Cerebral ischemic disorders are one of the main causes of death. In brain ischemia, blood flow disruptions limit the supply of oxygen and glucose to neurons, initiating excitotoxic events. These include activation of glutamate receptors and release of excess glutamate inducing neuron depolarization and significant increase of intracellular calcium, which activates multiple intracellular death pathways. 1 Accumulation of extracellular glutamate also inhibits cystine-glutamate exchanger, resulting in depletion of the intracellular antioxidant glutathione. 2,3 Under such conditions, reactive oxygen species are generated and implicated in neuronal cell death. 4 In this study, microbial metabolites were screened to find neuroprotective agents against glutamate toxicity.C6 rat glioma cells undergo cell death when exposed to 100 mM glutamate for 24 h. Thus, they provide a good model for evaluating neuroprotective activity against glutamate toxicity. Our screening using C6 cells resulted in the isolation of a new active compound designated as flaviogeranin (Figure 1) from Streptomyces sp. RAC226.The producing organism was cultivated in 500 ml Erlenmeyer flasks containing 100 ml of a medium consisting of glucose (2.5%), soybean meal (1.5%), dry yeast (0.2%) and calcium carbonate (0.4%) (pH 6.2, before autoclave) on a rotary shaker at 27 1C for 5 days. The culture broth (2 l) was centrifuged and the mycelium was extracted with acetone. After evaporation, the aqueous concentrate was adjusted to pH 3 and extracted with ethyl acetate. The extract was applied to preparative silica gel TLC plates, which was developed with hexaneethyl acetate-triethylamine (150:50:5). The crude material was subjected to HPLC (PEGASIL ODS, Senshu Scientific, Tokyo, Japan) with 87% methanol and 0.2% trifluoroacetic acid. The active fraction was further purified by XBridge C 18 HPLC (Waters, Milford, MA, USA) with 87% methanol and 0.2% triethylamine. The peak fraction was concentrated to dryness to give an orange powder of flaviogeranin (2.2 mg).The physicochemical properties of flaviogeranin are summarized as follows m.p. 160-165 1C; high-resolution FAB-MS m/z 371.1858 (MH + , calcd for C 22 H 27 O 5 , 371.1859); UV l max (e) 221 nm (252 000), 265 (125 000), 307 (64 200) and 429 (29 300) in methanol, 220 nm (250 000), 265 (125 000), 307 (64 000) and 429 (29 300) in 0.01 M HCl-methanol, 220 nm (sh, 246 000), 265 (109 000), 307 (59 500) and 429 (25 000) in 0.01 M NaOH-methanol; and IR (ATR) n max 3090, 3060, 1680 and 1630 per cm.The molecular formula of flaviogeranin was established as C 22 H 26 O 5 by high-resolution FAB-MS. The 13 C-and 1 H-NMR data for flaviogeranin are summarized in Table 1. All one-bond 1 H-13 C connectivities were confirmed by an HMQC 5 experiment. The HMBC 6 spectrum identified a naphthoquinone chromophore as shown in Figure 2. Long-range correlations from a phenolic hydroxyl (5-OH) to C-4a, C-5 and C-6 and from a singlet methyl (9-H 3 ) to C-5, C-6 and C-7 revealed the sequence of four aromatic carbons (C-4a, C-5, C-6 and C...

Flaviogeranin (I) is isolated from Streptomyces sp.