Abstract: To address the inherent limitations of conventional dry-heating methods (protracted reaction duration) and wet-heating approaches (excessive protein denaturation), an innovative strategy was developed for effectuating the Maillard reaction between whey protein isolate (WPI) and glucose within an ethanol-water binary system, thereby enabling efficient and precisely controlled preparation of glycoconjugates. Utilizing the degree of graft (DG) as the primary response variable, reaction parameters were systematically optimized through single-factor experimentation coupled with Box-Behnken design-based response surface methodology. The resultant conjugates were comprehensively characterized with respect to solubility profile, emulsifying properties, and antioxidant capacity. Optimal conditions were established as follows: ethanol-water volume concentration of 90%, ethanol-water volume of 3 mL, WPI-to-glucose mass ratio of 1:2, reaction duration of 3 h, and temperature of 70 ℃. Under these optimized parameters, the conjugates exhibited a DG of 80.74%±1.61%. Comprehensive physicochemical characterization revealed markedly enhanced functional attributes. Minimal browning intensity and negligible chromatic aberration were observed at reaction temperatures ≤70 ℃. Across the investigated pH range (2.0~10.0), the conjugates demonstrated exceptionally low turbidity without statistically significant variation (P>0.05), concomitant with superior solubility (>85.60%±1.20%). Furthermore, the emulsification activity index, emulsion stability index, DPPH radical scavenging activity, and ABTS⁺ radical scavenging activity attained values of 57.29±1.01 m²/g, 22.58±0.52 min, 32.19%±2.31%, and 59.65%±1.96%, respectively. These findings conclusively demonstrate that the ethanol-water system constitutes an efficient reaction medium for generating WPI-glucose Maillard reaction products. Moreover, judicious modulation of reaction time enables targeted optimization of solubility, emulsifying properties, and antioxidant activity, thereby establishing a theoretical foundation for potential applications within food processing systems.