The bacterium Pseudomonas aureofaciens 63-28 is antagonistic to several plant pathogenic fungi, including Pythium spp. The bacterium produced at least four antifungal metabolites active against Pythium ultimum and Phytophthora cryptogea when tested in culture for antifungal activity. Two of these compounds were identified as the novel butyrolactones (Z)-4-hydroxy-4-methyl-2-(l-hexenyl)-2-butenolide and (Z)-4-hydroxymethyl-2-(l-hexenyl)-2-butenolide, by using NMR and GC-MS. All compounds were different from other antibiotics produced by Pseudomonas spp., including pyoluteorin, pyrrolnitrin, and 2, 4-diacetylphloroglucinol, as determined by HPLC. This is the first report of butyrolactones with antifungal activity produced by a saprophytic Pseudomonas spp.