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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...
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Quantum-Dots particles as enhancers for chemiluminescence determinations in the SIA system
Bártová, Zuzana ; Sklenářová, Hana (advisor) ; Šatínský, Dalibor (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Analytical Chemistry Candidate: Zuzana Bártová Supervisor: Doc. PharmDr. Hana Sklenářová, Ph.D. Title of diploma thesis: Quantum-Dots particles as enhancers for chemiluminescence determinations in SIA system Quantum-Dots were tested as enhancers of chemiluminescence (CL) emission using sequential injection analysis (SIA). A CL reaction used for this testing was oxidation of luminol (5-aminophthalylhydrazide) by hydrogen peroxide in alkaline medium, QDs represented enhancer of CL signal. The following concentrations of the solutions were tested: hydrogen peroxide in the range 1.10-1 - 1.10-7 mol.l-1 , without QDs and with QDs having maximum emission at 520, 575, 603 a 636 nm and dilution of QDs in following levels 10 µl/10 ml (0.1% solution), 200 µl/10 ml (2% solution) and 200 µl/1 ml (20% solution). The CL emission was detected in two ways. The CL cell, which was constructed at the Department of Analytical Chemistry, FaF UK in HK, and photomultiplier Hamamatsu (Japan) as a detector, enabled monitoring of the CL emission including its time profile. The photomultiplier of the fluorimetric detector Schoeffel (Germany) with spiral flow cell were used for flow measurements in the SIA system. The CL emission was evaluated by...
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Evaluation of selected substances in fruit by HPLC-DAD-CAD method
Košková, Stanislava ; Sklenářová, Hana (advisor) ; Šatínský, Dalibor (referee)
Charles University Faculty of Pharmacy in Hradci Králové Deparment of Analytical Chemistry Candidate: Bc. Stanislava Košková Supervisor: Doc. PharmDr. Hana Sklenářová, Ph.D. Consultant: Ing. Radek Vávra, Ph.D. (VŠÚO Holovousy) Title of diploma thesis: Evaluation of selected substances in fruit by HPLC-DAD-CAD method Thisdiploma thesis was focused on determination of seven phenolic compounds, namely gallic acid, chlorogenic acid, epicatechin, rutin, phloridzin, quercetin, phloretin in twenty apple cultivars, that have been delivered by the Research and Breeding Institute of Pomology Holovousy s.r.o. For this purpose HPLC separation in reverse mode was used, using the column Omega Polar C 18 (150 mm x 4.6 mm; 5 µm). As the mobile phase the mixture of an ultrapure water acidified with the acetic acid to pH 2.8 and acetonitrile was used. A linear gradient was chosen with a gradual increase of the organic component from the initial 10 % to 50 % during 10 min of separation.This was followed with column equilibration - up to 12.50 min to 10 % organic component. The overall separation time was 12.50 min at a flow rate of 1 ml/min. Column temperature was set to 30 řC. Detection was performed using DAD (254, 280, 320 nm) and CAD detector (nebulizer temperature 25 řC). Part of this thesis is system...
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Determination of creatinine in urine using on-line SPE in SIA
Semerádová, Eva ; Chocholouš, Petr (advisor) ; Sklenářová, Hana (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Analytical chemistry Candidate: Bc. Eva Semerádová Supervisor: Assoc. Prof. PharmDr. Petr Chocholouš, Ph.D. Title of the diploma thesis: Determination of creatinine in urine using on-line SPE in SIA The aim of the diploma thesis was to determine creatinine in urine by a method whose results would be comparable to the routinely used method based on the Jaffé reaction. Three urine samples were used, the resulting values were statistically compared with the values measured on a clinical analyzer according to the Jaffé protocol. Urine creatinine was determined by online solid phase extraction (SPE) in sequence injection analysis (SIA). The column in the SIA system was filled with sorbent for extraction, a sorbent was used in the Plexa PCX material. The standard addition method was used to determine creatinine, each sample was measured twice and the average of the given values was used for evaluation. The experimental conditions were first optimized on standard solutions in dilute acetic acid, then applied to real urine samples. The method was evaluated for linearity, yield, repeatability and robustness. According to the value of the correlation coefficient, good linearity was achieved, yield around 100%. Repeatability was...
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HPLC Method Development and Validation for Determination of Active Substances and Impurity in Valetol Preparation
Maroušková, Alena ; Šatínský, Dalibor (advisor) ; Sklenářová, Hana (referee)
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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Monitoring of liberation tests.
Beran, Marek ; Sklenářová, Hana (advisor) ; Matysová, Ludmila (referee)
Charles University Faculty of Pharmacy in Hradci Králové Department of Analytical Chemistry Candidate: Marek Beran Supervisor: Doc. PharmDr. Hana Sklenářová, Ph.D. Consultant: Ing. Daniela Šmejkalová, Ph.D Title of Diploma Thesis: Monitoring of liberation tests The diploma thesis is focused on the development of an automated method for the monitoring of the dissolution profile of clotrimazole from topical dosage forms. The dissolution profile of formulations containing the antifungal agent clotrimazole are compared. Commercially available formulations Canesten and Clotrimazol AL in the form of creams, are compared with a new formulation called Delcore from Contipro a.s., which contains a complex matrix. Theoretical part addresses mycotic infections with focus on superficial mycoses and products available on the Czech market for the treatment of those infections. Furthermore, there is a description of selected non-separation flow methods. The analysis was carried out in a sequential injection analysis system coupled with three attached Franz dissolution cells with a membrane on which the sample was applied. The Franz cells were placed in a water bath at 32 řC. Phosphate buffer with pH 7.4 was chosen as the dissolution medium as it partially imitates the conditions of the human environment. Detection...
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Optimisation of HPLC clotrimazole determination
Žáková, Petra ; Sklenářová, Hana (advisor) ; Matysová, Ludmila (referee)
High-performance liquid chromatographic technique for the determination of clotrimazole and its degradation products (2-chlorophenyl)-diphenyl methanol and imidazole in the formulation Clotrimazol spray 1 % was developed. The presented thesis was focused to the selection of optimal column, composition of mobile phase, its pH and internal standard for the analysis of pharmaceutical formulation based on topically applied antifungal agent clotrimazole. Seven columns were tested in detail. The tested columns were: Discovery HS F5 (5 μm, 150 x 4.6 mm)Chromolith® Performance, RP-18e (100 x 3 mm)ZIC® HILIC (3.5 μm, 50 x 2.1 mm)XTerra® RP 18 (5 μm, 100 x 3 mm)Discovery RP Amide C 16 (5 μm, 250 x 3 mm)Zorbax Extend-C18 (3.5 μm, 75 x 4.6 mm)Discovery ZR-PBD (5 μm, 150 x 4.6 mm). HPLC was carried out using Discovery ZR- PBD column 5 μm, 150 x 4.6 mm. This column is based on polybutadiene-coated zirconium oxide. A zirconium oxide is stable in the whole pH range with the high pressure and temperature up to 200řC. Spectrophotometric detection at 210 nm was performed. The optimal mobile phase was a mixture of acetonitrile and water (pH 9.7) in the ratio of 50:50. The pH of the water component was finally adjusted to 9.7 with NH4OH 25 %. The aqueous solution was filtered through 0.45 m Millipore...
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