Tetrahedron Letters, Vol. 38, No. 40, pp. 6969-6972, 1997 Pergamon © 1997 Elsevier Science Ltd All rights reserved. Printed in Great Britain PH: S0040-4039(97)01666-3 0040-4039197 $17.00 + 0.00 Pitiamide A, a New Chlorinated Lipid from a Mixed Marine Cyanobacterial Assemblage Dale G. Nagle ~', Peter U. Park lb, and Valerie J. Paul* University of Guam Marine Laboratory, U.O.G. Station, Mangilao, Guam 96923, USA Abstract: Pitiamide A, a new chloro-lipid, was isolated from an extract of a mixed assemblage of Lyngbya majuscula and a Microcoleus sp. found growing on intact colonies of the hard coral Porites cylindrica on Guam. The structure was determined by interpretation of 2D-NMR spectra. © 1997 Elsevier Science Ltd. Black band disease, a cyanobacterial infection of scleractinian hard corals by Phormidium corallyticum has been documented to cause coral colony destruction in the western Atlantic and elsewhere.2 Cytotoxic metabolites, the nakienones A-C and nakitriol, have been isolated from a Synechocystis sp. found overgrowing (or infecting) an Okinawan species of hard coral) We have recently observed a thin blue-gray tuff-forming assemblage of Lyngbya majuscula and Microcoleus sp. (approx. 4:1) overgrowing the apical tips of otherwise healthy, intact branches of the yellow reef-building coral Porites cylindrica at various locations throughout Micronesia.4 Small cyanobacteria tufts (1-4 cm3) were carefully removed from coral tips and stored at -20* until extracted with CH2C12/MeOH (I:1). The crude extract strongly deterred feeding by the yellow-banded parrotfish (Scarus schlegeh), a common reef herbivore found throughout Micronesia: 2D-TLC analysis of the extracts showed the presence of several UV-active, orange and purple-charring (H2SO 4, heat) relatively non- polar secondary metabolites. A 752.4 mg portion of the crude extract (1.31 g, dark oil, 28.7 g dry marc) was fractionated by silica gel column chromatography with a gradient of hexanes to EtOAc to CH2C12 to MeOH. The fractions eluting with 40% EtOAc/hexanes to 100% EtOAc (v/v) were combined (242.0 mg) and a portion (210.9 mg) was separated by Sephadex LH-20 chromatography (50% (v/v) CH2C12 in MeOH), and further purified by repetitive NP-HPLC (75 and 65% EtOAc in hexanes). The two major compounds were decolorized with activated charcoal. Final purification by NP-HPLC (2% (v/v) MeOH in CH2C12provided pitiamide A (1, 8.3 rag) and pitiamide B (2, 15.1 mg). 6 Low resolution EIMS examination suggested the presence of a chlorine in the structure of 1 (m/z 381, 8.6% rel. abundance [M] +; 383, 3.1% rel. abundance [M+2]*). Analysis of 1 by t3C NMR and High resolution ElMS (381.2551; calc. for 381.2434; 70 eV) provided a molecular formula for [M]÷ of C22H36C1NO 2 (calc. for 5 ° unsaturation). Compound 1 was optically active [trip = -10.3" (C = 3.0, CHC13). Examination of 1 by IR (neat, x) = 3650-3100, 2959, 2930, 2873, 1709, 1645, 1553 cmt), UV (~ 223 nm, MeOH), and 'H-NMR data (Table 1) revealed the presence of a ketone, an amide-linkage, a conjugated diene, and two methyl branches. Three IH spin systems were assembled by tH-tH COSY (Figure I) and two carbonyl carbons were confirmed by ~3C-NMR. The ~H-~3CHMBC spectrum facilitated the attachment of spin systems a-c (Figure 1) and the carbonyl moieties. Specifically, 2 and 3-bond couplings from carbonyl carbon C8 (8 214 ppm) to H29 of partial structure b and H7 and H323 of a allowed for the attachment of a and b through a ketone moiety. The C14 carbonyl (~ 172.6 ppm) showed 2JcHand 3JcHcouplings to H215 and H216 of c and Hl2a and Hl2b of b. Attachment of a vinyl chloride to C 1 completed the planar structure of 1. Additionally, a pronounced 6969