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Optimization of a method for the determination of inorganic ions using capillary electrophoresis
Špačková, Simona ; Bezděková, Jaroslava (referee) ; Vaculovičová, Markéta (advisor)
Inorganic ions are one of the most important group chemicals. Their importance is crucial for all living organisms on the planet and monitoring of their content is essential for this purpose in clinical or environmental analysis but also in the food industry, agriculture or many other industries. Many analytical methods for ion detection have been designed and used, depending on the area of their use. Recently, numerous methods have been adapted to areas where not primarily intended. One such methods may be, for example, laser ablation with inductively coupled plasma mass spectrometry, which has recently begun to affect the analysis of biological samples. As a complementary method, able to detect also non-metallic ions is capillary electrophoresis. In this master thesis, the possibility of applying capillary electrophoresis with indirect photometric detection for the separation of inorganic ions in cell lysates of HeLa cells by capillary electrophoresis was verified. Main attention was paid to chloride ions. The ability to separate inorganic compounds in cellular material using salicylic acid as an absorbing anion in the base electrolyte was tested.
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Use of capillary electrophoresis to determine a selected group of flavonoids
Tran Xuan, Tiep ; Mravcová, Ludmila (referee) ; Řezáčová, Veronika (advisor)
In recent years, the use of natural medicinal properties of plants has again come to the forefront of interest of the public. Because of their therapeutic properties, these plants are commonly used in alternative treatments for minor ailments. This thesis deals with the determination of polyphenolic acids in extracts from aronia, echinacea, tetradenia and centella. One of the aims of the thesis was to compare the amount of polyphenolic acids in individual plants with their antioxidant properties. The theoretical part includes a general introduction to flavonoids, introduction to polyphenolic acids, introduction to the principles of capillary zone electrophoresis and a summary of selected foreign studies for the determination of flavonoids. An important chapter of the theoretical part is a description of individual plants with an explanation of their possible therapeutic use. The experimental part is devoted to the process of optimization of electrophoretic determination of selected polyphenolic acids and evaluation of their content. It was found that in the extracts six representatives of polyphenolic acids occur in minimal amounts, often below the limit of quantification or detection of the method.
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Electrophoretic determination of saccharides in flower nectar
Hurychová, Nikola ; Křížek, Tomáš (advisor) ; Sobotníková, Jana (referee)
This work deals with the development and basic validation of a method for determination of sucrose, glucose and fructose content in floral nectar. It uses capillary zone electrophoresis with contactless conductivity detection. The theoretical part describes the functions of carbohydrates in nectar, principles of capillary zone electrophoresis and contactless conductivity detection. There is also a brief overview of applications in the determination of carbohydrates. In the experimental part, sample injection at the short and at the long end of capillary were tested. In further experiments, the background electrolyte was optimized. The tested concetrations of NaOH were from 10 to 50 mmol l-1 . Optimal conditions for the experiments were injection at the short end of the capillary, 40mmol l-1 NaOH as background electrolyte and separation voltage of 15 kV. Then, the repeatability, limit of detection and quantification and recovery of the method with a real sample of nectar were verified. Lactose was used as an internal standard. Relative standard deviation was below 2 %. Limits of detection were below 0.003 mg ml-1 and limits of quantification were below 0.009 mg ml-1 . Recovery of the method was 97-105 %. Furthermore, the calibration dependences were measured, from which the concentration of...
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Electrophoretic Determination of Tantalum in Recycled Coolants
Gogulin, Semen ; Křížek, Tomáš (advisor) ; Coufal, Pavel (referee)
This master thesis deals with the development of electrophoretic methods for determination of tantalum in recycled coolants used in the production of tantalum capacitors, as well as the separation of tantalum from chemically similar niobium. Developed methods are based on the formation of negatively charged complexes of tantalum and niobium with a number of ligands added to the separation electrolyte, especially tartaric and citric acid and 4-(2-pyridylazo)resorcinol. The separations were performed in a fused silica capillary of the inner diameter of 50 µm and the effective length of 36.5 cm, a negative voltage of -25 kV was applied to the capillary. Using a separation electrolyte containing 0.1 mol·l-1 of tartaric and citric acid at pH = 2, separation of niobium and tantalum was achieved in less than 3 minutes; the absorption spectrometric detection was carried out in the ultraviolet region at a wavelength of 200 nm, the detection limit was 2.8 ppm of tantalum, and the quantification limit was 9.2 ppm of tantalum. Addition of 4-(2-pyridylazo)resorcinol reduced the detection limit to 0.8 ppm of tantalum and the quantification limit to 2.6 ppm of tantalum; the electrolyte contained 0.1 mmol·l-1 of 4-(2-pyridylazo)resorcinol, 10 mmol·l-1 of tartaric acid, 10 mmol·l-1 of citric acid, and 25 mmol·l-1...
