Construction And Gastrointestinal Environmental Protection Of Probiotics Delivery System

Probiotics are considered beneficial for human health,and consumers’interest in them is increasing with the deepening research on their positive effects on health.Probiotics play a role in the gastrointestinal tract by regulating intestinal microflora and releasing active substances to inhibit the growth of harmful bacteria and spoilage bacteria.Thus,realizing intestinal colonization and transfer of beneficial effects.This means that probiotics should keep metabolic stability and activity in foods,and survive in large numbers in the digestive tract.However,the p H value,dissolved oxygen content,temperature in the external environment,and the strongly acidic gastric juice,various digestive enzymes and bile salts in the internal environment will cause a large number of deaths of probiotics.Therefore,it is urgent to develop the novel effective encapsulation technology to improve the viability of probiotics in food and gastrointestinal tract.Firstly,based on the controllable structure and resistance to the external environment external environment,the probiotic delivery system of emulsion was constructed to investigate the protective effect of emulsion type,cell distribution and gel structure on the gastrointestinal environment of probiotics.Further,since probiotics are highly sensitive to water in the delivery system,a shell-core microbead was constructed to improve the stability of probiotics during storage using high-precision concentric dropping pill technology,with the dormant probiotics powder placed in an independent lipid environment.The influence of composition ratio and interaction of shell,and texture characteristics and gel structure of core on the stability of probiotics’internal and external environment was evaluated.It provides theoretical basis for the long-term storage,protection and controlled release on gastrointestinal tract of probiotics.Based on the research of this paper,various probiotic fresh breath products for preventing dental caries and probiotic crystal ball products for controlled delivery in gastrointestinal tract were developed.The main research findings and conclusions are as follows:（1）Preheated whey protein isolate（WPI）was used as emulsifier to construct anti-dilution oil-in-water high internal phase emulsions（O/W HIPEs）by adjusting the concentration of WPI and whether to include glucono-delta-lactone（GDL）.Results showed that the gel strength,viscosity,centrifugal stability,thermal stability and storage stability of HIPEs were improved with the increase of WPI concentration（2.5%-10%w/v）and the addition of GDL.The three-dimensional network formed by protein aggregation in the aqueous phase and the enhanced interaction of protein coated oil droplets can promote the gel formation of emulsion.It was worth noting that the gel network formed by the high concentration of WPI and the addition of GDL had good anti-dilution stability in the buffer solutions of p H 2.5,p H 4.8 and p H 7.0,which means that the constructed HIPE can maintain complete structure during the gastrointestinal digestion stage,and can be used as a potential carrier for the encapsulation of probiotics.Based on the above emulsion,low ester pectin and calcium carbonate（Ca CO₃）were added to the continuous phase to construct O/W HIPEs with structure stability to achieve high efficiency loading of Bifidobacterium lactis V9.It was demonstrated that pectin could effectively promote the formation of continuous phase double network gel,and the strength,stability and water holding capacity（WHC）of emulsion gel also increased with the increase of pectin concentration.Although probiotics were initially dispersed in the oil phase,they were found in the oil phase,water phase and interface after the preparation of HIPEs,which was attributed to the hydrophilic surface of bacteria.Moreover,the addition of pectin to HIPE can improve the thermal stability of probiotics and simulate gastrointestinal digestive activity,which is mainly related to the formation of double network gel structure,and the tight oil drop squeeze can effectively inhibit the contact of probiotics with gastric acid and bile salt.（2）The gastric acid-resisted water in oil（W/O）emulsion was constructed based on the water taxis of probiotics and the natural physical barrier effect of emulsion.The probiotics were encapsulated in the internal aqueous phase to investigate the effects of ordinary,aqueous,oil or biphasic gel on the physical and chemical properties of emulsion and probiotics encapsulation.Results suggested that water phase,oil phase or biphasic gel can improve the stability of W/O emulsion.The ordinary and aqueous gel W/O emulsions were fluid,while oil phase gel and biphasic gel W/O emulsions were solidified.Rheological analysis revealed that oil phase gel and biphasic gel W/O emulsions were weakly gelled.Furthermore,Cryo-SEM clearly indicated the gel network in the aqueous phase,and CLSM images showed that probiotics were completely encapsulated in the internal aqueous phase of all W/O emulsions.In addition,water phase gel and biphasic gel W/O emulsions improved the storage stability of probiotics at 4℃,but the long-term stability was poor.Most importantly,all W/O emulsions exhibited good probiotic activity retention during gastric digestion,while the oil phase gel and biphasic gel W/O emulsions significantly improved the stability of probiotics during intestinal digestion due to delayed lipolysis.（3）Since above-mentioned results indicated that probiotics were highly sensitive to water.The shell-core microbead delivery system was constructed by the high-precision concentric dropping pill technology to improve its long-term storage stability and gastrointestinal digestive activity maintenance.On the one hand,the effects of shell components and interactions on the physicochemical properties of microbead and the stability of probiotics were investigated.It was found that adding low ester pectin to the shell or/and carrying out gel treatment could improve the physical and chemical properties of the shell film,such as light transmittance,hydrophobicity and gel properties.The shell-core microbead loaded with probiotics had a spherical structure with uniform size and less than 5%water content.It was able to maintain high probiotic activity after long-term storage at 4℃,because probiotics were placed in an independent lipid environment to slow down the impact of water and oxygen.Besides,the shell gel microbeads（Cro-Pec-Mb）showed good thermal resistance and gastrointestinal activity,which was related to the integrity of the structure in the digestive stage,and part of the gel of the bead shell could effectively inhibit the direct contact of probiotics with gastric acid and bile salt.On the other hand,the effects of the texture characteristics of the microbead core on the protection of probiotics in the gastrointestinal tract were investigated by solid fat addition and oleogel technology.Results showed that the kernel lipids had more stable crystal structure and higher oil binding capacity（OBC）with the increase of the proportion of cocoa butter replacer and the concentration of gel agent.Additionally,no chemical interaction was existed between the components of the inner lipid formed by adding cocoa butter replacer,and the gel-like structure was constructed by molecular self-assembly or module.Comparatively,the microstructure of the core lipid is mainly petal-like crystalline aggregates,and intermolecular hydrogen bonds existed in core lipids prepared by oleogel technique withβ-sitosterol andγ-oryzanol as gels.Overall,both methods successfully encapsulated probiotics and had high initial probiotic activity,which had better long-term storage stability（6 months）under cold storage conditions,and the probiotic activity is higher than 6.0 log CFU/g.Among them,the oleogel technology was found to be more effective in constructing kernel lipids.Furthermore,in vitro simulated digestion showed that the microbead could maintain a complete spherical structure in the stomach and small intestine,and released the inner lipid in the colon.The increase in the proportion of cocoa butter replacer and the concentration of gels effectively improve the maintenance of intestinal probiotic activity.In addition,the study found that the microbeads prepared with corn oil and cocoa butter replacer at a1:9 ratio,and a compound gel concentration of 6%,maintained a complete spherical structure after 2 hours of oral administration in rats.Moreover,6%of the microbead still remained structurally intact after 4 hours,while the core of the 1-9 samples may have leaked due to the cracking of the fully swollen shell under the continuous peristalsis and stirring of the stomach for high core hardness.This study can provide insights for the construction of probiotics delivery system,and establish a theoretical basis for achieving the high gastrointestinal activity of probiotics.