Abstract: This study investigated the hydrothermal phosphorylation modification of whey protein isolate (WPI90) using mixtures of sodium hexametaphosphate (SHMP) and tetrasodium pyrophosphate decahydrate (SPP) at different mass ratios (1:1, 1:2, and 2:1) applied at varying mass concentrations (0.03, 0.06, and 0.09 g/100 mL). The secondary structure, thermal stability, solubility, foaming properties, emulsifying properties, and water-holding capacity of WPI were determined before and after phosphorylation. The results indicated that the negative charges introduced by phosphorylation altered the structural characteristics of WPI. This modification resulted in an increase in α-helix content in the secondary structure and induced noticeable microstructural changes. Phosphate addition enhanced the thermal stability and kinetic stability after heating to varying degrees; the phosphorylated WPI solutions remained clear and transparent after heating, showing a turbidity approximately 80% lower than that of unmodified WPI90. All phosphorylation modifications altered the foaming and emulsifying properties of WPI. Notably, the addition of the 2:1 mixture at 0.06 and 0.09 g/100 mL resulted in a 3- to 4-fold increase in the emulsion stability of WPI and increased the water-holding capacity from 20% to 50% and 90%, respectively, demonstrating a remarkable modification effect. This study provides a feasible approach to address the stability limitations of WPI in food processing.

Key words: phosphate mixtures; hydrothermal phosphorylation; whey protein isolate; functional properties