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Study of influence of mobile phase composition on selectivity and retention of the analytes on HILIC stationary phases
Štětková, Kateřina ; Kočová Vlčková, Hana (advisor) ; Chocholoušová Havlíková, Lucie (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of Analytical Chemistry Candidate: Kateřina Štětková Supervisor: RNDr. Hana Vlčková, Ph.D. Title of Diploma thesis: Study of influence of mobile phase composition on selectivity and retention of the analytes on HILIC stationary phases This graduation thesis deals with an influence of concentration of ACN, pH and buffer concentration in mobile phase on retention of selected analytes and on selectivity of Atlantis HILIC Silica column (3 μm, 2,1 mm x 100 mm) and Luna NH2 column (3 μm, 2,0 mm x 100 mm). A set of 35 analytes including neutral, basic and acidic compounds were chosen for this study. Detection was performed by PDA detector at the wave-length of 245 nm. Measurements were carried out using isocratic flow 0.4 ml.min-1 of mobile phase ACN/water (buffer). The column temperature was set up to 30 řC, auto sampler temperature to 4 řC and injection volume to 2 µl. For easier evaluation of results, measured data are presented in charts and tables. These were used to evaluation of columns in terms of selectivity and retention of the individual groups of analytes. The typical HILIC behavior, an increasing retention of analytes with increasing ACN concentration in mobile phase, was observed for Atlantis HILIC Silica column....
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Comparison of selectivity and retention of the analyties on two HILIC stationary phases
Hodovská, Nikola ; Vlčková, Hana (advisor) ; Chocholoušová Havlíková, Lucie (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of Analytical Chemistry Candidate: Nikola Klvanová Supervisor: RNDr. Hana Vlčková, Ph.D. Title of Diploma thesis: Comparison of selectivity and retention of the analytes on two HILIC stationary phases This diploma thesis deals with an influence of ACN concentration, pH and buffer concentration in mobile phase on the retention of tested analytes and selectivity of two analytical columns Luna HILIC (diol) column (3 μm, 3 mm x 100 mm) and Ascentis ES Cyano column (3 μm, 2.1 mm x 100 mm). A set of 28 analytes including neutral, basic and acidic compounds was chosen for this study. Detection was performed by PDA detector at the wave-length of 245 nm. Measurement was performed isocratically at the mobile phase flow rate of 0.7 ml/min and 0.3 ml/min. Mobile phase was consisted of a mixture of acetonitrile and water component (buffer or acetic acid). The column temperature was set up to 30 řC, autosampler temperature to 4 řC. Injection volume was 5 μl or 3 μl. For easier evaluation of results, measured data are presented in the charts and tables. The influence of the ACN content in mobile phase was evaluated based on the plots of the retention factor that showed increasing retention with increasing ACN content (typical HILIC...
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Development of MEPS method for UHPLC-MS/MS determination of entecavir in renal ultrafiltrate
Janák, Jaroslav ; Vlčková, Hana (advisor) ; Chocholoušová Havlíková, Lucie (referee)
1 Abstract Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of analytical chemistry Candidate: Jaroslav Janák Supervisor: RNDr. Hana Vlčková, Ph.D. Title of Thesis: Development of MEPS-UHPLC-MS/MS methods for the determination of entecavir in kidney ultrafiltrate. The thesis deals with the development and validation of sample preparation method for determination of entecavir in kidney ultrafiltrate, using techniques MEPS (microextraction by packed sorbent). The previously developed UHPLC-MS/MS method for the determination of entecavir in rat urine has been used for the analysis. Hydrophilic interaction chromatography by column Acquity BEH Amide and isocratic elution were employed. The composition of the mobile phase was acetonitrile and 5 mM ammonium acetate pH 4.0 in the ratio 75:25 (v:v). Entecavir C2 13 N15 was used as the internal standard. Ionization of the analyte was carried out by electrospray in the positive ion mode and triple quadrupole was employed as the detector. Quantification of the analyte was realized by two SRM transitions of entecavir and internal standard method. Based on the MEPS optimization method, porous graphitic carbon suitable for the analysis of polar compounds was selected as the most suitable sorbent. The mixture of acetonitrile and water in a...
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Analyses of Statins in Biological Samples
Vlčková, Hana ; Solich, Petr (advisor) ; Nobilis, Milan (referee) ; Šimek, Petr (referee)
5 ABSTRACT Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of analytical chemistry Candidate: Mgr. Hana Vlčková Supervisor: Prof. RNDr. Petr Solich, CSc. Supervisor specialist: Assoc. Prof. PharmDr. Lucie Nováková, Ph.D. Title of Doctoral Thesis: Analysis of statins in biological samples The doctoral thesis deals with analysis of statins in biological samples. Statins are the most widely used drugs for the treatment of Familiar hypercholesterolemia. Due to very low concentration of statins (ng/ml) in human plasma a choice of suitable analytical methods is limited. Liquid chromatography with mass spectrometry detection is the most suitable technique due to high selectivity and sensitivity. Sample preparation step has to be used before chromatographic determination of analytes in complex matrices, such as biological samples. Primarily theoretical part of doctoral thesis summarizes individual pharmacological properties and effects of statins. Another part is devoted to theoretical knowledge of UHPLC-MS/MS. The widest section deals with description of the individual sample preparation techniques. The practical part contends with development of UHPLC-MS/MS methods for the determination of statins in biological samples and selection and development of suitable sample preparation...
