|
|
New matrices for MALDI-MS analysis of lipids
Macháčková, Petra ; Cvačka, Josef (advisor) ; Volný, Michael (referee)
In this work focused on research of new matrices for MALDI-MS analysis of lipids several organic salts were tested as potential matrices for hydrocarbons and wax esters. It was shown previously that the effective ionization of lipids occurs in conjuction with a lithium cation to form adducts [M + Li] + . Therefore, lithium salts of aromatic acids were synthesized. Matrices were evaluated according to their MALDI properties and compared with lithium 2,5-dihydroxybenzoate, which is proven matrix for MALDI analysis of lipids. It was found that effectiveness of matrix depends on presence of hydroxyl group in matrix molecule. The best results were achieved with lithium vannilate, even in comparison with lithium 2,5-dihydroxybenzoate.
|
|
|
Analysis of antimicrobial peptides in venom glands of bumblebees.
Janechová, Daniela ; Cvačka, Josef (advisor) ; Monincová, Lenka (referee)
The growing resistance of bacteria to traditional antibiotics promotes the interest in finding new substances for their production. Antimicrobial peptides have comparable effect to conventional antibiotics, but a different mechanism of action and they do not provoke bacterial resistance. These peptides were characterized in all forms of multicellular organisms. Hymenoptera venom contains many biologically active substances including antimicrobial peptides. For this reason, this thesis focuses on the acquisition of antimicrobial peptide sequences from selected species of bumblebees (Bombus terrestris, B. hortorum, B. hypnorum, B. pratorum, B. lucorum, B. lapidarius, B. humilis and B. bohemicus). The isolation from the venom glands was performed by high performance liquid chromatography with reversed phases. Subsequent analysis was performed using the methods of mass spectrometry, matrix-assisted laser desorption/ionization with time of flight analyzer and electrospray ionization connected with hybrid linear ion trap analyzer with orbitrap. The sequences for the found peptides were determined by tandem mass spectrometry methods "de novo" and Edman degradation. In this work we characterized 17 sequences of peptides extracted from bumblebee venom glands for which antimicrobial activity was determined...
|
|
|
Hydrophilic Interaction Liquid Chromatography in the Analysis of Intact Glycopeptides
Molnárová, Katarína ; Kozlík, Petr (advisor) ; Coufal, Pavel (referee) ; Cvačka, Josef (referee)
The analysis of glycoproteins represents a significant challenge in glycoproteomics, primarily due to the macro- and microheterogeneity of protein glycosylation. Hydrophilic interaction liquid chromatography (HILIC) is a convenient alternative to reversed-phase chromatography, commonly used in glycoproteomic analysis. This dissertation thesis discusses the potential of HILIC in glycoproteomic analysis, ranging from the separation of glycopeptides on polar stationary phases to the use of HILIC in sample preparation processes. First, the effect of acetonitrile concentration on glycopeptide precipitation was investigated, depending on the type of glycan attached. Subsequently, three commercially available stationary phases were tested: a column containing a silica gel modified with five hydroxyl groups, an amide stationary phase, and a zwitterionic stationary phase. Their efficiency in separating glycopeptide isomers, differing only in branching and/or linkage position, was compared. Further research was devoted to the separation of human immunoglobulin G glycopeptides using relatively new columns that have not yet been characterized in glycoproteomic analysis. These columns, provided by Advanced Chromatography Technologies, included unmodified silica gel (HILIC-A), aminopropyl- modified sorbent...
|
|
|
Development of mass spectrometric methods for structural analysis of fatty acids and estolides
Cudlman, Lukáš ; Cvačka, Josef (advisor) ; Kozlík, Petr (referee) ; Lísa, Miroslav (referee)
(EN) This doctoral thesis summarizes the use of modern mass spectrometric methods for the structural identification of fatty acids and triacylglycerol estolides (TG-EST) and their subsequent characterization in the lipidome of vernix caseosa, i.e., the biofilm covering the skin of the human fetus during the last trimester of in-utero development. Vernix caseosa has mainly antimicrobial properties and the ability to heal wounds and burns. The first part of the doctoral thesis focuses on the diversity of fatty acids in the lipidome of vernix caseosa. These acids form the structural subunits of complex lipids, such as the abundant triacylglycerols and wax esters. By studying fatty acids and their structural features, this thesis has provided valuable insights into the structural variability of the vernix caseosa lipidome. Without a full understanding of the structure of these key subunits, it would be impossible to complete the analysis of this lipidome in the future. In this thesis, fatty acids using high-performance liquid chromatography (HPLC) and modern mass spectrometry methods, namely ultraviolet photodissociation (UVPD) and ozone-induced dissociation (OzID), were characterized. The coupling of HPLC and mass spectrometry (HPLC-MS) provided unambiguous information on the positions and geometry of...
|
|
| |
|
| |
|
| |
|
| |
|
| |
|
| |
guest ::
