Lactobacillus Plantarum High Density Fermentation,Freeze-Dried Protectant Screening And Microencapsulation Process Research

The global market for probiotics is growing year on year,with a market size of 56.6billion$in 2020 and expected to be close to 91.1 billion$by 2026.The development and use of probiotic products has considerable market prospects.The probiotic industry has taken shape and is gaining widespread attention in general food,health food as well as pharmaceuticals.Its role has also expanded from traditional gastrointestinal health to women’s health,oral health and emotional protection.Due to the late start of the development of probiotic products in China,there are still a series of problems that need to be solved in the product development process,such as low fermentation density and freeze-drying survival rate,less live bacteria reaching the human intestine and poor storage stability.Based on the above problems,this study investigates the key processes of high cell-density culture,freeze-drying and double-layer microencapsulation of Lactobacillus plantarum,and confirms them through in vitro and storage stability.The main findings were as follows:1.Taking growth performance as an index,a starting strain was screened from the lactic acid bacteria retained in the laboratory,identified as Lactobacillus plantarum,and named it Lactobacillus plantarum Y2.The body is round-ended straight rod-shaped,without flagella,and movable.Fermentation entered the logarithmic growth phase at 2 h and stable at 16 h.2.By combining single factor and response surface method,the formula of Lactobacillus plantarum Y2 high density fermentation medium was determined as follows:glucose 30 g/L,beef extract 25 g/L,Mg SO₄3.20 g/L,dipotassium hydrogen phosphate 2 g/L,sodium acetate 5g/L,diammonium hydrogen citrate 2 g/L,Tween 80 1 g/L.Applying this medium,the number of viable bacteria could reach 2.797×10¹¹CFU/m L after 12 h of anaerobic incubation at 37℃,which is 84 times higher than the original MRS liquid medium.3.A blend of skim milk,alginate and inulin significantly improved the survival of Lactobacillus plantarum Y2 during freeze-drying.The freeze-drying survival rate was 86.87%±0.9%when skim milk,alginate and inulin were mixed in equal proportions and blended with the bacterium in a 1:8 mass ratio.The low temperature environment is beneficial to the storage of the powder,when the powder was stored at 4℃for 203 days,the viable count could still be maintained at 7.44±0.36×10⁹CFU/g.Fluorescence detection and intracellular enzyme analysis showed that the mixture of skim milk,alginate and inulin could reduce the damage of the bacteria during the freeze-drying process,avoid the rupture of the cell membrane,protect the activity of lactate dehydrogenase and ATPase in the cells,thus improve cell viability.4.Lactobacillus plantarum Y2 sodium alginate microcapsules had an encapsulation rate of 88.10%±1.34%and sodium alginate/chitosan microcapsules had an encapsulation rate of59.39%±0.91%.The survival rate of sodium alginate microcapsules mimicking digestion in vitro ended up being 66.67%±3.20%;the survival rate of sodium alginate and chitosan double-coated microcapsules mimicking digestion in vitro ended up being 73.80%±7.95%,and the survival rate of free bacteria was only 16.09%±2.40%,which improved by 50.58%±0.80%and 57.71%±5.55%respectively after encapsulation.The survival rate of free bacteria was only 16.09%±2.40%.It is shown that suitable microencapsulated materials can effectively resist the damage caused by digestive enzymes,gastric acid and bile salts during digestion and improve the survival rate of Lactobacillus plantarum.