Modified Whey Protein Isolate Induced By Ethanol Treatment And Its Effect On Stabilities Of Oil-in-Water Emulsion

Whey protein isolate(WPI),a specific by-product resulting from the cheese production process.It has been widely used as an emulsifying agent,texturizing agent and gelling agent,etc.However,the compact molecular configuration of WPI may result in some of its excellent functional properties(e.g.,emulsification and foaming)or biological activities(e.g.,antioxidant activity)have not been fully developed,which greatly affects its application in practical processing.Therefore,it is necessary to modify WPI to comprehensively improve its functional properties and biological activity to further expand its application in food industry.This study examined how and to what extent protein denaturation by different concentrations of ethanol affected the structure,functional properties,antioxidant properties,and digestive properties of WPI.Meanwhile,the mechanism underlying the ability of ethanol-denatured WPI to improve the stability of an oil-in-water(O/W)emulsion system was further explored to provide a theoretical basis and suggest effective strategies for widening the application of WPI in the food industry.The main results were as follows:(1)With the increase in ethanol concentration(20-80%,v/v),the molecular structure of the WPI first gradually unfolds,and then aggregate through the formation of disulfide bonds and intramolecular hydrogen bonds,as well as hydrophobic interactions,which were specifically manifested in the secondary structure,intrinsic tryptophan fluorescence intensity and ultraviolet absorption intensity,which showed a significant trend of first increasing and then decreasing(P <0.05).The solubility of WPI decreased and the turbidity of WPI solution increased(P < 0.05).In addition,the surface hydrophobicity,emulsification activity,emulsification stability and foaming property of WPI were significantly improved after ethanol treatment,especially following treatment with 40%(v/v)ethanol(P < 0.05).(2)As the ethanol concentration was increased(20-80%,v/v),the antioxidant properties(ferrous reduction ability,free radical scavenging ability,metal ion chelating ability,and oxygen radical absorption capacity)all showed a trend of first increasing and then decreasing.Compared with native WPI,40%(v/v)ethanol-modified WPI had the highest antioxidant activity due to its most suitable expansion or aggregation state(P < 0.05).(3)The effects of different ethanol concentrations on WPI digestion characteristics and antioxidant activity of digestion products were studied.The results showed that with the increase of ethanol concentration(20-80%,v/v),the proteolytic rate of in vitro digestion of ethanol-modified WPI,as well as the antioxidant properties(same as mentioned above)of the digested product all showed a trend of first increasing and then decreasing,contrary to the particle size and Zeta potential were first decreased and then increased(P < 0.05).Denaturation with 40%(v/v)ethanol resulted in the unfolding of the compact spherical structure of WPI unfold to the best state that conferred the highest digestibility and highest antioxidant activity of the digested product(P < 0.05).(4)A comparison of the physical and oxidative stability of O/W emulsions prepared by native WPI and 40%(v/v)ethanol-modified WPI during storage revealed a greater distribution of protein in the interfacial layer after WPI denaturation,such that the emulsion always had a smaller particle size,Zeta potential,flocculation index,and coagulation index during storage(P < 0.05).At the same time,peroxidation value,conjugated dienes,thiobarbituric acid-reactive substances,tryptophan fluorescence intensity,and fluorescent protein oxidation products were lower during storage(P < 0.05)of the O/W emulsion prepared with denatured WPI than with native WPI.Above all,40%(v/v)ethanol modification changed the structure characteristic of WPI and significantly improved its functional properties(e.g.,emulsification and foaming ability)and biological activity(e.g.,oxidation resistance).These results show that ethanol-modified WPI could be used as a potential emulsifier and efficient antioxidant in emulsions to improve physical and oxidation stability during storage.