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Influence of plant biological protection on production of secondary metabolites of Papaver somniferum III.
Kaman, Petr ; Vytlačilová, Jitka (advisor) ; Sklenářová, Hana (referee)
Kaman, P.: Influence of plant biological protection on production of secondary metabolites of Papaver somniferum III. Diploma thesis. Charles University, Faculty of Pharmacy in Hradec Králové, Department of Pharmaceutical Botany, Hradec Králové, 2018, 77 pp. Diploma thesis was aimed to verify the impact of biological protection on the production of alkaloids of the opium poppy (Papaver somniferum, L.). Polyversum biofungicide was selected for this purpose, the active ingredient of which is oomycete Pythium oligandrum. The poppy tested were Orbis and Lazur high-morphine varieties. Extracts were prepared from the individual samples which were analyzed by HPLC. The following alkaloids were morphine, codeine, papaverine, noscapine and thebain. The average morphine content was around 0,5 %. The largest amount of 0,62 % was found in the poppyhead of untreated variant of the Orbis variety. Codeine for both varieties averaged 0,07 % in the poppyhead of untreated variant. Treated plants reached slightly lower values. The average papaverine content was around 0,01 % in the treated and also untreated variant, but only in poppyhead of the Lazur variety. Higher noscapine content was measured in the Lazur poppyheads, averaging 0,06 %. The content was only around 0,01 % for Orbis variety. The alkaloid content was...
Development of capillary electrophoresis method for the determination of three pharmacopoeial impurities in indomethacin
Doubková, Dagmar ; Jáč, Pavel (advisor) ; Sklenářová, Hana (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Analytical Chemistry Candidate: Mgr. Dagmar Doubková Supervisor: PharmDr. Pavel Jáč, Ph.D. Title of Thesis: Development of capillary electrophoresis method for the determination of three pharmacopoeial impurities in indomethacin. This thesis deals with the development of a micellar electrokinetic chromatography method for the determination of indomethacin impurities (4-chlorobenzoic acid, 5-methoxy-2-methyl-3-indolylacetic acid and 3,4-dichloroindomethacin). All separations were performed in a fused silica capillary with 50 µm id; total length 64.5 cm, effective length 56 cm and with extended light path (150 µm id) detection window. 1-naftylacetic acid (10 µg/ml) was used as an internal standard. The separations were carried out at 30 kV. The analytes were detected at the cathodic end of the capillary at 224 nm. A central composite design was applied for the optimization of the separation conditions. The effect of SDS concentration, content of methanol, concentration of phosphate buffer and buffer pH was studied at three levels. The optimized background electrolyte was: 20 mmol/L phosphate buffer (pH 7.57) containing 58 mmol/L SDS and 0% MeOH. The analysis time (with resolution of all compounds Rs ≥ 3.8) was shorter than 10...
Automation of toxicology tests
Žižková, Klára ; Sklenářová, Hana (advisor) ; Chocholouš, Petr (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Analytical Chemistry Candidate: Bc. Klára Žižková Supervisor: Doc. PharmDr. Hana Sklenářová, Ph.D. Title of Diploma Thesis: Automation of toxicology tests The aim of this diploma thesis was to find suitable method for determination of luciferase of the Ist and IInd types using the sequential injection method and optimize the assay conditions. The main aim was to automate analysis in a series of samples prepared for the testing of various substances, which, in particular, may affect the level of secreted luciferase (type II) upon the interaction of cells with tested substances in toxicological studies. The theoretical part describes luminescence, its types and applications. In addition, the types of luciferases and reagents with which a chemical reaction takes place are mentioned and principles of flow techniques are described. The review of luciferase determinations was carried out. Measurements were performed using the sequential injection method. Three instruments were used for the measurement. One flow injector analyser and one sequential injection analyser with two detection methods - batch-flow spiral detection cells were used. Measurement optimization was performed based on concentration and volume of samples and reagents,...
