To evaluate the effects of water deficit stress and growth regulator (Brassinosteroid) on some physiological characteristics cultivars of safflower, an split plot factorial experiment based on randomize complete block design with 3 replications was conducted at the Research Farm of Faculty of Agriculture, Mohaghegh Ardabili University in 2014. Water stress was in three levels: 80 mm evaporation (normal irrigation); 120 mm evaporation (low water stress); 160 mm evaporation (high water stress) from evaporation pan of class A which were assigned to main plots and three cultivars of safflower (Goldasht, Spiny Sina and Faraman) and two levels of Brassinosteroid, control and 10 -7 mol. in sub plots. The reasults showed that water stress decreased stomatal conductance, photosynthetic rate, sub-stomatal CO2, water-use efficiency, chlorophyll content index (SPAD), seed yield, leaf soluble protein and transpiration rate. The results also showed that use of brassinosteriod, increased photosynthetic parameters and reduced transpiration. Irrigation at evaporation of 80 mm from pan, resulted in the maximom amount of transpiration and stomatal conductance to the Faraman cultivar, sub-stomatal CO2 to Sina spiny cultivar and chlorophyll content index (SPAD) to Goldasht cultivar. Irrigation at 120 mm evaporation from class A pan resulted in the maximum amount of transpiration and stomatal conductance to Goldasht cultivar, while the highest sub-stomatal CO2, belonged to Sina spiny. Irrigation at 160 mm evaporation from class A pan resulted in the maximum amount of transpiration, stomatal conductance, chlorophyll content index (SPAD) and the sub-stomatal CO2 to Goldasht cultivar. It seems that Goldasht cultivar under both mild and severe drought stresses tolerates drought better than the other two cultivars in Ardabil rigion.