Phenolic Profile and Antioxidant Activity of Chocolates Supplemented with Bioactive Ingredients
- Posted
- Server
- Preprints.org
- DOI
- 10.20944/preprints202604.0565.v1
The growing demand for functional foods has stimulated the development of chocolate matrices enriched with bioactive ingredients. This study aimed to optimize the extraction of phenolic compounds using hydro-organic solvents of different polarities, characterize the phenolic and alkaloid profile of distinct chocolate formulations by HPLC-DAD, and evaluate whether fortification with microencapsulated vitamins E and D, zinc, and omega-3 fatty acids (EPA and DHA), either alone or combined with partial replacement of cocoa butter by a structuring oil, enhances the extractable bioactive compounds and in vitro antioxidant activity. Five formulations were evaluated: control chocolate (C), choc-olate containing vitamin microcapsules (T1), chocolate with DHA/EPA microcapsules (T2), lipid-modified chocolate with structuring oil (T3), and chocolate combining microcapsules with lipid modification (T4). Phenolic compounds were extracted using 50% ethanol, 70% methanol, and 70% acetone. Among the tested solvents, 70% methanol showed the highest extraction efficiency, enabling broader detection of bioactive compounds. HPLC-DAD analysis revealed compounds characteristic of cocoa-based matrices, including epicatechin, gallic acid, vanillin, and traces of procyanidins, as well as the methylxanthine alkaloids theobromine and caffeine. The T4 formulation showed a greater abundance of extractable compounds compared to the other formulations. Antioxidant activity was evaluated using DPPH radical scavenging and β-carotene/linoleic acid bleaching assays. T4 exhibited the highest antioxidant performance, with 21.8% DPPH inhibition (EC₅₀ = 2.5 mg/mL) and the highest optical density in the β-carotene assay, indicating enhanced protection against lipid peroxidation. These findings demonstrate that the combination of microencapsula-tion and lipid phase modification improves antioxidant functionality and supports the development of value-added functional chocolates.