| Rice proteins(RP),a by-product of rice starch production,are a potential alternative to animal proteins due to their large production,reasonable amino acid composition,and low allergenicity.However,the extensive disulfide bonds and hydrophobic interactions between rice protein molecules lead to high self-aggregation capacity and poor processing performance.Surface patch binding(SPB)is a process in which electrolytes with the same kind of charging properties search for sites on the surface of each other with opposite charges to their own and undergo electrostatic binding,which is also an important basis for the development of proteinpolysaccharide electrostatic complexes.Rice protein can achieve different self-assembly behaviors under different p H conditions because of its strong hydrophobicity;carboxymethyl cellulose(CMC)is a water-soluble polysaccharide whose charge density can be regulated by degree of substitution(DS).In this study,a spherical chain-like protein-polysaccharide supramolecular structure was constructed using SPB;the supramolecular structure was induced to self-assemble through liquid-liquid interfacial stacking action,forming an interfacial structure with skin-like properties(polysaccharide epidermis/protein dermis)which found important applications in intestinal delivery and 3D printing.In addition,the proteinpolysaccharide supramolecular structure is thermally induced to achieve the solid-liquid interface gel structure,which provides a new idea for the gel preparation of hydrophobic plant proteins.The details of the study are as follows.Formation mechanism of RP-CMC supramolecular structure mediated by the SPB effect.The results of water-dispersity measurement showed that the solubility of RP increased from1.7% to 38.3%～93.5%.Atomic force microscopy results confirmed that the presence of CMC altered the aggregation state of RP,and cryo-scanning electron microscopy results indicated that the RP-CMC composites were supramolecular co-assemblies composed of chain-like CMC with a width of 30 nm and spherical RP with a diameter of 200 nm.Analyzing the dynamic structural changes of the protein during p H cycle,it was found that alkali treatment puts RP in the molten globule(MG)state,and during neutralization,the positively charged regions on the surface of RP and the negatively charged CMC under electrostatic attractive force undergo SPB,and the CMC improves the anti-folding ability of RP from the level of the secondary structure,which inhibits the aggregation of the protein and maintains the unfolded conformation of the RP.The molecular weight and charge density of CMC as well as the mass ratio of RP/CMC had an influence on the formation of the RP-CMC composites,in which the small molecular weight of CMC possessed a stronger RP-binding ability,which was mainly reflected in the greater solubility and lower hydrophobicity,the study of the RP/CMC mass ratio found that when it was 10:1,the prepared composites exhibited maximum solubility.Besides,the structural stability,surface hydrophilicity,chargeability,charge shielding resistance,and emulsification of the RP-CMC composites were positively correlated with the CMC charge density.RP-CMC composites interfacial behavior and Pickering emulsion preparation.The CMC charge density-modulated RP-CMC composites constructed by p H-cycle induced hydrophilic modification of RP,and the three-phase contact angle was reduced from 160° to 91°～120°.Analysis of the behavior of the RP-CMC composites at the oil-water interface showed that the diffusion arrangement of the protein-polysaccharide composites at the oil-water interface had an obvious orientation,i.e.,the protein was located in the oil phase while the polysaccharide was in the outer layer facing the water phase,and the greater the charge density of the CMC,the greater the thickness of the polysaccharide adsorption layer.The emulsion microscopic morphology,particle size and rheological analyses showed that the emulsions prepared by the RP-CMC composites exhibited CMC charge density-dependent behavior,and the larger the charge density,the smaller the particle size and the larger the storage modulus(G’).This is because the larger the CMC charge density leads to the stronger the surface charge of the RPCMC composites,which generates strong electrostatic repulsion and spatial site resistance on the surface of the oil droplets,effectively inhibits the aggregation of the oil droplets,and improves the stability of the emulsion.The Cryo-SEM results of the emulsion showed that there were fibers and particles on the interface,which confirmed that the RP-CMC composites were a Pickering stabilizer.Meanwhile,the G’ of the Pickering emulsion was significantly improved compared to the emulsion prepared from RP.For example,the RP-CMC 1.2 emulsion showed no significant particle size change during 30 d of room temperature storage.In vitro digestion simulation experiments showed that the emulsion stabilized by RP-CMC composites was stable to resist the damage from gastrointestinal fluids.Using the oil-soluble molecule coumarin-6 as a delivery object,it was found that this Pickering emulsion could precisely control the retention time of coumarin-6 in the intestinal tract with a precision of 6～24 h mediated by the CMC charge density.Preparation and application of the high internal phase Pickering emulsion(HIPPEs)stabilized by RP-CMC composites.The preparation conditions of HIPPEs were optimized based on the results from the particle size,microstructure and rheological properties of the emulsions,with a continuous phase concentration of 2.5%,a p H of 7.0 and a φ of 0.85.The HIPPEs prepared under these conditions after treatment at 90°C for 30 min or storage at 4°C for 10 d could still maintain the self-supporting property.The mechanism of HIPPEs stabilization regulated by CMC charge density was analyzed,and it was found that the greater the CMC charge density,the greater the stability of HIPPEs,which was attributed to the stable and charged structure of the RP-CMC composites.As the RP-CMC composites with larger charge density had higher emulsification activity as well as stability,which endows the stronger anti-aggregation ability between oil droplets and resulted in enhanced the stability of HIPPEs.The HIPPEs prepared from RP-CMC composites were applied to 3D printing inks,and the textural analysis of the HIPPEs showed that the hardness and adhesiveness of HIPPEs were found to be positively correlated with the charge density of CMC,whereas adhesion was negatively correlated.As a result,the HIPPEs prepared by RP-CMC 1.2 had the strongest extrusion molding ability and the highest 3D printing resolution.With the improved solubility of RP,the effects of gel concentration,heating time,and RP/CMC mass ratio on the microstructure and rheological properties of thermotropic gels prepared from RP-CMC composites were investigated.The results showed that the minimum gel concentration of RP-CMC composites was 8%,and the gel strength was positively correlated with the concentration of RP-CMC composites and the mass ratio of RP/CMC.With the increase of heating time,G’ showed a trend of first increase and then decrease,in which the maximum gel strength could be obtained by heating RP-CMC 1.2 at 90°C for 4 h.With the increase of CMC charge density,the gel strength increases,which may be due to the fact that the structure of RP-CMC 1.2 is more unfolded,and the protein is more prone to conformational changes during heat treatment,exposing more positively charged regions to bind with polysaccharides under the electrostatic attraction,which improves the heat-resistance of polysaccharide chains and at the same time encourages more protein particles to self-aggregate under the force of hydrophobic interactions,disulfide bonds,hydrogen bonds,etc.,thus building an ordered protein gel network. |
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