Brussels sprouts are a source of polyphenolic compounds. However, their concentration is affected by many factors depending on the plant material, hydrothermal treatment methods and digestion in the gastrointestinal tract. The aim of this study was to determine the effect of hydrothermal treatment on the antioxidant activity, concentration and profile of polyphenols of Brassica oleracea var. gemmifera in an in vitro simulated gastrointestinal digestion model. The study showed a significant effect of the type of hydrothermal treatment on total polyphenol concentration, polyphenolic acid profile, flavonoid content and antioxidant activity. Traditional boiling in water was the least effective type of hydrothermal treatment with respect to bioactive components of Brussels sprouts. Sous-vide was the most effective hydrothermal treatment in terms of retention of polyphenolic compounds and high antioxidant activity, thus providing a better alternative to steam cooking. Using an in vitro model, a significant difference was demonstrated between the concentration of bioavailable polyphenolic compounds and the polyphenol content of the plant material before digestion. The influence of the type of hydrothermal treatment used on the concentration of bioavailable polyphenolic compounds was maintained in relation to material not subjected to in vitro digestion (except for antioxidant activity).