| High Performance Liquid Chromatography (HPLC) as an important modern measurement of analysis and separation is deveopled on the basis of gas Chromatography and classical Liquid Chromatography. Due to its good separation efficiency, high-speed analysis, high sensitivity, wide range of applications and simple operation, it has been widely used in the areas of environment, biology engineering, pharmacy, foodstuff and petroleum so far.Active components in traditional chinese medicine (TCM) are its substance foundations for use in curing diseases. The content of these active components in traditional chinese medicine are important for controlling product quality and evaluating its security.As an important procedure of chromatography analysis, sample preparation always draws much attention. A simple, rapid and effective sample preparation method for the complex matrix chromatographic analysis is always the target that pursued by the analyst.The thesis includes two sections-review and research report. In the review section, information about high performance liquid chromatography such as characteristic, the development of chromatography technology and the importance of sample preparation, the methods and application of sample preparation were summarized. In the next research report section, analysis methods of determination of chlorogenic acid, rutin and quercetin in Honeysuckle and isoflavones in Kudzuvin Root were reported by HPLC and Stability study of Vitamins and vitamins content in foods were repored by high-performance liquid chromatography.1. Micelle-mediated extraction for the analysis of chlorogenicacid, rutin and quercetin in Honeysuckle by HPLC-UVA MME-HPLC (UV) method was developed for the determination of chlorogenic acid, rutin and quercetin, nonionic surfactant Triton X-100 was used to extract and preconcentrate active ingredients from Honeysuckle prior to their determination by HPLC-UV. The experimental conditions were optimized by a L16 (45) orthogonal array design (OAD) with five factors at four levels using statistical analysis. Using a phenomenex C18 column (150 mm×4.6 mm,5μm), the mobile phase of methanol-0.2% acetic acid with elution, the flow rate was 0.8 mL/min, the detection wavelength was at 350 nm, the content of chlorogenic acid, rutin and quercetin were determined by external standard method.Under optimum conditions, i.e.5% Triton X-100 (w/v), liquid/solid ratio of 400:1 (mL.g-1), ultrasonic-assisted extraction for 30 min at 30℃, the extraction content of the active ingredients reached the highest value. When compared with commonly used solvents,5% Triton X-100 yielded in a way higher than the extraction efficiency of methanol. For the preconcentration of compounds by cloud-point extraction, sodium chloride was added to the solution to facilitate the phase separation. The preconcentration factor for each compound was about 12.5. The proposed method has been applied to the determination of active ingredients in different samples and a standard reference material (SRM).2. Simultaneous determination of isoflavonoids in Kudzuvin Root Starch using high performance liquid chromatography micelle mediated extractionA method based on micelle-mediated extraction was developed for the simultaneous determination of five isoflavonoids including puerarin, daidzin, daidzein, genistin and genistein from kudzuvin root starch by reverse-phase high performance liquid chromatography (RP-HPLC). Methods:The non-ionic surfactant Triton X-114 was chosen as the extract solvent. The samples were analyzed on a Phenomenex C18 column with methanol-0.25%(v/v) acetic acid glacial as a mobile phase using a gradient elution and detected with a UV detector set at 250 nm. Various experimental conditions were investigated to optimize the extraction and preconcentration process. Results:3% of Triton X-114 surfactant (w/v), liquid/solid ratio of 100:1 (mL/g), ultrasonic-assisted extraction at 40℃for 30 min, the extraction yield reached the highest value. For the preconcentration of compounds by cloud-point extraction, sodium chloride was added to the solution to facilitate the phase separation and increase the preconcentration factor. The limit of detection (LOD) was in the range of 0.016～0.079μg/mL (RSN=3) in kudzuvin root starch. The average recoveries were 93.7%～98.9%. Conclusion:This new method is low cost, high extraction efficiency, friendly-environment and simple operation, and suitable for use in the quality control of five isoflavonoids from kudzuvin root starch.3. Stability study of Vitamins A and E by high-performance liquid chromatographyThe stability of Vitamins A and E were investigated with HPLC.The study achieved by using a phenomenex C18 (150 mm×4.6 mm i.d) column with 100% methanal as mobile phase, accomplished by using SPD-20A UV detector. The results showed that Vitamin A is stable to heating, neutral to alkali, reducing agent, but unstable to acid solution and oxidizing agent and Vitamin E is stable in the condition of heat, neutral of faintly acid solution, reducing agent, but it was very sensitive to basic solution and oxidizing agent.4. Dispersive liquid-liquid microextraction coupled with high performance liquid chromatography for the determination of vitamins A and E in foodA simple, rapid and effective method, the dispersive liquid-liquid microextraction.coupled with high performance liquid chromatography (DLLM E-HPLC), has been developed for the extraction and determination of vitamins A and E in food. The six relevant factors, such as type and volume of dispersion agent and extraction solvents, the extraction time and ionic strength were investigated and optimized by the L18 (37) orthogonal design with the average extraction recoveries of vitamins A and E as the index determined by HPLC. Under the optimized extraction conditions, the reliability of the proposed method was evaluated. A good linear relationship was obtained in the range of 0.23～174μg/mL of both vitamin A and vitamin E. The limits of detection were 3.26 ng,7.6ng (S/N=3), respectively. The relative standard deviations were 0.68%-3.59%(n=6). The enrichment factors ranged from 75 to 90. The developed method was applied to the determination of vitamins A and E in food, the average recoveries were 90.6%-96.7% with RSD of 1.78%-3.2% (n=6). The optimized technology is stable, feasible and suitable for the analysis of vitamins A and E in food. |
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