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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,...
The testing of nanofiber polymers for stir bar sorptive extraction
Zatrochová, Slavomíra ; Šatínský, Dalibor (advisor) ; Chocholouš, Petr (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Analytical Chemistry Candidate: Slavomíra Zatrochová Supervisor: doc. RNDr. Dalibor Šatínský, Ph.D. Title of diploma thesis: Testing of nanofibrous polymers for extraction on a magnetic stirrer This thesis focuses on nanofibers polymers and their potential use for the sorption extraction on a magnetic stirrer (SBSE - stir bar sorptive extraction). The main aim of the work was to test selected types of polymer fibers and to prove their suitability as material for extraction on the magnetic stirrer. The presented work included the preparation of solutions to which we placed magnetic stirrers coated by nanofibers where the sorption extraction was carried out with selected substances. Afterwards chromatographic analysis and separation, the pre-concentrated substances on polymeric fibers was evaluated. After several measurements we found out that the extreme lipophilic analytes such as Sudan dyes were irreversibly adsorbed on polymeric fibers. Polystyrene fiber (PS) was not stable in used organic solvents and therefore was prepared again before every extraction. The highest theoretical enrichment factor (100) has not been achieved at any used nanofiber. In conclusion, we can say which selected kinds of polymeric fibers are suitable...
Purification of phlorizin from Malus domestica Borkh. leaves by solid-phase extraction and semi-preparative high performance liquid chromatography.
Prachařová, Eva ; Chlebek, Jakub (advisor) ; Chocholouš, Petr (referee)
DIPLOMA THESIS Purification of phlorizin from Malus domestica Borkh.by solid-phase extraction and semi-preparative high-performance liquid chromatography Eva Prachařová Charles University, Faculty of Pharmacy in Hradec Králové Department of Pharmaceutical Botany The aim of this diploma thesis was to find the best conditions for purification of a flavonoid extract from leaves of Malus domestica Borkh., and obtaining the purest fraction of phlorizin. Phlorizin may be used in the treatment of diabetes mellitus type 2 in the future, it has the ability to reduce glycemia by reducing the absorption of glucose in the small intestine and by increasing urinary glucose excretion. The first step was to find an SPE cartridge with a suitable sorbent and a suitable eluent for solid phase extraction. The DPA-6S cartridge and 100% methanol as an eluentwerefound to be the most suitable for SPE. The next step was to find the best possible conditions for semi-preparative HPLC using an ACE 5 C18 column (5 μm, C18, 150 x 10 mm i.d., 150 mm length). The mobile phase consisted of 1% (v/v) acetic acid in water (solvent A) and ethanol 100% (v/v) (solvent B), and a linear gradient elution was used (10-100% B), 0-60 min, the flow: 1mL/min. This method resulted in the 91.05% purity of phlorizin. Keywords: phlorizin, SPE,...
Extraction of zinc from water solutions using solid phase extraction in sequential injection analysis
Klinerová, Jana ; Chocholouš, Petr (advisor) ; Šatínský, Dalibor (referee)
v angličtině Charles University Faculty of Pharmacy in Hradec Králové Department of Analytical chemistry Candidate: Bc. Jana Klinerová Supervisor: PharmDr. Petr Chocholouš, Ph.D. Title of diploma thesis: Extraction of zinc from water solutions using solid phase extraction in sequential injection analysis This work is devoted to the extraction of zinc from aqueous samples by solid phase extraction in sequential injection analysis. Zinc extraction was tested with solutions of three concentrations (100 nmol/l, 200 nmol/l, 500 nmol/l) using four different chelating sorbents (Iontosorb OXIN, Iontosorb SALICYL, Iontosorb IDA a Toyopearl) at seven different conditions (pH of 3, 4, 5, 6, 7, 8 and 9) of the wash buffer and at pH 2 of the elution solution. During the detection was monitored absorbance of zinc complex with the reagent 4-(pyridyl-2-azo) resorcinol at wavelengths of 490, 510, 560 and 600 nm. In comparison of the four sorbents under different conditions of measurement was determined as the best sorbent Iontosorb OXIN and suitable conditions for the extraction - pH of the wash buffer and of the sample 6 and higher.
