Polychlorinated biphenyls (PCB) represent relevant persistent organopollutants of the environment and the estimated amount of PCB released into the environment is 750000 metric tons. White-rot fungi have been studied for long time due to their degradative potential toward various aromatic pollutants and it is known that these fungi are able to decompose PCB in vivo. Biodegradation of PCB by the fungus Pleurotus ostreatus was studied in the frame of this work. A high degradative efficiency of P. ostreatus was observed in the first set of experiments, even in the presence of relative high amount of added PCB. Fungus was able to transform 780±50 µg out of the intial amount 1000 µg in 20 ml of a cultivation media within 42 days. A decrease in toxicity was recorded during the degradation that suggests the suitability of this organism for a practical use in decontamination. In vitro experiments with purified laccase induced with Cu2+ from this fungus did not prove any participation of the enzyme in the first step of PCB transformation. The enzyme did not show an ability to degrade PCB even after purification from cultivation media containing PCB. It was found that the first step of PCB transformation can be performed by an intracellular process with microsomal fraction. A degradation of 44-67% was observed for...

Chlortoluron is a herbicide from the group of substituted ureas and it is used for weed control in production of cereals and poppy. Substituted ureas are widely used but these substances are known for their high mobility in soil, persistence and contamination of underground water. Therefore, the development of analytical methods suitable for residual concentrations determination is necessary. This thesis deals with the determination of chlortoluron using carbon paste electrode (CPE) by the methods of differential pulse voltammetry and high-performance liquid chromatography with electrochemical (HPLC-ED) and spectrophotometric (HPLC-UV) detection. The aim of the work was to find optimal conditions suitable for chlortoluron determination. Further, the possibility of using miniaturized carbon paste electrode (mCPE) and usability of tested methods for determination of chlortoluron in model samples of river water and soil were studied. Concentration dependences in deionized water were measured by the method of DPV in the Britton-Robinson (BR) buffer (pH = 3) and methanol (10 %, v/v). The limit of detection was 3,7∙10-7 mol∙dm-3 in the case of CPE and 8,7∙10-8 mol∙dm-3 in the case of mCPE. Also, concentration dependences were measured using HPLC with mobile phase containing BR buffer (pH = 4) and...

South American plant Clusia blattophila, growing on Nouraghes of French Guiana, produces relatively large amounts of volatile substances in the fragrance of its flowers. C. blattophila is dioecious plant with different flowering time of monosex flowers Male flowers bloom for one night, while female flowers bloom two nights. The volatiles were adsorbed to the adsorbent and washed out with hexane with internal standard. Analysis was performed by two-dimensional gas chromatography with mass spectrometric detection. The aim of my thesis was to determine the qualitative and quantitative characteristics of flower scents of both sexes. I also studied, whether there are differences in production of the major fragrance component, acetoin, during the first night of blooming. The flower fragrance is complex blend of volatiles. In total, 43 different chemicals were identified. Among them were monoterpenes (α-pinene, camphene, β-pinene, myrcene, limonene, (E)-β-ocimene, camphor) sesquiterpenes (α-copaene, β-caryophyllene, cis-α-bergamotene, trans-α-bergamotene, aromadendrene, (E,E)-α-farnesene, δ-cadinene), aldehydes (1-pentanal, 1-hexanal, 1-octanal, n-decanal), also four ketones (acetoin and its dimer, 2,5-hexanedione, 6-methylhept-5-en-2-one), alcohol butane-2,3-diol, carboxylic acid 2-methylpropanoic,...

Males of South American fruit fly Anastrepha fraterculus form leks and release sex pheromone to attract females during the reproductive behavior. The aim of my diploma thesis was to collect samples of volatiles released by variously old males, subsequently to analyze and to determine the changes in the quantitative composition of the sex pheromone depending on age. The volatiles were trapped on adsorbent, eluted with hexane containing internal standard and analyzed by gas chromatography with mass detection. Males release twenty volatiles, including fourteen terpenes: α-pinene, camphene, -pinene, myrcene, Δ3-carene, limonene, (Z)--ocimene, (E)--ocimene, aromadendrene, trans-α-bergamotene, (Z)-β-farnesene, (Z,E)-α-farnesene, germacrene D, (E,E)-α-farnesene; one aldehyde: nonanal; two alcohols: (3Z)-non-3-en-1-ol, (3Z,6Z)-nona-3,6-dien-1-ol and three lactones: suspensolide, anastrephin a epianastrephin. From the list of named compounds there were proved antenal activity of six compounds in previous work, namely trans-α-bergamotene, (Z,E)-α-farnesene, (E,E)-α-farnesene, (3Z)-non-3-en-1-ol, (3Z,6Z)-nona-3,6-dien-1-ol and epianastrephin.

