The flowers of Origanum sipyleum L., which is an endemic species in Turkey, are traditionally used as a beverage and medicinal tea. The extract was analyzed by reversed-phase high-performance liquid chromatography The main phenolic compounds of O. sipyleurn contained 6.81 +/- 1.95, 3.94 +/- 1.65, 6.78 +/- 1.53, 6.71 +/- 1.01, 4.95 +/- 3.30, 870.22 +/- 82.66, 19.47 +/- 1.77 and 1017.33 +/- 93.06 mg/100 g herb of apigenin, caffeic acid, carvacrol (phenolic monoterpene), hesperidin, naringenin, rosmarinic acid, rutin and vitexin, respectively. However acecetin, apigenin 7-glucoside, carnosic acid, eriodictyol, gallic acid, genistin, luteolin and naringin were not determined. In this study, the antioxidant and antimicrobial activities of O. sipyleurn extract were determined, and their phenolics identified. Total phenolics, scavenging of 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical and the antioxidant capacity by the formation of phosphomolybdenum complex of the extract were determined. The phenolic content was 226.66 +/- 1.18 mg gallic acid equivalents/g in the dried extract. At 100-ppm concentration, the DPPH radical scavenging activity of extract was 73.98 +/- 2.66%. The antioxidant capacity of the extract was 335.33 +/- 2.87 mg/g. This extract was investigated for its antibacterial activity by using agar diffusion method against 15 species of bacteria. Statistical differences within the bacteria were significant at P < 0.05. The extract was effective against all the bacteria tested, except for Yersinia enterocolitica.