A new procedure for the analysis and purification of naturally occurring anatoxin-a from the blue-green alga Anabaena flos-aquae

K. Harada, Y. Kimura, K. Ogawa, M. Suzuki, A. M. Dahlem, V. R. Beasley and W. W. Carmichael. A new procedure for the analysis and purification of naturally occurring anatoxin-a from the blue-green alga Anabaena flos-aquae. Toxicon 27, 1289-1296, 1989.-Anabaena flos-aquae produces at least two neurotoxins termed anatoxin-a (ANTX-a) and -a(s) [ANTX-a(s)]. ANTX-a is a potent postsynaptic depolarizing neuromuscular blocking agent. Its structure is the secondary amine, 2-acetyl-9-azabicyclo[4.2.1]non-2-ene. Known isolation and analysis methods have not always given satisfactory results. Therefore, an improved procedure was developed for both isolation and analysis of ANTX-a which involves four steps; extraction, clean-up, separation and determination. Extraction was efficiently done with 0.05 M acetic acid, and a reversed phase, ODS (octadecylsilanized) and a cation exchanger (COOH) organosilans bonded to silica gels were applied to a clean-up step. Separation and determination were then performed using two TLC and HPLC chromatographies.