A new compound, harzianum B (1), was isolated from the culture broth of a fungal strain, Hypocrea sp. F000527, together with a known trichothecene, harzianum A (2). Its structure was determined by spectroscopic analyses including HRFAB-MS and various 1H NMR and 13C NMR spectral data. Harzianum B (1) was characterised as (E,Z,E)-2', 4', 6'-octatriendioic acid esterified on the 4β-hydroxyl group of trichodermol. Harzianums A (2) and B (1) showed cytotoxicity against several human cancer cell lines.