Shellac-based PH-Responsive Emulsion System For Co-delivery Fish Oil And Curcumin For Ulcerative Colitis

Ulcerative colitis(UC)is a chronic relapsing intestinal disease with complex etiology that seriously affects human health and increases the risk of cancer.Currently,drugs,surgery,and microbiota transplantation have been used to treat UC,but these methods have disadvantages such as large side effects,high costs,and high risks.In contrast,dietary intervention is a lowcost,well-accepted,and less side-effect method for relieving UC that has emerged in recent years.The basis of dietary intervention to alleviate UC is that dietary active components can regulate intestinal microbiota,repair the mucosal barriers and enhance immune function.However,dietary active components often have disadvantages,such as poor water solubility,low bioavailability,and rapid metabolic elimination,which seriously restricts their practical application effect in alleviating UC.Obviously,the development of a food delivery system that can deliver dietary active components to the colon and even to the inflammatory site of UC is of great significance to improve its bioactivity in relieving UC.As a food colloid with a multihierarchical structure,Pickering emulsion can not only provide a variety of choices for the loading of hydrophobic active substances but also select bioactive functional oils as its oil phase,which is expected to achieve the purpose of synergistically alleviating UC with multiple dietary active components.However,conventional Pickering emulsions will be digested in the stomach and small intestine,which cannot achieve colon-targeted delivery of active substances.Therefore,the rational design of Pickering emulsions is required to enhance its colon-targeting ability.This research took full advantage of the colon targeting properties of shellac nanoparticles(SNPs)and chitosan(Chitosan,CS),the effects of p H-induced gelation transition of CS on enhancing emulsion stability,and the Cur-encapsulation properties and UC inflammatory site targeting properties of SNPs.The SNPs-based Pickering emulsions that codelivered Cur and fish oil were constructed under the synergy of CS,and the colon-targeted delivery of fish oil and the inflammatory site-targeted delivery of Cur were realized,which effectively relieved UC in mice.The main research results of this paper are as follows:(1)Physicochemical properties of SNPs and their potential as functional Pickering particlesThe SNPs were successfully prepared by anti-solvent precipitation.The particle size of SNPs increased with the concentration of the shellac ethanol solution,and the SNPs had good polydispersity and surface potential up to-30 m V in distilled water.Moreover,the SNPs had significantly increased negative surface potentials in neutral and alkaline environments.SNPs were spherical particles with a compact structure formed by the self-assembly of shellac molecules under the hydrophobic interaction and had strong hydrophobicity.Depending on the hydrophobic interaction,Cur could be encapsulated in SNPs,and the encapsulation rate and encapsulation efficiency of Cur could reach 10% and 90%,respectively.Cur showed no obvious effect on the particle size,PDI,surface potential,and microscopic morphology of SNPs,but could significantly enhance the antioxidant capacity of SNPs.When CS was present in the system,SNPs could stabilize the emulsion and exhibited the potential as Pickering particles.(2)Construction,properties,and formation mechanism of Pickering emulsions synergistically stabilized by SNPs and CSThe introduction of CS could significantly enhance the ability of SNPs to stabilize the oilwater interface.When using SNPs-7 and 2% CS,while maintaining the SNPs: CS volume ratio at 3:7 or 5:5 and the oil volume fraction at 20% or 30%,CS could synergize with SNPs to construct stable Pickering emulsions.Both the decrease of the SNPs: CS volume ratio and the increase of the oil volume fraction increased the droplet size of the emulsions but had no significant effect on the surface potential of the emulsion.In addition,both CS and oil volume fractions increased the shear viscosity of the emulsions.The G’’ of all emulsions was larger than G’,and showed sol-like fluid characteristics.When CS cooperated with SNPs to stabilize emulsions,less CS in the emulsion could lead to weak bridging flocculation of emulsion droplets,resulting in a slight increase in G’.The increase in viscosity or bridging flocculation caused by CS could also reduce the migration ability of emulsion droplets.Meanwhile,the emulsion could significantly attenuate the oxidation of fish oil during storage,and the SNPs and CS formed a good interfacial layer at the oil-water interface.The emulsion showed good stability at p H 1.2,and the droplets aggregated but did not break at p H 6.8 and 7.4.Based on the above properties,the formation mechanism of the emulsion was revealed.In simple terms,CS interacted with SNPs electrostatically during the emulsion preparation,transforming the SNPs into amphiphilic particles with both hydrophilic and hydrophobic surfaces,thereby effectively stabilizing the oil-water interface to form Pickering emulsions.(3)Mechanism of structural transition of Pickering emulsions synergistically stabilized by SNPs and CS in response to intestinal p HAfter in vitro digestion,only a very small amount(< 2.5%)of free fatty acids was released from the above Pickering emulsions,indicating the emulsion can resist gastrointestinal digestion.The droplets became slightly larger in the gastric digestion stage but did not break,and the droplets aggregated significantly after entering the small intestine digestion stage.To explain the role of CS for emulsion resisting the stomach and small intestine digestion,the existing state of CS-SNPs composite sol and CS sol under different p H conditions was explored by rheological test and scanning electron microscope.It was found that when p H < 6,CS-SNPs and CS existed in the form of a sol,while when p H > 6.4,both CS-SNPs and CS started to gel.In addition,the rheological behavior and microscopic morphology of the Pickering emulsions synergistically stabilized by SNPs and CS in different digestive fluids were explored,demonstrating that the transformation of the emulsions in response to intestinal p H was dependent on the CS in the emulsions.CS gelled in small intestinal fluid,thus forming a more compact CS gel film on the surface of the droplets,and aggregated the droplets into the CS gel network.This structure facilitated the emulsion’s resistance to digestion in the gastric and small intestine,enabling colon-targeted delivery.(4)SNPs and CS synergistically stabilized Pickering emulsion co-delivered fish oil and Cur to relieve UC in miceThe alleviating effects of Cur-loaded fish oil Pickering emulsions(CCSEs)synergistically stabilized by Cur-loaded SNPs(CLSNPs)and CS on UC were investigated in a UC mouse model.The results demonstrated that CLSNPs could be enriched in the inflamed colon after gavage.CCSEs intervention attenuated DSS-induced weight loss in mice,reduced disease activity index(DAI),attenuated colon shortening,and decreased the levels of inflammatory markers.Meanwhile,CCSEs intervention favored the maintenance of normal colon tissue structure with reduced goblet cell depletion and increased MUC2 and tight junction protein secretion.At the same time,CCSEs intervention could also down-regulate the levels of proinflammatory cytokines and up-regulate the levels of anti-inflammatory cytokines.In addition,CCSEs emulsion reversed the imbalance of gut microbiota in mice caused by DSS treatment and promoted the proliferation of probiotics such as Lactobacillus and Bifidobacterium.Therefore,CCSEs may alleviate UC by repairing the intestinal barrier,modulating the immune response,and regulating the gut microbiota.