Cladospirone bisepoxide

The biosynthesis of cladospirone bisepoxide (1) was investigated by feeding 13C-labeled acetate to growing cultures of the fungus Sphaeropsidales sp. (strain F-24'707). 13C NMR spectral analysis demonstrated the polyketide origin of both naphthalene units. The origin of two epoxide oxygens was confirmed as from air by cultivation of the strain in an 18O2-enriched atmosphere. The [18O]incorporation pattern into palmarumycin C12 (11), the putative precursor of 1 led to the hypothesis that the carbonyl oxygen of 1 is derived from water by exchange of an oxygen atom. Inhibition of the biosynthesis of 1 with tricyclazole, an inhibitor of the 1,8-dihydroxynaphthalene (DHN) melanin biosynthesis, confirmed the connection of both biosynthetic pathways.

Cladospirone bisepoxide (1), a novel metabolite, was isolated from cultures of a fungus which was characterized as a coelomycete by the formation of pycnidia. By optimization of media and fermentation conditions, a titer of up to 1.5 g/liter on shake level and 1.16 g/liter on bioreactor scale could be achieved. The isolation of the compound was performed by solvent extraction of the culture broth and subsequent crystallization. Cladospirone bisepoxide displays selective antibiotic activity against several bacteria and fungi and inhibits germinations of Lepidium sativum at low concentrations.

The metabolite pattern of UV mutants of the spirobisnaphthalene producing fungus F-24'707 by TLC and HPLC analysis has been investigated. Mutants with differences in colony morphology or colour compared to the parent strain were isolated. Cultivation in shaking flasks and P flasks showed differences in the metabolite pattern of some of the strains. Furthermore, enzyme inhibitors were used to block the spirobisnaphthalene biosynthesis of the parent strain at different steps. Feeding of precursors and intermediates of cladospirone bisepoxide (15) led to a two-fold increase of the production of 15. From these data and preceding biosynthetic studies we deduced a general pathway for the biosynthesis of all spirobisnaphthalenes of the fungus F-24'707. This enables us to present the hypothesis that all bisnaphthalenes described so far are produced using a common pathway with only a few intermediates as central branching points.