Chemical investigations of fungicolous and endophytic fungi

Fungi are rich sources of structurally diverse secondary metabolites, some of which possess biological activities of importance to medicine and agriculture. Research in our group focuses on application of ecological considerations to the selection of underexplored fungi for chemical analysis in search of new natural products with potentially important bioactivities. The work described in this thesis involved studies of members of two different fungal ecological groups, fungicolous and endophytic fungi. Fungicolous fungi colonize other fungi and are viewed as potential sources of antifungal agents due to the negative effects often exerted on the host organisms. Our prior studies of such fungi have proven them to be productive sources of new bioactive natural products. Fractionation of extracts produced from eleven fungicolous fungal isolates resulted in the isolation and identification of 24 different, structurally-diverse natural products, many of which were bioactive. Most of these were previously known, but six were new compounds, with one being a distinctive new peptaibol-type metabolite. Endophytic fungi colonize the inner tissues of host plants, often asymptomatically, and occur widely in most plant species. Some endophytes may benefit the host, possibly through production of protective secondary metabolites, but in most cases, their roles are not well understood. Stenocarpella maydis is a widely occurring fungal endophyte and pathogen of corn and is associated with diplodiosis (a neuromycotoxicosis) of cattle. Investigations of extracts of S. maydis and S. macrospora cultures led to the identification of seven compounds, including diplodiatoxin, diplosporin, chaetoglobosins K, L, M, and O, and (all-E) trideca-4,6,10,12-tetraene-2,8-diol, none of which were previously known from U.S. Stenocarpella isolates. Diplodiatoxin was detected as a major component of S. maydis-rotted maize grain, stalks, and stalk residues, and chaetoglobosin M was detected in extracts of naturally-diseased S. maydis-rotted maize seeds collected in the field. Chaetoglobosin K displayed potent antifungal and antiinsectan activity, while diplodiatoxin displayed phytotoxicity. Because of their well-known cytotoxic effects, we proposed that mixtures of chaetoglobosins are responsible, at least in part, for inducing diplodiosis in livestock.

Our prior investigations of another common corn endophyte, Acremonium zeae, led to isolation of the pyrrocidines, which display potent antifungal and antibacterial activities against a range of corn pathogens. Further studies of these extracts led to the isolation and identification of three resorcylic acid lactone (RAL)-type compounds, one of which was new. These RAL's, did not exhibit antifungal activity, but demonstrated mild phytotoxicity to maize leaves in assays. They are also members of a compound class we have encountered among other endophytes that inhibit the heat shock protein Hsp90, and have been proposed to play a role in the colonization process. These chemical investigations provide further knowledge that may be useful in determining the roles that endophytes, in particular S. maydis and A. zeae, play in maize. The metabolites encountered during these investigations represented different biosynthetic pathways, and included polyketides, terpenoids, and compounds of mixed biogenetic origin. Structures were characterized by analysis of 1D and 2D NMR data, mass spectrometry, chemical degradation or derivatization reactions, and/or X-ray diffraction analysis. Absolute configuration assignments were determined using Mosher's method, Marfey's method, or chiral amino acid analysis methods.