Optimization of fermentation conditions of milk with blend probiotic strains based on high viable count

Abstract: The viable probiotic count is the key factor to functionality of fermented milk. It is very important for the milk industry to improve the number of viable bacteria in its final products. Lactobacillus casei and Bifidobacterium were blended using them as a starter culture for fermented milk to achieve a high viable probiotic count. Lactobacillus casei and Bifidobacterium were studied and a high density culture technology of Lactobacillus was applied to increase the number of viable bacteria in the fermentation system. Determination of the absorbance method was used to assess the viable count of probiotics in fermented milk in which the calcium ion would combine with salt under alkaline conditions to make fermented milk dispersed evenly in a solution state. Ethylenediamine Tetraacetic Acid (EDTA) was selected as a chelating agent to chelate the calcium ion in the fermented milk, and to make the solution clarify. The OD value of the bacteria did not change significantly after entering the stable phase, and the number of viable probiotics count in the fermented milk achieved the maximum. The OD value of fermented milk in this study did not change significantly from the second day of fermentation. On this basis, OD value was measured for three consecutive days from the second day, and the average OD value was used as indicator of the viable probiotic count.Inoculation volume、ratio of probiotics strains、added glucose,、added soybean peptide, and fermentation temperature were studied as a single factor affecting the number of viable bacterium in the fermented milk. Results show that the average OD value at different inoculation volume was not significant (P<0.01) while the other four factors were highly significant (P>0.01). The ratio of probiotics strains in blend culture、glucose level、soybean peptide level, and fermentation temperature were further studied using a Box-Benhnken design to optimize the fermentation technology. The results were analyzed with the software Design Expert to 8.0. Results indicated that the fermentation system model had an extremely significant effect on predicting the result of the test. The studied factors had significant effects on the average OD value of fermented milk. Results of the response surface optimization test showed that the optimal fermentation condition was: inoculation volume was 6%, the mass fraction of kim milk solid was 12%, the ratio of Lactobacillus casei and Bifidobacterium in blend culture was 3:1, glucose level was 2.9%, soybean peptide level was 0.8%, and fermentation temperature was at 34℃. The predicated average OD value was 1.076 with the optimal fermentation condition, and the verification tests demonstrated that the actual value was 1.087, the relative error was 1.0% compared with the predicated value. The verification tests also showed that the coagulation state of fermented milk was uniform, with less whey separation, and that there was a rich fermented milk flavor, pure without a peculiar smell. The maximum number of viable probiotics count was 4.1×1011 cfu/mL., which is 78.3% higher than the maximum number of viable count (2.3×1011 cfu/mL) before the optimization process. When stored under 4℃ after 21 d, the number of viable probiotics count remained at 4.7×1010 cfu/mL. The research result could be used as a probiotic fermentation technology applied to production of probiotic beverages. In addition, it was applied to the industrialized production of probiotic strains used in common acid milk production. Thus, the result greatly increased the viable count of probiotic in the fermented milk and improved functional properties of the product.