Analysis of free oligosaccharides by negative-ion electrospray ion trap tandem mass spectrometry in the presence of H2PO4− anions

Positive-ion mode-tandem mass spectrometry has become a frequently used tool for the structural analysis of neutral saccharides. However, their behavior in the positive-ion mode is not always adequate for oligosaccharides structure identification. Thus, fragmentation of [M−H]− or anion adducts produced in the negative-ion mode MS, offers an alternative as their fragmentation patterns provide more structure-related information. The main drawback of the negative-ion mode MS of neutral saccharides is the low sensitivity. In this study, we tested several salts as additives of saccharide samples for sensitivity improvement in the negative-ion mode. The addition of NH4H2PO4 forming [M+H2PO4]− seemed to be the best choice due to a relatively high sensitivity increase in both MS and MSn spectra. Moreover, the salt content could be monitored by the presence of phosphate clusters at high salt-saccharide ratios and salting effects could thereby be avoided. Compared to other tested salts, fragmentation spectra of phosphate adducts also contained additional fragments which improved both the certainty and accuracy of the structural identification. The NH4H2PO4 addition was applied to collected fractions from brewing samples separation where oligosaccharides derived from starch, fructans and oligosaccharides of the raffinose family were detected and identified. Our suggested approach is a promising alternative to a more common mass spectrometric analysis of neutral oligosaccharides in the positive-ion mode or chloride or nitrate additions in the negative-ion mode.

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► Addition of NH4H2PO4 increases the sensitivity of negative-ion MS of oligosaccharides.
► NH4H2PO4 clusters indicate high amounts of salt.
► Adduct-derived cross-ring fragments appeared during fragmentation.
► Dextrins, fructans and raffinose oligosaccharides were identified in plant samples.