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Determination of proteinogenic amino acids by high-performance separation techniques
Hodek, Ondřej
(EN) Proteinogenic amino acids are key components of living organisms. Thus, the latest metabolomics research has focused on developing fast and sensitive methods for the determination of amino acids. In this context, this thesis contains two studies describing development of high-performance separation techniques for the quantification of amino acids. In the first study, a capillary electrophoresis method was developed for the determination of free amino acids in tobacco plants, particularly focusing on optimizing the extraction of amino acids from solid plant materials. The extraction procedure was optimized using design of experiments (DoE) to obtain the highest possible extraction yield of amino acids. Factors such as volume and concentration of the extraction solvent (hydrochloric acid) were assessed as the most significant. Subsequently, the optimal values of these factors were determined using response surface methodology (RSM). Lastly, proteinogenic amino acids were quantified using capillary electrophoresis with contactless conductivity detection and calibration with internal standard, which improved the precision of the method. The second study aimed at developing a supercritical fluid chromatography method for the determination of free proteinogenic amino acids in human plasma. The most...
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Determination of selected components in human urine with electrophoresis in short capillary.
Makrlíková, Anna
Capillary zone electrophoresis is frequently used in various analyses. In this diploma thesis a hydrodynamic sample introduction method controlled by pressure pulse has been proposed for short-capillary electrophoresis. The base electrolyte flushes sample from the loop of a six-way sampling valve and is carried to the injection end of the capillary. At the time when the sample zone reached the capillary, a short pressure impulse is generated in the electrolyte stream, which provides injection of the sample into the capillary. Then the electrolyte flow is stopped and the separation voltage is turned on. The amount of sample introduced to the capillary is controlled by the duration of the pressure pulse. This new sample introduction method was tested in the determination of ammonia, histidine, creatinine, uric acid and hippuric acid in human urine and for rapid screening of the contents of the inorganic ions in cerebrospinal fluid and blood plasma. The determination was performed in a capillary with an overall length of 10,5 cm and two base electrolytes was tested - 50 mM MES + 5 mM NaOH (pH 5,10) and 1 M acetic acid + 1,5 mM crown ether 18-crown-6 (pH 2,40). Using dual detection techniques contactless conductivity and UV spectrometric detection, anorganic and organic substances in the sample could...
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The application of high-efficiency separation methods for chiral and achiral separations
Šubová, Martina
In the first part of the doctoral thesis, the capillary electrophoresis was used to test the potential chiral separation properties of monosubstituted cyclodextrin derivatives, namely PEMEDA- and PEMPDA-β-cyclodextrins for the group of selected analytes. Both selectors exhibited excellent enantioseparation properties for N-boc-D,L-tryptophan, where the enantiomers were completely separated even at 0.5 mmol·l-1 concentration of the cyclodextrin derivative in the background electrolyte. However, the differences between the enantiodiscrimination properties of individual derivatives were minimal. The second test group consisted of two cyclodextrin derivatives, namely 2-O- and 3-O- cinnamyl-α-cyclodextrins. These derivatives are able to form supramolecular polymers in aqueous solutions that disintegrate at elevated temperature. The formation of these polymers was tested by NMR and DLS experiments. None of the tested cyclodextrin derivatives showed enantiodiscrimination properties towards a group of selected analytes. In the frame of antipredatory study, HPLC-MS/MS method working in HILIC mode was used for separation of ten pterin derivatives and riboflavin, which can be present as pigments in insects, reptiles or amphibians as a part of their warning coloration. The developed methodology was applied for...
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Electrophoretic determination of rivaroxaban
Petrák, Ondřej ; Křížek, Tomáš (advisor) ; Kubíčková, Anna (referee)
Capillary electrophoresis is a method used in pharmaceutical analysis because of its low cost, speed and environmental friendliness. This diploma thesis deals with development of electrophoretic method for rivaroxaban determination. After several optimizations, a method suitable for rivaroxaban determination inside its dosage forms was developed. Optimizations involved change of background electrolyte's composition from aqueous solution of low molecular weight organic acids to non-aqueous solution of acetic acid and cetyltrimethylammoniumbromide in acetonitrile in multiple steps. Final conditions of analysis included background electrolyte composed of 1M acetic acid and 40mM cetyltrimethylammoniumbromide in acetonitrile, sample injection carried out hydrodynamically by pressure of 5,0 kPa for period of 3 s and subsequent insertion of separation voltage of 30,0 kV for entire duration of analysis. Capillary content was mobilized by pressure of 0,50 kPa for entire duration of analysis. With aforementioned optimizations a selective method for determination of dosage forms of rivaroxaban was obtained. This method provides limit of detection 0,0056 mg/ml and limit of quantification 0,019 mg/ml and is linear in 0,01 - 0,40 mg/ml range with a recovery of 93,2 %. Keywords capillary electrophoresis,...
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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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