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Chiral separation of acyclic nucleoside phosphonates and determination of association constants of their complexes with cyclodextrins by capillary electrophoresis
Mikysková, Hana ; Jelínek, Ivan (referee) ; Coufal, Pavel (advisor)
Capillary electrophoresis (CE) has been applied to the chiral separation of acyclic nucleoside phosphonates, which belong toof the most important already registered or potential antiviral drugs. For enantioseparation of acyclic nucleoside phosphonates, the following native and derivatized cyclodextrin were used as chiral selectors: cyclodextrin, cyclodextrin, cyclodextrin, heptakis(2,6dimethyl) cyclodextrin, 2hydroxypropylcyclodextrin and quaternary ammonium cyclodextrine). The best resolution of (R,S)enantiomers of acyclic nucleoside phosphonates was achieved in basic backgroung electrolyte composed of 35 mM sodium tetraborate; pH 10, with addition of 17.6 mM cyclodextrin or 10 mM quaternary ammonium cyclodextrin chiral selectors. Capillary electrophoresis has been employed for determination of association constants of complexes of acyclic nucleoside phosphonate enantiomers with above mentioned chiral selectors. The association constants were determined by nonlinear regression analysis from the experimentally measured dependence of effective electrophoretic mobilities of the analytes on concentration of the chiral selector in the background electrolyte. Prior to association constant calculation, the effective electrophoretic mobilities were corrected to reference temperature,...
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Cytokine estimation in liver homogenate supernatant in rats with liver steatosis
Burešová Nedvídková, Jaroslava ; Živný, Pavel (referee) ; Dršata, Jaroslav (advisor)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Bc. Jaroslava Burešová Nedvídková Supervisor: Doc. MUDr. Pavel Živný, CSc. Title of diploma thesis: Cytokine estimation in liver homogenate supernatant in rats with liver steatosis The aim of this work was the determination of cytokines - interleukin 6, tumor necrosis factor-alpha and leptin levels in liver homogenate supernatant in rats with hepatic steatosis. These tests were performed within extensive experimental work intented on the ability of liver regeneration after induction of steatosis. The determination was performed using enzyme-linked immunosorbent assay. The results were compared with cytokine concentrations of Wistar and PHHP control groups, who were fed a standard laboratory diet. The concentrations of IL-6, leptin and TNF-alpha in liver tissue supernatant were higher in the control group of Wistar rats than in PHHP rats control group. Receiving steatosis diets led to a decrease in IL-6 for all rat files. Leptin concentrations were significant lower in Wistar rats than Wistar rats controls were. Orotic acid administration statistically significant increased leptin level in rats PHHP. The consumption of diets causing steatosis reduced TNF-alpha concentrations in liver...
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Analytical evaluation of drugs using chromatographic methods V
Raindlová, Martina ; Sochor, Jaroslav (referee) ; Mokrý, Milan (advisor)
Charles University in Prague, Pharmaceutical Faculty in Hradec Králové Department of pharmaceutical chemistry and drug control Analytical drug evaluation by chromatographical methods V Diploma paper Martina Raindlová In this diploma paper optimal conditions for HPLC analysis of risperidon in tablets were elaborated. One part of the task was finding optimal mobile phase and inner standard, other part was validation of the method and its application in analysis of remedy in pharmaceutical preparation. For the mobile phase a mixture was used in composition : (NH4)2HPO4 (0,1 mol/l ; pH 6,0 - adjusted by H3PO4 0,1 mol/l) : methanol 30:70 (v/v) and with flow rate 0,7 ml/min. LiChroCART 250-4 column with bonded reversed phase C 18 and average particle size of 5 μm. Detection was performed by UV detector at 280 nm wave lenght. Fenacetine was used as an inner standard. Validation of method was confirmed by linearity and precision. For linearity a calibration curve was used, which was constructed from results of five solutions of increasing concentration. Its parameter is equation : y = 4,38x + 0,191. Correlation coefficient is 0,9998. Method is linear. Quantification of risperidone in tablets was determined by HPLC analysis. Declared amount in tablets was 2 mg, real amount was 1,9885 mg/ml which corresponds...
