Gastrointestinal Tolerance and Gut Microbiota Modulation of Encapsulated and Free Forms of <i>Lactobacillus acidophilus</i> and <i>Bifidobacterium animalis</i> subsp. <i>lactis</i>

The viability and efficacy of probiotics are strongly influenced by their delivery form, especially under harsh gastrointestinal conditions. This study assessed the gastrointestinal resistance of encapsulated and free from <i>Lactobacillus acidophilus</i> and <i>Bifidobacterium animalis</i> subsp. <i>lactis</i>, and evaluated the specific effects of digested free from strains on the gut microbiome. An <i>in vitro</i> digestion model simulating gastric and intestinal conditions was used to assess probiotic survival, followed by fecal fermentation to examine microbial and metabolic responses. The encapsulated probiotics, formulated with a multilayer matrix, demonstrated significantly higher viability and preserved membrane integrity than the free forms. Despite reduced viability, free from probiotics modulated the gut microbiota. Both strains promoted colonization of beneficial genera such as <i>Bifidobacterium</i>, <i>Lactobacillus</i>, and <i>Prevotella</i>, while maintaining microbial balance without inducing dysbiosis. Additionally, probiotic supplementation enhanced short-chain fatty acid production, particularly acetate and butyrate, indicating increased microbial fermentation activity. Notably, distinct ecological patterns emerged between the two strains, with <i>L. acidophilus</i> inducing dynamic shifts and recovery, and <i>B. lactis</i> contributing to structural stability. These findings underscore the importance of strain-specific selection and encapsulation strategies in developing targeted and effective probiotic interventions.