Radiobiosynthesis of paralytic shellfish toxins in the dinoflagellate alexandrium tamarense

Paralytic shellfish toxins (PSTs) are a family of neurotoxic alkaloids produced by certain dinoflagellates including strains of Alexandrium tamarense that are found in Hong Kong and South China coastal waters. The frequent occurrence of these toxins in commercial shellfish and other seafood is a serious food safety, economic and environmental problem worldwide. There is an urgent need for sufficient quantities of 14C-labeled PSTs for toxicological studies and other research and development uses. In the present study, we set out to 1) produce 14C-labeled PSTs using appropriate 14C-labled precursors in an optimized culture of a local strain of Alexandrium tamarense (ATCI01) that produced mostly C2 toxin (C2) and to 2) optimize the post-column reaction conditions for different PSTs to maximize the sensitivity of their detection using high performance liquid chromatography (HPLC). Screwed flasks were used as culture vessels to incorporate 14C from bicarbonate, acetate or arginine into C2. Nitrate, but not ammonium, was a good nitrogen source for the culture and was required for both growth and toxin productivity. Bicarbonate was essential for both algal growth and toxin biosynthesis. Acetate was toxic to the culture at relatively high concentrations. Arginine significantly enhanced both toxin yield and cellular toxin content, suggesting that it is a committed precursor in PST biosynthesis in dinoflagellates. The 14C-C2 produced was purified by column chromatography, which could be converted to obtain 14C-labeled C 1 , gonaytoxin (GTX) 2, and GTX3. Under the post column reaction conditions optimized for each of the different toxins, the specific activity of each was precisely determined by HPLC and scintillation counting. Several million dpm of14C-C2 toxin were produced and purified in the present study. These products are useful for metabolism studies and for calibrating the analytical PST standards available from commercial sources