Polar flavonoids

Normal phases (unmodified silica gel) are rarely employed, except for the occasional separation of weakly polar flavonoid aglycones, polymethoxylated flavones, flavanones, or isoflavones. The polymethoxylated flavones present in citrus fruits can, for example, be separated on silica gel columns. The big drawback is that solvent gradients cannot normally be run with normal phases. 

The purpose of the MS techniques is to detect charged molecular ions and fragments separated according to their molecular masses. Most flavonoid glycosides are polar, nonvolatile, and often thermally labile. Conventional MS ionization methods like electron impact (El) and chemical ionization (Cl) have not been suitable for MS analyses of these compounds because they require the flavonoid to be in the gas phase for ionization. To increase volatility, derivatization of the flavonoids may be performed. However, derivatization often leads to difficulties with respect to interpretation of the fragmentation patterns. Analysis of flavonoid glycosides without derivatization became possible with the introduction of desorption ionization techniques. Field desorption, which was the first technique employed for the direct analysis of polar flavonoid glycosides, has provided molecular mass data and little structural information. The technique has, however, been described as notorious for the transient... 

Williams, C.A., Greenham, J., and Harbome, J.B., The role of lipophilic and polar flavonoids in the classification of temperate members of the Anthemideae, Biochem. Syst. Ecol., 29, 929, 2001. 

Apart from these techniques, supercritical fluids extraction (SEE) is the preferred technique in many areas of active compounds extraction. However, in what concerns to flavonoids, its utility is highly influenced by the matrix s composition. For example, in the case of isolation of non-polar flavonoid aglycones by SEE, when compared with classical techniques, SEE provided identical or sometimes better results. Nevertheless, when water-soluble glycosides are considered, the use of SEE results in considerably different yields.Still, the use of aqueous methanol constitutes a useful and balanced extraction solvent that allows the extraction of both aglycones and flavonoid glycosides, depending on the conditions used, such as time and temperature, among others. Other reported solvents used for extraction are acetone, ethanol, and dimethyl sulfoxide (DMSO), with... 

The more polar flavonoids occur in the alcohol-soluble fraction of the wood, whereas lipophilic flavonoids, if present, appear in the chloroform or ether extract. Flavonoids may crystallize directly from such extracts, but are more usually obtainable in pure form after some suitable chromatographic separation. They are then characterized by standard spectroscopic procedures (14, 15) and, where appropriate, by comparison with authentic markers. Some wood flavonoids are labile in solution, undergoing oxidation or polymerisation during handling. It is possible during the processes of extraction and purification that interconversions of one type to another (e.g. of dihydroflavonol to flavonol) may occur. Care must therefore be exercised in flavonoid analyses of wood tissues to avoid artifact production in this way. Also, some structural analyses have not been entirely unambiguous examples will be mentioned in later sections where is has not proved possible to re-isolate and confirm the presence of a particular wood flavonoid. 

In order to remove chlorophylls, stilbenoids, less polar flavonoids and other non-polar compoimds, the extract of Cornus alba fruits was partitioued with ethyl acetate [Bjoroy et al., 2007]. 

A wide range of stationary and mobile phase combinations has been reported in the literature to obtain adequate resolution, since this is considered to be the main difficulty for the separation of flavonoids in a complex mixture. Normal phases (i.e., silica gel columns), yet seldom used, can be considered appropriate for the separation of nonpolar or weakly polar flavonoid aglycones, such as polymethoxylated fla-vones, fiavanones, or isofiavones. Usually, no gradient can be employed. Thus, most of the different classes of plant flavonoids and their metabolites are separated by RP-HPLC (Table 3.7). 

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