Abstract
This study was conducted to assess the biotechnological potential of the Bacillus subtilis A-12 strain for the development of probiotic feed additives. Lyophilisation was employed as the primary storage method to ensure the stability of the probiotic biomass. During the study, the lyophilisation tolerance of the B. subtilis A-12 strain was assessed using various cryoprotective medium compositions. Specifically, a gelatin-sucrose-milk medium, a sucrose medium, a milk medium, a gelatin-sucrose medium, and a gelatin medium were tested. The results showed that the post-lyophilisation viability of B. subtilis A-12 cells was significantly dependent on the composition of the protective medium. Among the tested formulations, the gelatin- and sucrose-based medium provided the highest cell viability after lyophilisation and showed a statistically significant advantage compared with carbohydrate-free media (p < 0.05). Although the number of viable cells decreased by approximately 25-35% following lyophilisation, further losses during storage for up to six months at 4°C were only 2-5%. This demonstrates the long-term stability of the obtained lyophilised biomass. Furthermore, the sporulating nature of the B. subtilis A-12 strain was found to enhance its resistance to dehydration and thermal stress, positively affecting the stability of the lyophilised preparation. As a result of the study, an experimental probiotic feed additive based on lyophilised B. subtilis A-12 biomass was developed. The obtained data indicate that the developed technology is promising for practical application and justify the need to assess its in vivo efficacy in animal models in the future.