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Determination of propyl gallate on carbon paste electrode
Vysoká, Marie ; Fischer, Jan (referee) ; Zima, Jiří (advisor)
Propyl gallate (PG) is a significant synthetic antioxidant and preservative. Its determination has been studied at carbon paste electrode (CPE) using differential pulse voltammetry (DPV) and high performance liquid chromatography with electrochemical detection (HPLC-ED) and with UV spectrometric detection (HPLC-UV). Concentration dependences were measured in the media of Britton-Robinson buffer (pH 5) and methanol (20 %, v/v) by DPV and the limit of detection 0,6110-7 moldm-3 was obtained. Using HPLC with a mobile phase consisting of 0,01 moldm-3 phosphate buffer (pH 4) and methanol (50 %, v/v) with potential of working electrode E = +0,8 V and detection wavelength λ = 280 nm, concentration dependences were measured. Limit of detection was determined to 0,39 moldm-3 for HPLC-ED and 4,95 moldm-3 for HPLC-UV. After verification of the extraction procedure PG was determined in vegetable oil. The resulting value of 3,2 mgkg-1 corresponds with permited limits.
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HPLC Method Development and Validation for Determination of Active Substances and Impurity in Valetol Preparation
Maroušková, Alena ; Sklenářová, Hana (referee) ; Šatínský, Dalibor (advisor)
This work extends the graduation thesis which found optimal chromatographic conditions for a separation of paracetamol, caffeine and propyphenazon, and the impurity p-aminophenol in the pharmaceutical preparation Valetol by HPLC method. At first, the benzoic acid was found as a suitable internal standard for the quantification. Next, the analytical method validation was carried out, including the suitability test of the chromatographic system. Within the suitability test of the chromatographic system the following attributes were tested: column efficiency expressed as a number of theoretic plates N assymetry of chromatographic peaks expressed as factor T resolution of chromatographic peaks reproducibility expressed as a relative standard error Within the analytic method validation were tested: precision linearity accuracy selectivity robustness stability The robustness test confirmed the analysis sensitivity to the change in the character of the mobil phase. As a part of the robustness test two parameters of mobile phase were tested: pH and triethylamine concentration changes. It was confirmed that the change of pH of the mobile phase influences mostly the retention times of p-aminophenol and benzoic acid. The change of triethylamine amount in the mobile phase influences mostly...
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HPLC Method Development for Determination of Vitamin E in Geladrink Preparation
Švestková, Petra ; Chocholouš, Petr (referee) ; Šatínský, Dalibor (advisor)
A HPLC method was developed for the determination of content of vitamine E acetate in preparation Geladrink Forte pulverized drink - pineapple. The method is based on using Chromolith Performance RP-18e column (100 x 4.6 mm) and UV detection at 290 nm. The vitamine E acetate was separated in the presence of internal standard tocol from ballast matrix. It was used isocratic elution of the mobile phase methanol:water (98:2) at a flow-rate of 1.5 ml/min. There was temperature 30 řC during the measurement. The system enabled successful separation in time less than 5 min. The retention time of tocol was 2.337 min and the retention time of vitamine E acetate was 4.271 min. The chromatographic resolution between tocol and vitamine E acetate was 7.515. The method was applied to analysis of vitamine E acetate in preparation Chondrotin - MSM 2600. Developed concentrations of vitamine E acetate were re-count on content of vitamine E that the producer declared. Keywords: vitamine E acetate, HPLC, Geladrink, Chondrotin - MSM 2600
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Evaluation of an Influence of Electrolytes on CMC by Ion-Selective Electrode
Syrovátková, Jana ; Kubíček, Vladimír (advisor) ; Šnejdrová, Eva (referee)
In the rigorous thesis an ion-selective electrode has been utilized to measure CMC of five cationic tensides and to evaluation of an influence of electrolytes on CMC values. Commonly supplied cationic tensides have been used, i. e. benzalkonium chloride, benzethonium chloride, cetyltrimethylamonium bromide, cetylpyridinium chloride and carbethopendecinium bromide. Sodium chloride has been chosen as a suitable electrolyte for increasing ionic strength of the tenside solutions. Repeatability of the CMC determinations has been evaluated and the measured dependences of the CMC values on the concentration (ionic strength) of sodium chloride have been mathematically correlated.
