The effects of bioactive fractions from Brachyglottis monroi on biological activity were investigated. this bioactive subfraction 6-5 is the most cytotoxic to P388 murine leukaemia cell lines. A comparison of IC₅₀ values of these subfraction in cancer cell lines showed that their susceptibility to these subfractions decreased in the following order; Fr. 6-5 > Fr. 6-3 > Fr. 6-6 > Fr. 6-1 > Fr. 6-2 > Fr. 6-4 by the MTT method. Silica gel flash column chromatography concentrated the cytotoxic activity in subfraction 6-5 which eluted with 30% and 40% ethyl acetate : hexane gave a major bioactive (51 mg, P388 IC₅₀ 8,286 ng/mL at 7.5 ㎍/disc).