This research was performed to evaluate the effect of substitution wheat flour by ascending levels (4, 8 and 12%) of defatted chia flour (DCF) on the physicochemical properties, nutritional value, quality criteria and organoleptic evaluation of produced hard sweet biscuits. The results indicated that the crude protein and fiber contents of defatted chia flour were higher than that of wheat flour. The results revealed that biscuit samples prepared by using defatted chia flour at all substitution levels had the lowest caloric value and the highest contents of protein, lipid, ash, and crude fiber contents as compared to the control sample. Additionally, as the substitution level of wheat flour with DCF, the contents of Ca, Mg, P, Zn, and Fe were considerably (p <0.05) increased. Regarding, the sensory attributes, biscuits samples particularly, those that were prepared by substituting wheat flour with 4 and 8 %were more acceptable as compared to the control which was made of wheat flour only. Based on the information provided above, it can be inferred that using defatted chia flour in bakery goods like biscuits will enhance its nutritional value and physicochemical characteristics. The created biscuits enhanced with defatted chia flour might live up to consumer expectations for wholesome and useful food.