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The aqueous ethanol extract of Helicteres isora root were partitioned using various solvents like petroleum ether, chloroform, ethyl acetate and butanol. Of the 10 tested microbial strains, all fractions exhibited antimicrobial activity against 9 microbial strains at concentrations of 10, 5, 2.5 mg/ml. Among tested organisms, Micrococcus luteus, Aspergillus niger and Candida albicans were most sensitive and Salmonella typhimurium was most resistant . Butanol extract was found to possess most potent antimicrobial activity.