Beyond the water activity and T_g: water usability can govern xerophilic fungal and bacterial proliferation in sucrose/whey protein isolate solid matrices

High-solid food matrices are often unstable; how water behaves in them dictates molecule mobility and microbial proliferation. Here, we employ lyophilized sucrose/whey protein isolate (WPI) mixtures as a model system to link water sorption isotherms, glass-transition temperatures (T_g), structural relaxation dynamics, and relaxation times to microbial growth of Aspergillus flavus and Bacillus subtilis. Although the sorption isotherms, T_g, and relaxation processes of the studied matrices were affected by a_w and WPI, the microbial growth showed a high dependence on water mobility rather than a_w (0.75-0.92 a_w at 30 °C). Therefore, this study used water usability (U_w), a thermodynamic parameter (ranging from 0 to 1), where higher values indicate greater amounts of freely mobile water. Moreover, U_w quantifies the full plasticizing effect of bound and free water and the kinetic immobilization imposed by protein networks. As U_w declines with increasing WPI content, extensive hydrogen-bonding and microphase domains sequester water into tightly bound shells, reducing free volume and slowing segmental motions; this reduction in U_w predicts slower microbial growth behavior more reliably than a_w or T_g alone. By uniting thermodynamic and dynamic facets of water behavior in a single parameter, U_w offers a quantitative framework to anticipate and control nutrient transport, textural stability, and microbial safety in complex, non-equilibrium solid foods, with broad implications for mitigating spoilage and ensuring product quality under shifting climatic conditions.