This work aimed to produce supplemeted yogurt with using three different concentrations of 1, 3, and 5% from raw and germinated (R&G) barley flour. The alterations in the chemical structure, antioxidant activity, color value, microbiological and sensory evaluation of yogurt samples were determined through two weeks of storage at 4 °C. Result obtained indicated that increasing amounts of added barley flour (R&G) led to a significant increase in the total solids (TS), protein, ash contents, and acidity. On the other hand, TS, protein, fat, ash, acidity, and syneresis increased during storage at 4 °C for 2 weeks. Moreover, DPPH antioxidant activity was higher in the yogurt samples supplemented with barley flour from (R&G) than in control yogurt samples, slightly increased with the storage, and declined on the 14th day of storage. The lightness (L) of yogurt supplemented with barley flour was lower, and the values of (a) and (b) were high compared with control samples yogurt. The value of (L) and (a) increased slightly with the storage and declined on the 14th day in all stored yogurt samples. By increasing the percentage of barley flour R&G in yogurt, the total bacterial count and Streptococci, Lactobacilli, yeast and mould were significantly elevated. In sensory evaluation, yogurt containing 1% germinated barley flour scored the highest total acceptability score. Thus the quality of stored barley yogurt was comparatively reasonable. Based on the current findings, it can be established that supplementing yogurt with barley flour from R&G enhanced the yogurt's quality and antioxidant activity.