Studies On High Cell Density Culture Technology Of Probiotic L. Casei Zhang

High cell density culture is an important link of probiotic starter prepares, and plays important role in developing probiotic yoghurt or fermented dairy products. In this paper, L. casei Zhang, a potential probiotic lactic bacteria, was choosen as original culture to prepare starter culture. Optimization of medium and culture condition, substrate inhibition, nutrient supply, et al. were studied to complete the high cell density culture and probiotic starter prepare.Single factor test, orthogonality test, response surface analysis were used to design the Multi-Regression Equation with cell dry weight of L. casei Zhang(Y), carbon source(x1), nitrogen source(x2) and buffer salt(x3): Y= 2.30+0.47x1+0.59x2+0.22x3+0.18x1x2+0.16x1x3- 0.047x2x3- 0.25x12- 0.27x22-0.084x32 + 0.100x1x2x3-0.41x12x2-0.039x12x3-0.20x1x22Optimal medium was determined by the Multi-Regression Equation: glucose 20.9 g/L, soy peptone 10.45g/L, yeast extract 10.45g/L, K2HPO4 3.5g/L, sodium acetate 14.6g/L, citrate sodium 2.35g/L, MgSO4?7H2O 1.0g/L, MnSO4?5H2O 54mg/L, Tween 80 1.0 g/L. Cultivated in the optimal medium for 18h at 37℃, the living cells of L. casei Zhang were increased to 4.78×109cfu/mL.Optimal culture condition was obtained by Single factor test: temperature 37℃, revolution speed 0rpm, initial pH6.5, inoculation amount 1×106 cfu/mL. On the basis of optimal medium and culture condition, the growth kinetics model was obtained:Studied on the growth of L. casei Zhang in a 3 L fermentor, result showed the optimum fermented condition was: culture temperature 37℃, inoculation amount 7% after seed liquid was cultured for 10h(1.5×108 cfu/mL), revolution speed 300rpm, 17.5% ammonia water as neutralizer, pH5.9(initial pH6.5), added nitrogen 200mL/2h, supplied all the nutrient at one time. The final medium components were: glucose 8.08g/L, soy peptone 12.8g/L, yeast extract 8.02g/L, K2HPO4 3.18g/L, sodium acetate 13.27g/L, citrate sodium 2.12g/L, MgSO4?7H2O 0.88g/L, MnSO4?5H2O 47.77mg/L, Tween 80 0.88g/L. Cultivated under the optimum fermented condition for 12h, the living cells of L. casei Zhang were increased to 3.0×1010cfu/mL.As the basis of optimum fermented condition in 3L fermentor, cultivated the L. casei Zhang in a 30L fermentor, the living cells were 2.89×1010cfu/mL, made the fermentation broth to powder, the living cells were 4.0×1011cfu/mL, and the cell vitality was 5.16u.