Cocoa Bean Shell as a Functional Food By-Product: Antioxidant Potential and Toxicological Thresholds in Drosophila...

Agro-industrial fruit processing generates by-products with potential for the development of health-promoting supplements. One such by-product is the cocoa bean shell, a residue from chocolate production that is rich in polyphenols and other bioactive compounds. This study evaluated the chemical composition and biological effects of a combined hot and cold aqueous extract of cocoa bean shell (Cascau), using Drosophila melanogaster as an experimental model. Cascau was analyzed for its content of total polyphenols, flavonoids, tannins, and alkaloids, and its main constituents were identified by UPLC-QToF-MS. In vitro assays confirmed moderate antioxidant activity and suggested potential genomodulatory effects, with evidence of DNA protection and no detectable DNA damage in the applied cell-free assay. In in vivo tests, flies were exposed to diets supplemented with different concentrations of Cascau for up to 7 days. No acute toxicity was observed after 48 h (LC₅₀ = 7%, with significant effects observed at concentrations ≥ 3%). However, chronic exposure to concentrations ≥ 3% significantly reduced survival and increased oxidative stress markers and acetylcholinesterase activity, whereas locomotor function impairment was observed at higher doses. These findings indicate that Cascau exerts dose-dependent biological effects: lower concentrations exhibit antioxidant and genoprotective potential, whereas higher concentrations induce neurotoxicity and redox imbalance. The results suggest a narrow therapeutic window and underscore the importance of dose optimization. Although Cascau displays promising bioactivity, its narrow therapeutic window suggests the need for careful evaluation prior to its application in functional foods or dietary supplements.