The neuronal degeneration which results from cerebral ischemia is thought to be due to an overexcretion of the excitatory amino acid, L-glutamic acid, which acts as a neurotransmitter in the major part of brain. Brain ischemia injury maybe expected to be overcome by L-glutamate toxicity suppressors. In the course of our screening for substances that protect neuronal hybridoma N18-RE-105 cells from L-glutamate toxicity, we isolated carquinostatin A, lavanduquinocin, aestivophoenins A and B, and 4-demethoxymichigazone. Further investigation has resulted in the isolation of a novel neuronal cell protecting substance, naphthomycinol (1). We report herein the fermentation, isolation and structure determination of 1. The naphthomycinol producing organism, identified as Streptomyces sp. PF7, was cultivated in a 50-liter jar fermenter under specific conditions. The mycelial acetone extract was processed through solvent extraction and various chromatographic methods to purify 1. The molecular formula of 1 was established as C40H49NO9 by high-resolution FAB-MS. The 1H and 13C NMR spectral data indicate that the structure of 1 is very similar to those of naphthomycins except for the presence of an additional oxymethine. Naphthomycinol, a member of the naphthomycins series, is the first compound which has a hydroxyl function at C-ll so far reported. In the evaluation system we employed, 1 decreased the L-glutamate toxicity in N18-RE-105 cells with EC50 value 400 nM. Since the L-glutamate toxicity in N18-RE-105 cells was thought to be caused by glutathione depletion, we assessed buthionine sulfoximine (BSO) toxicity which directly inhibits gliutathione synthesis. Antioxidants such as vitamin E suppress both the L-glutamate and the BSO toxicities in N18-RE-105 cells. Naphthomycinol, however, did not suppress the BSO toxicity. This result strongly suggests that the mode of action of naphthomycinol is not based on the antioxidative activity.