Evaluation of phenolic compounds content in fruits
Baďurová, Kristýna ; Sklenářová, Hana (advisor) ; Šatínský, Dalibor (referee)
Charles University Faculty of Pharmacy in Hradci Králové Department of Analytical Chemistry Candidate: Bc. Kristýna Bortlová Supervisor: Doc. PharmDr. Hana Sklenářová, Ph.D. Title of Diploma Thesis: Evaluation of phenolic compounds content in fruits This diploma thesis is focused on the analysis of selected phenolic compounds (gallic acid, chlorogenic acid, epicatechin, rutin, quercitrin, phloridzin and phloretin). The content of these phenolic substances was evaluated in ten selected varieties of apples using an optimized HPLC method. A general description of phenolic substances is included in the thesis. Further, the thesis contains a short review of scientific publications devoted to the content of phenolic substances in fruits and a short description of the HPLC method for determination of phenolic substances. HPLC analysis was performed on the guard column Ascentis Express C18 (5 x 4.6 mm x 5 μm), and separation columns Kinetex C18 (150 x 4.6 mm x 5 μm) with core- shell particles and Luna Omega Polar C18 column (150 x 4.6 mm x 5 μm) with fully porous particles. Detection was performed by a UV detector at wavelengths of 255, 280, 320 and 365 nm. The injected volume was 10 μl and a flow rate of 1 ml/min at column temperature of 30 řC was used. A gradient elution of the mobile phase was applied...
Influence of plant biological protection on production of secondary metabolites of Papaver somniferum III.
Kaman, Petr ; Vytlačilová, Jitka (advisor) ; Sklenářová, Hana (referee)
Kaman, P.: Influence of plant biological protection on production of secondary metabolites of Papaver somniferum III. Diploma thesis. Charles University, Faculty of Pharmacy in Hradec Králové, Department of Pharmaceutical Botany, Hradec Králové, 2018, 77 pp. Diploma thesis was aimed to verify the impact of biological protection on the production of alkaloids of the opium poppy (Papaver somniferum, L.). Polyversum biofungicide was selected for this purpose, the active ingredient of which is oomycete Pythium oligandrum. The poppy tested were Orbis and Lazur high-morphine varieties. Extracts were prepared from the individual samples which were analyzed by HPLC. The following alkaloids were morphine, codeine, papaverine, noscapine and thebain. The average morphine content was around 0,5 %. The largest amount of 0,62 % was found in the poppyhead of untreated variant of the Orbis variety. Codeine for both varieties averaged 0,07 % in the poppyhead of untreated variant. Treated plants reached slightly lower values. The average papaverine content was around 0,01 % in the treated and also untreated variant, but only in poppyhead of the Lazur variety. Higher noscapine content was measured in the Lazur poppyheads, averaging 0,06 %. The content was only around 0,01 % for Orbis variety. The alkaloid content was...
Evaluation of antioxidant activity of natural compounds
Kunovská, Klára ; Sklenářová, Hana (advisor) ; Horstkotte Šrámková, Ivana (referee)
Charles University, Faculty of Pharmacy in Hradec Králové Department of Analytical Chemistry Candidate: Klára Kunovská Supervisor: doc. PharmDr. Hana Sklenářová, PhD. Consultant: doc. RNDr. Miroslav Polášek, CSc. Title of Diploma Thesis: Evaluation of antioxidant activity of natural compounds The aim of this thesis was to evaluate antioxidant activity of natural compounds by chemiluminescence detection in flow system based on sequential injection analysis (SIA). The advantages of this system are high sensitivity, simplicity and high repeatability of individual analyses. The antioxidant activity was evaluated like dependence of chemiluminiscent signal on the concentration of tested antioxidant and then index Q was derived. Luminol was used as chemiluminiscent reagent, which showed chemiluminiscent emission in alkaline enviroment during oxidation of hydrogen peroxide together with a catalyst. As the catalyst, solution of potassium ferricyanide was used and whole measurement was conducted during working voltage of 435 mV. Chemiluminescent signal was monitored for a period of 60 seconds. For each tested substance, 3 cycles of measurement were conducted. The results of testing were evaluated using the area and height of the chemiluminescent signal and compared with the experiment without the tested...
