Analysis of Anatoxin-a by GCIECD*

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Abstract
Anatoxin-a (ANTX-a), a potent cholinergic alkaloid produced by some toxigenic strains of the cyanobacterium Anabaena flos-aquae, has been responsible for death of livestock, pets, and wildlife. An increase in the frequency of cyanobacterial blooms in fresh waters of temperate North America has produced a concomitant increase in the threat posed by toxic strains of some of these microorganisms. Presently, a mouse bioassay is the general procedure used for testing the toxicity of a bloom—approximately 5 µg/mL ANTX-a sensitivity. A method that is over three orders of magnitude more sensitive than that bloassay is described here. It is readily capable of detecting and quantitating sublethal levels of ANTX-a. ANTX-a is isolated via solid-phase extraction (SPE), derivatized with trichloroacetic anhydride, isolated a second time via SPE, and analyzed by GC/ECD. Sensitivity to 5 ng is achieved. Sample size can be as large as 10 mL. “Nontoxic” samples are shown to contain ANTX-a.