Batrachotoxins

Biosynthetically, little is known of the origin of these alkaloids. In a preliminary study, radioactive acetate and mevalonate were incorporated into skin steroids of frogs, while incorporation of these potential precursors into skin alkaloids was not detected (750). Neither radioactive cholesterol nor serine was significantly incorporated into the batrachotoxins. Three species, Phyllobates aurotaenia, Dendrobates pumilio and Dendrobates auratus were used in these exploratory biosynthetic studies. 

The present review summarizes current knowledge of the properties, occurrence, synthesis and biological activity of the dendrobatid alkaloids. 

Briefly, the most satisfactory isolation procedure was as follows 1) extraction of minced skins twice with 70% aqueous methanol, followed by evaporation in vacuo to yield a dry tan residue 2) trituration with 0.9% NaCl, adjustment of pH to 2 with HCl, followed by extraction of lipids into chloroform 3) adjustment of pH to 8.5 with aqueous NH3, followed by extraction of toxic principles into chloroform 4) after evaporation to dryness in vacuo the active principles were purified by thin-layer chromatography on silica gel with chloroform-methanol (6 1). Two iodoplatinate-positive alkaloid spots were detected the higher Rf material accounted for virtually all of the toxicity. In view of subsequent studies this higher Rf 

Chemical properties of the batrachotoxin fraction were assessed on a microscale. In retrospect it appears that most reactions resulted in loss of the pyrrole entity and hence a product which did not afford a positive Ehrlich reaction. These chemical reactions included catalytic hydrogenation with palladium on charcoal, reduction with lithium aluminium hydride, treatment with acidic methanol, oxidation with manganese dioxide, treatment with acid, reaction with 2,4-dinitrophenylhydrazine, and exhaustive methylation with methyl iodide. An Ehrlich-positive methiodide could be obtained under milder conditions with methyl iodide. Acetylation of the batrachotoxin fraction with acetic anhydride and pyridine afforded two Ehrlich-positive 0-acetyl derivatives. Reaction with methoxyamine afforded an Ehrlich-positive 0-methyloxime. Reduction with sodium borohydride afforded an Ehrlich-positive dihydro-derivative. This product apparently isomerizes to other dihydro-compounds (257). Autoxidation, a serious problem during isolation of batrachotoxin, led to Ehrlich-negative products. 

A knowledge of the structure of batrachotoxinin A led to a re-evaluation of the spectral properties of the much more toxic batrachotoxin and homobatrachotoxin. It soon became apparent that the ultraviolet absorption peaks (A.max 234, 264 nm), the infrared red absorption band at 1690 cm S and additional nuclear magnetic resonance absorption peaks in 

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Barrellene oxide flash thermolysis oxirenes in, 7, 125 Basic Violet 10 (C.I. 45170), 3, 879 Batrachotoxin, 4, 374 Baumgarten method cinnoline synthesis, 3, 44... 

Figure 3. The chemical structures of two classical alkaloid activators (batrachotoxin and veratridine) and of a recently characterized marine toxin [brevetoxin B (BvTX-B)], that acts at a different site on the Na channel.

Figure 11. Comparative potencies of several saxitoxins and tetrodotoxin in the mouse bioassay, displacement of radiolabelled saxitoxin from rabbit brain, and block of single batrachotoxin-treated rat sarcolemmal sodium channels incorporated into planar lipid bilayers. The horizontal axis is the log of the potency relative to saxitoxin. Compound numbering corresponds to that in Figure 1. Data from Ref. 10, 94, 95.

Figure 12. Records of current through single batrachotoxin-treated rat sarcolemmal sodium channels incorporated into planar lipid bilayers, before and after treatment with saxitoxin and derivatives. Data from Ref. 91.

We wish to express our appreciation to J. Daly, NIH, for generously supplying the batrachotoxin essential for the bilayer experiments to S. D. Darling, University of Akron, for his work on the crystal structures of C2 and C4 and to T. Chambers, FDA, for molecular graphics. 

The lipid-soluble toxins (veratridine, batrachotoxin, aconitine, grayanotoxins). These toxins cause persistent activation of Na channels, i.e., their permanent opening and hence membrane depolarization 56-58). 

A growing number of other diverse compounds have also been shown to bind to an allosteric site on the muscarinic receptors. Among them are pirenzepine (highly selective for Mi receptor), lidocaine and verapamil (ion channel blockers), tacrine (anticholinesterase compound), batrachotoxin, and strychnine (glycine receptor antagonist) [25,31-35],... 

Cohen-Armon M, Kloog Y, Henis YI, Sokolovsky M. Batrachotoxin changes the properties of the muscarinic receptor in rat brain and heart possible interaction(s) between muscarinic receptors and sodium channels. Proc Natl Acad Sci USA 1985 82 3524-3527. 

Holz, K.W. and Coyle, J.T., The effects of various salts, temperature and the alkaloids veratridine and batrachotoxin on the uptake of pH]-dopamine into synatosomes from rat striatum, Mol. Pharmacol., 10, 746, 1974. 

SAM, samandarines BTX, batrachotoxins HTX, histrionicotoxins PTX, pumiliotoxins aPTX, allopumiliotoxins hPTX, homopumiliotoxins DHQ, 2,5-disubstituted decahydroquinolines 3,5-P, 3,5-d [substituted pyrrolizidines 3,5-1 and 5,8-1, disubstituted indolizidines 1,4-Q, 1,4-disubstituted quinolizidines Epi, epibatidine Pseudophry, pseudophrynamines. With the exception of 3,5-P and 3,5-1, these alkaloids are not known to occur in arthropods (see text). Histrionicotoxins may occur in Minyobates and Mantella, but the evidence is not conclusive. 

FIGURE 1 Structures of lipophilic amphibian alkaloids. Alkaloids indicated by asterisks represent structural classes that have not been detected in nature except in amphibians and, in the case of batrachotoxins, in one species of bird (9). 

The batrachotoxins were the first class of unique alkaloids to be characterized from skin extracts of frogs of the family Dendrobatidae (see ref. 23 for a review of amphibian alkaloids). Batrachotoxin was detected in only five species of dendrobatid frogs and these frogs were then classified as the monophyletic genus Phyllobates, based in part on the presence of batrachotoxins (24). However, levels of batrachotoxins differ considerably, with the Colombian Phyllobates terribilis containing nearly 1 mg of batrachotoxins per frog, while the somewhat smaller Phyllobates bicolor and Phyllobates aurotaenia, also from the rain forests of the Pacific versant in Colombia, contain 10-fold lower skin levels (8). The two... 

Batrachotoxin South American frog Cardiovascular system... 

FIGURE 10.5 Batrachotoxin from dart-poison frogs. 

The active principle of dart frog poisons is alkaloids. The study of the den-drobatid poisons led to the discovery of over 200 new alkaloids, including batrachotoxins Pig. 10.5), pumiliotoxins, histrionicotoxins, gephyrotoxins, and decahydroquinolines (Daly et al., 1994). The most common compounds have the basic structure of piperidine and include histrionotoxin. In Phyllobates, the synthesis of other alkaloids is suppressed in favor of batrachotoxins. These are... 

Frogs of the genus Dendrobates are ant specialists. Of the more than 20 structural classes of lipophilic alkaloids found in the frogs, six occur in myrmicine ants. However, many dendrobatid alkaloids such as the batrachotoxins, histri-onicotoxins, and pumiliotoxins, have not yet been found in insects and other leaf-litter prey such as beetles and millipedes (Daly et al., 2000). The snake Lio-phis epinephelus feeds on Dendrobates and may further bioaccumulate alkaloids. 

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