In order to study the effects of irrigation regimes and humic acid application on some of physiological and biochemical characteristics of Portulaca oleracea L., a greenhouse experiment was conducted as factorial based on completely randomized design with three replications at research greenhouse of plant production faculty of Gorgan University of Agricultural Sciences and Natural Resources, during of 2014-15. The main factors were irrigation regimes including four levels: (25, 50, 75 and 100 percent of field capacity) and four concentrations of humic acid: (0, 200, 400 and 600mg/L). Traits evaluated were proline, RWC, total phenol, flavonoid, antioxidant, soluble carbohydrate and carotenoid content. The results showed the treatments had significant effects on the traits. As, Irrigation regimes had significant effect on proline, total phenol, flavonoid, antioxidant, soluble carbohydrate. The highest level of irrigation regimes (25 percent FC) increased proline and decreased RWC and carotenoid. Humic acid had significant effect on all traits except of carotenoid content that were the highest in 400 mg/L humic acid. Interaction effect of humic acid and irrigation regimes was significant for all traits except of total phenol. Irrigation regimes increased antioxidant, soluble carbohydrate, in 25% and 50% FC, respectively. Generally the best of treatments is 400mg/L humic acid and 25percent FC, economically. Because the highest yield achieved with low water and humic acid.