Preparation,Characterization,and Fruit Preservation Of Pickering Emulsion Based On Hollow SSPS-Zein Nanoparticles

Microbial food infection is one of the major threats to human health.Natural antibacterial compounds in plants have functional characteristics such as antioxidation,antibacterial,anti-inflammatory,nerve protection and blood pressure reduction,but volatility and low water solubility limit their development.To expand the application of natural bioactive compounds,many strategies have been built through different encapsulation methods,such as nanoliposomes,nanoparticles,nanoemulsions and nanofibers.Among them,the excellent stability of Pickering emulsion can protect the core materials in the process of micro encapsulation,which is considered to be an effective strategy for essential oil packaging and antibacterial applications.Due to its unique amphiphilic properties,zein can be used to prepare Pickering emulsion.However,Pickering emulsion stabilized by a single protein is structurally unstable and easy to aggregate.Consequently,using the interaction between protein and polysaccharide,polysaccharide adheres to the interface by protein as an anchor,which can promote the formation of Pickering emulsion with excellent stability.Based on the amphiphilic properties of zein,zein-soluble soybean polysaccharide（SSPS）composite nanoparticles（SSPS-Zein）with Na₂CO₃ or Na HCO₃ as sacrificial templates were constructed by hollow method in this study.And the formation mechanism of nanoparticles was discussed by the characterizing of interaction and particle characteristics.Then,the effect of encapsulated thymol on the structure of hollow nanoparticles was investigated,and thymol release control in different matrices was compared.Finally,hollow composite nanoparticles were used as stabilizer to form a high internal phase Pickering emulsion,and the fresh-keeping effect of Pickering emulsion on cantaloupe and blueberries was explored.The main research contents and results are as follows:（1）Hollow zein composite nanoparticles used Na₂CO₃ or NaHCO₃ as sacrificial templates.Compared with solid nanoparticles（120-270 nm）,the hollow method can significantly reduce the particle size（72-92nm）and the turbidity of the dispersion.The tertiary structure,secondary structure,and interaction force of hollow nanoparticles prepared by Na HCO₃ are similar to those prepared by Na₂CO₃.Moreover,the small size brought by it does not decline the encapsulation efficiency and antioxidant activity（encapsulation efficiency remained above 64%）.Because the hollow method expands the specific surface area of the nanoparticles,which is probably created by changes in hydrogen bonds and electrostatic interactions,the fluorescence emission peak is notably enhanced.The hydrophobicity is also strengthened to change the secondary structure,increase the content ofα-helix,and decrease the content ofβ-sheet.The SSPS coating can stabilize the particle characteristics and improve the storage stability,which is primarily completed by hydrogen bonds.The results show that the hollow nanoparticles prepared by Na₂CO₃ have smaller particle size and higher encapsulation efficiency than those treated with Na HCO₃,but the yield of hollow particles is less and miscellaneous proteins are more.And the strong alkalinity of sodium carbonate can not be neutralized by SSPS,which is not friendly to the biological activity of thymol and causes the adhesion of solid nanoparticles.In contrast,SSPS can neutralize the alkalinity of NaHCO₃,decrease the miscellaneous proteins of hollow nanoparticles prepared by Na HCO₃,make the hollow structure clearer,and stabilize its particle characteristics.Thus,NaHCO₃ may be a more suitable sacrificial template for hollow zein composite nanoparticles.