benzofurans and fomannoxin

The secondary metabolome of an undescribed stereaceous basidiomycete (BY1) was investigated for bioactive compounds. Along with a known fomannoxin derivative and two known vibralactones, we here describe three new compounds of these natural product families, whose structures were elucidated using 1D and 2D NMR spectroscopy and high-resolution mass spectrometry. The new compound vibralactone S (4) shows a 3,6-substituted oxepin-2(7H)-one ring system, which is unprecedented for the vibralactone/fomannoxin class of compounds. Stable isotope labeling established a biosynthetic route that is dissimilar to the two published cascades of oxepinone formation. Another new compound, the antifungal methyl seco-fomannoxinate (6), features a 2-methylprop-1-enyl ether moiety, which is only rarely observed with natural products. The structure of 6 was confirmed by total synthesis. (13)C-labeling experiments revealed that the unusual 2-methylprop-1-enyl ether residue derives from an isoprene unit. The diversity of BY1's combined fomannoxin/vibralactone metabolism is remarkable in that these compound families, although biosynthetically related, usually occur in different organisms.

Topics: Antifungal Agents; Arthrodermataceae; Aspergillus fumigatus; Basidiomycota; Benzofurans; Candida albicans; Lactones; Molecular Structure; Nuclear Magnetic Resonance, Biomolecular; Penicillium chrysogenum

Rhizosphere-associated Streptomyces sp. AcH 505 (AcH 505) promotes infection of Norway spruce (Picea abies) with the pathogenic fungus Heterobasidion abietinum 331, while Streptomyces sp. GB 4-2 (GB 4-2) enhances spruce defense against the fungus. To identify whether these bacteria influence the availability of the fungal phytotoxin fomannoxin and hence spruce infection, we analyzed the fomannoxin yield in H. abietinum 331-AcH 505 dual cultures. Further, the fate of fomannoxin was studied by adding the compound to cultures of AcH 505, GB 4-2 and nine other soil streptomycetes. Culture filtrates were extracted with ethyl acetate and analyzed by HPLC. Structures of novel compounds were elucidated by HPLC-HR-ESI-Orbitrap-MS and NMR spectroscopy. Phytotoxicity of the compounds was determined by in vivo measurement of maximum photosystem II efficiency of Arabidopsis thaliana seedlings. The amount of fomannoxin in H. abietinum 331-AcH 505 dual cultures was reduced compared to axenic fungus cultures and fungus-plant dual cultures. Following addition of fomannoxin to AcH 505 cultures, the compound disappeared and three novel fomannoxin derivatives without phytotoxic activity were detected. Another novel compound, fomannoxin amide, was discovered following fomannoxin addition to GB 4-2 cultures. Nine other streptomycetes converted fomannoxin into fomannoxin acid or fomannoxin amide. Both compounds exhibit the same phytotoxicity as fomannoxin. We, thus, conclude that the streptomycete-mediated modulation of spruce infection with H. abietinum 331 does not depend on the availability of fomannoxin. We further add evidence to the observation that the lipophilic side chain of fomannoxin is an important structural element for its phytotoxicity.

Topics: Anti-Bacterial Agents; Arabidopsis; Basidiomycota; Benzofurans; Biotransformation; Streptomyces

Fomannoxin is a biologically active benzohydrofuran, which has been suggested to be involved in the pathogenicity of the root rotting fungus Heterobasidion annosum sensu lato. The biosynthesis of fomannoxin was investigated through an isotopic enrichment study utilizing [1-¹³C]glucose as metabolic tracer. ¹³C NMR spectroscopic analysis revealed the labeling pattern and showed that the isoprene building block originates from the mevalonic acid pathway, whereas the aromatic motif is formed via the shikimic acid route by elimination of pyruvate from chorismic acid. A natural product, 4-hydroxy-3-(3-methylbut-2-enyl)benzaldehyde, was isolated and characterized, and was suggested to be a key intermediate in the biosynthesis of fomannoxin and related secondary metabolites previously identified from the H. annosum fungal species complex.

Topics: Basidiomycota; Benzofurans; Molecular Structure; Plant Roots; Tracheophyta

Fomannoxin [(+/-)-5-Formyl-2-isopropenyl-2,3-dihydrobenzofurane] is a phytotoxic secondary metabolite, which is produced by the forest pathogenic basidiomycete Heterobasidion annosum during the infection process. Fomannoxin shows growth-inhibiting effects on callus and suspension cultures of conifer cells. By investigating the interaction of the phytotoxin with Pinus sylvestris cells a detoxification of fomannoxin was detected, presumably as a defense reaction of the plant cells. Undifferentiated green cell lines of Pinus sylvestris were used as target cells. To provide rac-fomannoxin as a substrate a simple method for the chemical synthesis was developed. It was found that the aromatic aldehyde group is reduced by the plant cells producing the non-toxic fomannoxin alcohol which was isolated and identified by spectroscopy. After longer incubation times, also fomannoxin acid-beta-glucoside could be isolated as another detoxification metabolite. For comparison this glucoside was also synthesized.

Topics: Antifungal Agents; Basidiomycota; Benzofurans; Biotransformation; Cell Survival; Cells, Cultured; Cycadopsida; Isomerism; Kinetics; Magnetic Resonance Spectroscopy; Oxidation-Reduction; Plant Diseases; Temperature

In addition to a preceding publication (Sonnenbichler, J., Bliestle, I.M., Peipp, H. & Holdenrieder, O. (1989) Biol. Chem. Hoppe-Seyler 370, 1295-1303), six further metabolites formed in dual cultures of the antagonistic fungi Heterobasidion annosum and Gloeophyllum abietinum are described; three of them represent unknown structures. The producing organisms have been identified. The metabolites fommanoxin from H. annosum and oosponol from G. abietinum show that the toxins can be detoxified by metabolization by the antagonistic fungus. The newly-described compounds have been tested for their fungicidal properties.

Topics: Basidiomycota; Benzofurans; Chromatography, High Pressure Liquid; Coumarins; Magnetic Resonance Spectroscopy; Mass Spectrometry; Spectrophotometry, Ultraviolet

A series of synthetic analogues of fomannoxin and fomajorin S, the toxic metabolites of the basidiomycete Heterobasidion annosum, was prepared in order to investigate which partial structures of the molecules are responsible for their biological activities. Of the compounds synthesized the five indane-2-acid derivatives 5-acetylindane-2-carboxylic acid (2), (+/-)5-formylindane-2-carboxylic acid (3), (+/-)indane-2,5-dicarboxylic acid (4), (+/-)formyl-2-methylindane-2-carboxylic acid (7) and 5-acetyl-indane-2,2-dicarboxylic acid (10) and 4-(2'-ethylbutyloxy)benzaldehyde (18) have not yet been described. Different biological test systems were employed to investigate the antibiotic activities of the synthesized analogues. The experimental results suggest that in the case of fomajorin S, the structural element essential for its toxicity is the 2-H- or 2-methylindane 2-acid. From the results obtained with the synthetic analogues of fomannoxin, it is now evident that the toxicity of this fungal metabolite can be attributed to an aromatic ring with a carbonyl or carboxyl substituent combined with a branched lipophilic side chain in p-substitution.

Topics: Antifungal Agents; Basidiomycota; Benzofurans; Computer Simulation; Culture Media; Indans; Microbial Sensitivity Tests