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Chemical compositions of hydrodistilled essential oil from flowering aerial parts of Chenopodium botrys L. were analyzed by GC and GC-MS. Antimicrobial activity of essential oil was screened against different kinds of microorganisms by disc diffusion and micro broth dilution assays. Forty-three components were identified accounted for over 98 % of the total oil. 2,3-dehydro-4-oxo-β-lonone (22.4 %), (+)-7-epi-amiteol (11.5 %) were found as the major components of the oil. The essential oil showed strong antimicrobial activity against Staphylococcus saprophyticus followed by Klebsiella pneumoniae, Bacillus cereus, Staphylococcus epidermidis, Streptococcus mutans, Listeria monocytogenes and Salmonella typhimurium. The oil had slight effect on Candida albicans and showed inhibitory effect on Aspergillus species and Bacillus subtilis.Introduction: Nowadays, the use of medicinal plants has become popular as alternatives for treatment of infectious diseases and as functional ingredients in food and drinks because of increasing concern about potentially harmful traditional antibiotics and synthetic additives.Chenopodium botrys L. is a member of Chenopodiaceae family which is known as Jerusalem oak 1 . Whole organs of plant have aromatic odors. In Iranian traditional medicines, the flowering aerial parts of C. botrys has long been used as expectorant, antiasthmatic, anticatarrh, anticonvulsant agent and as tonic 2 . In many cases, C. botrys is used instead of lavenders to keep away moths 3 .Chemical compositions of C. botrys essential oil have been reported by other researchers 3-7 . C. botrys essential oil from Greece contained elemol acetate (16.3 %), elemol (14.1 %), botrydiol (11.1 %), α-chenopodiol (9.5 %), β-eudesmol (7.0 %) and selina-3-11-dien-6 α-ol (6.1 %) 3 ; from two different localities of Iran; juniper camphor (16.5 and 25.7 %), elemol (14.3 % and 13.4 %) and αcadinol (8.2 % and 11.6 %) 4 , from Saudi Arabia; αand β-eudesmol 6 , from North America; αchenopodiol, β-chenopodiol, eudesma 3,11-dien-6α-ol 7 were as the major components of the oil. Other research from Iran revealed the presence of γ-terpineol (52.8 %), p-cymene (19.0 %) and isoascaridole (7.0 %) as the main components of the oil 5 .There are some studies on antimicrobial activity of C. botrys essential oil 3,8 . The oil has not Journal of Essential Oil Bearing Plants