Improved Methods for the Extraction and Purification of Corynetoxins for Preparative or Analytical Purposes using Water Solubilizing Additives

摘要

One per cent and 2% (w/v) solutions of methyl-p-cyclodextrin in water have proven to be useful solvents, comparable to the 80% methanol in water benchmark, for extracting CTs from bacterium-infected nematode galls in annual ryegrass seed heads. The aqueous cyclodextrin CT extracts are suitable for analysis by HPLC and ELISA for the presence of CTs. The aqueous cyclodextrin CT extracts are also suitable for determining the presence of the CT-producing bacterium since the bacterium-specific antigen recognized by the ELISA is water soluble (Andrew Gregory, Department of Agriculture, Western Australia, personal communication). The use of an ampipathic quaternary ammonium bromide provides a simple, efficient, safe and relatively inexpensive method to rapidly enhance the purity of crude CT extracts. Toxic and flammable methanol has been replaced, in assays and extraction protocols for CTs, by non-flammable, aqueous solutions of cyclodextrin and cetyltrimetimethylammonium bromide, both registered for human use. The use of the commercially available and inexpensive cyclodextrins and/or quaternary ammonium phase transfer reagents could have value in the extraction and purification of other lipophilic or ampipathic natural toxins.