Immobilized Lactobacillus Fermentation L-Lactic Acid Study And Application

In this paper, the Lactobacillus casei was focused on. Fistly, the quantification for L-lactic acid was studied and the capillary electrophoresis method was established. The mutant strain ZZ-06 was bred by the complex radiation of UV & laser on the Lactobacillus casei protoplasts, followed with metabolic regulation. Subsqently, physiological and biochemical properties, key enzyme activity and genetic stability of ZZ-06 were investigated respectively.The response surface methodology (RSM) was used to optimize nutritional complements and culture conditions. Also, the embeding method for ZZ-06 were studied and immobilization conditions was optimized by RSM. The scanning electron microscopy was used to observe the micro-strcture of immobilized ZZ-06 cell for verifying the embeding effect. Finally, the immobilized ZZ-06 cells were applied to fermentation experiments to research its actual acid-producing capability for the future soy sauce yielding.The content of L-lactic acid was determinated by capillary electrophoresis method under the optimized operating conditions and buffer system:21KV of separation voltage,254 nm of detection wavelength, pH 7.5 of buffer system (5 mM of potassium hydrogen phthalate and 0.6 mM of CTAB), at 20℃. At 50～1000μg·mL-1 concentration range, the linear equation was Y= 02389X+2.778, R2= 0.9996, which suggest a good linear relationship.The high-yielding mutant strain ZZ-06 was obtained by the UV & laser complex radiation and combined with metabolic regulation on the Lactobacillus casei protoplasts. The optimal conditons for acid-producing was achieved by RSM:110g/L of glucose,0.279831g/ L of MnSO4·4H2O,17.9882 g/L of yeast extract,33.9029 ml/L of wort 0.6g/L of MgSO4·7H2O,50g/L of CaCO3, and 8% of 14-hour inoculum. The optimum growth temperature for Lactobacillus casei ZZ-06 was 38℃, which was not in agree with 43℃of fermentation temperature.Sodium alginate was chosed as the most suitable embedded material to immobilize Lactobacillus ZZ-06, The RSM was used to optimize immobilization conditions, and the optimal preparation conditions was as following:3.445g/L of alginate,1.4549g/L of calcium chloride 13.917 h of cross-linking time. The predictive value of L-lactic acid production was 107.8434g/L, which was in agree with 6 times of verified experiments and indicating that the model can predict the actual Lactobacillus casei immobilized ZZ-06. SEM observation on the immobilized small plastic beads ball showed the embedding effect was remarkable. Simultaneity, the fenmentation stability of immobilization gel beads was studied with several times of acutal yielding, which was unconspicuous on the decrease of L-lactic acid.The soy sauce production experiment showed that adding of the immobilized lactic acid bacteria can producing acid in soy oil without pasteurism and even at lower temperauter, and the 10% of inoculum was proper, which should be instructional for the future yielding of soy sauce.