High-pressure liquid chromatography coupled with mass spectrometry is a technique which is becoming indispensable in the pharmaceutical field. It combines two analytical techniques: high-performance liquid chromatography and mass spectrometry. Liquid chromatography separates the components of the compound according to their affinity with the stationary and mobile phase. These components can be detected by UV light. The basis of mass spectrometry is separating ionized components in a vacuum, according to their mass-to-charge ratio.
In the development of active substances, compound libraries are one of the key compnents of research. They are tested on various biological targets, using high-throughput screening. They need to be organised and well-formed, to test the largest number of compounds in the shortest possible time. It is of key importance, that the compounds, that are being kept in the library, are of adequate quality and well preserved for future use.
The aim of the master's thesis was to assess the library of compounds, which were synthesised at the Faculty of Pharmacy with the HPLC-UV-MS technique. The number of samples in the compound library was 1145. Reverse-phase chromatography was used for the separation and electrospray ionization was used for ionization on the mass spectrometer. Many difficulties were also encountered in the evaluation of results, namely poor mass spectra, decarboxylation of compounds, mistakes in preparation of samples, hydrolysis of compounds, fragmentation of compounds, and instability of solutions. However, despite the errors, the instrumental method was suitable for most of the compounds (90,1%). We organised the library and collected all data, that were gathered when interpreting results, in an Excel table. We determined the retention time of the compounds, assessed the purity and reactions in UV spectroscopy. We confirmed the structure of the compounds with mass spectrometry, attained the MS chromatogram and mass spectra of the compounds. We determined the formation of adducts. This allowed us to quantitatively and qualitatively evaluate the compound library, and also to obtain data on the behaviour of the lead compounds in an HPLC-UV-MS chromatographic system on a representative sample.
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