Anatoxins toxicity

Daphnia assay, the brine shrimps are exposed to different concentrations of toxicant, and the toxicity is expressed as the LCjo value. Extracts of cyanobacterial blooms and laboratory cultures, containing microcystins or anatoxin-a, have been found to be toxic towards brine shrimp," and fractionation of such extracts resulted in brine shrimp fatalities only with fractions containing microcystins." " ... 

The ability to identify and quantify cyanobacterial toxins in animal and human clinical material following (suspected) intoxications or illnesses associated with contact with toxic cyanobacteria is an increasing requirement. The recoveries of anatoxin-a from animal stomach material and of microcystins from sheep rumen contents are relatively straightforward. However, the recovery of microcystin from liver and tissue samples cannot be expected to be complete without the application of proteolytic digestion and extraction procedures. This is likely because microcystins bind covalently to a cysteine residue in protein phosphatase. Unless an effective procedure is applied for the extraction of covalently bound microcystins (and nodiilarins), then a negative result in analysis cannot be taken to indicate the absence of toxins in clinical specimens. Furthermore, any positive result may be an underestimate of the true amount of microcystin in the material and would only represent free toxin, not bound to the protein phosphatases. Optimized procedures for the extraction of bound microcystins and nodiilarins from organ and tissue samples are needed. 

Alteromonas species producing sodium channel blockers, 80r,82 Anabaena flos-aquae neurotoxins, 88 toxic principle, 108 Analyses, definition, 43 Anatoxin(s) isolation, 88 types, 88,91 Anatoxin a... 

Neurotoxins, such as saxitoxin and anatoxin-a, have been implicated in mediating competitive interactions between toxic cyanobacteria and other photoautotrophs, but few studies have explicitly examined the allelopathic effects of these compounds (e g., Kearns and Hunter 2001). Although it is reasonable to assume that these compounds bind to algal and cyanobacterial sodium channels in a similar fashion as in vertebrate neurons, support for this hypothesis is currently lacking. 

One role of cyanobacterial allelochemicals may be to alter the motility and distribution of competing photoautotrophs. In a recent study, Kearns and Hunter (2001) examined the effects of toxic metabolites from the filamentous cyanobacterium A. flos-aquae on a unicellular phytoplankton species, Chlamydomonas rein-hardtii. A. flos-aquae synthesizes both microcystins as well as anatoxins, providing the authors with an ecologically relevant opportunity to assess the individual and combinatorial effects of these toxins on an alga. 

Cyanobacteria toxins are toxins produced by certain species of blue-green algae that have become a major environmental and public health concern. The behavior of cyanotoxins during chlorination treatment has been recently reviewed by Merel et al. [129]. Chlorination DBFs have been reported only for the hepatotoxins microcystin-LR and cylindrospermopsin. Other cyanotoxins, such as nodularins, saxitoxins, and anatoxins, have yet to be investigated. Different isomers of six chlorination products of microcystin-LR have been characterized dihydroxy-microcystin, monochloro-microcystin, monochloro-hydroxy-microcystin, monochloro-dihydroxy-microcystin, dichloro-dihydroxy-microcystin, and trichloro-hydroxy-microcystin. Only two chlorination DBFs have been reported so far for cylindrospermopsin 5-chloro-cylindros-permopsin and cylindrospermopsic acid [129]. Chlorination of microcystin, cylindrospermopsin, and nodularins seems to reduce the mixture toxicity however, this aspect has not been extensively studied [129]. 

The main genera responsible for freshwater toxic blooms are Microcystis, Anabaena, Aphanizomenon and Oscillatoria. Toxins produced include 1. anatoxins, alkaloids and peptides of Anabaena 2. the peptide microcystin and related peptides of Microcystis 3. aphantoxins, compounds of Aphanizomenon with properties similar to some paralytic shellfish poisons. Properties of Oscillatoria toxin suggest they are peptides similar to those of Microcystis. Microcystis toxins are peptides (M.W. approx. 1200) which contain three invariant D-amino acids, alanine, erythro-3-methyl aspartic and glutamic acids, two variant L-amino acids, N-methyl dehydro alanine and a 3-amino acid. Individual toxic strains have one or more multiples of this peptide toxin. The one anatoxin characterized is a bicylic secondary amine called anatoxin-a (M.W. 165). The aphantoxin isolated in our laboratory contains two main toxic fractions. On TLC and HPLC the fractions have the same characteristics as saxitoxin and neosaxitoxin. 

