Unraveling gelation in ultra-high-temperature sterilized milk: compositional changes and proteomic characterization

Gelation is the most serious quality issue during storage of UHT milk. While endogenous enzymatic activity is acknowledged as a contributing factor, the precise molecular mechanisms governing casein micelle destabilization and gel network formation remain incompletely elucidated. This study provides the first physicochemical characterization and proteomic identification of potential biomarkers associated with gelation in UHT milk. Plasmin system dysregulation was found to accelerate casein hydrolysis, concomitant with physicochemical alterations including elevated acidity, particle size, and viscosity. Proteomics identified 247 proteins, with 113 differentially expressed. These proteins primarily included plasmin system-associated proteins, calcium-binding proteins, and lipid metabolism regulators. Dysregulation of the plasmin system (including reduced activity of plasmin inhibitors such as α-2-antiplasmin, α-2-macroglobulin, and serpin A3-8) was identified as a core mechanism promoting casein hydrolysis and gel network formation. Calcium-binding proteins (e.g., S100-A12, annexin A1, and calreticulin) significantly promoted protein aggregation and gelation by mediating calcium ion release and recapture; S100-A12, in particular, may play a key role in gel network construction. Lipid metabolism disorders (such as apolipoprotein A-II/E upregulation) exacerbate fat globule aggregation, forming irreversible gelation with precipitated proteins. These insights not only deepen understanding of protein instability during storage but also provide potential biomarkers for early detection of gelation risk.