Elsevier

Toxicon

Volume 36, Issue 7, July 1998, Pages 953-962
Toxicon

Review Article
the toxicology of microcystins

https://doi.org/10.1016/S0041-0101(97)00102-5Get rights and content

Abstract

Microcystins are a family of more than 50 structurally similar hepatotoxins produced by species of freshwater cyanobacteria, primarily Microcystis aeruginosa. They are monocyclic heptapeptides, characterised by some invariant amino acids, including one of unusual structure which is essential for expression of toxicity. Microcystins are chemically stable, but suffer biodegradation in reservoir waters. The most common member of the family, microcystin-LR (L and R identifying the 2 variable amino acids, in this case leucine and arginine respectively) has an ld50 in mice and rats of 36–122 μg/kg by various routes, including aerosol inhalation. Although human illnesses attributed to microcystins include gastroenteritis and allergic/irritation reactions, the primary target of the toxin is the liver, where disruption of the cytoskeleton, consequent on inhibition of protein phosphatases 1 and 2A, causes massive hepatic haemorrhage. Microcystins are tight-binding inhibitors of these protein phosphatases, with inhibition constants in the nanomolar range or lower. Uptake of microcystins into the liver occurs via a carrier-mediated transport system, and several inhibitors of uptake can antagonise the toxic effects of microcystins. The most effective of these is the antibiotic rifampin (a drug approved for clinical use), which protects mice and rats against microcystin-induced lethality when given prophylactically and, in some cases, therapeutically.

Introduction

Microcystins (also known as cyanoginosins) are a family of toxins produced by species of freshwater cyanobacteria, primarily Microcystis aeruginosa, but also other Microcystis species and other genera, namely Anabaena, Oscillatoria and Nostoc. The microcystins are monocyclic heptapeptides composed of d-alanine at position 1, two variable l-amino acids at positions 2 and 4, γ-linked d-glutamic acid at position 6, and 3 unusual amino acids: β-linked d-erythro-β-methylaspartic acid (MeAsp) at position 3; (2S,3S,8S,9S)-3-amino-9-methoxy-2,6,8-trimethyl-10-phenyldeca-4,6-dienoic acid (Adda) at position 5; and N-methyl dehydroalanine (MDha) at position 7. There are over 50 different microcystins which differ primarily in the two l-amino acids at positions 2 and 4 and methylation/demethylation on MeAsp and MDha. The unusual amino acid Adda is essential for expression of biological activity, and a different stereochemistry about the conjugated double bond, for example, results in abolition of toxicity (An and Carmichael, 1994; Luukainen et al., 1994; Trogen et al., 1996). The most common microcystin is microcystin-LR, where the variable l-amino acids are leucine (L) and arginine (R). Its structure is shown below. Another cyanobacterial toxin, nodularin, from Nodularia spumigena, has a similar toxicity and structure to microcystins, but is a cyclic pentapeptide rather than a heptapeptide (An and Carmichael, 1994).

There is some dispute about the 3-dimensional structure of microcystin-LR, as deduced by NMR spectroscopy and mathematical calculations. Rudolph-Böhner et al. (1994) proposed a compact boat-like ring structure with the Adda chain bent into a position above the ring, whereas Trogen et al. (1996)deduced a saddle-shaped ring with Adda directed away from the cyclic backbone.

Section snippets

Analysis

Four sensitive methods or potential methods of analysis of microcystins have been published, based on reaction with a fluorescent probe, enzyme-linked immunosorbent assays (ELISAs), inhibition of protein phosphatase, and mass spectrometry, respectively.

Shimizu et al. (1995)synthesized a fluorogenic reagent to target conjugated dienes and called it DMEQ-TAD. This reagent reacted quantitatively with vitamin D metabolites and synthetic analogues, and the fluorescent products could be quantified

Stability

In laboratory experiments using low levels of microcystin-LR (10 μg/l) in reservoir water, Cousins et al. (1996)found that primary degradation of the toxin occurred in less than 1 week. It was stable over 27 days in deionised water, and over 12 days in sterilised reservoir water, indicating that the instability in normal reservoir water is due to biodegradation. The mechanism of inactivation is probably by modification of the Adda side chain (Cousins et al., 1996).

Purified microcystins are also

Toxicity

Poisoning of livestock from drinking water containing blue–green algae (cyanobacteria) was first observed at Lake Alexandrina in South Australia in 1878. In this case, the toxin was nodularin (Falconer, 1992). Since then there have been many reports of intoxications of birds, fish and other animals by cyanobacterial toxins, including microcystins. Growth of cyanobacterial blooms is encouraged by an increase of nutrients such as nitrates and phosphates in the water; these nutrients may be

Mechanism of action

It appears that microcystins mediate their toxicity by uptake into hepatocytes, via a carrier-mediated transport system, followed by inhibition of serine/threonine protein phosphatases 1 and 2A. The consequent protein phosphorylation imbalance causes disruption of the liver cytoskeleton, which leads to the massive hepatic haemorrhage which is the cause of death (Honkanen et al., 1990; Eriksson et al., 1990a; Falconer et al., 1981).

Protection against microcystin toxicity

Because of the rapid, irreversible and severe damage to the liver caused by microcystins, therapy is likely to have little or no value, and effective prophylaxis is critical.

The basis of the toxicity of the microcystins (inhibition of protein phosphatases) was not known until recently (1990; above). Prior to this, Adams et al. (1989)investigated chemicals known to affect macrophage function as potential prophylactic agents. This rationale was based on the known protective effect of

Future directions

Further research on microcystins could well focus on a characterisation of the transport mechanism of the toxin into hepatocytes (Runnegar et al., 1995a), and clarification of the specificity of microcystins as inhibitors of protein phosphatases (Honkanen et al., 1994). In searching for more effective medical countermeasures to the microcystins, primarily as prophylactic drugs, investigations into the role of glutathione as an antidote, and into potent inhibitors of the binding of microcystins

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