In the course of a screening program for new antitumor antibiotics, Streptomyces flavidovirens strain K3619 was isolated from a Brazilian soil sample. This strain produces a new cyclic hexadepsipeptide antibiotic designated citropeptin, whose structure—elucidated by NMR spectral analysis (including two-dimensional techniques)—is closely related to azinothricin and A83586C. This paper describes the taxonomy of the producing organism, as well as the fermentation, isolation, and physicochemical properties of citropeptin. Citropeptin exhibits potent cytotoxicity with IC50 values of 0.02 μg/ml against murine P388 leukemia cells and 0.1 μg/ml against B16 melanoma cells. In in vivo studies, it prolonged the survival of CDF1 mice bearing P388 leukemia (T/C 123%) when administered intraperitoneally at 2 mg/kg/day on days 1 and 5, but caused toxic death at 4 mg/kg/day.