Biosynthesis of 3-acetyldeoxynivalenol and sambucinol - Identification of the two oxygenation steps after trichodiene

The first two oxygenation steps post-trichodiene in the biosyntheses of the trichothecenes 3-acetyldeoxynivalenol and sambucinol were investigated. Th e plausible intermediates 2-hydroxytrichodiene (2 alpha- and 2 beta-) and 1 2,13-epoxytrichodiene and the dioxygenated compounds 12,13-epoxy-9,10-trich oene-2-ol (2 alpha- and 2 beta-) were prepared specifically labeled with st able isotopes. They were then fed separately and/or together to Fusarium cu lmorum cultures, and the derived trichothecenes were isolated, purified, an d analyzed. The stable isotopes enable easy localization of the labels in t he products by H-2 NMR, C-13 MMR, and mass spectrometry. We found that 2 al pha-hydroxytrichodiene is the first oxygenated step in the biosynthesis of both 3-acetyldeoxynivalenol and sambucinol. The stereoisomer 2 beta-hydroxy trichodiene and 12,13-epoxytrichodiene are not biosynthetic intermediates a nd have not been isolated as metabolites. We also demonstrated that the dio xygenated 12,13-epoxy-9,10-trichoene-2 alpha-ol is a biosynthetic precursor to trichothecenes as had been suggested in a preliminary work. Its stereoi somer was not found in the pathway. A further confirmation of our results w as the isolation of both oxygenated trichodiene derivatives 2 alpha-hydroxy trichodiene and 12,13-epoxy-9,10-trichoene-2 alpha-ol as natural metabolite s in F. culmorum cultures.