1. Neuroactive Compounds From Mollusk-Associated Bacteria Zhenjian Lin 1, Ma. Diarey B. Tianero 1,2, Rowena Antemano 1,2, Olivier Peraud 1, Margo Haygood 3, Gisela P. Concepcion 2, Baldomero M. Olivera 5, Alan Light 5, Eric W. Schmidt 1 Cone snails are an excellent source of neuroactive natural products, but to our knowledge no study of natural compounds from cone snail symbionts has been reported. 1 We have been studying the potential of symbiotic bacteria to contribute to the neuroactivity of their host snails, using calcium imaging of dorsal root ganglion (DRG) cells as the primary assay. DRG is a collection of cell bodies of sensory neurons monitoring touch, stretch, temperature, and pain. Here we used cultured DRG cells to screen for neuroactive compounds. The structures of pulicamides A-J Acknowledgment: This work was financially supported by ICBG (NIH). Response of intracellular [Ca 2+ ] to various additives 1 KCl/wash extract/drug KCl+drug/wash CP32 Streptomyces sp. CP32 was isolated from the hepatopancreas of Conus pulicarius, which was collected in the Philippines. DRG assay showed the crude extract increased the second KCl response at 25 ug/mL. 1 th KCl/wash drug 2 th KCl+drug/wash Strain CP32 was cultured both in 2.8 L Fernbach flasks and 10 L fermentor, each containing ISP2 medium. The cultures were grown for 8 days at 30 ℃ while shaking at 200 rpm. The culture was extracted with HP20 resin (20–30 gL -1 ) and the resin was eluted with MeOH and the solvents were dried. The resulting fraction was extracted with EtOAc and concentrated. The organic extract was subjected to pressure column chromatography over C18 using increasing amounts of MeOH in H 2 O. The residue from the 60~70% MeOH in H 2 O fraction was further purified on reverse phase HPLC to afford pulicamides A-J (1-10). HPLC showed CP32 produced a series of compounds with similar UV absorption. Lanthanide-induced shifts of (S)- and (R)-Mosher esters defined the absolute configurations of the primary alcohol in compounds 1 and 2. 2 Δδ OCH3 Configuration of C-4 MTPA esters (S)-(R)- 1+0.05+0.34S 2+0.01+0.05S CD mdeg Wavelength (nm) KCl KCl drug drug+KCl 200mMCap 100mM KCl Pulicamide B ( 2) decreased the 2 nd KCl response Introduction Fermentation and Purification Structure Elucidation The C4 chiral center dominated the CD spectrum, giving positive Cotton effects at 260~280 nm for S configuration. Conclusion: 60ug/ml NOESY correlations and relative configurations of compounds 1,5, 6 and 7. References: 1. Peraud, O. et al. Applied and Environmental Microbiology, 2009, 6820-6826. 2. Len, C. et al. Tetrahedron 61, 2005,10583–10595. Ten new hydroxamate compounds, pulicamides A-J (1-10), were isolated from the neuroactive strain CP32. The structures of these pulicamides were determined by NMR analyses and ESIMS experiments. Pulicamides are a new class of secondary metabolites. DRG assay showed that pulicamides A and C decreased the second KCl response, while pulicamide B increased the second KCl response. Their slight structural differences lead to opposite activities. To explain the structure-activity relationship, other analogs will be assayed in further studies. Departments of Medicinal Chemistry, 1 Anesthesiology, 5 and Biology, 4 University of Utah, Salt Lake City, Utah 84112, USA; Marine Science Institute, University of the Philippines, Diliman, Quezon City 1101, Philippines, 2 Department of Environmental and Biomolecular Systems, OGI School of Science & Engineering, Oregon Health & Science University, Beaverton, Oregon 97006, USA. 3 Pulicamide C ( 3) increased the 2 nd KCl response. 20ug/mL. Pulicamide A ( 1) decreased the 2 nd KCl response. 10ug/mL Neuroactivities KCl drug drug+KCl 200mMCap 100mM KCl KCl drug drug+KCl http://www.conchology.be