Methanol extract of roots of Amblytropis pauciflora Kitagawa showed the antimicrobial activity to three test strains. Antimicrobial spectra of various extracts of Amblytropis pauciflora Kitagawa were tested against 24 strains of bacteria and fungi. The crude methanol extract inhibited the growth of 12 strains of bacteria and Asp. fumigatus with the exception of yeasts. The properties of the antimicrobial substance were very stable under heat(at 12$0^{circ}C$), acid(pH 3.0) and alkali(pH 11.0) treatment. Only the root harvested in spring showed the antimicrobial activity. Among the components extracted by butanol, ginseng saponin Rg1 and various saponin-like materials were detected by TLC analysis using a plate of silica gel 60F254. The antimicrobial compound was purified by methanol extraction, activated charcoal column chromatography, Sep-pak(C18) pretreatment and reverse phase HPLC. The purified compound was detected at 13.520 min as a single peak(about 98% purity) through the HPLC analysis.