| Capillary electrophoresis (CE) is considered as one of the most powerful separation techniques with advantages of fast, simple, high separation resolution, little sample consumption and high sensitiviy, it has been used to separate a variety of complex samples in varies fields such as medicine, environment and food. Laser induced fluorescence (LIF) detection is one of the most sensitive method of CE which is suitable for trace components in complex samples. Currently, CE-LIF is often used in the analytes which have fluorescence property or can be labeled and colored by fluorescent reagents, while the traditional fluorescence probes have some disadvantages, such as fewer kinds or longer label time. Quantum dots (QDs) are considered as new fluorescent probes which have many unique size-tunable optical properties, such as broad excitation spectrum, narrow emission spectrum, high photobleaching threshold and excellent photostability, they have board application in life analytical chemistry, biochemistry, molecular biology and so on. The biological samples are precious, complicated and diverse, the target is also at low concentration, so the development of a rapid and sensitive method for the quantitative analysis of the target in biological samples is required. In addition, it is meaningful to do the research on the analysis of trace components in biological samples.Based on the previous studies, In this paper, there innovative works were carried out in this paper.(1) A novel micellar electrokinetic capillary chromatography (MECC) method had been developed for the separation and detection of cortisol,17β-estradiol and testosterone in fish blood. The factors affecting separation efficiency such as concentration of buffer solution and surfactant, additives, pH, separation voltage and temperature were investigated. The optimal conditions were as follows:12mmol/L borax+60mmol/L sodium dodecyl sulfate (SDS)+6mmol/L β-cyclodextrin (β-CD)+8%(v/v) acetonitrile, pH9.5, detection wavelength of225nm, separation voltage and temperature of25kV and25℃, respectively. Under the optimal conditions, the analytes could be base-line separated within4min using reverse separation mode with capillary length of10cm. Good linear relationship were obtained in the range of1.0～100.0μg/mL with correlation coefficient larger than0.9932. The proposed method had been successfully applied for the determination of steroid hormones in fish blood with the recoveries in the range of84.60～113.30%, it meet the demand of biological sample detection.(2) A novel CE method had been developed for the separation and analysis of glutathione-capped CdTe quantum dots and Lysozyme bioconjugates. The GSH-QDs were coupled to Lysozyme using EDC and NHS as bifunctional linkage reagent. The factors affecting coupling efficiency such as the concentration of bifunctional linkage reagent, types of buffer solutions, pH. In the50mmol/L KH2PO4-Na2HPO4buffer solution with pH7.4,20μL2.8×10-3mol/L GSH-CdTe,30μL25mg/mL EDC,30μL1.5mg/mL NHS and40μL lmg/mL Lysozyme coupled effectively. In addition, the optimal separated conditions were as follows:12mmol/L borax with pH9.25, separated voltage and temperature of20kV and25℃, respectively. The QDs bioconjugates were efficiently separated with free QDs under the optimization. This method can be used for detecting the lysozyme in the biological samples.(3) A simple and sensitive method for the determination of diprophylline in human serum was developed based on the fluorescence quenching of GSH-CdTe QDs, which were prepared in the previous work. Parameters affecting the fluorescence intensity of QDs were investigated such as types of buffer solutions and temperature. In addition, parameters affecting the quenching efficiency including pH of buffer solution, reaction time, adding sequence, interfering substances were investigated and optimized. The optimal conditions were as follows:50mmol/L buffer solution with pH8.4, reaction temperature was25℃, reaction time was8min. Under optimum conditions, fixing the concentration of QDs was2.8×10-6mol/L, the calibration plot of quenched fluorescence intensity Fo/F with concentration of diprophylline ranged from1.67×10-6to1.33×10-5mol/L was linear. The detection limit (signal to noise of3) for diprophylline was found to be2.24×10-7mol/L, the relative standard deviation was3.05%(n=3). The proposed method had been successfully applied for the determination of diprophylline in human serum samples. The recovery of the method was in the range of87.41～117.94%. Furthermore, the possible quenching mechanism of the GSH-QDs and diprophylline was also discussed. |
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