Effects Of Dynamic High Pressure Microfluidization On The Formation Mechanism,Antigenicity,and Functional Properties Of β-lactoglobulin/Fatty Acidc Omplexes

The reduction of milk protein allergy by vary treatments was an important research at food processing,and has accumulated a large number of high-quality results.Our previous study found that the antigenicity and functional properties ofβ-lactoglobulin could be improved by dynamic high pressure microfluidization treatment,and have relevance with its structural changes.β-lactoglobulin belongs to lipocalin family and can binds with fatty acid,retinol and other small hydrophobic molecules,but there is little research about the antigenicity and functional properties ofβ-lactoglobulin/fatty acid complexes.In addition,the effect of dynamic high pressure microfluidization on the formation,antigenicity and functional properties ofβ-lactoglobulin/fatty acid complexes are also unclear in milk processing.Therefore,in this study,effects of dynamic high pressure microfluidization treatment on the formation mechanism,antigenicity,functional properties and structural changes ofβ-lactoglobulin/fatty acid complexes were studied.The main results are the following three aspects:1.Effect of dynamic high pressure microfluidization treatment on antigenicity and functional properties ofβ-lactoglobulin/fatty acid complex.The mixture ofβ-lactoglobulin and oleic acid solution were treated by dynamic high pressure microfluidization treatment(0.1,40,80,120,160 MPa)to formβ-lactoglobulin/fatty acid complex.The antigenicity ofβ-lactoglobulin/fatty acid complex was gradually decreased with pressure increasing.The solubility,emulsifying and foaming properties of complex were improved.The solubility and foaming properties improved best at 80MPa,were 95.36±1.37%and 32.00±2.00%,respectively.The emulsifying properties improved best at 120 MPa,was 1.09±0.06.2.Effect of dynamic high pressure microfluidization treatment on formation mechanism and structural changes ofβ-lactoglobulin/fatty acid complex.Under the 0-80 MPa conditions,the slightly unfolding ofβ-lactoglobulin result in the free sulfhydryl group and hydrophobic groups exposed,some binding sites destroyed,but the association constant K_a increased.This results might cause functional properties improved and antigenicity decreased due to the epitope masked by oleic acid molecule.Under 120-160 MPa conditions,the reaggreation of protein molecules led to both total sulfhydryl content and surface hydrophobicity decreased,resulting in reduced functional properties,and the association constant K_a increased significantly made antigenicity further reduced.3.The interaction betweenβ-lactoglobulin and oleic acid based on the molecular simulations.By Autodock Vina molecular docking and molecular dynamic simulation Gromacs to simulate the interaction betweenβ-lactoglobulin and the oleic acid.The results showed that,β-lactoglobulin has two binding sites-the main binding site 1 and the secondary binding site 2:site 1 within theβ-barrel,site 2 located in molecular surface between helix turn 153-157 andβ-stand B.Combined with the previous results,the unfolding of protein caused by DHPM treatment destroyed site and led to site 1exposure.In the process of oleic acid molecule binding with site 1 and site 2,the structure of the CD loop,EF loop and AA156-159 changed remarkably,and interact with Lys69 and Ser21,His161 by hydrogen bonds,respectively.This results might cause the tertiary structure of the protein changed.The solvent accessible surface area(SASA)of protein molecule increased,wherein the solvent accessible surface area of Trp 61 decreased,but the SASA of Tyr20 and Tyr42 increased.These changes might cause the fluorescence intensity of tryptophan and tyrosine residues increased.