In the course of screening for new antibiotics, a Chromobacterium strain Y-T0678H isolated from a soil sample collected at Lake Kamakita in Saitama Prefecture, Japan, was found to produce a new antibiotic. The strain was identified as Chromobacterium violaceum based on morphological, cultural and physiological characteristics. The antibiotic Y-T0678H was isolated from the fermentation broth via extraction, concentration and preparative thin-layer chromatography. Its physicochemical properties and spectroscopic data (HR-MS, UV, IR, NMR) suggested the structure was 6-hydroxy-3-oxo-1,2-benzisoxazolin, which was confirmed by synthesis. Y-T0678H exhibits selective activity against Gram-negative bacteria, and the acute toxicity (LD50) in mice is approximately 1,560 mg/kg (intravenous).