Study On Preparation Technology Of Lactobacillus Salivarius Microcapsule

Lactobacillus salivarius is a gram-positive bacteria that can produce lactic acid,and it is one of the food processing bacteria approved by China.As a probiotic possessed great potential,L.salivarius has gradually been a research hotspot in recent years.L.salivarius plays a key role in host when two conditions requirement are satisfied below:Enough L.salivarius can reach the gut through the gastric juice;enough L.salivarius colonizes the gut.However,L.salivarius presents usage limitation due to its poor resistance to acid and inconvenient preservation.Therefore,surviving L.salivarius is not enough,limiting their normal function after entering the body andusage.The main purpose of this study is to ferment and culture the isolated L.salivarius and microencapsulate them,so as to increase the storage resistance of L.salivarius and enhance resistance togastric acid.This study focuses on biological characteristics of L.salivarius strains isolated from gut of chickens.This study take the isolated L.salivarius as the object,the MRS medium as the material,using single factor combined with orthogonal experiments and other method to optimize the culture conditions,medium of L.salivarius were preliminarily explored.In addition,the best preparation technology of microcapsules was determined by using the single factor combined with orthogonal test and internal emulsification methods,and the storage resistance of microcapsules and their resistance to gastric acid were explored.The results showed that Calcium carbonate-containing MRS medium was employed to isolate bacteria that can inhibit Staphylococcus aureus from the gut of chickens.The isolated bacteria identified as L.salivarius show significant inhibitory effects for pathogenic bacteria such as S.aureus and Pseudomonas aeruginosa.The suitable medium and culture conditions for L.salivarius were:2%glucose,1.2%peptone,1.2%beef extract powder,0.6%yeast powder,1%sodium acetate,0.2%diammonium hydrogen citrate,0.2%p Hosp Horic acid Potassium hydrogen,0.2%Tween-80,0.04%magnesium sulfate,0.002%manganese sulfate,p H 6.4-6.8,optimum culture temperature 37?.The fermentation conditions were explored in a 20 L fermenter with a controlled temperature of 37?,dissolved oxygen(DO)of10%and rotation speed of 50 rpm.The p H was adjusted to a constant p H of5.5 using ammonia.The proliferation medium was added as total nutrient supplement after 14 h.Bacterial content reached the peak after 26 h,with OD₆₀₀value of 2.25.Microencapsulation was carried out in a 5 L fermenter.In a 3 L reaction system,1 L of the aqueous phase contained sodium alginate and calcium carbonate at a concentration of 1.8%,2 L of the oil phase was rapeseed oil,6 m L of spirulina-85 was added as an emulsifier,the speed and emulsification time were set to 400 rpm and 5 min respectively,after which glacial acetic acid was added for pre-fixation for 10 min,stirring was stopped,and then Calcium chloride solution was added for fixation,filtered and then added to 5%chitosan solution for secondary embedding for 10 min.The embedding rate of the prepared microcapsules was 87.1%,the highest viable cell count was 9.03 lg CFU/g,with good granulation,low adhesion,and uniform particle size.After 5 weeks of storage at 28°C,the viable count of L.salivarius fermentation broth decreased from 7.02 lg CFU/m L to4.64 lg CFU/m L.When protective agents such as algae sugar and diatomaceous earth were added during the preparation of microcapsules,the viable count of microcapsules with the added protective agents decreased from 6.35 lg CFU/g to 5.95 lg CFU/g.After 5 weeks of storage at 4°C,the viable count decreased from 6.31 lg CFU/g to 6.02 lg CFU/g.Not only that,the free L.salivarius at a starting concentration of 7.22 lg CFU/m L had a viable count of 7.068 lg CFU/m L after 2 h of treatment with artificial gastric juice and 7.1 lg CFU/g in L.salivarius microcapsules at a starting concentration of 7.23 lg CFU/g after 2 h of treatment with gastric acid under the same conditions,indicating that microencapsulation embedding was effective in protecting L.salivarius.In contrast,the microcapsules released a large amount of L.salivarius at 3 h after treatment with artificial enteric fluid,indicating that L.salivarius microcapsules have good enteric solubility.In conclusion,L.salivarius has good antibacterial activity.Fermentation culture and microencapsulation in fermenters can effectively increase the concentration of L.salivarius.This study establishes a microencapsulation system to enhance its stability and provides a reference for the preparation and subsequent use of L.salivarius microcapsules.