6.0 before sterilization), in a 500-ml Erlenmeyer flask. The inoculated flask was incubated in a rotary shaker (210 r.p.m.) at 271 Cf or 3d ays. For production of 1, 2, and the other known trichosporins (3–7), a 1-ml portion of the seed culture was transferred to each of nine 500-ml Erlenmeyer flasks containing 100 ml of the production medium, consisting of 3.0% soluble starch, 2.0% soybean meal, 1.0% glycerol, 0.3% dry yeast, 0.3% KCl, 0.2% CaCO3 ,0 .05% MgSO4� 7H2O, and 0.05% KH2PO4 (adjusted to pH 6.5 before sterilization), fermentation taking place on a rotary shaker (210 r.p.m.) at 271 Cf or 6d ays. To the whole culture broth (1.0 l) was added 1.0 l of ethanol, followed by filtration. The filtrate was concentrated under reduced pressure to remove ethanol and then extracted with 1.0 l of ethyl acetate (pH 2). The ethyl acetate layer was concentrated under reduced pressure to afford a crude extract (745 mg). The ethyl acetate extract (426 mg) was applied to an ODS column (Pegasil Prep ODS-751512A, 20f� 120 mm, Senshu Scientific Co., Tokyo, Japan) pre-equilibrated with 20% methanol. The column was eluted with 20, 40 and 60% methanol stepwise (120 ml each) and the active principles were eluted with 80% methanol (120 ml), followed by concentration in vacuo to yield a brown material (91.0 mg). The material was purified by HPLC using a Pegasil ODS column (20f� 250 mm, Senshu Scientific Co.) with 70% CH3CN at 5 ml min � 1 detected at UV 210 nm. The retention times of the active fractions 1, 2, 3, and 4 were 16, 18, 20, and 24 min, respectively. The active fractions 1 and 2 were concentrated in vacuo to dryness to afford trichosporins B-V (3, 9.9 mg) and B-VIb (4, 10.0 mg), respectively, as white powders. The active fraction 3 (8.4 mg) was purified by HPLC using an XBridge C8 column (10f� 250 mm, Waters Co., Milford, MA, USA) with 50%