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Qualitative analysis of selected glycosphingolipids subclasses isolated from human blood plasma using HILIC-ESI-MS/MS
ŠIKLOVÁ, Michaela
This bachelor thesis deals with the qualitative analysis of selected subclasses of glycosphingolipids isolated from human blood plasma using hydrophilic interaction liquid chromatography coupled with electrospray ionization tandem mass spectrometry (HILIC-ESI-MS/MS). Glycosphingolipids are ubiquitous and structurally very diverse components of cell membranes involved in many cellular functions. It has been shown that changes in the structures of glycosphingolipids and their distinct expression can pose a key factor in the development of numerous diseases and different types of cancer.The theoretical part of this thesis is dedicated to a summary of fundamental knowledge about glycosphingolipids, their biosynthesis, classification, biological functions and their association with various types of diseases and cancers. Furthermore, methods of isolation, purification and analytical possibilities for the determination of glycosphingolipids in biological samples are described here.In the experimental part, blood samples were taken and processed together with the preparation of the necessary solutions, plasma samples, and standards. Moreover, a test analysis, optimization of mobile phase gradient and comparison of the most frequently used extraction methods according to Folch, Bligh and Dyer, Matyash, and Alshehry alongside fractionation of lipid extract is described in this part. An integral part of the experimental section was the identification and structural characterization of glycosphingolipids within selected subclasses, including the generation of GSL profiles in human blood plasma. In conclusion, the results of the work and answers to the stated research questions are summarized, including the evaluation of possible improvements in this field.