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Pharmaceutical application of high performance ion chromatography in analysis of organic anions and cations
Čujová, Sabína ; Čabala, Radomír (referee) ; Pacáková, Věra (advisor)
The thesis is focused on application of ion chromatography in pharmaceutical analyses of organic ions. Ion chromatography is increasingly used in the field of pharmaceutical analysis. This includes the analysis of impurities and metabolites. In the first part of this thesis, ion chromatography is compared with common separation techniques used in pharmacy, such as gas chromatography and high performance liquid chromatography. In the second part development and validation of methods of ion chromatography for purity evaluation and quality control of active pharmaceutical substances Rivastigmine hemitartrate and Pramipexole hydrochloride were carried out. Key words: ion chromatography, reversed-phase chromatography, ion-pair chromatography, ion-exclusion chromatography, ion-exchange chromatography, GC, HPLC
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Optimization of conditions for derivatization of fatty acids and perfluorinated organic acids with methylchloroformate
Hložek, Tomáš ; Feltl, Ladislav (referee) ; Čabala, Radomír (advisor)
Statistical methods designed for optimization were applied for selection of optimal experimental conditions of derivatization process for perfluorinated acids, PFA (C8, C10 ,C12), fatty acids, FA (C8, C12, C14) and their mixtures. The calibration curves and the extraction efficiences for system acetonitrile/heptane were measured for all studied acids. The Plackett-Burman screening was performed for obtaining the significant experimental parameters from their original series and significant ones were remitted to optimization via central composite design, which takes a second order effects and mutual interactions in between parameters into account. Optimal conditions of derivatization were found for PFA, FA and their mixtures. The principle of the search of optimal selective derivatization conditions for one group of acids in the presence of the other is based on the search of optimum on the response surface where the dependent variable is either sum or difference of acid group responses normalized to common internal standard. It was found that FA influence total response more than PFA and that in the case of demand for selective derivatization of PFA in the presence of FA in the sample, the most significant parameter is the volume of methylchloroformate added. The complete selectivity of...
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Optimization of experimental conditions for measurements of perfluorinated organic acids by GC
Ston, Martin ; Coufal, Pavel (referee) ; Čabala, Radomír (advisor)
Commercial software Pro ezGC was used for calculation of optimal temperature program for analysis of isobutylesters of perfluorinated organic acids (C6 - C12) by gas chromatography with flame ionization detector. Optimization was done with respect to the complete resolution of all peaks of analytes. The calculated temperature program was also applied to the analysis of substances of interest on GC-MS. The calculated and measured values have been compared. The measured resolutions were higher than the calculated ones in all cases. Keywords: perfluorinated organic acids, isobutylesters, peak resolution Subject words: gas chromatography, numerical optimization
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Epoxide hydrolases expressed from environmental DNA: characteristics of soluble and immobilized enzyme forms
Grulich, Michal ; Bezouška, Karel (advisor) ; Čabala, Radomír (referee)
8 Abstract Epoxide hydrolases (EHs) demonstrating high degree of enantioselectivity or enantioconvergence are useful biocatalysts for the production of optically active epoxides and vicinal diols, which can serve as chiral building blocks for syntheses of biologically active drugs. EHs can play an important role also in degradations of xenobiotics. Genes encoding EHs Kau2 and Kau8 were expressed in E. coli host strains TOP10 and RE3. Enantioselectivities and regioselectivities of Kau2 and Kau8 in supernatants of desintegrated cells were determined for four substrates: tert-butylglycidyl ether, para-chlorostyrene oxide, para-nitrostyrene oxide, α-methylstyrene oxide. The highest values of enantioselectivity and regioselectivity were achieved with Kau2 and para-nitrostyrene oxide as a substrate. The Kau2 was chosen for further experiments on the basis of these results. Kau2 was overexpressed in the recombinant strain RE3(pSEKau2). We performed two batch cultures and one fed-batch culture in stirred bioreactor. The highest volumetric activity of 4500 U/l was obtained in the case of fed-batch culture. Two phase system consisting of polyethylenglycole 6000 and sodium citrate (pH 7.7) was used for Kau2 purification from the supernatant of desintegrated cells. Purification factor 2.6 +/- 0.3 was achieved and...
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Toxicity of Organic Mixtures
Reitmajer, Josef ; Feltl, Ladislav (advisor) ; Čabala, Radomír (referee) ; Ventura, Karel (referee) ; Mejstřík, Viktor (referee)
6. CONCLUSION The aim of this study was to test additiviý of chemical compounds in distribution process between two phases and to compare distribution coefficients K-;^ for different molar ratio of mixture with acute toxicity index EC50 gained on oligochaeta Tubifex tubifex. Mixtures of organic solvents ethanol, benzene, aniline and nitrobenzene were studied. Deviations from additivity were found for distribution coefficients of all tested mixtures. The most significant deviation exhibited benzene - ethanol mixture. The lowest deviations were observed for mixture ethanol - aniline. The results obtained indicate that the partition coefficient K.;* behave similarly as acute toxiciý. Definitely it is for benzene - ethanol mixture. It is possible to suppose that this parameter might be used for estimation of changes in acute toxicity of mixtures. The aim of the second part of the thesis was to study how distribution of chemicals changes with concentration and composition of mixture. Variations in distribution of mixture components beťween gaseous and aqueous phase were found. The highest deviations were found for benzene - ethanol and benzene - eniline mixtures. No deviation was found for mixture aniline - nitrobenzene. Deviations were not as significant as in first part of study. Dependences of distribution...
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