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- Encapsulation of Akkermansia muciniphila in calcium-alginate matrix: a live biotherapeutic strategy for gut microbiota modulation and intestinal infection mitigationPublication . Machado, Daniela; Vedor, Rita; Fonseca, Mariana; Sousa, Sérgio; Barbosa, Joana Cristina; Carvalho, Nelson Mota de; Costa, Célia Maria; Madureira, Ana Raquel; Gomes, Ana MariaObjective: Akkermansia muciniphila is a next-generation probiotic recognized for improving gut barrier function and metabolic health. However, ensuring its viability during processing, storage, and gastrointestinal transit remains a challenge. This study investigated calcium-alginate encapsulation as a strategy for enhancing the stability of A. muciniphila during refrigerated aerobic storage and gastrointestinal passage as well as biological validation in an in vitro fecal model. Methods: A. muciniphila was encapsulated using an extrusion technique with 2% (m/v) sodium alginate and 4% (m/v) calcium chloride. Viability of free and encapsulated cells was assessed over 28-days of refrigerated aerobic storage. Gastrointestinal stability was evaluated using the INFOGEST protocol. In vitro colonic fermentation was conducted in healthy and Escherichia coli O157:H7-infected fecal samples. Microbial composition was analyzed via metagenomic sequencing, while microbial metabolism was assessed by ammonia quantification and short- chain fatty acids (SCFA) profile analysis. Results: Encapsulation achieved a 60% yield and preserved high probiotic viability (at levels around 8-log CFU/g) during 28- days of storage. Moreover, encapsulated cells exhibited higher survival when exposed to gastrointestinal conditions compared to free cells. In fecal fermentation assays, encapsulated A. muciniphila enhanced microbial diversity, reduced Escherichia genus abundance, and modulated SCFA profile. Furthermore, an increasing trend in ammonia production until the 30-hour mark, followed by a decline suggests potential ammonia assimilation by the probiotic. Conclusions: Calcium-alginate encapsulation is a promising delivery strategy to improve the viability and functionality of A. muciniphila, supporting its use as a live biotherapeutic agent for modulating gut microbiota and intestinal infections mitigation.
- Evaluation of the stability of Akkermansia muciniphila in a plant-based ice cream with germinated quinoa flour during storagePublication . Torres, Silvia Melissa Garcia; Teixeira, José A.; Encina-Zelada, Christian R.; Machado, Daniela; Silva, Cristina L. M.; Gomes, Ana M.Objective: The growing demand for plant-based diets and interest in ancestral crops have driven the development of innovative functional foods, such as plant-based ice creams. Akkermansia muciniphila, a next-generation probiotic, is notable for its therapeutic potential and its ability to modulate metabolic and inflammatory disorders, including obesity and metabolic syndrome. However, its incorporation into frozen food matrices has been scarcely explored. This study evaluated a plant-based ice cream formulated with germinated quinoa flour (Rosada de Huancayo variety, Peru) as a potential vehicle for delivering A. muciniphila. Methods: The formulation also included coconut and sunflower oils, along with sugars such as dextrose, glucose, and sucrose. A. muciniphila was added in free form during the mixing stage. As a control, free cells frozen in saline solution were used. Both sample types were stored at −20 °C for 60 days. Microbiological quality (enterobacteria, mesophilic bacteria, yeasts, and molds), overrun (day 0), hardness (every 15 days), and the viability of A. muciniphila (days 1, 7, 14, 21, 28, 40, and 60) were evaluated. Results: The ice cream exhibited an overrun of 11.9%, with hardness ranging from 35.9 N to 237.3 N, and maintained good microbiological quality throughout the storage period. The viability of A. muciniphila remained above 10⁸ CFU/g in both treatments, with no significant differences from the initial counts (p > 0.05). Conclusions: These results demonstrate that quinoa-based ice cream is a viable matrix for maintaining A. muciniphila stability during frozen storage, reinforcing its potential in functional plant- based foods and enabling new strategies for incorporating next- generation probiotics.