OPTIMIZATION OF THE FERMENTATION CONDITIONS AND SURVIVAL OF Bacillus licheniformis AS FREEZE-DRIED POWDER FOR ANIMAL PROBIOTIC APPLICATIONS

Preparations containing probiotic bacteria have a beneficial effect on animal health. The probiotics benefits translate into an increased interest in techniques for the preservation of microorganisms. In this study, the viability of Bacillus licheniformis (BL) ATCC 21424 strain, was evaluated in shake flask culture (Erlenmeyer 100 mL on shaking incubator) and batch 7-L stirred bioreactor under submerged fermentation (SMF), respectively. The inoculum was grown in a nutrient medium (37 degrees C, 24 h +/- 2 h, 200 rpm) and the viability was evaluated by 10-fold dilutions. The fermentation process in the bioreactor was examined at 37 degrees C for 24 h under constant agitation (200 rpm). During SMF under controlled pH and oxygen availability, the cell growth rate was measured by optical density (OD 600 nm) at different interval times (6, 12, 18, 22 and 24 h). The maximum specific rate of BL in the exponential phase was calculated 0.524 h-1. When the stationary phase was reached, the OD in SMF increased, which was 2.01 times higher than that in flask culture. Without any cryoprotectant, the cell suspension was subjected to cold shock first and then freeze-dried. The proven survival rate of cells after freeze-drying was 90.65%. The viability of BL powder decreased only by 1.09 log (CFU/mL) vs. SMF, this resistance being also due to Bacillus spp. ability to sporulate. These results convincingly demonstrated that freeze-drying could be used in the preparation of BL ATCC 21424 strain as a lyophilized probiotic product with applicability in animal nutrition.