When cultures of Fusarium culmorum CMI 14764 were treated with the furanocoumarin xanthotoxin, trichothecene biosynthesis was suppressed, and large amounts of trichodiene accumulated, along with smaller amounts of 12,13-epoxytrichothec-9-ene (EPT) and isotrichodermin. Trichodiene was shown to be a precursor of the trichothecene mycotoxins in F. culmorum, and when large amounts of trichodiene were supplied, a new trichodiene metabolite isotrichodiol was found to accumulate. This compound, 12,13-epoxy-2-alpha,11-alpha-dihydroxytrichodiene, was shown to be a biosynthetic precursor of the trichothecenes, and represents the first demonstrated post-trichodiene intermediate in the pathway to trichothecenes. Slow, acid-catalysed cyclization of isotrichodiol to EPT was demonstrated, but the rapid in vivo incorporation into EPT points to an enzyme-catalysed process in the fungus. The biosynthesis of oxygenated trichothecenes such as 3-acetyldeoxynivalenol in F. culmorum appears to require further 3-hydroxylation to isotrichotriol prior to cyclization.