| At present, high performance liquid chromatography (HPLC) has mostly been used to enhance the selectivity of analytical methods, because of its high efficiency for separation of analytes from various interfering substances. One of the major research areas about HPLC involves the development sensitive detection methods. More sensitive detection can be realized by HPLC when the analytes are subjected to the use of chemiluminescence (CL) detection. The chemiluminescence (CL) detection system combined with HPLC separation method can offer excellent analytical selectivity and sensitivity. As an important analytical method, CL has the merits of great sensitivity, wide dynamic range, simple equipment and easy automatization. However, the disadvantage of poor selectivity limites its application in many areas. So, the combination between HPLC and CL becomes a necessary consideration. Compounds are separated on the HPLC column and next mixed with CL reaction reagents, the lights emissed from the substances involved in this reaction course are recorded and conducted. Thus, the frame of HPLC-CL comes into being. This analytical method has both excellences of CL and HPLC, at the same time, succeeds in conquering the disadvantage of CL. Compare to UVdetector used in HPLC, CL detector could achieve higher sensitivity for determination of many substances. Since the first report of HPLC-CL technique by Hartkopf etal in 1974, this technique has developed rapidly. It has become a powerful tool for analysis of trace and ultra-trace ingredients and is widely used in the fields of food, chemical industry, pharmacy, environment, life science, biology and clinical medical science.This thesis includes two sections: review and research report. In the review section, informations about high performance liquid chromatography chemiluminescence detection (HPLC-CL) such as principles, HPLC-CL equipment and the main chemiluminescence reaction systems used in HPLC-CL are summarized. In the next research report section, applied reports about HPLC-CL for determination of vanillin, CL for determination of colchicine, a primary study of bubble microinjection system are reported, respectively.1. Determination of Vanillin by High Performance Liquid Chromatography with Chemiluminescence DetectionAn analytical method consisting of HPLC separation and chemiluminescence detection for determination of Vanillin is described. It is based on the chemiluminescence reaction of luminol-K3Fe(CN)6-vanillin. The chemiluminescence intensity is correlated linearly with concentration of vanillin in the range of 6.0×10-7-4.0×10-5mol/L, and the determination limit is 2.0×10-7mol/L (3σ), Its relative standard deviation is 3.9% for 9 measurements (c=4.0×10-6mol/L,n=9). This method has been applied to the determination of vanillin in food samples, the analysis results agrees well with those from HPLC-UVmethod. The average recoveries of vanillin (spiked at the levels of 3.0×10-6-2.0×10-5mol L-1) in food samples ranges from 95 to 104%, and the RSDsof the quantitative results are from 0.9 to 4.5%.2. A Chemiluminescence Flow Injection Method for the Determination of ColchicineA chemiluminescence method for colchicine determination with flow injection is described. It is based on the chemiluminescence emitted during the oxidation of colchicine by potassium permanganate in acidic medium in the presence of formaldehyde. The linear range of determinationfor colchicine was 1.0×10-7 mol/L-6.0×10-5mol/L with the relative standard deviation of 1.9%(c=1.0×10-6mol/L, n=11). The detection limit was 3.0×10-8mol/L (3σ). The recommended method has been used for determination of colchicine in pharmaceutical formulation and the analysis results agreed well with those from official method.3. A Primary Study of Bubble Microinjection SystemWe design a bubble microinjection system . The system is composed of syringes, injection pump, surfactant component (SDS). It produces continuous, stable, homogeneous bubbles. Reagents and samples pass through the detector in the form of air bubbles. The characteristics of above microjectionin system have been evaluated in the presence of reactions and have shown that the proposed approach can be employed to chemilunimescence analysis for determine CL from excited state reaction product. The use of microinjection system can significantly decrease the sample and reagent consumption, promise high throughput, approach simple automation scheme. |
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