Preparation,Properties And Stability Mechanism Of Perfume Aldehyde/Protein Emulsion

Emulsion is widely used in the food industry,and the physical and chemical properties of the emulsion can be regulated by the modification of the emulsifier.Protein is a natural emulsifier with excellent emulsification and high biocompatibility,which meets consumers’requirements for safety,health and environmental friendliness.More and more attention has been paid to protein stabilized emulsion system.However,protein emulsions are easily affected by environmental factors（such as p H,temperature,ionic strength）during production,processing,storage,transportation,and digestion,resulting in emulsion instability,including gravity separation,flocculation,coalescence,and Ostwald ripening.Previous studies have shown that the use of dynamic imine chemical reaction between cinnamaldehyde and protein at the interface can improve the properties of the interface layer and the stability of protein emulsions.On this basis,this study first systematically compared the effects of different structures of perfume aldehydes and proteins on the formation and stability of emulsions to explore the universality of the interaction between aldehydes and proteins in the preparation of protein emulsions;processing（resistance to salt ions）and digestion stability of protein emulsions after protein interface interaction,and finally by studying the interfacial rheological behavior and chemical reaction kinetics between aldehydes and proteins to reveal the stabilization mechanism of aldehyde-protein interface interactions on emulsions,the safety and nutrition of the obtained emulsion were also investigated.The main findings of this study are as follows:1.The effect of perfume aldehyde/protein structure on emulsion formation and stabilityPerfume aldehydes with different structures（aromatic aldehydes:cinnamaldehyde CA,vanillin VL;branched aliphatic aldehydes:citral CT,citronellal CN;linear aliphatic aldehydes:valeraldehyde VA,octanal OT,decanal DC）and proteins with different structures（globular protein:whey protein isolate WPI,soy protein isolate SPI;flexible protein:sodium caseinate SC）were used to construct emulsions,and the effects of different aldehydes and different proteins on the formation and stability of emulsions were explored.The initial particle sizes of the emulsions prepared by different perfume aldehydes and whey protein isolate were all between 150 and 200 nm and the droplets were evenly distributed.The absolute values of the zeta potential were all higher than 30 m V and the stability of droplets mainly depended on electrostatic repulsion force.Only DC emulsion and blank emulsion appeared phase separation.The long-term stability of emulsions was predicted by backscattering spectroscopy,and the results showed that the emulsions prepared by CA and VA had higher stability under p H conditions（p H2 and p H7）far from the isoelectric point of proteins.Select CA and VA with the best stabilization effect from perfume aldehydes to prepare emulsions with different proteins.SC and SPI emulsion droplets aggregated to form clusters,while WPI emulsion droplets remain dispersed and evenly distributed,which had advantages in long-term stability.Compared with VA,the synergistic effect of CA in improving emulsion stability was more significant.The microstructure（SEM）of the WPI/CA emulsion showed that after a sufficient reaction time,more proteins were coated on the surface of the droplets.The interfacial proteins and the water-phase proteins were cross-linked to form an ordered network structure that embedded the droplets into in the network.2.Stability of perfume aldehyde/protein emulsions in high salt and gastric environmentsTaking CA as the aldehyde component,to study the tolerance of different protein emulsions to salt ions(Na⁺,K⁺,Ca²⁺,Mg²⁺and Fe²⁺)in the presence of CA,as well as the stability of emulsions during gastric digestion.For blank protein emulsions,monovalent ions promoted droplet aggregation mainly through their ability to shield electrostatic repulsion.The presence of divalent ions led to larger droplet size than monovalent ions and the formation of a viscoelastic network,which was attributed to electrostatic shielding,ionic binding and ionic bridging of divalent ions.The addition of CA significantly reduced the sensitivity of the emulsion to monovalent ions and increased the critical concentration of divalent ions to induce droplet aggregation.In vitro gastric simulated digestion of WPI emulsions in the presence of different types and concentrations of ions showed that the emulsion droplets gradually aggregated into agglomerates with reduced electrostatic repulsion,increased bridging effect,and interfacial adsorption proteolysis,while CA significantly reduced the emulsion droplets agglomerate particle size in the stomach,slowing down the magnitude of particle size increase and increasing the resistance of droplets to aggregation during gastric digestion.For the SPI emulsions and SC emulsions,CA significantly improved their tolerance to monovalent ions,while the ability of the emulsions to resist aggregation in the presence of divalent ions was not as effective as the WPI emulsions.3.Stability mechanism and safety and nutritional evaluation of perfume aldehyde/protein emulsionThe interfacial properties of perfume aldehydes and WPI and the chemical reaction kinetics in the homogeneous system were studied to explore the mechanism of emulsion stabilization.In addition,the safety and nutritional properties of WPI/CA emulsions were investigated by in vitro simulated gastrointestinal digestion and Caco-2 cell transport assays.The interfacial rheological results showed that the ability of perfume aldehyde to reduce the interfacial tension was in the order of CA>VA>CT>CN>VL.In the test,the interfaces of OT oil droplet and DC oil droplet were very fragile and collapsed in a short time.More proteins were adsorbed on the interface of the WPI-CA system,and a solid-like interfacial film was formed,whose interfacial property were regulated by p H values.The results of chemical reaction kinetics showed that the maximum solubility of CA in water was about 1.57 mg/m L.With the progress of the reaction,the concentration of free CA decreased rapidly in the first 2hours,and basically reached an equilibrium state after 8 hours of sufficient reaction.The progress of the Schiff base reaction between CA and WPI was controlled by the aldehyde content and there was a reaction limit.During in vitro digestion,the emulsion structure changed.The blank protein emulsion aggregated in the stomach and exposed a large number of free oil droplets,while most of the oil droplets of the WPI/CA emulsion were still encapsulated by proteins,which can be attributed to the interfacial cross-linking of CA with WPI.After digestion in the small intestine,the free fatty acid（FFA）release rate of the WPI/CA emulsion was only about half of that of the blank protein emulsion,indicating that CA could play a role in reducing the release rate to protect oil droplets.Caco-2 cytotoxicity experiments showed that the emulsion was nontoxic in the range of protein concentration of 0.00-5.94μg/m L.A small amount of peptides in the emulsion digest could completely penetrate the Caco-2 cell monolayer,while most peptides were further hydrolyzed,and more peptides in M70C30 could be absorbed.