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In vitro cultures of medicinal of plants - VII
Machačková, Jana ; Martin, Jan (referee) ; Tůmová, Lenka (advisor)
In vitro cultures of medicinal of plants - VII Elicitation is one of the methods that can be used to increase production or accumulation of secondary metabolites in explants cultures. There were used callus and suspensions cultures of Silybum marianum (L.) Gaertn. Derivate N-phenylpyrazin-2-carboxamid was used as abiotic elicitor in concentrations c1=1.159x10-3 mol/l, c2=1.159x10-4 mol/l and c3=1.159x10- 5 mol/l. The effect of elicitors on the production (flavonolignans of silymarin complex and flavonoid of taxifolin) was evaluated after 6, 12, 24, 48, 72 and 168 hours. Cultures were cultivated on Murashige and Skoog medium with addition 10 mg/ml α-naphtylacetic acid (α- NAA). HPLC method for analysis was used. The maximal content of flavonolignans was reached after elicitor application in concentration c2 after 168 h in callus culture (0,08%). However the highest content of silydianin was detected after application of elicitor in concentration c1 (0.11 %) in suspensions culture. The maximal content of flavonoid taxifolin was noted in of nutritient medium of suspensions cultures.
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Application of sequential extraction procedure for determination of antimony speciation in contaminated soils
Tejnecký, Václav
The city of Příbram (Czech Republic) is located in a traditionally industrial, mining and metallurgical district. During the hundred years of these activities, the area has been severely contaminated by toxic elements, including antimony (Sb). Generally, the acceptable Sb concentration limit in soils is supposed to be 5 mg.kg-1 . In the Příbram area, the Sb concentrations in soils are exceeding more than 200x this limit. The main source of Sb in the studied environment is related to emissions from primary and secondary Pb smelting. This study is focused on application of sequential extraction procedure (initially used by Wenzel et al., 2001 for As speciation) to forest and tilled soil profiles heavily polluted by Sb and located in the vicinity of the Příbram smelter (1 km). Maximum concentrations were 1720 mg Sb.kg-1 and 467 mg As.kg-1 in horizon B of forest soil. In the tilled soil, significantly lower concentrations of Sb and As were determined (100 mg Sb.kg-1 and 170 mg As.kg-1 in horizons A and B, respectively). These results can be related to higher interception of smelter emissions by tree canopies in the forest system and/or to intense agricultural practice in tilled soils (tillage, biomass loss,...). In the tissues of plants growing on these contaminated soils, the Sb concentrations reached...
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Possibilities of affecting of secondary metabolites production in culture Silybum marianum in vitro
Kunzová, Lucie ; Martin, Jan (referee) ; Tůmová, Lenka (advisor)
1 ABSTRAKT Mgr. Lucie Kunzová Possibilities of affecting of secondary metabolites production in culture Silybum marianum in vitro Elicitation is the method making use defensive mechanism of plants to increasing poduction of secondary metabolites in plants and cultures in vitro. The effect of 6, 12, 24, 48, 72 and 168 hours influence by three concentrations of the abiotic elicitor (3-jodo-4-methylfenyl)amide 5-methylpyrazine-2-karboxylic acid on the flavonolignans production in Silybum marianum callus and suspension culture was monitored in this study. The in vitro culture was cultivated on Murashige-Skoog medium with the addition of 10 mg/l of α-naphtylacetic acid as a growth regulator. The content of flavonolignans was determined by HPLC. The maximum content of flavonolignans (0,03 %) in callus culture was demostrated after 24 hours of elicitation (c1 = 2,83 · 10-3 mol/l). The maximum content of flavonolignans (0,04 %) in suspension culture was demonstrated after 48 hours of elicitation (c3 = 2,83 · 10-5 mol/l).
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