On-line solid phase extraction using sequential injection analysis
Hadysová, Zuzana ; Chocholouš, Petr (advisor) ; Sklenářová, Hana (referee)
Department: Department of Analytical chemistry Candidate: Bc. Zuzana Hadysová Supervisor: PharmDr. Petr Chocholouš, Ph.D. Title of the diploma thesis: On-line solid phase extraction using sequential injection analysis This diploma thesis is focused on the comparison of three sorbents (Iontosorb OXIN 50, Iontosorb Oxin 100 and TOYOpearl AF-chelate-650M) with the aim to find the most suitable one for analysis of Zn2+ using on-line solid phase extraction based on sequential injection analysis. Zinc sulphate solutions at concentrations of 25 nmol/l, 50 nmol/l, 100 nmol/l, 200 nmol/l, 500 nmol/l, 1000 nmol/l, 1500 nmol/l and 2000 nmol/l were used in this work. During the analysis was used the buffer at pH 6, the elution solution HCl at pH 1 and the reagent 4-(2-Pyridylazo)resorcinol (PAR). The effort was to extract Zn2+ at the lowest possible concentrations. Tested parameters were calibration range and repeatability. The best sorbent in comparison was Iontosorb OXIN 100.
Development and optimization of new chromatographic method for determination of retinol in human urine
Mikanová, Zuzana ; Kujovská Krčmová, Lenka (advisor) ; Sklenářová, Hana (referee)
Charles University, Faculty of Pharmacy in Hradec Králové Department of Analytical Chemistry Candidate: Zuzana Mikanová Supervisor: assoc. prof. RNDr. Lenka Kujovská Krčmová, Ph.D. Title of the Diploma Thesis: Development and optimization of new chromatographic method for determination of retinol in human urine This diploma thesis is based on the method development for determination of retinol and creatinine using HPLC. The aim of this project was to find optimal HPLC-MS/MS conditions for clinical research. Experiment was carried out using UHPLC set Nexera with mass spectrometer LCMS- 8030, (Shimadzu, Japan). Three stationary phases were tested. The chromatographic separation was achieved using a Kinetex 2.6 µm PFP 100A 100x4.6 mm (Phenomenex, USA). The used mobile phase consisted of acetonitrile (with addition of formic acid 0.005 M) and ammonium acetate (pH 6.69 ) in the ratio 78:22 (v/v). The temperature was maintained at 25řC, flow rate was set at 0.5 ml/min and 3 µl of sample was injected. After optimization of separation conditions, the method was applied to urine samples, and simple sample preparation procedures were tested In these days is the work on method finishing and the new method will be used for determination of renal damage in clinical research and practice. The inclusion of...
Automation of toxicology tests
Žižková, Klára ; Sklenářová, Hana (advisor) ; Chocholouš, Petr (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Analytical Chemistry Candidate: Bc. Klára Žižková Supervisor: Doc. PharmDr. Hana Sklenářová, Ph.D. Title of Diploma Thesis: Automation of toxicology tests The aim of this diploma thesis was to find suitable method for determination of luciferase of the Ist and IInd types using the sequential injection method and optimize the assay conditions. The main aim was to automate analysis in a series of samples prepared for the testing of various substances, which, in particular, may affect the level of secreted luciferase (type II) upon the interaction of cells with tested substances in toxicological studies. The theoretical part describes luminescence, its types and applications. In addition, the types of luciferases and reagents with which a chemical reaction takes place are mentioned and principles of flow techniques are described. The review of luciferase determinations was carried out. Measurements were performed using the sequential injection method. Three instruments were used for the measurement. One flow injector analyser and one sequential injection analyser with two detection methods - batch-flow spiral detection cells were used. Measurement optimization was performed based on concentration and volume of samples and reagents,...

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