Optimized Food-Grade Semi-Synthetic Culture Medium and Freeze-Drying Approach with Antidiabetic Cryoprotectants for Akkermansia Muciniphila

Akkermansia muciniphila is a mucin-degrading intestinal bacterium known for its beneficial effects, including antidiabetic and antiobesity effects.It typically grows on media supplemented with animalderived mucin.However, the use of such substrates in cultivation is unsuitable for food applications due to ethical and dietary concerns.To address this, we developed a plant-based, semi-synthetic medium using soy and pea peptones along with L-threonine as nitrogen sources, and Nacetylglucosamine with glucose as carbon substrates.The strain reached high cell density (4.8510 CFU/mL) on this medium, exceeding yields typically observed in mucin-based media.The medium composition was optimized using response surface methodology, resulting in the following formulation: 1.5 g/L threonine, 10 g/L soy peptone, 15 g/L pea peptone, 5.0 g/L glucose, 7.0 g/L GlcNAc, pH 7.0.The obtained biomass was lyophilized with cryoprotectants of camel milk and Dallulose.Cell viability in the final freeze-dried powder was 84.2%, with minimal membrane damage confirmed by flow cytometry.Our results support the use of this animal-free system for the development of functional foods containing A. muciniphila.