as derivatization reagent by nonaqueous capillary electrophoresis has been proposed. The separation conditions are as follows:formamide as solvent, ammonium acetate-acetic acid as buffer, a 58.5 cm×50μm i.d. (50 cm effective length) untreated fused-silica capillary, temperature 18℃,voltage 30 kV, and DAD detection at 290nm. The separation conditions have been studied and optimized. The results indicated that 15 derivatized amino acids achieve baseline resolution.Chapter ThreeA simple and rapid method for the separation of derivatized small peptides using 2-[2-(7H-dibenzo[a,g]carbazole-ethoxy)]-ethyl chloroformate (DBCEC-C1) as derivatization reagent by nonaqueous capillary electrophoresis has been developed. With formamide as the elution solvent, a satisfactory separation for the 5 small peptide derivatives could be obtained when ammonium acetate and acetic acid were used as electrolytes. The effects of several key factors were also investigated in this experimentation.Chapter FourThe principal contents of this chapter consist of two parts:the synthesis of novel fluorescent lableling reagent 2-(12-benzo[b]acridin-5-(12H)-yl)-acetohydrazide (BAAH) and its application to fatty acids in high perfornance liquid chromatography-mass spectra (HPLC-MS).4.1 With benzo[b]acridone used as maternal ring, a novel fluorescent lableling reagent 2-(12-benzo[b]acridin-5-(12H)-yl)-acetohydrazide (BAAH) was designed and synthesized. Its structure and spectroscopic properties were also characterized and researched.4.2 Pre-column derivatization methods for the sensitive determination of fatty acids with 2-(12-benzo[b]acridin-5-(12H)-yl)-acetohydrazide (BAAH) as labeling reagents followed by HPLC-MS analysis have been developed. The optimization of derivatization and chromatographic separation conditions were evaluated. The proposed method applied to fatty acid analysis from oils to give the most satisfactory results. |
PDF Full Text Request