Bioactive phenols in algae: the application of pressurized-liquid and solid-phase extraction techniques

A new extraction technique based on the off-line combination of pressurized-liquid with solid-phase extraction (PLE-SPE) is described. The method was used for the extraction of bioactive phenolic acids (protocatechuic, p-hydroxybenzoic, 2,3-dihydroxybenzoic, chlorogenic, vanillic, caffeic, p-coumaric, salicylic acid), cinnamic acid and hydroxybenzaldehydes (p-hydroxybenzaldehyde, 3,4-dihydroxybenzaldehyde, vanillin) from in vitro culture of two freshwater algae (Anabaena doliolum and Spongiochloris spongiosa) and from food products of marine macroalgae Porphyra tenera (nori) and Undaria pinnatifida (wakame). For the identification and quantification of the compounds the molecular ions [M-H](-) and specific fragments were analyzed by quadrupole mass spectrometry analyzer connected on-line with a reversed-phase HPLC system. Our analysis showed that the freshwater algae and marine algal products contained submicrogram or microgram level of above-mentioned phenols per gram of lyophilized sample. In addition, the total phenol content (Folin-Ciocalteu assay) and antioxidant activity (TEAC assay, Trolox equivalent antioxidant capacity assay) of the PLE-SPE extracts were determined and discussed.