Dual-Phosphorylated Pumpkin Seed Protein Pickering Emulsions: Structural Evolution, Interfacial Regulation, and Coffee Creamer Application Quality

Abstract Pumpkin seed protein (PSP) was modified with sodium tripolyphosphate (STP) and sodium trimetaphosphate (STMP) to improve Pickering emulsions. Phosphorylation increased with STMP content but plateaued at high ratios. Mass spectrometry identified phosphorylated Ser and Thr residues, while 31P NMR indicated multiple phosphorus environments. Modification altered protein conformation and surface charge, reduced particle size, and improved dispersion without major subunit degradation. Although emulsifying activity decreased, emulsion stability increased; mixed-phosphate treatments generally produced finer droplets and better resistance to centrifugal destabilization than single-phosphate treatments. A 4:1 STP/STMP ratio combined small droplet size with favorable wettability and interfacial adsorption, whereas higher STMP content did not consistently improve overall performance. In coffee beverages, increasing emulsion incorporation intensified phase separation and altered color and flavor. At a 1:9 emulsion-to-coffee ratio, phosphorylated PSP emulsion improved perceived body and smoothness while retaining coffee aroma, outperforming commercial creamer in balance and supporting its potential use as a plant-based food.