Figure 5. HPLC profile of Anatoxin-a(s) toxic peak (far-left ...

Our research group is working on the development of electrochemical biosensors for the detection of microcystin and anatoxin-a(s), based on the inhibition of protein phosphatase and acetylcholinesterase, respectively. These enzyme biosensors represent useful bioanalytical tools, suitable to be used as screening techniques for the preliminary yes/no detection of the toxicity of a sample. Additionally, due to the versatility of the electrochemical approach, the strategy can be applied to the detection of other cyanobacterial toxins. 

Anatoxin-a is a toxic alkaloid occurring in the filamentous blue-green alga Anabaena flos-aquae and has been responsible for fatalities amongst livestock and wildlife. Mass and n.m.r. spectroscopy, and X-ray diffraction analysis of its N-acetyl derivative, pointed to structure and absolute stereochemistry (12) for ana-... 

A variety of other tropane alkaloids have been isolated of which the most important is anatoxin-A, a highly toxic nACh-R agonist and depolarizing neuromuscular blocking agent deriving from Anabaena cyanobacterium species that can contaminate inland waters. 

Anatoxin-a(s) inhibits acetylcholinesterase by acting as an irreversible active-site-directed inhibitor [61]. This prevents degradation of ACh and leads to over-stimulation of the muscle cells (Figure 6.1) [56,62]. Thus, although the mechanism of action of anatoxin-a(s) is quite different from that of anatoxin-a, the observed toxicity is similar. In addition, it was the first irreversible acetylcholinesterase inhibitor to be found in a cyanobacterium. 

The synthetic aspects of anatoxin-a (AN) and analogues are discussed in a separate chapter of this text, and the analytical methods for anatoxins are the subject of a separate review to be published elsewhere. A number of reviews have demonstrated the importance of understanding toxic cyanobacteria as potential environmental and health hazards as well as a resource of bioactive molecules (Harada 1999 Skulberg 2000 Briand et al. 2003). AN was one of the first cyanobacterial toxins to be chemically and functionally characterized and its high neurotoxicity has attracted extensive research activity. 

Table 8.1. Distribution of anatoxin-a and analogues and toxic incidents...

In Canada, between 1961 and 1975, suspected AN poisoning of cattle and dogs occurred in six locations. While Anabaena flos-aquae was found to be the predominant bloom in each case, other toxins were reported to be present in addition to AN, including anatoxin-a(s) (Gorham 1964 Carmichael et al. 1975 Carmichael and Gorham 1978 Juday et al. 1981). While the earliest confirmation that AN was implicated in animal poisonings was reported in Canada (Carmichael and Gorham 1978), the majority of locations where AN was detected, or toxic incidents have occurred, have been in Europe (Codd et al. 2005). 

Namikoshi, M., Murakami, T, Watanabe, M.E, Oda, T, Yamada, I, Tsujimura, S., Nagai, H., and Oishi, S. 2003. Simultaneous production of homoanatoxin-a, anatoxin-a, and a new non-toxic 4-hydroxyhomoanatoxin-a by the cyanobacterium Raphidiopsis mediterranea Skuja. Toxicon 42, 533-538. 

Rapala, J., Sivonen, K., Luukkainen, R., and Niemela, S. 1993. Anatoxin-a concentration mAnabaena andAphanizomenon under different environmental conditions and comparison of growth by toxic and non-toxic Anabaena-strains -a laboratory study. JAppl Phycol 5, 581. 

Smith, R.A., and Lewis, D. 1987. A rapid analysis of water for anatoxin-a, the imstable toxic alkaloid from Anabaena flos-aquae, the stable non-toxic alkaloids left after bioreduction and a related amine which may be nature s precursor to... 

An extensive work has been developed for the control of microcystins but not much effort has been made for the development of methods for the detection of other cyanobacterial toxins sneh as anatoxins or cylindrospermopsins despite their important acute toxicity as occurs in the case of the... 

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