Effects of exogenous spermidine (Spd) (10µM) application on the biochemical enzymes and molecular effects in the Burakbey cultivar of barley were investigated both normal (distilled water) and drought (-6,30 mPA PEG 6000) conditions. It is obvious that drought stress applied to barley cultivars negatively affected plant metabolism and inhibited plant growth when all other parameters were taken into account. Under normal conditions, Spd showed an impact as if drought stress were applied to the plant at a low rate. Exogenous Spd application under drought stress alleviated the harmful effects of drought stress by showing encouraging effects on cell cycle, total protein content, malondialdehyde (MDA), and ascorbate peroxidase (APX) levels. Superoxide dismutase (SOD) and catalase (CAT) enzyme levels in the same group increased further after exogenous Spd application and formed a response to drought stress. As a result, it has been revealed that the application of exogenous Spd in barley can increase endogenous polyamine, antioxidant capacity, promotion of cell division, and protein content, as well as ameliorate the detrimental effects of drought stress on plant metabolism.