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Two novel compounds, stachyflin and acetylstachyflin, have been isolated by solid-state fermentation of Stachybotrys sp. RF-7260. The structures of both metabolites, determined by detailed NMR analyses and X-ray crystallographic analysis, are novel with a pentacyclic moiety including cis-fused decalin. The absolute stereochemistry of stachyflins was determined by circular dichroism analysis. Stachyflin showed antiviral activity against influenza A virus (H1N1) in vitro with an IC50 value of 0.003 microM. Acetylstachyflin was about 77-fold less active than stachyflin.

In the course of screening for yeast squalene synthase inhibitors, bisabosqual A was isolated from the culture broth of Stachybotrys sp. RF-7260. The related compounds bisabosquals B, C and D were also isolated from Stachybotrys ruwenzoriensis RF-6853. Bisabosquals inhibited squalene synthases. IC50 values of bisabosqual A against the microsomal squalene synthases from Saccharomyces cerevisiae, Candida albicans, HepG2 cell and rat liver were 0.43, 0.25, 0.95 and 2.5 microg/ml, respectively. Bisabosqual C exhibited inhibitory activities similar to bisabosqual A. Bisabosqual A showed broad spectrum antifungal activity in vitro.

A practical method for the production of calcium 2-keto- l -gulonate (an intermediate in the Reichstein synthesis of l -ascorbic acid) from d -glucose has been established by using a two-stage fermentation system. d -Glucose was first converted to calcium 2,5-diketo- d -gluconate by a mutant strain of Erwinia sp. in a medium containing d -glucose, corn steep liquor, (NH 4 ) 2 HPO 4 , and CaCO 3 . After a 26-h cultivation, 328.6 mg of calcium 2,5-diketo- d -gluconate per ml was obtained, with a 94.5% yield from d -glucose. This broth was used directly for the next conversion without removal of cells by treatment with sodium dodecyl sulfate. The stereospecific reduction of calcium 2,5-diketo- d -gluconate to calcium 2-keto- l -gulonate was performed with a mutant strain of Corynebacterium sp. When the cell growth reached a maximum (about 16 h) in a medium containing d -glucose, corn steep liquor, NaNO 3 , KH 2 PO 4 , and trace elements, NaNO 3 was added to the culture, and then the calcium 2,5-diketo- d -gluconate broth was fed over a period of about 50 h. Since the mutant strain requires a hydrogen donor for reduction, the calcium 2,5-diketo- d -gluconate broth was mixed with d -glucose before being fed. The results of four two-stage fermentations in 10-m 3 conventional fermentors showed that an average of 106.3 mg of calcium 2-keto- l -gulonate per ml was obtained, with a 84.6% yield from d -glucose, the starting material of calcium 2,5-diketo- d -gluconate production. Calcium 2-keto- l -gulonate was stable in the broth. Neither 2-keto- d -gluconic acid nor 5-keto- d -gluconic acid was detected in the final broth.