（2）The hollow zein nanocomplexes prepared with NaHCO₃ as a sacrificial template were selected to load thymol,and the effect of loaded antibacterial active substances on the structure,the interaction of each component,the release and antibacterial properties of hollow nanoparticles were investigated.The results show that the overall size of the nanoparticles increases after encapsulation of thymol（～30%）,but still less than 100 nm（H-ZHS/T 95.2±1.3 nm,H-ZH/T 92.7±2.0 nm）.The hollow method decreases the particle size distribution index of nanoparticles and increases the absolute value of zeta potential,which may be that the hollow provides more electrostatic repulsion.The adsorption of SSPS also improves the stability of the nanoparticles loaded with thymol.Furthermore,the fluorescence intensity of nanoparticles enlarges about 1 time,indicating that the tertiary structure of protein has changed and more exposure of hydrophobic surface area.The change of hydrophobicity may be due to the hydrogen bond and methyl group caused by encapsulation of thymol,and the hydrophobic interaction and electrostatic interaction caused by hollowing,which alter the secondary structure of protein and promoteα-helix content together.The FTIR and XRD characteristic peaks of thymol are evidently weakened,suggesting thymol interacts with zein and SSPS molecules,and thymol is successfully encapsulated in nanoparticles from crystal to amorphous solid solution.Besides,the hollow method does not weaken the thermal stability of the nanoparticles,but the temperature of the maximum mass loss rate enhances slightly from about 299℃ to 314℃.And NaHCO₃ deteriorates the thermal stability of zein nanoparticles,but the SSPS coating can upgrade this problem.With thymol encapsulated,hollow nanoparticles show a 72-hour sustained bactericidal effect,while solid nanoparticles continue to release for only 56 hours,which may be related to the magnification of protein hydrophobic area with hollow method.（3）To further expand the application of hollow nanoparticles,hollow nanoparticles loaded with thymol were used as stabilizer to prepare Pickering emulsion,and the effect of Pickering emulsion on the preservation of fresh-cut cantaloupe and blueberries was investigated.The results show that the high internal phase Pickering emulsion（φ=0.7）stabilized by hollow zein composite nanoparticles is an oil-in-water emulsion.The oil-water interface adsorption layer of the emulsion has a gel structure formed by hollow zein composite nanoparticles and oil droplets.In the preservation of fresh-cut cantaloupe and blueberries,the hardness of blueberries（26.48%）and cantaloupe（20.80%）coated with Pickering emulsion decreases much less than that of the blank group（60.67%,63.54%）,which probably results from the excellent dispersity and continuous release control of Pickering emulsion combine low temperature environment,inhibiting microbial growth and enzyme activity,thereby reducing water loss and bacterial infection.After 15 days of storage,the appearance of blueberries coated with Pickering emulsion does not change visibly,and the shape is still plump and spherical.The decrease of L*,a*and the change of b*from negative to positive may be related to the transform of surface reflection characteristics of blueberries by the coating.The brightness and saturation of cantaloupe coated with thymol and Pickering emulsion are higher than those of the blank group,probably because the coating inhibits the production of ethylene.And the b*value is lower than that of the blank group,indicating that the effective delay of cantaloupe browning.In addition,the Pickering emulsion coating of cantaloupe has the most significant antibacterial effect within 6 days（MY 0 lg CFU/g,B 1.28 lg CFU/g）.Overall,the hollow method can control the size of nanoparticles by adjusting the sacrificial template.The size of hollow nanoparticles prepared in this paper is less than 100 nm,and more thymol is encapsulated due to the large surface area and volume ratio.Compared with sodium carbonate,sodium bicarbonate is more suitable as a sacrificial template to prepare hollow nanoparticles,and SSPS can improve its stability markedly.Furthermore,thymol transforms crystalline into amorphous solid solution,and changes the protein structure through hydrogen bonding,methyl,electrostatic and hydrophobic interactions to be encapsulated in nanoparticles.Hollow nanoparticles extend the sustained release ability of nanoparticles and show excellent antibacterial properties.Besides,hollow SSPS-Zein nanoparticles can be used to stabilize the Pickering emulsion,and the Pickering emulsion can improve the preservation of fresh-cut cantaloupe and fresh blueberries from color,hardness and microbial colonies.The microbial population less than the maximum safety limit of fresh-cut cantaloupe is from1 day to 6 days,and that of cleaned blueberries is extended from 1 day to 15 days.