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Development of electroanalytical methods for detection of cholesterol, 7-dehydrocholesterol and cholecalciferol
Benešová, Lenka
Steroidival substances are an integral part of the human organism, such as bile acids, steroid hormones, vitamins. The most important sterol in whole organism is cholesterol, which is cornerstone of the all cell membranes. It is involved in important processes in the body, but its storage in blood vessels in the formo f low-density lipoprotein (LDL) is crucial of reason serious health diseases as atherosclerosis, myocardial infarction and others. 7-dehydrocholesterol is a precursor of cholesterol and another sterol compound studied by me, cholekalciferol, also known as a vitamin D3. Deficiency of 7-dehydrocholesterol reductase is the main indicatior of the possible genesis of Smith-Lemli-Opitz syndrome. One of the goals of this thesis was developed a suitable electrochemical method to facilitate the diagnosis of this syndrome from plasma and amniotic fluid in the prenatal and postnatal phases of pregnancy. Within the framework of this thesis were developer electrochemical methods in a batch and flow arrangement for detection of the above-mentioned sterols, namely cholesterol, 7-dehydrocholesterol and cholekalciferol. It was wokered in the medium contain perchloric acid or its salt sodium perchlorate in acetonitrile as supporting electrolyte. It was worked in the wide range of working disc...
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Development of electroanalytical methods for detection of cholesterol, 7-dehydrocholesterol and cholecalciferol
Benešová, Lenka ; Schwarzová, Karolina (advisor) ; Navrátil, Tomáš (referee) ; Šelešovská, Renáta (referee)
Steroidival substances are an integral part of the human organism, such as bile acids, steroid hormones, vitamins. The most important sterol in whole organism is cholesterol, which is cornerstone of the all cell membranes. It is involved in important processes in the body, but its storage in blood vessels in the formo f low-density lipoprotein (LDL) is crucial of reason serious health diseases as atherosclerosis, myocardial infarction and others. 7-dehydrocholesterol is a precursor of cholesterol and another sterol compound studied by me, cholekalciferol, also known as a vitamin D3. Deficiency of 7-dehydrocholesterol reductase is the main indicatior of the possible genesis of Smith-Lemli-Opitz syndrome. One of the goals of this thesis was developed a suitable electrochemical method to facilitate the diagnosis of this syndrome from plasma and amniotic fluid in the prenatal and postnatal phases of pregnancy. Within the framework of this thesis were developer electrochemical methods in a batch and flow arrangement for detection of the above-mentioned sterols, namely cholesterol, 7-dehydrocholesterol and cholekalciferol. It was wokered in the medium contain perchloric acid or its salt sodium perchlorate in acetonitrile as supporting electrolyte. It was worked in the wide range of working disc...
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Determination of cholesterol, 7-dehydrocholesterol and cholecalciferol by HPLC with spectrophotometric and electrochemical detection
Dostálková, Lucie ; Schwarzová, Karolina (advisor) ; Vyskočil, Vlastimil (referee)
The aim of this thesis is to verify the possibility of simultaneous determination of three structurally related biologically active substances (cholesterol, 7-dehydrocholesterol and cholecalciferol) after previous separation by high-performance liquid chromatography followed by electrochemical (on a boron-doped diamond electrode) and spectrophotometric detection, which were compared in this work. The influence of the water content in the mobile phase (in the range of 0-10 %) on the retention factor of the analytes and their response for both types of detectors was investigated. Using all mentioted water contents, the analytes baseline-separated. As the water content increased, the retention factor of the analytes increased as well as the background current for higher positive detection potentials. Based on the recorded hydrodynamic voltammograms, the optimum detection potential + 1.8 V was chosen. The optimum wavelength for spectrophotometric detection was 200 nm. The concentration dependence was measured in an of 50 mmol l−1 sodium perchlorate in acetonitrile mixed with water (95:5, v/v). The spectrophotometric detector provided lower detection limits compared with the electrochemical detector, which had detection limits of 2.3 μmol l−1 for cholesterol, 1.4 μmol l−1 for 7-dehydrocholesterol and...
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Ion Transfer Voltammetry across the Polarized Ionic Liquid/Water Interface: Base for Electrochemical Sensors
Langmaier, Jan ; Samec, Zdeněk
Some electrochemical techniques such as voltammetry at the polarized interfaces between two\nimmiscible electrolyte solutions (ITIES) represent interesting alternatives to classical\nelectrochemistry. The benefit of the ion transfer voltammetry lies in the possibility of detection\nof ionic species which are not otherwise redox active. The methodology enables apart of direct\ndetermination of ionic samples (including pharmaceutical and clinical ones) also to monitor\nreaction processes (acido-basic, enzymatic, etc.), determination of reaction substrates and\nproducts in one experimental step, evaluation of reaction and transport kinetics, and estimation\nof lipophilicity of involved species.
