Novel toxins produced by the dinoflagellate Karenia brevisulcata

► Novel toxins have been isolated from cultures of the very toxic dinoflagellate Karenia brevisulcata. ► K. Brevisulcata caused widespread devastation of marine life when it bloomed in New Zealand during the summer of 1998. ► KBT-F and KBT-G are large polyether toxins MW > 2000 that are strongly cytotoxic, haemolytic and very toxic to mice. ► Brevisulcatic acids are less toxic but have activity at voltage-gated sodium channels and have some structural similarity to brevetoxin-A. ► BSXs are found in cultures mainly as the lactone-ring opened hydrolysis products BSX-1 and BSX-2. Karenia brevisulcata (Chang), a new toxic dinoflagellate of the genus Karenia was isolated from a harmful algal bloom that occurred in Wellington Harbour, New Zealand in 1998. The bloom severely affected most marine biota resulting in long-term ecological damage and causing respiratory distress in harbour bystanders. Cultures of K. brevisulcata produced a range of novel toxins including ten lipid-soluble K. brevisulcata toxins (KBTs) and six water-soluble brevisulcatic acids (BSXs). Brevetoxins were not detected. KBT-F, KBT-G, BSX-1 and BSX-2 were isolated from 1450 L of bulk cultures and purified in mg quantities. Preliminary chemical and toxicological investigations show that KBT-F (M 2054 C 107H 160O 38) and KBT-G (M 2084 C 108H 162O 39) are complex polycyclic ethers with UVmax at 227 nm. NMR data gave characteristics of ladder frame polyether structures and a 2-methylbut-2-enal side chain, similar to gymnocins. The mouse i.p. LD 50s for KBT-F and -G were 0.032 and 0.040 mg kg −1, respectively. These KBTs were also highly cytotoxic and haemolytic. BSX-1 (M 916 C 49H 72O 16) and BSX-2 (M 872 C 47H 68O 15) are polycyclic ether dicarboxylates with UVmax 196 nm. BSX-4 and BSX-5, the lactone ring-closed analogues and the presumed primary toxins in the algal cells, were isolated in smaller quantities. Preliminary structural information from NMR and MS showed a carboxylated side chain and some similarities to brevetoxin-A. However, the structures have not yet been fully elucidated due to conformers confounding the NMR. The mouse i.p. LD 50 for BSX-1 was 3.9 mg kg −1 while no deaths were seen in mice injected with BSX-2 at 6.6 mg kg −1. The LD 50s for the lactones BSX-4 and -5 were 1.4 and 1.6 mg kg −1 respectively. BSX-4 and -5 were agonists of voltage-gated sodium channels but only weakly haemolytic. Activities in the Neuro-2a cytotoxicity assay were ca 10% of dihydrobrevetoxin-2 and we