Hydroxytyrosol bioaccessibility and absorption by the intestinal cells were studied using an in vitro digestion model and Caco-2 TC7 monolayers cells in culture in the presence and absence of beta-cyclodextrin and foods. Hydroxytyrosol was either provided as a pure standard or in an alperujo powder. The presence of foods significantly decreased hydroxytyrosol bioaccessibility and absorption (-20 and -10%, respectively), while beta-cyclodextrin had no effect. Moreover, the presence of other compounds from alperujo in the intestinal compartment reduced hydroxytyrosol absorption by Caco-2 cells compared to pure standard (-60%). The final bioavailability of hydroxytyrosol, defined as its quantity at the basolateral side of cultured cell monolayers compared to the initial amount in the test meal, was 6.9 +/- 0.4, 31.1 +/- 1.1, and 40.9 +/- 1.5% when hydroxytyrosol was from alperujo or a standard administered with or without food, respectively. Our results show that conversely to foods, beta-cyclodextrin does not alter hydroxytyrosol bioavailability.