Capillary electrophoresis often uses complexing agents since the interaction between the analyte and the complexing agent can result in achieving or improving the separation. Examples of such methods can be electrokinetic chromatography or affinity capillary electrophoresis (ACE). ACE is used to determine the complexing parameters. In case of chiral separation, this issue gets complicated, since the parameters of the two analytes (enantiomers) are not completely independent to one another. Therefore, a procedure has been proposed in this thesis, that should always be used to evaluate the complexing parameters of two enantiomers. Statistical evaluation of these parameters was assessed as well. This work also proposes a method that allows to determine the relative migration order of two enantiomers in two different complexing separation systems. The mathematical description of electrophoresis is based on continuity equations, that are inherently nonlinear. However, these equations can be linearized to obtain an approximate analytical solution. There was recently presented a generalized model, that enables inclusion of complete complexing equilibria in the theoretical description of electromigration. Thus, various phenomena, including nonlinear ones, associated with complexation can be predicted. This...

This diploma thesis deals with the determination of creatinine using a combination of flow injection analysis (FIA) or high-performance liquid chromatography (HPLC) with pulse amperometry, an electrochemical technique based on the application of potential pulses on a gold working electrode. The determination was performed in a basic environment of borate buffer with creatinine concentration of 1∙10-4 mol∙l-1 . The lenght of the cleaning and activation pulse was optimized as well as the pH of the running buffer. A cleaning pulse of +1.8 V was first applied to the electrode for 100 ms, then an activation potential of -0.5 V was applied for 150 ms and then a measuring potential of +0.2 V for 300 ms. The optimal pH was selected as pH=9,4. Methanol and acetonitrile were added to the borate buffer to test whether creatinine could be determined in presence of these organic solvents and whether flow injection analysis could be transformed into HPLC. Methanol in the system caused peak deformation, acetonitrile did not cause the peak deformation in the system, at higher contents the baseline was destabilized. Furthermore, the calibration dependence in the range of concentrations from 2.5∙10-4 mol∙l-1 to 5∙10-6 mol ∙ l-1 was measured using PAD in combination with FIA. At higher concentrations, peaks splitted....

Charles University Faculty of Pharmacy in Hradec Králové Department of Biophysics and Physical Chemistry Candidate: Kamila Kynclová Supervisor of Diploma Thesis: Mgr. Monika Kuchařová, Ph.D. Title of Diploma Thesis: Determination of fatty acids levels in selected human tissues Fatty acid are an essential part of many metabolic processes in the body. The theoretical part of the the thesis presents primarily omega-3 and omega-6 fatty acids, their general characteristics, metabolism and their effect on various disease. The roles of fatty acid in cancer, neuropsychiatric and inflammatory diseases are described here. The thesis also mentions the method of gas chromatography, which was used to determine the levels of fatty acids. The experimental part of the thesis includes method, chromatographic analysis and results. 13 fatty acids in 7 different tissues were analyzed: subendocardial tissue, liver parenchymal tissue, kidney and adrenal tissue, skeletal muscle tissue, adipose and brain tissue. Tissues were collected from 20 healthy, suddenly deceased, cadavers. Donors were divided into two groups according to age. The obtained data were statistically processed and divided according to individual fatty acids. In the end the results between the two groups were evaluated with a possible explanation of the...

This diploma thesis is based on the current issue of the presence of pharmaceuticals in various components of the environment. Concerning the contamination by residues of pharmaceuticals the most affected environment is the aquatic environment where these substances leaks especially from wastewater treatment plants, which eliminate them during the cleaning process only partially. This work is focused on the selected group of pharmaceuticals, non-steroidal anti-inflammatory drugs (salicylic acid, ibuprofen, caffeine, naproxen, ketoprofen, diclofenac) in waste water. For analysis purposes two types of sampling were used and compared: the conventional spot sampling of wastewater and the sampling using passive samplers POCIS. The sampling took place at the inflow and outflow of the wastewater treatment plant in Brno Modřice. The solid phase extraction (SPE) using Oasis HLB columns was used as the extraction method. Extracted sample was derivatized then. Derivatization agents were: MSTFA (N-methyl-N-(trimethylsilyl)trifluoroacetamid) and BSTFA (N, O-bis(trimethylsilyl) trifluoroacetamid). The final analysis was performed using gas chromatography with mass spectrometric detection Time-of-Flight (GC/TOF-MS).

Master’s thesis deals with a determination problem of ignitable liquids from fire debris. The aim of this work is to introduce the properties of used fire accelerants and to give an overview and evaluation of the various techniques which can be conducive to the fire investigator. Determination of fire accelerants from fire debris was made by the technique of solid phase microextraction (SPME) with subsequent chemical analysis by GC/MS. Based on the chromatographic results were established the target compounds and reconstructed ion chromatograms which are typical for some kinds of flammable liquids. There were used gasoline, diesel, kerosene and technical gasoline (white spirit) as the fire accelerants. This work also deals with the influence of interfering products in fire debris analysis, including their identification and characterization. Different kinds of substrates were burned, extracted and analyzed in order to identify all the interfering products that they may release.

