Development of functional chocolate incorporating Akkermansia muciniphila: the role of cacao powder

Objective: This study evaluated the feasibility of using chocolate as a food matrix for delivering Akkermansia muciniphila, a next- generation probiotic known for improving gut barrier function and metabolic health. The research assessed the viability of A. muciniphila in three different chocolate formulations, characterised the chocolate’s biological characteristics, and evaluated the potential prebiotic and biological activities of cocoa powder. Methods: The viability and stability of A. muciniphila DSM 22959 in three chocolate matrices (33.6%, 54.5% and 70.5% w/w cocoa) were evaluated over 28 days of aerobic storage. Total phenolic content was measured (Folin-Ciocalteu assay), and antioxidant (ABTS) and antidiabetic properties were assessed. Cocoa powders samples (30%, 90% and 100%) were also evaluated for antioxidant and antidiabetic properties, as well as prebiotic potential for supporting A. muciniphila (via viable cell counts) and acidification through pH monitoring and HPLC analysis of organic acids. Results: Chocolate containing 54.5% (w/w) cocoa emerged as the most suitable matrix for preserving A. muciniphila viability (above 106 CFU/g) during storage. The 70.5% (w/w) cocoa chocolate exhibited higher levels of total phenolic compounds, as along with good antioxidant (349916.87 μmol of Trolox equivalent per g of sample) and antidiabetic (100%) properties. Cocoa powders with 90% and 100% (w/w) cocoa not only showed elevated phenolic content and antioxidant activity but also promoted A. muciniphila growth, as evidenced by a decrease in pH and increased production of short-chain fatty acids after 48 hours of fermentation. Conclusions: Chocolate matrixes demonstrated to have favourable biological characteristics acting as a suitable vector for A. muciniphila DSM 22959.