Ionomycin, a new polyether antibiotic with a high affinity for Ca ions, is obtained in pure form from fermentation broths of Streptomyces conglobatus by solvent extraction. It is unique among known polyether antibiotics in that it has a UV absorption maximum at 300 nm, distinguishing it from other antibiotics of its class. The Ca salt has the molecular formula C41H70O9Ca. Ionomycin is a narrow spectrum antibiotic being active against gram-positive bacteria.