Olea europaea L. leaves are an attractive and readily available source of biologically active compounds. In particular, olive leaf polyphenols are a complex mixture of different compounds, among which secoiridoids, phenolic alcohols and flavonoids. Oleuropein and hydroxytyrosol are generally the most represented phenolic compound in leaves of different olive cultivars. It has been reported that olive leaves have the highest antioxidant and scavenging power among the different parts of the olive tree. Beyond the antioxidant capacity, olive leaf extract has been reported to possess antihypertensive, antiatherogenic, anti-inflammatory, hypoglycemic, and hypocholesterolemic properties (El and Karakaya, 2009). These biologically active compounds can be used for the preparation of dietary supplements, nutraceuticals, functional food ingredients or cosmeceuticals. However, the chemical composition of olive leaves varies depending on several factors among which cultivar, geographical origin, leaf age, climatic conditions, moisture content and storage conditions (Papoti and Tsimidou, 2009). The objective of the present work was to characterize the phenolic compounds and the antioxidant capacity in leaves from some Olea europaea L. cultivars, harvested in different times of the year. First of all, optimal conditions for extraction of olive leaves were determined using the Response Surface Methodology (RSM) and the considered parameters were: solvent composition, solid to solvent ratio and temperature. The antioxidant activity was assessed using 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) radical scavenging assays. Total phenol content was determined using Folin-Ciocalteu assay while the analysis of individual phenol compounds was performed by HPLC-DAD-MS. The results showed some significant differences between the selected cultivars both considering the phenol compounds and the antioxidant activity. During olive harvesting period, the highest values of polyphenol content and antioxidant activity were found in the samples from Dolce Agogia cultivar while the lowest values in Frantoio cultivar samples. Some relationship were observed between phenol compounds and antioxidant properties of the extracts.