Oregano (Origanum vulgare L.) is an important medicinal, culinary, and essential oil plant. Oregano essential oil is extracted from either leaves or shoots through steam distillation. Researchers and industry in various countries reported different distillation times (DTs) for oregano; however, there are no reports on optimum DT. This study evaluated the effect of DT (1.25, 2.5, 5, 10, 20, 40, 80, 160, 240, 360 min) on essential oil yield, composition, and antioxidant activity of the oregano essential oil. In general, the concentration of the low boiling essential oil constituents (alpha-thujene, alpha-pinene, camphene, l-octen-3-ol, myrcene, alpha-terpinene, paracymene, beta-phellandrene/limonene, gamma-terpinene, cis-sabinene hydrate, terpinolene) were highest at the shortest DT (1.25 or 2.5 min), reduced with increasing DT up to 40 min, and then stayed the same. However, the concentration of the major oil constituent, carvacrol, was lowest at the shortest DT of 1.25 min (18%) and increased steadily with increasing DT up to 40 min, where it leveled at 80% to 82%. The concentration of other higher boiling constituents (borneol, 4-terpineol, beta-bisabolene, beta-caryophylenne) was maximum at 5 to 20 min DT. Maximum yield of the low boiling constituents was achieved at relatively short DT, at ≈20 min DT, and peaked again at 240 min DT. Maximum yields of alpha-terpinene, beta-phellandrene/limonene, and gamma-terpinene were reached at 240 min DT. Maximum yields of paracymene cis-sabinene hydrate, terpinolene, and transsabinene hydrate were also achieved at 240 min DT, but yields at 20 min DT were not different. Yields of borneol, 4-terpinenol, carvacrol, beta-caryophyllene, and beta-bisabolene also were highest at 240 min DT. Distillation time at 20, 80, or 360 min did not alter antioxidant or antimicrobial activity of oregano oil. The relationship between the concentration and yield of the constituents with DT was adequately modeled by the asymptotic and Michaelis-Menten nonlinear regression models, respectively. Results demonstrated that 1) DT can be used to obtain oregano essential oil with differential composition; 2) maximum essential oil yield of steam-distilled oregano leaves could be obtained at 240 min DT; and 3) reports on oregano essential oil yield and composition using different DTs may not be comparable. Results from this study will aid in comparing published reports on oregano essential oil that used different lengths of DT.