The endophytic fungus Trichothecium roseum LZ93 from Maytenus hookeri was found to antagonize other pathogenic fungi in vitro. To identify which compound contributed substantially to the antagonism, we fermented the strain and purified its fermentation products. Eleven compounds were obtained, including two trichothecenes, five rosenonolactones, two cardiotonic cyclodepsipeptides, and two sterols. Compound 11β-hydroxyrosenonolactone (1) was assigned according to 1D and 2D-NMR data for the first time. At the same time, the (1)H and (13)C-NMR assignments for 6β-hydroxyrosenonolactone (2) were revised. Of all of them, only trichothecin (6) showed strong antifungal activity. Based on our observations of the antagonistic activity and the other experimental results, we suggest that the antifungal compound trichothecin was the main contributor to the antagonistic action of T. roseum LZ93.