Antibacterial activity of Zanthoxylum rhetsa seed essential oils steam distilled from aqueous, alkaline and acidic media against four pathogenic bacteria namely Staphylococcus aureus, Escherichia coli, Proteus vulgaris and Klebsiella pneumoniae were statistically evaluated. The acid distilled oil showed significant activity against all the tested microorganisms, and its activity is quite comparable with the standard antibiotic gentamicin screened under similar conditions. So, the acid distilled seed essential oil of Zanthoxylum rhetsa can be used as a potential external antiseptic and can be incorporated into drug formulations. The activity of aqueous and alkaline extract was found to be moderate. Zanthoxylum rhetsa seed oil showed significant activity against these organisms probably due to the synergic effect of the components present in the oil.