Spectrophotometric determination of chlorhexidine in mouthwash employing Lab-In-Syringe automated ion-pair extraction and back-extraction
Řeháková, Pavla ; Horstkotte, Burkhard (advisor) ; Chocholouš, Petr (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Analytical Chemistry Candidate: Pavla Řeháková Supervisor: Dr. Burkhard Horstkotte, Ph.D., M.Sc. Consultant: doc. PharmDr. Hana Sklenářová, Ph.D. Title of the diploma thesis: Spectrophotometric determination of chlorhexidine in mouthwash employing Lab-In-Syringe automated ion-pair extraction and back-extraction In this work, a magnetic stirring-assisted dispersive liquid-liquid micro-extraction (MSA-DLLME) automated by the Lab-In-Syringe technique is presented. MSA-DLLME is based on mixing the sample and an immiscible solvent by the action of a magnetic stirrer for the dispersion of the solvent into fine droplet to enhance the extraction process. In Lab-In-Syringe, the magnetic stirrer is placed inside a syringe void, which is used as extraction chamber. To allow the use of an extraction solvent lighter than water, the syringe was turned upside down in this work. This method was designed for the determination of chlorhexidine in commercial mouthwash samples. To allow the extraction of chlorhexidine, an ion-pair complex with the reagent methyl orange needed to be formed. After the extraction into an organic solvent, the aqueous solution was exchanged and the analyte was back-extracted into an acidic aqueous acceptor to yield...
Optimization of a method for iodine determination in urine in a flow system using solid phase extraction
Lipenská, Zuzana ; Horstkotte Šrámková, Ivana (advisor) ; Chocholouš, Petr (referee)
Department: Department of Analytical Chemistry Candidate: Bc. Zuzana Lipenská Supervisor: PharmDr. Ivana Horstkotte Šrámková, Ph.D. Consultant Doc. PharmDr. Hana Sklenářová, Ph.D. Title of diploma thesis: Optimization of a method for iodine determination in urine in a flow system using solid phase extraction Iodine is an important element for the human body to ensure proper functioning of thyroid gland. This organ produces the hormones thyroxine and triiodothyronine necessary for proper brain development and correct function of other hormones (catecholamines). The human body is unable to produce iodine, so the only income is via food. Incorrect level of this element can cause serious health complications, such as congenital mental retardation. Body level of iodine is most commonly determined from urine, which requires a reliable and sensitive method. The most widely used method is spectrophotometric analysis based on the Sandell-Kolthoff reaction. For urinary iodine determination, it is advantageous to use flow methods to ensure sufficient measurement sensitivity and automation. Most methods of iodine monitoring use toxic reagents or generate toxic waste. The aim of this diploma thesis was to optimize the hydrodynamic and chemical parameters of a method for urine iodine analysis using...
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.
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,...
Use of silica sorbents for on-line solid phase extraction in flow methods
Chrástková, Hana ; Chocholouš, Petr (advisor) ; Horstkotte Šrámková, Ivana (referee)
v anglickém jazyce Charles University Faculty of Pharmacy in Hradec Králové Department of Analytical Chemistry Student: Bc. Hana Chrástková Supervisor: PharmDr. Petr Chocholouš, Ph.D. Title of diploma thesis: Use of silica sorbents for on-line solid phase extraction in flow methods. The diploma thesis is devoted to the extractions of substances such as Methylparaben, Butylparaben, α-tocopherol, Cholecalcipherol, Sudan I, II and IV, β-estradiol, Estriol, Estrone using on-line solid phase extraction with sorbent for Supported Liquid Extraction automated by Sequential Injection Analysis method. Substances of different chemical properties were selected, and extraction parameters were optimized to achieve the highest possible extraction recovery. The following parameters were tested: the volume of sample loaded in the column, sample loading flow rate, analyte concentration, effects of pH and kind of solvent of sample, time interval for sample adsorption. Under optimized conditions, the repeatability of the method and calibration curves of selected analytes were measured

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