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New procedures in metabolomic analysis of biofluids
Řimnáčová, Lucie ; Čabala, Radomír (advisor) ; Feltl, Ladislav (referee) ; Sýkora, David (referee)
The aim of this thesis has been the research on reactivity of protic metabolites with chloroformates and its application for GC-MS analysis of biofluids. The research was conducted in three separate studies and the results are three new, original methods for GC-MS determination of low- molecular protic metabolites in biological material especially in biofluids. The first study explores the discovery of fast derivatization of alicyclic hydroxyl groups by fluoroalkyl chloroformates (FCFs) under anhydrous conditions [1]. FCF fully converts the hydroxyl group into a corresponding carbonate and the step can easily be coupled with liquid-liquid microextraction (LLME) of the arising derivatives into organic phase. The reaction of the alicyclic OH group with FCFs was tested on 12 clinically relevant steroids and 4 tocopherols. The analytical properties of determined analytes were described and the method was validated for the GC-MS determination of 6 diagnostic sterols and 4 tocopherols in human serum and amniotic fluid. The new method was further successfully used for determination of sterols and tocopherols in tissues of the bug Pyrrhocoris apterus [2]. The second study was focused on the reactions of protic, particularly urinary metabolites with FCFs, mainly heptafluorobutyl chloroformate (HFBCF)....
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