Liquid chromatographic/electrospray ionization tandem mass spectrometric study of the phenolic composition of cocoa (Theobroma cacao)

Liquid chromatography coupled with ionspray mass spectrometry in the tandem mode (LC/MS/MS) with negative ion detection was used for the identification of a variety of phenolic compounds in a cocoa sample. Gradient elution with water and acetonitrile, both containing 0.1% HCOOH, was used. Standard solutions of 31 phenolic compounds, including benzoic and cinnamic acids and flavonoid compounds, were studied in the negative ion mode using MS/MS product ion scans. At low collisional activation, the deprotonated molecule [M − H]− was observed for all the compounds studied. For cinnamic and benzoic acids, losses of CO2 or formation of [M − CH3]−· in the case of methoxylated compounds were observed. However, for flavonol and flavone glycosides, the spectra present both the deprotonated molecule [M − H]− of the glycoside and the ion corresponding to the deprotonated aglycone [A − H]−. The latter ion is formed by loss of the rhamnose, glucose, galactose or arabinose residue from the glycosides. Different fragmentation patterns were observed in MS/MS experiments for flavone‐C‐glycosides which showed fragmentation in the sugar part. Fragmentation of aglycones provided characteristic ions for each family of flavonoids. The optimum LC/MS/MS conditions were applied to the characterization of a cocoa sample that had been subjected to an extraction/clean‐up procedure which involved chromatography on Sephadex LH20 and thin‐layer chromatographic monitoring. In addition to compounds described in the literature, such as epicatechin and catechin, quercetin, isoquercitrin (quercetin‐3‐O‐glucoside) and quercetin‐3‐O‐arabinose, other compounds were identified for the first time in cocoa samples, such as hyperoside (quercetin‐3‐O‐galactoside), naringenin, luteolin, apigenin and some O‐glucosides and C‐glucosides of these compounds. Copyright © 2003 John Wiley & Sons, Ltd.