Transport phenomenon of gas and vapors through polymer barrier materials as well as the factors affecting permeability of gaseous and liquid toxic compounds were investigated and results are presented in this thesis. Permeation characteristics investigation methods were elaborated and verified with focusing on utilization of gas chromatographic method and its instrumental modifications.

5 Abstract Genetic mechanisms of regulation of gene expression form the basis for proper development, function of organisms and their responses to variable life conditions. However, they are relatively slow. Life processes that require a fast response to the changing environmental and metabolic conditions are mostly executed on the level of proteins especially their posttranslational modifications and protein- protein interactions.The goal of the experimental work that led to the presented thesis consisted in exploitation of the model organism Caenorhabditis elegans for analysis of regulation of gene expression by transcription factors from the protein family of nuclear receptors. The model system C. elegans enables very efficient experimental procedures in the field of genetics, genomics and functional analysis of phenotypes. In the experimental work connected with this thesis, I studied the regulation of gene expression under specific experimental conditions from the perspective of advanced functional proteomics and I focused on the employment of separation methods and methods of advanced proteomics, especially by mass spectrometry.In the first part of the work, I characterized the nuclear receptor NHR-60 on the protein level. This nuclear receptor is expressed as two protein forms with a mass of 50 kDa...

Backgroun and aims: Pharmacokinetic variability is of paramount importance for sucessfull pharmacotherapy. The main purpose of this work was to study variability of pharmacokinetics in clinical and non-clinical setting with the aim to predict variability in target population. Specifically, three drugs were chosen, sufentanil, with relativelly narrow therapeutic index, and nabumeton and abirateron, both with known high variability. Methods: The study of pharmacokinetic variability of sufentanil was based on clinical samples taken from patients undergoing surgical cardiac procedure, where the sufentanil was used as a part of the drug coctail used during the procedure. New analytical method was necessary to prepare and validate to measure sufentanil concentrations and obtain pharmacokinetic parameters. These were compared between determined genotype groups of MDR1 and OPRM1. Similarly, clinical study was executed with nabumetone, in which nabumetone was administered in a group of 24 subjects on two separate occassions. Plazma samples were obtained and concentrations of nabumetone and its active metabolite, 6-methoxynaphtylacetic acid (6-MNA), were determined. Obtained pharmacokinetic profiles were compared between female and male volunteers, and genotypes for MDR1 and CYP2D6. Finaly for abiraterone,...

The Ceratitis capitata is a very important agricultural pest, whose reproduction behaviour is controled by chemical signals. Males initiate mating by creating leks, where they release sexual pheromones to attract females. The main goal of this diploma thesis was to determine the influence of host plants on the composition of male sex-pheromones C. capitata and to compare emanations of wild males with those originating from laboratory population. We studied the chemical composition of volatiles, released by calling males C. capitata from laboratory and two wild populations, using two-dimensional gas chromatography with time-of-flight mass spectrometric detection (GC×GC-TOFMS), gas chromatography with electroantennographic and flame ionization detection (GC-EAD-FID). All data were statistically analyzed by multivariate data analyses. Significant differences were observed in the quantitative and qualitative composition of the chemical emanations among males from the three populations. The GC-EAD-FID analyses revealed fourteen antenally active compounds with a possible behavioral function. Isomenthone, geraniol, bornyl acetate, geranyl acetone and ethyl octanoate were newly identified antenally active compounds of C. capitata male sex pheromone. Statistical analyses indicated that males and females of...

This diploma thesis deals with the optimization of processes for extraction of bioactive lipophilic compounds from fruits of sea buckthorn (Hippophae Rhamnoides) into various types of plant oil bases using simple maceration. The theoretical part of this thesis deals with the characterization of this herb, its botanical classification, traditional use, chemical composition and medicinal effects. Increased attention is also paid to the characterization of individual types of plant oils used, namely camellia, camellia organic, passionflower, kukui and kiwi oil. The experimental part of the thesis deals with application of theoretical knowledge. The profile of total and free fatty acids for individual plant oil bases was determined by GC/FID, furthet the basic fat numbers were also determined. When optimizing the extraction, emphasis was placed not only on the effect of the extraction agent used, but also on the extraction time (1, 3, 5, 7, 10, 14, 21 and 66 days). The macerates were continuously subjected to the determination of selected parameters (total amount of carotenoids, total amount of phytosterols, lutein, neoxanthin, astaxanthin, stigmasterol, -sitosterol and vitamin E) using UV-VIS spectroscopy and HPLC/PDA. Likewise, the peroxide number was monitored during maceration to assess the degree of oxidative degradation of macerates. The recovery of selected total parameters in individual oils did not differ significantly in most cases. On the contrary, the yield of individual monitored parameters differed significantly. At the same time, static maceration under our conditions was not very suitable for the extraction of vitamin E, stigmasterol and total phytosterols.