HPLC separation and comparative toxicity of saxitoxin and its reaction products

A chromatographic method was developed that was used to purify saxitoxin and separate it from its chemically modified products and the reagents used in the reactions. The separation time is about 10 minutes. Using differential-refractive-index detection, quantitation of the products (+/- 10%) can be done on 30-100 microgram of toxin. A simple bioassay with crab leg nerves in vitro was used in conjunction with the chromatography to determine, within a factor of two, the inhibition binding constants of saxitoxin and its products. The binding constant for saxitoxin at ambient temperature, 18-21 degree C, is Ki approximately 80 nM. The acid-hydrolysis product has Ki approximately 8 microM under the same conditions. The chemistry of saxitoxin was investigated using the chemical and bioassays.