MALDI-MS of flavonoids: a systematic investigation of ionization and in-source dissociation mechanisms

Abstract

Matrix assisted laser desorption ionization (MALDI) is a technique widely employed in the analysis of proteins and peptides, and nowadays it has also been applied to small molecules. There is little significant information regarding the in-source dissociation processes on MALDI for natural products. Twenty-six flavonoids (flavanones, flavones and flavonols) were analyzed by MALDI using different methods (with different matrices) and without matrix to comprehend the in-source reactions and establish good analysis methods for these compounds. Depending on the class, structure and the laser intensity applied, methoxylated flavonoid aglycones can eliminate methyl radicals (˙CH3) in the source, such as flavonols, but lithium 2,4-dihydroxybenzoate matrix suppresses the ˙CH3 eliminations and retro-Diels-Alder cleavages in the source. All of the flavonoid O-glycosides evaluated herein eliminated the sugar in source, even in the presence of the matrix, and its product radical ions ([M-H-sugar](-)˙) were observed in the negative mode. The flavone C-glycosides suffered intense dissociation, which was reduced by the addition of a matrix and the application of low laser intensity, mainly in the negative mode. Depending on the hydroxyl substituents, the [M-H-H](-)˙ ion was observed with variable relative intensity in the spectra.

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“…LDI analyses without a matrix have also been described, mainly for conjugated compounds, which is possible for ionization without a matrix since they can absorb the laser radiation. Some natural products were already analyzed by LDI, such as flavonoids and carotenoids, but the role of the matrix is fundamental in reducing the in-source dissociation as well as increasing the efficiency of ionization and thereby increasing sensitivity [22,23], as confirmed in the investigation of ionization and in-source dissociation of twenty-six flavonoids [22] and others studies (section Application -for more examples and details).…”

“…Some alternatives have been described to reduce this background, for example, carbon nanotubes, ionic liquids, graphene, surfactants with traditional matrices, and a low proportion of matrix (matrix : analyte) [24][25][26]. In addition, there have been great efforts to reduce the fragmentation of analytes in the source [22]. This information is relevant for improving MALDI results and applications in many study areas, such as metabolomics, dereplication, quantification, biological fingerprinting analyses, and IMS, stimulating its utility in natural products research.…”

“…Several glycosylated and non-glycosylated secondary metabolites have been analyzed, such as hydrolysable and condensed tannins, anthocyanins, alkaloids, flavonoids, saponins, rotenoids, carotenoids, xanthophylls, glycosylated triterpenes, theaflavins, thearubigins, phenolics, sesquiterpene lactones, steroids, diterpenes, sesterterpenes, cyanogenic glycosides, and others (l " Fig. 3, Table 2) [22,23,36,37,[44][45][46][47][48][49], demonstrating its huge applicability and ability to analyze secondary metabolites. Among glycosides analyzed by MALDI-MS from the period 1999 to 2010 [44][45][46][47][48][49], saponins, steroids, triterpenes, and flavonoids are the glycosylated metabolites predominantly described (l " Fig.…”

“…6 A). The compounds suffered in-source dissociations [80], which were likely intensified due to the matrix absence; therefore, more studies are required to understand the matrix role for secondary metabolites analyses and their chemical reactions in the source, such as the evaluations performed with aromatic carboxylic acids and flavonoids [22,81,82]. The in-source dissociations of twenty-six flavonoids (glycosylated and non-glycosylated), including flavanones, flavones and flavonols (l " Fig.…”

“…The MALDI conventional matrix application reduced the in-source fragmentation of C-and O-glycosylated flavonoids, but they were not eliminated. In addition, the methyl radical losses from methoxylated flavonoids were completely eliminated only with the LiDHB matrix; besides this, all the retro Diels-Alder fragmentations in the source were also eliminated [22]. Thus, the use of a specific matrix may be required, depending on the drawn objectives and the type of molecular target.…”