| Summary: | Aqueous enzymatic extracts of edible mushroom Agaricus bisporus (AbAEE) have been studied as a key source of biological active compounds, among which those with antiviral activity against HCV are of special interest. Searching for a better knowledge of AbAEE, a new approach based on sequential fractionation with solvents of different polarity is here proposed to improve the tentative identification of AbAEE components by liquid chromatography–tandem mass spectrometry (LC–QTOF MS/MS) in high resolution mode. Fifty five out of one hundred twenty-three total metabolites tentatively identified by combination of MS and MS/MS information were identified for the first time. The newly identified compounds include amino acids, sugars, carboxylic, fatty and cinnamic acids, mono- and disaccharides, phospholipids, and purines as the most outstanding, thus demonstrating that fractionation based on sequential liquid–liquid extraction followed by LC–QTOF MS/MS is a suitable option to obtain a wide, representative snapshot of AbAEE composition.
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