Basil ( Ocimum basilicum L.) is a highly important medicinal plant. In this study, the interactional eff ects of water defi cit and melatonin application on growth and physio-biochemical characteristics of basil were investigated. The treatments included three levels of irrigation (100, 75 and 50% of fi eld capacity) and three melatonin concentrations (0, 50 and 100 μmol). Drought stress decreased the dry weight of plants but application of 100 μmol melatonin markedly increased dry weight under these conditions. Melatonin profoundly enhanced plant drought tolerance by stimulating the antioxidant enzymes (catalase, phenylalanine ammonia-lyase, ascorbate peroxidase and polyphenol oxidase) activities and secondary metabolites (total phenol, fl avonoid and anthocyanin contents). Furthermore, melatonin application reduced lipid peroxi dation and increased the relative water content and photosynthetic pigments, resulting in an improvement of plant dry matter. It is concluded that foliar spraying with melatonin induced the pre-adaptive responses to drought stress followed by regulating antioxidant defense mechanisms.