| Heat treatment is an indispensable means of food processing,which has become the key technology in the dairy production.Different degrees of heat treatment can lead to the changes of physical and chemical properties of milk in different degrees,influencing the quality of dairy products.In this paper,the thermal denatured polymers of whey protein and casein during heat treatment were analyzed,the difference of polymer formation of different heat-treated milk was explored,and the process and mechanism of thermoagglutination reaction between milk proteins were discussed.Meanwhile,the mathematic model of milk protein thermal denaturation under different heat treatment was established to provide a more intuitive description of the changes of milk protein during heat treatment,and the theoretical basis for the selection of milk heat-treatment conditions.Firstly,the microstructures,secondary structure,tertiary structure,surface hydrophobicity and thermal properties of whey protein and casein were studied under the thermal conditons of 65 ?/30 min,80 ?/15 s,95 ?/5 min and 137 ?/5 s,separately.The results showed that heat treatment had a significant effect on the thermal aggregation of milk protein,and with the degree of heat treatment increasing,the aggregation degree of milk protein was more obvious.The heat treatment resulted in the reduction of a-helix structure and addition of random coil significantly.Moreover,?-sheet structure and/?-turn structure showed a trend of increased firstly and then decreased.The surface hydrophobicity of the heat-treated milk protein was enhanced,but the surface hydrophobicity decreased slightly with the increasing of heat treatment.It was indicated that heat treatment led to casein and whey protein forming casein-whey protein polymers.Secondly,we studied the species and contents of protein in polymers by reduced and non-reduced SDS-PAGE,further using nano-liquid chromatography-electrospray ionization-tandem mass spectrometry(Nano-LC-ESI-MS/MS).The results showed that the main components of the polymers formed by thermal denatured milk protein were a-casein(a-S1-CN and ?-S2-CN),?-casein,?-LG and a-LA through disulfide bonds.In the process of thermal denaturation of milk,?-LG hydrophobic group was exposed,then exchanged by mercapto and disulfide bonds to form heat-denatured aggregates.At the same time,the denatured a-LA were combined with each other to form a larger whey protein aggregates with the formation of ?-LG aggregates.Furthermore,the whey protein aggregates combined with the ?-casein covered on the surface of the casein micelles by the disulfide bonds.And the stronger of heat-treated intensity,the significant increase binding to the ?-casein.Finally,the effects of different heat treatment time on the denaturation degree of whey protein and the combination degree of whey protein and casein were studied by Native-PAGE and SDS-PAGE electrophoresis.Matlab and SPSS software were used to fit the polynomial model.The denaturation degree of whey protein had a quadratic regression relationship with heat treatment temperature as follows:Zi=-320.101-31.532X+5.877Y+0.112X2+0.770XY-0.021Y2+0.006X3-0.0054X2Y-0.0037XY2(R2=0.985,P?<0.001);the relationship between denaturation degree of whey protein with the heat-treated time at different temperatures was established as follows:Z2=238.116-25.244X-5.039Y+0.430X2+0.446XY+0.027Y2-0.0032X3-0.003X2Y-0.0019XY2(R2=0.993,P<0.001),where X is the heat-treated time(min),and Y is the heat-treated temperature(° C).The model was very significant and low determinative error.It was able to be effective and intuitive to measure the denaturation degree of whey protein and the combination degree of whey protein and casein through the fitting of the model.Therefore,the equations proposed was suitable for the theoretical analysis of milk heat processing conditions in this study.Compared to 95 ?/5 min heat-treated yogurt,75 ?/15 min heat-treated yogurt could produce the best quality of yogurt and ensure the quality of yogurt,saving energy consumption. |
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