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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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Biofyzikální studium malých RNA
Šmerková, Kristýna
Thanks to the prove of connection between the aberrant occurrence of small RNA and various diseases and their potential in diagnostics and treatment led to discovery of new methods and materials facilitating their detection and targeted transport during gene therapy. This work summarizes present knowledge about chosen groups of small RNA, their significance in medical science and the possibilities of their detection. This work primarily concentrates on combination of magnetic separation with electrochemical detection. Magnetic particles (MPs) with different surface modifications were used for isolation. Non-specific isolation was carried out using silanol-coated MPs; streptavidin-coated MPs modified with specific biotinylated probe were used for specific separation. Square wave voltammetry (SWV) was used as a very sensitive electrochemical detection method. Optimized method based on specific magnetic separation with SWV was able to reach nanomolar detection limit (4 nM) with microRNA. The method was applied on human embryonic cells for specific isolation and detection of miR-124. The CdTe quantum dots (QDs) were studied as a nanomaterial tool for nucleic acid detection. The QDs were modified with streptavidin for their bioconjugation with biotinylated molecules were used. Interaction of QDs with nucleic acids was studied using capillary electrophoresis.
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Moderní analytické metody pro detekci aminokyselin asociovaných se zhoubnými nádory prostaty
Cernei, Natalia
In the present thesis called "The modern analytical methods for detection of aminoacids asociated with malignant prostate tumours" sarcosine has been studied as a potential tumor marker which has significance for the diagnosis of aggressive forms of prostate cancer. The effect of addition of sarcosine prostate line PC3 has been also studied as well. Next studied area were the the interactions with amino acid sarcosine, sodium chloride, uric acid and uracil. We have primarily focused also on the development of methods for the detection of sarcosine in urine samples of patients with Cap and prostate cells.
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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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Štúdium možností elektrochemickej detekcie špecifických sekvencií nukleových kyselín
Vaňová, Veronika
This work deals with the study of the possibility of creating an electrochemical biosensor for the detection of specific nucleic acid sequences. The theoretical part describes various types of RNA, with a special focus on miRNAs and possibilities of its detection. It also describes various electrochemical detection approaches, both nucleic acids, and more specifically miRNAs. miRNA is a potential biomarker that can be used for early and non-invasive diagnosis of cancer. The experimental part is aimed at designing, optimizing and preparing a sensitive biosensor for the detection of miRNA-21. A biosensor was prepared to detect the lowest concentration of miRNA-21 in the sample. A linear concentration range for the calibration curve from 1 nM to 1 µM concentration was measured. We measured LOD of 1 fM and by calculation from the regression equation to 3.2 zM and LOQ was 10.8 zM. Subsequently, samples were measured in artificial urine to verify the functioning of the sensor in real conditions. The results showed a minimal effect of the matrix on the determination of the target miRNA.
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Vývoj elektrochemických metod využívajících nanotechnologie pro environmentální mobilní detekční systémy
Kudr, Jiří
During the 21st century, pollution of environment was steadily increasing. Continuous monitoring of pollutants levels represents important part of effective environment protection. Demands for fast analysis of a large numbers of samples caused introduction of automation to analytical chemistry. Techniques based on flow analysis attracted big attention. They are able to decrease need for manual treatment of samples and decrease analysis duration. The advantage of flow-based analytical devices is also compatibility with several detection systems. Among others, electrochemical detectors are very attractive due to their high sensitivity, good selectivity and possibility of miniaturization. The presented thesis focuses on methods for detection of heavy metal and bacterial contamination. In order to it, electrochemical automatized methods and methods which utilized nanotechnology were developed.
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