Analytical chemistry plays a crucial role in studies on chemical ecology and only the development of sophisticated methods enables the detection of biologically active compounds that are usually present in minute quantities and often in very complex mixtures. My thesis is dedicated to the application of modern analytical techniques and instrumentation to unravel the identity, chemical diversity and function of semiochemicals and defensive compounds used by various species of termites. The first section of this thesis aims at the identity of chemicals used in communication, the pheromones. I studied the chemistry of the trail-following communication in three selected species of termites. Besides the identification of (3Z,6Z,8E)-dodeca-3,6,8-trien-1-ol, the most frequent termite trail-following pheromone, as the trail-following pheromone in Psammotermes hybostoma (Rhinotermitidae), I participated in the description of two new structures, (10Z,13Z)-nonadeca-10,13-dien-2-one in Glossotermes oculatus (Serritermitidae) and syn-4,6-dimethylundecan-1-ol in Hodotermopsis sjoestedti (Archotermopsidae). We identified (3Z,6Z,8E)-dodeca-3,6,8-trien-1-ol to be the female sex pheromone in Psammotermes hybostoma, and syn-4,6-dimethylundecanal to be the male sex pheromone in Hodotermopsis sjoestedti. I also...

Lipidomics as a part of metabolomics is a fast-growing area of research due to the improvement in analytical techniques. This master thesis is focused on lipid extraction techniques optimization, using liquid liquid extraction and solid phase extraction as pre-separation methods and ultra performance liquid chromatography coupled with mass spectrometry for extraction and subsequent identification of branched-chain fatty acid esters (FAHFA - branched-chain Fatty Acid esters of Hydroxy Fatty Acids). This newly discovered class of lipid molecules is associated with insulin secretion, which could improve whole body and local glucose metabolism, providing potential for diabetes 2 type treatment. Solid phase extraction of biological samples was optimized on columns regarding to sorbent composition using reversed phase columns with modified styrene divinylbenzene polymer or octadecyl-bonded polymer and normal phase columns packed with silica gel. Column Strata SI-1 Silica was the most effective for FAHFA separation from biological samples. Chromatographic separation of FAHFA was performed on UPLC Ultimate 3000 RSLC equipped with Kinetex C18 1,7 µm, 2,1 x 150 mm column using gradient program. UPLC was coupled to QTRAP 5500/SelexION, a hybrid, triple quadrupole, linear ion trap mass spectrometer equipped...

(EN) This dissertation thesis is focused on physicochemical and chromatographic characterization of polyaniline-coated stationary phases. In the first part, surfaces of bare silica and octadecyl silica sorbents were modified by in-situ chemical polymerization of aniline hydrochloride and their subsequent systematic characterization was performed by using the linear solvation energy relationship approach in the HILIC mode of capillary LC. In addition, several common physicochemical techniques were used to characterize properties of these altered materials. The modified sorbents were then packed into capillary columns. The retention interactions taking place between solute and the separation system were evaluated on the basis of retention data of a number of various solutes. The results showed that polyaniline coating had a significant effect on the retention promoting interactions of both polyaniline-coated stationary phases. The assumed mixed-mode retention mechanism was proven for both the stationary phases. The second part dealt with investigation of the separation potential of polyaniline- coated silica stationary phase in different chromatographic modes. The retention factor curves of structurally similar solutes were constructed as a function of organic modifier portion in the mobile phase....

(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...

Scaffolds are proteins with high conformational stability, allowing us to implement multiple mutations into specific parts of the protein. Even with these mutations, the structural integrity of the protein is maintained as well as its physical-chemical properties. These mutations give the specific scaffold new properties. In most cases it is the binding specificity towards previously chosen target. The biggest advantages of scaffolds are their small size, stability, low-cost manufacturing, and easiness of preparation. Scaffold utilized in this thesis is unique for having two binging surfaces designed on which it can be mutated. Each of those two surfaces can be separately mutated to develop a binging site for two different proteins. In our case these mutations led to binding two nonidentical receptors of a human cytokine. Mutations are made with a use of yeast display, one of the methods of directed evolution. The main focus of this thesis is changing an expression system of the binding proteins from the yeast system to a bacterial one, their production and purification followed by characterization of those binding proteins using biophysical methods. These methods were used to evaluate structural and thermal stability, and binding affinity to both receptors of the beforementioned binding proteins....

Previously, selenium was known as an element with negative properties. However, in the last century, the significant positive effects on human health were detected. Currently, the function, behavior and toxicity of selenium are still not well known. The key to understand it is to do speciation analysis. The aim of this diploma thesis is to develop method for determination inorganic (sodium selenite and selenate) and organic (selenourea, selenocystine, selenomethionine) selenium compounds. Parameters of apparatus for electrochemical hydride generation with atomic absorption spectrometry detector were optimized, final conditions were set and optimal conditions for separation process using HPLC were investigated. Calibration dependences for selenium compounds were measured and analytical figures of merit were investigated. In conclusion, a coupled method HPLC- EcHG- QFAAS for determination of individual selenium compounds was proposed, and it was tested on urea samples. Calibrations for these measurements were investigated and analytic characteristics were calculated. Based on these comparisons it can be said that proposed method allows the determination of selected selenium compounds in both aqueous and urea matrices. Subject words: Spectroscopy, analytical chemistry Key words: Atomic absorption...