Structural and Viscoelastic Changes in Fresh and Reconstituted Skim Milk Induced by Microbial Transglutaminase

ABSTRACT This study investigated the time‐dependent structural and viscoelastic responses of fresh and reconstituted skim milk to microbial transglutaminase (MTGase) using dynamic oscillatory rheology and intrinsic fluorescence spectroscopy. Samples were incubated with or without MTGase at 40°C for 120 min. Untreated samples showed only minor rheological changes, whereas MTGase markedly increased the storage modulus ( G ′) and loss modulus ( G ″) in both milk systems. Reconstituted skim milk exhibited a rapid initial increase in viscoelasticity followed by stabilization, while fresh skim milk showed a more sustained increase and developed a higher final G ′. Intrinsic fluorescence intensity decreased progressively during incubation, with a greater reduction in reconstituted milk, while no pronounced or systematic shift in the emission maximum was observed. These spectral changes were therefore interpreted as reflecting MTGase‐associated reorganization of the molecular and colloidal protein environment rather than as a direct measure of covalent crosslinking. PCA clearly differentiated the major treatment‐dependent trajectories and captured 99.93% and 99.41% of the total variance in the rheological and fluorescence datasets, respectively. The differing temporal patterns observed by the two techniques indicate that molecular‐scale spectral changes were not necessarily proportional to macroscopic viscoelastic development. Because the fresh and reconstituted milks differed in both composition and processing history, the observed differences cannot be attributed specifically to either factor. Overall, the exploratory measurements revealed different temporal patterns of MTGase‐associated structural and viscoelastic change in the fresh and reconstituted skim milk systems examined. Because independent rheological replicates were unavailable and the two milk systems differed somewhat in composition, these patterns may be regarded as hypothesis‐generating rather than as evidence of reproducible differences attributable specifically to processing history.