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Determination of selected terpenoids by HPLC with electrochemical detection
Mužíková, Jana ; Dejmková, Hana (advisor) ; Schwarzová, Karolina (referee)
This thesis deals with the determination of carvacrol, thymol, and eugenol by HPLC with electrochemical detection. Carbon paste electrode and boron doped diamond film electrode were used as the working electrodes. For the comparison, UV spectrophotometric detection at 275 nm was used besides the electrochemical detection. The separation was performed on LiChroCART 125-4, RP-18e (5 µm) column. Optimum separation conditions were found: mobile phase consisting of acetonitrile and acetate buffer in ratio 50:50, the optimum buffer pH was pH 5. The optimum potential of working electrode during electrochemical detection was +0,8 V and +1,2 V for carbon paste electrode and boron doped diamond film electrode, respectively. For both electrodes, repeatability of the measurement was examined; the surface of both electrodes had to be renewed between the measurements. Under the obtained optimum conditions, calibration dependences were measured. The studied substances were determined in real samples, in Thymus vulgaris L. and Achillea millefolium L. and in thyme-containing tea and syrup.
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Determination of carvacrol using HPLC with electrochemical detection
Mužíková, Jana ; Dejmková, Hana (advisor) ; Schwarzová, Karolina (referee)
This thesis deals with determination of a mixture of carvacrol with thymol and eugenol by HPLC with electrochemical detection. Carbon paste electrode was used as the working electrode. The separation was performed on Kromasil-C18, 250x4,6 mm column. For the comparison, UV spectrophotometric detection at 275 nm was used besides electrochemical detection. Optimal separation conditions were found: mobile phase consisting of acetonitrile and phosphate buffer in ratio 60:40, the optimal buffer pH was pH 4. As the optimum potential of working electrode during electrochemical detection, potential +1.1 V was found. Under the obtained optimal conditions, calibration dependences were measured. Limit of quantification for carvacrol was found to be 9,6·10-7 mol dm-3 using UV spectrophotometric detection and 4,0·10-8 mol dm-3 using electrochemical detection.
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Study of redox reactions and adsorption of 4-nitrophenyl triazole labeled nucleosides and cresols on boron doped diamond electrodes for development of electroanalytical methods
Vosáhlová, Jana ; Schwarzová, Karolina (advisor) ; Havran, Luděk (referee) ; Navrátil, Tomáš (referee)
In this work, the application of boron-doped diamond electrodes with various surface pre- treatment were tested on selected oxidizable and reducible model compounds, specifically para and ortho cresols and 2'-deoxycytidine and 7-deazaadenosine labeled by 4-nitrophenyl triazole. The aim of the study was the investigation of electrochemical and adsorptive processes with respect to the development of electroanalytical methods of detection of the selected compounds, or utilization of boron-doped diamond electrodes as detectors in liquid-flow systems. Cresols were used as the model oxidizable compound. On O-terminated and polished boron- doped diamond electrodes, cresols provide oxidation signal owing to their oxidation to methylphenoxy radical at comparable potentials as on other carbon-based electrodes used as a reference in this work, i.e., glassy carbon electrode, pyrolytic graphite electrode, and carbon paste electrode. These carbon electrode materials show relatively high propensity to adsorption of reaction products, while such adsorption is minimal on boron-doped diamond. In situ anodic activation allows for a rapid regeneration of boron-doped diamond surface prior to each scan. A differential pulse voltammetric method that was developed with a detection limit of 0.61 mol∙l-1 to 2.97 mol∙l-1...
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HPLC-ED/UV with Solid Phase Extraction for the Determination of 5-Hydroxyindole-3-acetic Acid
Němečková-Makrlíková, Anna ; Dejmková, H. ; Navrátil, Tomáš ; Barek, J. ; Vyskočil, V.
Determination of 5-HIAA in human urine was successfully performed using HPLC-ED/UV with SPE. HPLC-ED is a very sensitive technique, presented method does not need any sample pre-treatment apart from SPE procedure, which replaced filtration of samples of human urine to avoid problems with clogging a HPLC column. The found native\nconcentrations of 5-HIAA in human urine using HPLC-ED with SPE fully corresponds with published normal concentrations in urine (17.8 - 58.3 micromol l-1). The found native concentrations of 5-HIAA were 18.4 micromol L-1 (calculated from peak height). The presented method is fast and could be used for simple\nscreening of human urine, because HPLC-ED/UV allows simultaneous determination of 5-HIAA, HVA, and VMA as important tumour biomarkers.
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Determination of selected terpenoids by HPLC with electrochemical detection
Mužíková, Jana ; Dejmková, Hana (advisor) ; Schwarzová, Karolina (referee)
This thesis deals with the determination of carvacrol, thymol, and eugenol by HPLC with electrochemical detection. Carbon paste electrode and boron doped diamond film electrode were used as the working electrodes. For the comparison, UV spectrophotometric detection at 275 nm was used besides the electrochemical detection. The separation was performed on LiChroCART 125-4, RP-18e (5 µm) column. Optimum separation conditions were found: mobile phase consisting of acetonitrile and acetate buffer in ratio 50:50, the optimum buffer pH was pH 5. The optimum potential of working electrode during electrochemical detection was +0,8 V and +1,2 V for carbon paste electrode and boron doped diamond film electrode, respectively. For both electrodes, repeatability of the measurement was examined; the surface of both electrodes had to be renewed between the measurements. Under the obtained optimum conditions, calibration dependences were measured. The studied substances were determined in real samples, in Thymus vulgaris L. and Achillea millefolium L. and in thyme-containing tea and syrup.
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