Study On Co-microencapsulation Of Highland Barley β-glucan And Probiotics And Its Application In Cheese Product

Probiotics are living microorganisms that,when administered at adequate levels,confer health benefits to the host,including enhancing immunity,regulating intestinal flora balance,treating inflammatory bowel disease,lowering cholesterol and blood lipids,and even inhibiting tumors and cancer.However,probiotics are susceptible to stress from harmful environmental factors,which can easily lead to loss of viability and death when exposed to high temperature,low p H,and high osmotic pressure.In this paper,probiotic microcapsules were prepared by spray drying method using soybean isolate protein,pectin(PEC),highland barley β-glucan(HBBG),and transglutaminase(TGase)as wall materials,and the protective effects of different formulations of microcapsules on probiotics were evaluated via in vitro simulated gastrointestinal digestion,storage stability,and heat resistance,to determine a better microcapsule formulation.On this basis,animal experiments were conducted to investigate further and analyze the digestive properties and colonization potential of probiotic microcapsules in the gastrointestinal tract of mice,as well as their effects on intestinal microecology.Finally,probiotic microcapsules were added to the food matrix to develop a probiotic cheese product.The main experimental contents and results are as follows:Firstly,four soybean isolate protein matrix probiotic microcapsules(SPI,SPIPEC,SPI-TGase,SPI-PEC-TGase)were prepared by spray drying to investigate the protective effects of microcapsules with different formulations on probiotics in terms of in vitro simulated digestion,storage stability,and thermal stability.It was found that all the microcapsule samples obtained by spray drying had a live bacterial count greater than 9 Log CFU/g,moisture content lower than 7%,and better powder quality.In the tests of in vitro gastrointestinal digestion,storage stability,heat resistance,etc.,it was found that the survival rate of microencapsulated probiotics were significantly higher compared with free probiotics,indicating that microencapsulation can effectively protect probiotics from damage by environmental harmful factors.Among the tested microcapsule samples,SPI-PEC microcapsules provided the best protection for probiotics,with a probiotic spray drying survival rate of 50.52% and only 0.43 Log CFU/g loss in vitro digestion,mainly attributed to the good thermal stability and digestion resistance properties of pectin,which improved the structural morphology of SPI matrix microcapsules and provided an excellent microenvironment for probiotic survival.In addition,SPI-TGase microcapsules also significantly enhance the antidigestive,heat,and UV resistance of probiotics,mainly due to the covalent crosslinking of SPI induced by TGase,which gives the microcapsules a stronger and denser structure to increase the physical barrier effect of the microcapsules.Then,HBBG was added to SPI-PEC and SPI-TGase matrix at different concentrations(0.1%,0.5%,1.0%)to prepare probiotic microcapsules by composite coalescence and enzymatic cross-linking,respectively,to investigate the effect of HBBG on the resistance performance of probiotics.For the composite coagulation microcapsule,the addition of HBBG significantly improved the structure and physicochemical properties of microcapsules,the moisture content of microcapsules decreased with the increase of HBBG,and the microcapsules of corresponding formulations showed better storage stability.The scanning electron microscopy images showed that the addition of HBBG significantly improved the apparent morphology of microcapsules,reduced the cracks and voids on the surface of particles,with a smooth surface,and enhanced the structural density and physical barrier effect of microcapsules.At the same time,the appropriate amount of HBBG significantly improved the protection effect of composite cohesive microcapsules on probiotics.When HBBG was added at 0.5%,the survival rates of spray-dried and in vitro simulated digestion of encapsulated probiotics increased to 57.23 ± 2.39% and 40.00 ±3.30%,respectively,which may be attributed to the prebiotic properties of HBBG and the enhanced barrier effect of microcapsules.For the enzymatic cross-linked microcapsule,the protective effect of different concentrations of HBBG on probiotics was similar to that of composite cohesive microcapsules,and when the additional amount of HBBG was 0.5%,it had the best protection effect on probiotics,with the survival rates of spray drying and gastrointestinal digestion up to 46.05 ± 0.75% and64.35 ± 3.94%,respectively.Subsequently,the digestive properties and colonization potential of highland barley β-glucan and probiotic co-microcapsules in mice were investigated,as well as their effects on intestinal microecology.In vivo gastrointestinal digestion experiments showed that all the microcapsules significantly improved the in vivo digestive survival of probiotics compared with free probiotics,with viable counts higher than 7.0 Log CFU/g.In addition,the addition of HBBG to the microcapsules further enhanced the viability of probiotics,attributed to the good physicochemical properties and prebiotic efficacy of HBBG,consistent with the in vitro results.By incubation the gastrointestinal contents(stomach,duodenum,jejunum,ileum,cecum,and colon),it was found that the vitality of probiotics in the HBBG microcapsules groups(S-L-0.5%H-P,S-L-0.5% H-T)were significantly higher than the other groups,and viable bacteria could still be detected in the cecum and colon contents at 32 h posed gavage,indicating that HBBG significantly increased retention time of microencapsulated probiotics in the intestine and had the potential to promote probiotic colonization in the cecum or colon.High-throughput sequencing results showed that HBBG significantly increased the abundance of Lactobacillus and Bifidobacterium compared with the control group;HBBG microcapsules significantly up-regulated the abundance of Lactobacillus,Bifidobacterium,Akkermansia,Parabacteroides,and Clostridium,etc.,to promote the proliferation of beneficial bacteria and improve the intestinal microecology.In addition,by measuring the organic acid content in feces,it was found that HBBG could be degraded by intestinal flora and promote the production of shortchain fatty acids,especially propionic acid,and butyric acid;Lactobacillus plantarum promoted the production of acetic acid and butyric acid.Co-microencapsulation of highland barley β-glucan with Lactobacillus plantarum further enhanced the content of short-chain fatty acids such as acetic acid,propionic acid,and butyric acid,which has the potential for a synergistic effect.Finally,probiotic microcapsules are added to the food matrix to develop probiotic cheese products with the best formula of 7% white sugar,4% gelatin chips,20% cheese slices,and 1.5% probiotic microcapsules(S-L-0.5% H-P).The product is light yellow in color,with uniform tissue texture,sufficient milk flavor and pleasant aroma,delicate,lubricious and moderate sweetness.Compared with commercially available cheese products,there was no significant difference in color and tissue state,and the sensory score was slightly lower in taste and flavor because only basic ingredients were added.The results of in vitro digestion simulation showed that compared with the commercially available probiotic cheese products,the self-made probiotic microcapsules cheese products had ideal gastric acid resistance and bile salt resistance,and the survival rate was further increased to 80% with high bioavailability.The selfmade probiotic cheese products showed good storage stability with no significant decrease in the number of viable bacteria when stored at 4 ℃ for 8 weeks.