Phosphorylation of macadamia protein isolate: Unveiling the mechanism behind enhanced solubility and high internal phase emulsion stability

Macadamia protein isolate (MPI) was phosphorylated using sodium tripolyphosphate (STPP) and sodium trimetaphosphate (STMP), and the effects of phosphorylation on its solubility, structure, and high internal phase emulsion (HIPE) stabilization of MPI were investigated. Both STPP and STMP reached the highest phosphorylation degree at 5% addition, with phosphorus contents of 3.95 ± 0.05 and 3.22 ± 0.02 mg/g, respectively. Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy confirmed the successful introduction of phosphate groups into MPI through the formation of phosphorus-containing bonds. Phosphorylation markedly altered the aggregation state and molecular conformation of MPI. In particular, STMP-treated MPI exhibited its highest solubility at 4% addition, whereas 5% STPP-treated MPI showed the best overall performance, with particle size decreasing from 197.9 ± 0.99 to 165.2 ± 0.57 nm, absolute zeta potential increasing from 25.77 ± 0.06 to 70.67 ± 0.41 mV, and solubility increasing from 14.01% ± 0.72% to 96.78% ± 0.24%. In addition, phosphorylation increased surface hydrophobicity and free sulfhydryl content, and promoted the transformation of β-sheet and β-turn structures into α-helix and random coil, suggesting pronounced conformational rearrangement of MPI. These changes improved the aqueous dispersion and interfacial adsorption behavior of MPI. Accordingly, 5% STPP-treated MPI exhibited superior ability to stabilize HIPEs, producing smaller droplet sizes, a more uniform and compact microstructure, while native MPI failed to stabilize HIPEs at an internal phase fraction of 80%. Overall, phosphorylation is an effective strategy for improving MPI functionality and broadening its application in HIPE-based food systems.