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Voltammetry and liquid chromatography of cholesterol, its precursors, and oxidation products
 Kubešová, Jana ; Schwarzová, Karolina (advisor) ; Baluchová, Simona (referee)
Electrochemical detection of sterols is difficult. This thesis aims to study the voltammetric behaviour of selected oxysterols, i.e. 25-hydroxycholesterol and 7-ketocholesterol on the boron-doped diamond electrode. Further, an HPLC method with spectrophotometric and electrochemical detection was developed for separation of selected sterols. The study was performed using cyclic and differential pulse voltammetry. 0.1 mol l−1 perchloric acid in acetonitrile (water content 0.55 %) was chosen as the most suitable electrolyte. 25- hydroxycholesterol provides anodic signal at the potential of +1500 mV and 7-ketocholesterol at +1900 mV in that media. Electrochemical reduction of 7-ketocholesterol was also studied. However, the observed cathodic signal isn't reproducible. The stability of oxysterols in the presence of perchloric acid was confirmed by UV-VIS spectroscopy. Calibration dependence of oxysterols was measured using differential pulse voltammetry and resulted in the limit of detection of 0.82 µmol l−1 for 25-hydroxycholesterol and 0.66 µmol l−1 for 7-ketocholesterol. In the second part of the thesis, an HPLC method was developed for the separation of a sterol mixture on a reversed-phase C18 column using a mobile phase containing 50 mmol l−1 sodium perchlorate in acetonitrile with 0% or 6% water...
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