Palytoxin (1, C129H223N3054, Figure 1) is one of the most potent and complex marine toxins.' Though first isolated from the zoanthid Palythoa toxica, its biogenetic origin was questioned because of marked seasonal and regional variations,2 but the question was never pursued. Sporadic occurrence of palytoxin in an alga,3 crabs," and a herbivorous fish5 also suggested that the toxin might be produced by a microorganism and transmitted to the marine food chain, as is the case with ciguatera, where the epiphytic dinoflagellate Gambierdiscus toxicus produces ciguatoxins and maitotoxin that are isolated from fish.6 Ostreopsis siamensis drew our attention because of its close taxonomical relationship to G. toxicus and the potency of its toxins, named ostre~cins.~ In this Communication, we report the major ostreocin as a palytoxin analog. 0. siamensis (SOA1 strain) was collected at Aka Island, Okinawa, Japan, and grown for 30 days under the same culture conditions as G. toxic~s.'~ Cells harvested by filtration were extracted three times with MeOWH20 (1 : 1) at room temperature and then with MeOH/H20/HOAc (50:50:0.2) under reflux. Purification of ostreocins from the combined extracts was carried out by solvent partition and column chromatography.8 Ostreocins were resolved into five constituents. Purification was monitored by mouse assay following characteristic W maxima. The major constituent, ostreocin D, accounted for 70% of total toxicity. From 936 L of culture, 3.8 mg of ostreocin D was recovered. (2) Moore, R. E.; Helfrich, P.; Patterson, G. M. L. Oceanus 1982, 25, 54-62. (3) Maeda, M.; Kodama, R.; Tanaka, T.; Yoshizumi, H.; Nomoto, K.; Takemoto, T.; Fujita, M. 27th Symposium on the Chemistry of Natural Products; Tanaka, O., Ed.; Symposium papers, Hiroshima, Japan, 1985; Yamawaki Publishing: Hiroshima, 1985; pp 616-623. (4) (a) Yasumoto, T.; Yasumura, D.; Ohizumi, Y.; Takahashi, M.; Alcala, A. C.; Alcala, L. C.(8) The algal extracts were suspended in CHzCl2 and extracted with MeOWH20 (1:l). The aqueous phase, after being concentrated, was extracted with 1-BuOH. The 1-BuOH extract was successively chromatographed on the following columns with solvents as indicated: Toyopearl HW60-fine, MeOWH20 (3:2), Inertsil C8, H20, H20iPrOH (4:1,3:Z); CM Toyopearl 650M. 1% HOAC. The eluates were monitored by mouse assay. Resolution of ostreocins into five constituents and final purification of ostreocin D were achieved on Develosil TMS-5 with H20/CH3CN (3: 1) containing 0.1% HOAc. Eluates were monitored with a Hitachi C-6500 J3D Chromatosystem at 235 and 263 nm. All columns were washed with 10 mM aqueous EDTA-2Na before use. 47, 1029-1033. 0002-7863/95/15 17-5389$09.00/0 on Figure 1. Planar structure of palytoxin (1) and ostreocin D (3,26bisdesmethyldeoxypalytoxin). Bold lines denote connectivities elucidated by DQF-COSY, TOCSY, HSQC, and HMBC data. W chromophores are bracketed.12 Palytoxin, Rl = RZ = Me. Ostreocin D, R1 = Rz = H; O H -H (position not located). Ostreocin D (OST-D) was a colorless amorphous solid: positive to ninhyd...

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Negative-ion fast-atom bombardment collision-induced dissociation tandem mass spectrometric (FAB-CID-MS/MS) methodology was successfully applied to verify the highly complex structure of ostreocin-D (MW 2633), a new palytoxin analog isolated from the marine dinoflagellate Ostreopsis siamensis and proposed to be 42-hydroxy-3,26-didemethyl-19,44-dideoxypalytoxin based on NMR data. The charge-remote fragmentations were facilitated by a negative charge introduced to a terminal amino group or to a hydroxyl group at the other terminus by a reaction with 2-sulfobenzoic acid cyclic anhydride. Product ions generated from the [M - H](-) ions provided information on the structural details of ostreocin-D. Comparisons between the spectral data for ostreocin-D and palytoxin also provided a rational basis for the assignments of product ions.