Calophyllum inophyllum Linn. (Guttiferae) is a tree common in Southeast Asia, whose seed, bark and leaf are reputed to be a fish poison. Previous studies isolated three 4-phenylcoumarin derivatives from the nuts and four triterpenes of the friedelin group from the leaves. This work reports the isolation and identification of the piscicidal constituents of the leaves, guided by the killie-fish bioassay. Ether extracts from dried leaves (1.4 kg) were chromatographed on a silicic acid - celite column (eluted stepwise with benzene containing increasing ethyl acetate), followed by Florisil chromatography, preparative thin layer chromatography or recrystallizations, yielding three active compounds I (112 mg), II (19 mg) and III (631 mg). Compound I (C25H22O5, m.p. 188-191°, [α]D + 130°) was identified as (+)-inophyllolide by spectral data (UV, IR, MS, NMR) matching (*)-inophyllolide reported by Polonsky, representing the first isolation of inophyllolide in an optically active form (vs. racemic from nuts). Compound II (C25H22O5, m.p. 149-151°, [α]D +70°), an isomer of I, was the cis isomer (J10,11 = 3.7 cps). Compound III (C25H24O5, m.p. 200-202°, [α]D +43°), a dihydro derivative with a hydroxyl group at C-12 (confirmed by oxidation to II), is the first natural 4-phenylcoumarin possessing a 2,3-dimethylchromanol ring. Bioactivity assays showed I and II were four times more potent than III but one-fifth as strong as pentachlorophenol. Studies on the absolute configuration of the dimethylchromanol ring are in progress.