Zooxanthellatoxin-B was isolated from a symbiotic marine alga, Symbiodinium sp. and the structure of zooxanthellatoxin-B was determined to be a 62-membered lactone by comparing spectral data and degradation products with those of zooxanthellatoxin-A. Marine microalgae produce various types of compounds including nitrogenous neurotoxins, polyether sea food toxins, sulfonium compounds of dimethylsulfide precursors and antineoplastic macrolides. Zooxanthellatoxin-A (ZT-A) and ZT-B were isolated as vasoconstrictive substances from a symbiotic marine dinoflagellate Symbiodinium sp. (strain No. Y-6) belonging to zooxanthellae, well known symbionts distributed in a wide range of marine invertebrates. The structure of ZT-A has recently been determined as a 62-membered lactone (1) on the basis of the spectral data and the extensive degradation experiments. Here we would like to report the structure of ZT-B, a congener of ZT-A. ZT-B (2) was isolated as amorphous solids and showed a pseudo molecular ion at m/z 2829.9 in the negative FAB-MS spectrum, suggesting 42 mass unit loss from 1. The molecule contained one nitrogen (elemental analysis) and one sulfate ester (ion chromatography) like ZT-A. Lactone structure of 2 was evident by formation of a seco-acid (3) upon treatment with KOH like 1. The lowest oxymethine signal of 2 at δ 5.42 was disappeared in 3. Comparison of the TOCSY and 13C NMR spectra of ZT-B with those of ZT-A revealed the presence of common characteristic functionalities including two conjugated dienes, an exomethylene, a bisepoxide, two acetals (one spiroacetal δ C 95.6 and one hemiacetal δ C 99.6), and two carbonyls (one ester, δ C 173.5, and one amide, δ C 175.2) found in ZT-A. The structural similarity including localization of the common functional groups between 2 and 1 was established by the following degradation experiments: Hydrolysis of 2 with LiOH afforded a terminal acid fragment (4, C1'-C25') which is identical with one from 1 in the 1D and 2D 1H NMR and negative FAB-MS spectra. Periodate oxidation of 1 followed by sodium borohydride reduction gave a dienediol and a bisepoxide-containing diol corresponding to C7-C13 and C14-C56, respectively, which were identical with those from ZT-A in their 1H NMR spectra. Based on these results, all of proton signals for the partial structures from C7 to C56 and from C1' to C25' were able to be assigned in the 1H NMR spectrum of 2 at the almost same chemical shift with the same splitting pattern to those of 1. Pulse field gradient (PFG) COSY and TOCSY spectra of 2 allowed us to follow a proton sequence from H7 to H2, the latter correlated to C1 carbonyl carbon by the HMBC spectrum. Among them the lowered chemical shift of H3 (δ 4.54) suggested that oxygenation occurred at C3. On the other hand, oxymethine signal (H56) could be connected to a series of three oxymethine proton signals and then to the lowest oxymethine proton signal (H61, δ 5.42) through H60 methylene signals, indicated the presence of a 62-membered lactone structure. The partial structure could be extended to C62 and further connected to C68 by long range couplings of exomethylene signals to both H66 oxymethine and H68 methylene signals, indicating loss of two olefinic carbons in the middle part of the structure of 2. The structure could be extended to C74 via a characteristic triplet signal for H72 based on the TOCSY spectrum. Proton network at the high field region of TOCSY spectrum of 2 was almost superimposable to that of 1, suggesting the presence of a spiroacetal structure in 2. The carbon connectivity from the spiroacetal to C79 oxymethine was revealed by TOCSY and PFG COSY spectra. The structure from C74 to C79 was rather difficult to be determined because of serious signal overlapping of the repeated 1,3-diol structure. The structure was confirmed by a fact that a heptaacetate (5) was isolated after acetylation from a mixture of the above mentioned periodate oxidation reaction. Under these conditions no 1,2-diol structures could survive. In the COSY and TOCSY spectra of 5, proton connectivities from H66 to H71 and from H72 to H75 was established. Although three oxymethines (H75, H77, and H79) were not well resolved, their correlation to two isolated methylenes (H76 and H78), verified the existence of 1,3-diol structure rather than 1,2-diol structure. The proton sequence from oxymethine H91 of the spiroacetal to H102 oxymethine was able to be traced by 1H-1H cross peaks in the TOCSY spectrum. Chemical shifts of isolated methylene proton H104 validated the amide structure in 2. The molecular formula C138H200NO56SNa (m/z 2829.9) implied the existence of one more ring structure in addition to the six ring structures, 14 olefins and two carbonyls established by the above chemical degradation experiments. Thus the tetrahydropyrane hemiacetal structure was assigned for a structure from C99 to C103 on the basis of a carbon signal at δ 99.6, resulted in a hydroxyl group at C3 carbon. Symbiodinium sp. (strain No. Y-6) produced not only ZT-A and ZT-B as vasoconstrictive substances but also the corresponding seco-acids having no vasoconstrictive activity and their production depended on the culture conditions. It is noteworthy to mention that subsistence of the common ring structures in zooxanthellatoxins in spite of difference of two carbon unit in the middle of the structures may suggest the importance of the ring structures for biological activities as well as their convergent biogenetic pathway. Spectral similarity of the ring portions in 1 and 2 may suggest that the same stereochemistry for 2 and 1. Further studies on related compounds and detailed investigation of their chemical structure and biological activities are in progress in our laboratory.