Supercritical CO₂ extraction of oil, fatty acids and flavonolignans from milk thistle seeds: Evaluation of their antioxidant and cytotoxic activities in Caco-2 cells

The optimal conditions of supercritical carbon dioxide (SC-CO₂) (160-220 bars, 40-80 °C) technology combined with co-solvent (ethanol), to recover oil, flavonolignans (silychristin, silydianin and silybinin) and fatty acids from milk thistle seeds, to be used as food additives and/or nutraceuticals, were studied. Moreover, the antioxidant and cytotoxic activities of the SC-CO₂ oil seeds extracts were evaluated in Caco-2 carcinoma cells. Pressure and temperature had a significant effect on oil and flavonolignans recovery, although there was not observed a clear trend. SC-CO₂ with co-solvent extraction at 220 bars, 40 °C was the optimum treatment to recover oil (30.8%) and flavonolignans from milk thistle seeds. Moreover, linoleic (47.64-66.70%), and oleic (19.68-24.83%) acids were the predominant fatty acids in the oil extracts recovered from milk thistle under SC-CO₂. In addition, SC-CO₂ extract showed a high antioxidant activity determined by DPPH and ABTS tests. Cytotoxic activities of silychristin, silydianin and silybinin and the obtained SC-CO₂ extract (220 bars, 40 °C) were evaluated against Caco-2 cells. The SC-CO₂ extract inhibited the proliferation of Caco-2 cells in a dose-responsive manner and induced the highest percentage of mortality of Caco-2 cells (from 43 to 71% for concentrations from 10 up to 100 μg/ml of SC-CO₂ oil seeds).