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Two new isochromans, panowamycins A and B, were purified by solvent extraction, silica gel and octadecylsilyl silica gel (ODS) column chromatography followed by preparative HPLC, from a culture broth of Streptomyces sp. K07-0010, together with the known compounds NFAT-133, conglobatin, piericidin C series and dinactin. Structures of panowamycins were elucidated as new analogs of NFAT-133 by spectroscopic studies including various NMR experiments. Panowamycins A and B showed moderate antitrypanosomal activity, with IC 50 values of 0.40 and 3.30 lg ml -1 , respectively.

In the course of screening for new antitrypanosomal agents, several novel bioactive natural compounds have been discovered and reported by our group. 1-3 Recently, we have isolated two new trichosporin analogs, designated as trichosporins B-VIIa (1) and B-VIIb (2), together with five known trichosporins (3-7), 4 from the culture broth of Trichoderma sp. FKI-4452. These have proved to exhibit antitrypanosomal activity (Figure 1). In this paper, the fermentation, isolation, structure elucidation and biological activity of these novel trichosporins are described.Fungal strain FKI-4452 was isolated from a soil sample collected in Yakushima-Island, Kagoshima, Japan by the dilution plating method. The ITS sequence of strain FKI-4452 was determined and deposited at the DNA Data Bank of Japan, with the accession number AB517619, using The TrichOKEY program of ITSH (International Subcommission on Trichoderma and Hypocrea Taxonomy), which suggested that the strain FKI-4452 belongs to Trichoderma polysporum. 5 Moreover, FKI-4452 was a 98% match to the nucleotide sequences of T. polysporum CBS 820.68 according to the TrichoBLAST database of ITSH. 5 Thus, the strain FKI-4452 was identified as T. polysporum and designated Trichoderma polysporum FKI-4452. There was no report of trichosporin production by T. polysporum CBS 820.68.The strain FKI-4452 was grown and maintained on an LcA agar slant consisting of 0.1% glycerol, 0.08% KH 2 PO 4 , 0.02% K 2 HPO 4 , 0.02% MgSO 4 Á7H 2 O, 0.02% KCl, 0.2% NaNO 3 , 0.02% yeast extract and 1.5% agar (adjusted to pH 6.0 before sterilization). A loop of spores of Trichoderma sp. FKI-4452 was inoculated into 100 ml of the seed medium, which consisted of 2.0% glucose, 0.2% yeast extract, 0.5% Polypepton (Wako Pure Chemical Industries, Osaka, Japan), 0.05% MgSO 4 Á7H 2 O, 0.1% KH 2 PO 4 and 0.1% agar (adjusted to pH 6.0 before sterilization), in a 500-ml Erlenmeyer flask. The inoculated flask was incubated in a rotary shaker (210 r.p.m.) at 27 1C for 3 days.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% CaCO 3 , 0.05% MgSO 4 Á7H 2 O, and 0.05% KH 2 PO 4 (adjusted to pH 6.5 before sterilization), fermentation taking place on a rotary shaker (210 r.p.m.) at 27 1C for 6 days.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-7515-12A, 20fÂ120 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 princ...

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