Study On Preparation And Sustained Release Of VD₃ By Ovalbumin And Polysaccharide Complexes

Vitamin D₃ is mainly synthesized by the human body after receiving sunlight,and it is not enough to take VD₃ through food in general.As a fat-soluble vitamin,VD₃ can be easily destroyed under acidic conditions,which greatly reduces its bioavailability.The lack of VD₃has become a worldwide problem.Proteins and polysaccharides are two important macromolecules in food,which not only affect the nutritional properties of food,but also affect the texture and structure of food owing to their own properties?such as gelation,emulsifying,etc.?.The protection and controlled release of functional factors have become one of the research hotspots in the food field.In this thesis,ovalbumin?OVA?and high methoxyl pectin?PEC,DM?74%?were used to prepare OVA-PEC complexes to encapsulate and protect vitamin VD₃ in order to achieve sustained release in human gastrointestinal tract.And OVA-PEC complexes were adjusted by adding sodium alginate?SA?to improve the stability of the complexes system.The main findings of this thesis are as follows.?1?The effects of pH,mass ratio,salt ionic strength and additives on the interactions between OVA and PEC were studied.When the mass ratio of OVA:PEC is 2:1,the pectin has enough sites to bind OVA molecules.At pH=2⁴,the interactions between OVA and PEC are strong,and the results show that the main driving force of the interactions between OVA and PEC is electrostatic interaction.Hydrophobic and hydrogen bonding interactions also have a certain stabilizing effect on the systems.Moreover,it was found by differential scanning calorimetry?DSC?that the thermal denaturation temperature is significantly increased after the formation of OVA-PEC complexes,and the interactions between OVA and PEC increase the thermal stability of protein.From the SEM observation,when the mass ratio of OVA:PEC is 2:1,the microstructure of OVA-PEC complexes appears as small particles with a regular shape.?2?By comparing the VD₃ encapsulation efficiency and the particle size of complexes formed by OVA and PEC at different pH,the optimal preparation conditions were obtained.The mass ratio of OVA and PEC is 2:1,and the pH value is equal to 2.8.Endogenous fluorescence analysis indicated that the binding constant of VD₃ to OVA-PEC is found to be10¹²-10¹⁴ L·mol^-1·s^-1,suggesting that the OVA-PEC complex nanoparticles can be used as carrier for VD₃.?H<0 is obtained by thermodynamic analysis,indicating that the electrostatic interaction is the main force of the system;?S>0,indicates that hydrophobic interactions may exist in the system.Synchronous fluorescence showed a slight red shift in the maximum emission wavelength after complex formation compared to OVA alone,indicating the decrease of hydrophobicity in the microenvironmental around tyrosine and tryptophan residues.Circular dichroism analysis showed that the?-helix content of OVA decrease and the?-sheet increase after the addition of PEC.The results showed that the steric hindrance of the polysaccharide causes the protein secondary structure stretched after the introduction of polysaccharide molecules into the protein peptide chain.After adding VD₃into OVA,the content of?-helix remains unchanged,and the content of?-sheet is increased and the random curl is decreased.After the formation of OVA-PEC-VD₃ complex nanoparticles,the?-helix decrease and the?-sheet increase.The random curl drops to 15.3%,and it can be seen that the interactions among OVA,PEC and VD₃ change the direction and interaction modes of hydrogen bonds,leading to the stretch of the secondary structure of OVA,then the content of the ordered secondary structure is increased.?3?Dynamic light scattering?DLS?explored the optimal conditions to prepare the OVA-PEC-SA complexes,pH=4,SA:PEC=7:3.Compared to the OVA-PEC complexes,the encapsulation efficiency?EE?and loading efficiency?LC?of OVA-PEC-SA complexes for VD₃ are obviously increased,and the particle size of complex nanoparticles is significantly reduced.The microstructure of both complex nanoparticles from TEM are all spherical.From low temperature storage experiments,it was found that the stability of complex nanoparticles after the addition of SA is increased,indicating that SA has regulatory effect on the stability of OVA-PEC complexes.?4?The in vitro simulated gastrointestinal experiments combined with SEM were used to observe the digestive behavior of binary OVA-PEC complex nanoparticles and ternary OVA-PEC-SA complex nanoparticles loaded with VD₃.It can be seen that when the complex nanoparticles enter into the simulated gastric fluid?SGF?,a small amount of VD₃ are released from both complex nanoparticles.When the complex nanoparticles continue to enter into the simulated intestinal fluid?SIF?,the microstructure of both complex nanoparticles disintegrate,and a large amount of VD₃ are released.And through kinetic model fitting,it was concluded that the sustained release kinetics of both complex nanoparticles in the SGF and SIF are closer to Higuchi equation and belong to Fick release.Our researches demonstrated that the VD₃ can be effectively encapsulated by protein-polysaccharide complexes and could achieve the goal of sustained release in the simulated gastrointestinal tract.