Development and application of LC/MS and MS/MS methods for the structural characterization of flavonoid glycosides

Flavonoids are one of the most diverse and widespread groups of natural products. They belong to the class of plant phenolics and generally occur as glycosides in plants. They act in plants as antimicrobials, photoreceptors, visual attractors, feeding repellants, pigments, etc. They have a wide range of biological and physiological activities, are of interest for chemotaxonomy, as tracers in medicinal plant preparations and as antioxidants in food. Therefore, they have been extensively investigated in the past and during recent years, and the interest in them is still increasing.; In the search for new compounds as well as in quality control of plant extracts there is a need to have reliable methodology for the analysis of flavonoids. Mass spectrometry can make an invaluable contribution herein. The use of fast atom bombardment (FAB) and electrospray ionization (ESI) enable the direct ionization of polar molecules, while the coupling of MS with liquid chromatography represents a very useful tool for the analysis of natural products since it combines a powerful separation technique with a very sensitive and universal detector. Additional structural information can be obtained by tandem mass spectrometry (MS/MS) in combination with collision-induced dissociation.; All these currently available techniques were employed in this thesis for the development and application of methods for the analysis and structural characterization of flavonoids. Chapters 1 and 2 are introductory and provide information on the instrumentation used and the state-of-the-art of the mass spectrometric analysis of flavonoids. The original research is described in the following six chapters and consists of three main parts. (1) An LC/MS method based on simultaneous ESI-MS and UV-photodiode array detection (DAD) has been optimized for analysis of flavonoid glycosides (Chapter 3). (2) MS/MS methodology was developed in order to characterize the aglycone part of the most commonly found flavonoids (Chapter 4), the glycan sequence and the interglycosidic linkages (Chapter 5 and 6) as well as to allow the direct stereochemical assignment of the terminal monosaccharide residues (Chapter 7). (3) Finally, the LC/MS and MS/MS methods developed have been applied to characterize the flavonoid fraction of the seeds of Carrichtera annua (Chapter 8).