The ethanol extract from the root bark of Morus alba showed the strongest antimicrobial activity on the growth of almost all the tested microorganisms which were food-borne pathogens and food-related microorganisms. 1) In order to isolate and purify of antimicrobial substance extracted from the root bark of Morus alba, the antimicrobial substance from the ethanol extract which exhibited a strong antimicrobial activity was purified by solvent fractionation, silica gel column chromatography, TLC and HPLC. Among the fractions fractionated by 4 kind of solvents from the ethanol extract, the antimicrobial activity of ethyl acetate fraction had the strongest antimicrobial activity against B. subtilis. Unknown compound were isolated from the ethyl acetate fraction by silica gel column chromatography, TLC and HPLC and the compounds showed strong absorbance at 207, 217 and 285 nm, therefore, it was supposed to be a kinds of aromatic compound.