It is well documented that biostimulants could play an important role in agriculture. Additionally, increased fertilizer use efficiency is essential for maintaining both yield and grain quality, especially for bread wheat, which is a major global crop. In the present study, we explored the effects of mixing urea-ammonium-nitrate fertilizer with Glutacetine® on the physiological responses, agronomic traits and grain quality of winter wheat. Grain proteome analysis revealed that Glutacetine strongly reduced 11 proteins including storage proteins. Indeed, 2 alpha-gliadins and 2 avenin-like proteins decreased after Glutacetine application, which were good for celiac disease patients. Moreover, 2 glutenin HMW subunit were reduced, changing the gliadin/glutenin ratio and the HMW/LMW ratio, thus modifying the wheat flour dough quality. Our investigation reveals the important role of these formulations in achieving significant increases in seed yield and grain quality.