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Solid-phase extraction and its miniaturization by method Lab-On-Valve for determination of pharmaceutical substances
Jelínková, Iva ; Chocholouš, Petr (advisor) ; Vlčková, Hana (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Analytical chemistry Candidate: Iva Jelínková Supervisor: PharmDr. Petr Chocholouš, Ph.D. Title of diploma thesis: Solid-phase extraction and its miniaturization by method Lab-On- Valve for determination of pharmaceutical substances This thesis deals with the development of the solid-phase extraction (SPE) method with the use of microextraction by packed sorbent (MEPS) for the determination of vitamins A, D, E and the possibility to automate this method. The determination was based on absorption of vitamins on microcolumn MEPS, interfering components were removed by washing solution (dilute acetic acid of pH 3) and extraction was performed by eluent solution (100% ACN). Detection was made by UV spectrophotometer at wave lengths of absorbing maximum for vitamin A - 325 nm, for vitamin D - 265 nm, for vitamin E - 295 nm. The method was optimized. There was developed a program, where concentrations of individual vitamins, dosing volumes, flow speed and composition solvents were tested. After the optimization method there was performed the extraction and the subsequent determination of blood plasma fortified with vitamins A, D, E by the sequential injection chromatography method. Recovery of the method for vitamin A was...
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A development of UHPLC-MS/MS method for the determination of pravastatin, rosuvastatin and their metabolites
Mikšová, Alena ; Nováková, Lucie (advisor) ; Vlčková, Hana (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of Analytical Chemistry Candidate: Alena Mikšová Supervisor: Doc. PharmDr. Lucie Nováková, PhD. Title of Graduation Thesis: A development of UHPLC-MS/MS method for the determination of pravastatin, rosuvastatin and their metabolites This graduation thesis is dealing with the development of UHPLC/MS-MS method for the determination of pravastatin, rosuvastatin and their metabolites. The basic parameters were optimized for MS detection - desolvation temperature, ion optics (RF lens and extractor), nitrogen flow at the sample cone, capillary voltage, nitrogen flow and cone voltage, which is specific for each substance. Separation of analytes was performed on BEH C18 analytical column (2.1 x 100 mm, 1.7µm). Triple quadrupole was used for detection. Electrospray ionization was performed in both negative and positive mode. Quantification of analytes was performed using the SRM (selected reaction monitoring). Ion [M+H]+ was chosen for all statins determined in positive ion mode. This ion was the most intense in full scan spectrum. In positive ion mode pravastatin lactone offered also ammonium adduct [M+NH4]+ . All statins determined in negative mode provided a precursor ion [M-H]- . Pravastatin lactone and rosuvastatin lactone...
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Novel stationary phasis in HPLC for the analysis of biologically active compounds for clinical applications-determination of retinoic acid
Vlčková, Hana ; Solich, Petr (advisor) ; Nováková, Lucie (referee)
All-trans retinoic acid (ATRA) and 13-cis-retinoic acid (13-cis RA) are naturally occurring retinoids. All-trans-retinoic acid plays multiple roles in reproduction, embryogenesis, cellular differention, vision and immune response and is also used therapeutically in acute promyelocytic leukemia. Tocopherols are major serum antioxidants. Both retinoids and tocopherols are studied as chemopreventive and therapeutic agents in cancer patients. In the present study, a simple and rapid HPLC method was developed for simultaneous determination of fat-soluble vitamins and retinoic acids (retinol, all-trans retinoic acid, 13-cis retinoic acid, gamma- and alpha-tocopherol) in human serum using monolithic technology. The monolithic column Chromolith Performance RP-18e (100 mm × 4.6 mm) was operated at ambient temperature. Mobile phase (MF) consisted of mixture Acetonitrile (ACN) : 1% ammonium acetate in water (AMC) at pH 7.0 starting with ACN : AMC 98:2 (column pre- treatment) at flow rate 2ml/min following 4,2 minutes ACN:AMC 95:5 at the flow rate 1.5 ml/min than 3 minutes flow rate 3,2ml/min with the same mobile phase. Detection and identification was performed using photodiode array detector. Retinol, 13-cis- and all-trans- retinoic acid were monitored at 325 nm. Alpha- and gamma-tocopherol were carried out...
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MIDWIFE´S ROLE IN EDUCATION OF WOMEN WITH PELVIC INFLAMMATORY DISEASE
VLČKOVÁ, Hana
MIDWIFE´S ROLE IN EDUCATION OF WOMEN WITH PELVIC INFLAMMATORY DISEASE Nowadays, the number of women who have underestimated the symptoms of pelvic inflammatory disease and who have been coping with its consequences such as chronic pain or infertility is increasing. Therefore the following topic was selected for the bachelor thesis: "Midwife´s Role in Education of Women with Pelvic Inflammatory Disease". The theoretical part of the bachelor thesis deals with the educational activity provided by a midwife, basic aetiology of pelvic inflammatory disease, its treatment and nursing process for women with pelvic inflammatory disease. A quantitative research method using anonymous questionnaires was selected for the practical part of the bachelor thesis. The research was conducted in the period of 1 January - 31 March 2011 and 93 responders participated in it. Three goals were stated in the thesis and they were met. The first goal of the research was to find out whether women are informed about the symptoms, treatment and subsequent prevention of pelvic inflammatory disease. The second goal was to find out how satisfied women are with the level of education provided by a midwife in the field of pelvic inflammatory disease. The third goal was to create educational material on pelvic inflammatory disease. Further, for the purpose of the research, 3 hypotheses were stated. Hypothesis No. 1: "Women are aware of the symptoms of pelvic inflammatory disease and they pay respective attention to them," was confirmed. Hypothesis No. 2: "Women lack information on causative agents of pelvic inflammatory disease causing post-inflammatory changes such as infertility," was confirmed. Hypothesis No. 3: !Women have enough information on causes of occurrence of gynaecological inflammation, its symptoms, treatment and subsequent prevention," was not confirmed. The results of the research investigation of this thesis could be useful for midwives and their educational activity. Educational material in form of a leaflet was created for women´s needs.
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