Azonazine, a unique hexacyclic dipeptide was isolated from a Hawaiian marine sediment-derived fungus eventually identified as, Aspergillus insulicola. Its absolute configurations, 2R, 10R, 11S, 19R were established using NMR, HRESIMS, and CD data plus insights derived from molecular models. A possible route for its biogenesis is proposed and biological properties were explored against cancer cell lines and in an NFκB inhibition assay.We believe that scientific study of fungi (Ascomycota) within the genus Aspergillus (Class: Eurotiomycetes, Family: Trichocomaceae) is important. Though more than 200 Aspergillus strains have been isolated from a host of terrestrial ecological niches, they provide a steady stream of diverse small molecules.1 Recent understanding the genomics of chemically Correspondence to: Phillip Crews, phil@chemistry.ucsc.edu. Supporting Information Available: Extraction scheme and bioactivity data, NMR data, possible configurations and biosynthetic pathway of azonazine. This material is available free of charge via the Internet at http://pubs.acs.org. We began a campaign to emphasize chemical study on the >20 marine-derived Aspergillus strains (from sponges and sediments) housed in the UCSC repository. The prospect of encountering orphan 13 biosynthetic pathways was part of the rationale for this investigation. The results reported below represent a first step and involve the characterization and biological evaluation of a novel hexacyclic dipeptide, azonazine from A. insulicola related to ochraceopetaliformis (syn: A. flocculosisi). 14 This project was initiated by scanning our collection of Aspergillus cultures to identfy those possessing strain uniqueness accompanied by activity in a cytotoxicity soft agar-based disk diffusion assay. 15 One top candidate was A. insulicola (strain no. 088708a, identifed by molecular taxonomy evaluations shown in SI Table S5) obtained from a Hawiaan shallow water sediment. The EtOAc crude extract of the initial small-scale culture (125 mL, in Czapek-Dox liquid medium, pH adusted to 7.0, prepared with artificial seawater) exhibited potent activity (See Supporting Information (SI) Table S2) against murine Colon 38 cell lines and selectivity against human prostate adenocarcinoma (LNCaP) vs human leukemia cell lines (CEM). A scale up culture (10 L, same media) was harvested at 21 days (shaking at 150 rpm) and worked up by passing it through HP-20 resin. The resin was subsequently washed using water (fraction coded: Hp1, 362 mg), 50% methanol/water (Hp2, 1327 mg), methanol (Hp3, 443 mg) and isopropanol (Hp4, 12.8 mg). Each fraction was subjected to LCMS analysis and the Hp3 was selected for further purification via RP-HPLC (30-50% acetonitrile / 0.1% formic acid-water, 50 mins), and 42 fractions (F1-F42) were collected. Further purification of fraction F11 (3.5mg) via RP-HPLC yielded azonazine (1.1mg) and, insulicolide A (2.2 mg). 16 NIH Public AccessEarly on we recognized that azonazine, 17 obtained as a colorless powder and possessing molecular formula C 2...

Sampling of California nearshore sediments resulted in the isolation of a Gram-negative bacterium, Photobacterium halotolerans, capable of producing unusual biosynthetic products. Liquid culture in artificial seawater-based media provided cyclic depsipeptides including four known compounds, kailuins B–E (2–5), and two new analogues, kailuins G and H (7 and 8). The structures of the new and known compounds were confirmed through extensive spectroscopic and Marfey's analyses. During the course of these studies, a correction was made to the previously reported double-bond geometry of kailuin D (4). Additionally, through the application of a combination of derivatization with Mosher's reagent and extensive 13C NMR shift analysis, the previously unassigned chiral center at position C-3 of the β-acyloxy group of all compounds was determined. To evaluate bioactivity and structure–activity relationships, the kailuin core (13) and kailuin lactam (14) were prepared by chiral synthesis using an Fmoc solid-phase peptide strategy followed by solution-phase cyclization. All isolated compounds and synthetic cores were assayed for solid tumor cell cytotoxicity and showed only minimal activity, contrary to other published reports. Additional phenotypic screenings were done on 4 and 5, with little evidence of activity.