Purification and characterisation of large and small maitotoxins from cultured gambierdiscus toxicus

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Abstract
Three strains of cultured Gambierdiscus toxicus yielded distinct maitotoxins (maitotoxin‐1, ‐2, and ‐3) which were purified to homogeneity by high pressure liquid chromatography. Maitotoxins‐1 and ‐2 are large toxins (molecular weights for the sodium salts = 3,422 and 3,298, respectively), whereas maito‐toxin‐3 is relatively small (molecular weight = 1,060 for the sodium salt). The contractile actions on isolated guinea‐pig left atria, vas deferens and ilea of maitotoxins‐1 and ‐2 were compared with those of the small maitotoxin, maitotoxin‐3. Maitotoxin‐1, ‐2, and ‐3 each produced quantitatively similar, concentration‐dependent patterns of positive and negative inotropy in atria when compared on a mouse unit/ml basis (one mouse unit is the intraperitoneal LD50 dose for a 20 g mouse; the LD50 for maitotoxin‐2 = 0.08 μ g/kg). Concentrations of maitotoxin‐2 greater than 5 × 10−13 M caused positive inotropy. The three maitotoxins produced patterns of contractions in vas deferens and ilea that were qualitatively similar, including a period of prominent spike activity in vas deferens. On a mouse unit/ml basis, the order of potency on smooth muscle was maitotoxin‐1> maitotoxin‐3 > maitotoxin‐2. The contractile responses of smooth muscle to the maitotoxins were followed by an inhibitory phase where control agonist responses could not be elicited. The maitotoxin‐induced contractile responses of vas deferens were inhibited by nicardipine but not phentolamine, indicating that in this tissue, each maitotoxin has mainly a direct contractile effect that requires calcium influx through voltage‐sensitive calcium channels. The qualitatively similar in vitro (and in vivo) actions of the three toxins suggests that each maitotoxin has a similar mode of action, despite the considerable difference in their molecular size. The reduction in potency upon solvolysis of these maitotoxins suggests that sulphate moities are important for the biological activity of each maitotoxin.