Isolation of pectenotoxin-2 from Dinophysis acuta and its conversion to pectenotoxin-2 seco acid, and preliminary assessment of their acute toxicities

We have developed a simple and effective method for isolating pectenotoxin-2 (PTX-2) from Dinophysis cells collected from a natural bloom. A two-step extraction procedure followed by two column chromatography steps produced PTX-2 in high purity suitable for use as an analytical standard and for toxicological studies. Incubation of purified PTX-2 with the supernatant from ultracentrifuged blue (Mytilus edulis) or Greenshell(R) (Perna canaliculus) mussel hepatopancreas homogenate caused rapid conversion to pectenotoxin-2 seco acid (PTX-2 SA). Purification of PTX-2 SA was achieved by solvent extraction followed by column chromatography. PTX-2 and PTX-2 SA were fully characterized by LC-MS and NMR, and full H-1 and C-13 NMR assignments were obtained. Okadaic acid C-8-diol ester was isolated during the purification of PTX-2, and its identity confirmed by NMR and LC-MS analyses. Pectenotoxin-2 seco acid methyl ester, identified by LC-MS, was also produced during the hydrolytic procedure due to the presence of methanol. PTX-2 was acutely toxic to mice by i.p. injection (LD50 = 219 mug/kg) but no effects were seen with PTX-2 SA at 5000 mug/kg. Neither PTX-2 nor PTX-2 SA was overtly toxic to mice by the oral route at doses up to 5000 mug/kg. No diarrhea was observed in mice dosed with either compound, suggesting that pectenotoxims do not belong in the diarrhetic shellfish poison group. (C) 2003 Elsevier Ltd. All rights reserved.