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Colorectal cancer (CRC) is the third most common worldwide. Its treatment includes adjuvant chemotherapy with 5-fluorouracil (5FU) administered intravenously. 5FU is an antineoplastic drug of the fluoropyrimidines group, widely used in the treatment of solid tumors, mainly CRC. Nevertheless, it causes several adverse effects and poor effectiveness due to its short half-life. This work aimed to employ bacterial nanocellulose (BNC) as an encapsulation material for the oral administration of 5FU. First, the adsorption phenomena were analyzed by isotherms, thermodynamic parameters, and kinetic models. Then, encapsulation was carried out using spray-drying, and encapsulated 5FU desorption profiles were assessed in simulated fluids. The biological behavior was evaluated on colon cancer SW480 and SW620 cell lines. As result, it was found that at 25 °C a monolayer of 5FU was formed and the process showed to be the most spontaneous one. In the characterization of the nanocapsules, important changes were detected by the presence of 5FU. The delivery in the colon corresponded to a controlled release behavior. The in vitro assay indicated an improvement in the toxicity effect of the drug and its mechanism of action. Accordingly, BNC is a promising biomaterial for the development of a colon drug delivery platform of 5FU., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022. Published by Elsevier B.V.)