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Analysis and characterization of liposomes using capillary electrophoresis
Orgoníková, Andrea ; Křížek, Tomáš (advisor) ; Coufal, Pavel (referee)
4 Abstract This diploma thesis deals with the analysis of three different types of liposomes using capillary electrophoresis with UV-VIS and laser induced fluorescence detection and the use of liposomes as a pseudostationary phase for analyte separations by liposome electrokinetic chromatography method. In experiments with DSPC-DSPG-PEG2000-DMPE liposomes encapsulating 5-fluoruracil, the peak of liposomes with encapsulated 5-fluoruracil and the peak of free 5-fluoruracil were successfully identified in the electropherograms. Both peaks showed the same absorption spectrum in the UV region, thus confirming their identity. It was proved that capillary electrophoresis with UV-VIS detection is useful for the separation and detection of free and encapsulated drug, which is necessary to determine the efficiency of encapsulation. By monitoring the change of effective analyte mobility after the addition of liposomes to the background electrolyte the applicability of the investigated liposomes in liposome electrokinetic chromatography was evaluated. A change in mobility was observed for negatively charged 5-fluoruracil and positively charged tryptamine and p-toluidine. The absolute value of the effective mobility of negatively charged 5-fluoruracil decreased by 18.2 % due to interactions with liposomes and effective...
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Monitoring of myrosinase enzymatic activity by capillary electrophoresis in off-line and on-line setting
Orgoníková, Andrea ; Křížek, Tomáš (advisor) ; Kubíčková, Anna (referee)
4 Abstract Glucosinolates are secondary products of plants, that are hydrolysed by the enzyme myrosinase. Hydrolysis of glucosinolates leads to a production of toxic compounds, such as isothiocyanates. These degradation products are involved in the defense system of the plant against external pathogens. This bachelor thesis deals with monitoring of myrosinase enzymatic activity by capillary electrophoresis. As a substrate, 3-indolylmethylglucosinolate known by trivial name glucobrassicin, was selected. The enzymatic reaction was tested in both off-line and on-line setting by capillary electrophoresis. As the background eletrolyte a solution containing 9 mM sodium tetraborate, 15 mM phosphoric acid and 10 mM hexadecyltrimethylammonium chloride (pH = 7.02) was used. The sample was injected hydrodynamically by pressure (5 kPa, 3 s). A voltage of -20 kV was applied to the capillary and solution in the capillary was simultaneously mobilized by a pressure 5 kPa. The optimized method was evaluated by measuring calibration curve, limit of detection, limit of quantification, repeatability of injection of glucobrassicin in off-line and on-line setting and also repeatability of on-line enzymatic reaction. The limit of detection was 0.011 mg ml-1 and the limit of quantification was 0.035 mg ml-1 . In both off-line and...
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