Antifungal metabolites produced by Bacillus pumilus in Potato Dextrose Broth (PDB) were isolated from culture supernatant fluid by precipitation with ammonium sulphate. The antifungal metabolites inhibited mycelial growth of many species of Aspergillus, Penicillium and Fusarium. They also inhibited production of aflatoxins, cyclopiazonic acid, ochratoxin A and patulin. The metabolites were heat-stable and remained active after sterilization at 121 degrees C for 15 min. Their activity was stable over a wide range of pH (2-10). The metabolites were resistant to hydrolysis by various proteases, peptidases and other enzymes. They were also resistant to denaturation by many protein-denaturing detergents except Nonidet P-40. The metabolites were soluble in water and relatively polar organic solvents. Chromatographic bioassay revealed that a crude precipitate of the metabolites contained only one compound with antifungal activity. The active compound did not form a fluorescent derivative with fluorescamine suggesting that the compound is either a cyclic polypeptide or a non-peptidic compound.