Study On The High Cell-Density Freeze-Dried Powder Of Lactobacillus Rhamnosus (ATTC53103) Under Insustrialization

Lactobacillus rhamnosus (ATTC 53103) is currently one of the most well known probiotics around the world, it has rich and remarkable features to the human body. The strain is resistant to acid and bile, and it has a prominent colonization in the human intestinal, during the shelf-life period, the numbers of the strain maintain a constant quantity and so on. Industrialization of high-density Lactobacillus rhamnosus (ATTC 53103) freeze-dried powder under the conditions of mass production possess large economic value and social benefits. From the following seven aspects of this subject , a systematic study had been conducted to improve the process of proliferation and the survival rate after freeze-drying, and providing a basic application case for dairy plant. The main findings are as follows:Carbon and nitrogen source were determined through single factor experiments and orthogonal design. The optimal basic culture medium included:12% skim milk powder, 10% glucose, 1% yeast extract, 2% casein hydrolyzate, 0.15% Tween 80.At the same time single factor experiment confirmed three better growing factors of either, on the basis, orthogonal design was used to study the interact influence, Lactobacillus rhamnosus (ATTC 53103) grows well in 8% mushroom juice,5% tomato juice,8% beer. In addition, Mg2+ and Mn2+ can promote the strains amount; the best add were both 0.025%. The max viable count could reach 3.02*109 cfu/mL in the above high cell-density culture medium.37℃and 3% were determined the best temperature and inoculums for the stain growing. Under the above conditions, on the base of studying the growth curve,pH value and changes in glucose concentration of 500mL salt water bottles and 10L automatic fermentor,it was determined that pH value maintaining at 6 and adding glucose from the concentration of 4g/L were best for Lactobacillus rhamnosus (ATTC 53103) fed-batch high density culure. As mentioned above, the strains were in the logarithmic phase during 6-20 hours, the max viable count could reach 3.75*1010cfu/mL at the end of logarithmic phase. However, the max viable count were only 3.8*109 cfu/mL via automatic fermentor batch culture, the former increased by 10times.The result showed that the optimal centrifuge condition of Lactobacillus rhamnosus (ATTC 53103) is 8000rpm×20min via single factor experiments at 4℃. Skim milk, trehalose, glycerol and sodium glutamate were determined to be basic cryoprotective agents via single factor experiments and orthogonal design . Add volume were respectively 12%, 8%,1%, 0.5%. In addition to improve the solubility of freeze-dried powder,the experiment added 3% maltodextrin specially. Under these conditions the survival rate of freeze-dried strains reached 78.8%, each gram contained 5.6*1010 survival stain,instant capability were satisfactory.It was determined 10mm was the best thickness for freeze-drying, it took the shortest time for unit thickness under the above condition.Lactobacillus rhamnosus (ATTC 53103) was used to make sugar-free yogurt together with different strains of Streptococcus thermophilus and Lactobacillus bulgaricus .The screening criteria were that the yogurt has satisfactory aroma and viscosity, the fermentation speed grows rapidly and the postacid keeps weak.The selected stains were made into direct vat starter culture. Linear scaling law was applied in the blind comment on the developed sugar-free yogurt and similar market products by 12 people, the former scoring was higher than the latter by 23 points, the result proved the developed yogurt were better. The probiotic yogurt contained more than 107cfu/mL alive probiotics during the shelf life at 25℃.This study ascertains that Lactobacillus rhamnosus (ATTC 53103) can produce bacteriocin.The experiment excluded interference of hydrogen peroxide and organic acids through alkali and enzymolysis.The cell-free supernatant (CFSs) of the strain exhibited a broad spectrum of antagonistic activity against various species of gram-positive. CFSs of the strain was heat-stable (30 min at 121℃) and remained active after incubation at pH 2.0 to 10.0. CFSs of the strain was found to be sensitive to proteolytic enzymes (pepsin, trypsin, papain,α-chymotrypsin and proteinase K). Treatment with catalase did not result in any change of antimicrobial activity.