Bacillaene, a novel polyene antibiotic, was discovered and isolated from fermentation broths of a strain of Bacillus subtilis. The novel antibiotic has a nominal molecular weight of 580 and an empirical formula of C35H48O7. Bacillaene is active against a broad spectrum of bacteria in agar-plate diffusion assays. Studies in vitro indicate that the antibiotic inhibits prokaryotic protein synthesis but not eukaryotic protein synthesis. Cell survival studies performed with strains of Escherichia coli indicate that the antibiotic is a bacteriostatic agent.