An organosulfur compound was isolated from oil-macerated garlic extract by silica gel column chromatography and preparative TLC. From the results of NMR, IR, and MS analyses, its structure was determined as E-4,5,9-trithiadeca-1,7-diene-9-oxide (iso-E-10-devinylajoene, iso-E-10-DA). This compound was different from E-4,5,9-trithiadeca-1,6-diene-9-oxide (E-10-devinylajoene, E-10-DA) only in the position of a double bond. Iso-E-10-DA had antimicrobial activity against Gram-positive bacteria, such as Bacillus cereus, B. subtilis, and Staphylococcus aureus, and yeasts at the concentration lower than 100 micrograms/ml, but Gram-negative bacteria were not inhibited at the same concentration. The antimicrobial activity of iso-E-10-DA was inferior to those of similar oil-macerated garlic extract compounds such as E-ajoene, Z-ajoene, and Z-10-DA. From these results, it was suggested that trans structure and/or the position of double bond of iso-E-10-DA reduce the antimicrobial activity.