Microcystins concentrations

Lehman PW, Boyer G, Satchwell M, Waller S (2008) The influence of environmental conditions on the seasonal variation of Microcystis cell density and microcystins concentration in San Erancisco Estuary. Hydrobiologia 600 187-204... 

Fig. 16.5. Colorimetric standard curves for the inhibition of PP2A from GTP Technology immobilised on different supports by (a) microcystin-LR (MC-LR) and (b) microcystin-RR (MC-RR). Inhibition is expressed as percentage of the control (no microcystin). Concentrations refer to those in the well. Reprinted from Campas et at. [86], with permission from Elsevier.

Vezie, C. et al.. Effect of nitrogen and phosphorus on growth of toxic and nontoxic Microcystis strains and on intracellular microcystin concentrations, Microb. Ecol, 43, 443, 2002. 

Kardinaal, W.E.A. and Visser, P.M., Dynamics of cyanohacterial toxins sources of variability in microcystin concentrations, in Harmful Cyanobacteria, Huisman, J. et al., Eds., Springer, Berlin, 2005, 41. 

In order to be able to detect very low concentrations of toxins in the water, volumes of about a liter of water can be concentrated employing disposable SPE cartridges. N-Octadecyl silica (CIS) is by far the most common sorbent for the extraction of neutral, mildly polar, or hydrophobic organic compounds, and is excellent for microcystins concentration. However, it does not allow for an efficient extraction of polar toxins such as anatoxin-a, saxitoxins, or even cylindrospermopsin. 

In the Slimmer of 1989, Rutland Water, the largest man-made lake in Western Europe and which supplies potable water to approximately 500 000 people in the East of England, contained a heavy bloom of Microcystis aeruginosa. By the end of the summer, a number of sheep and dogs had died after drinking from the bloom and concentrated scum. Analysis revealed that the cyanobacterial bloom material was toxic to laboratory mice, and that rumen contents from a poisoned sheep contained fivemicrocystin variants.Microcystins were detected in waters used for recreation in Australia at concentrations greater than 1 mg per... 

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." " ... 

It is obvious from the provisional risk assessment values for microcystins, and, being of the same order of magnitude of mammalian toxicity, similar values may be calculated for the cyanobacterial neurotoxins, that sensitive detection methods are required to detect these low concentrations of toxins. Of the biological methods of detection discussed earlier, the mouse and invertebrate bioassays are not sensitive enough without concentration of water samples, in that they are only able to detect mg of microcystins per litre. Only the immunoassays (ng-/rg 1 and the protein phosphatase inhibition assays (ng O... 

Significant concentrations of cyanotoxins have been found to accumulate in the tissues of macroinvertebrates such as mollusks and crustaceans, presenting an indirect route of exposure for invertebrates, fish, and aquatic mammals at higher trophic levels (Negri and Jones 1995). In natural systems, mortality among benthic invertebrate herbivores is probably low because most bloom-forming bacteria are planktonic and only periodically come into contact with the benthos. Nevertheless, Kotak et al. (1996) determined that enhanced mortality of snails at the end of a bloom cycle in Canadian lakes was due to consumption of Microcystis cells that had formed a scum on the surface of macrophytes. Oberemm et al. (1999) found that aqueous microcystins, saxitoxins, and anatoxin-a all resulted in developmental delays in fish and salamander embryos. Interestingly, more severe malformations and enhanced mortality were observed when larvae were exposed to crude cyanobacterial extracts than to pure toxins applied at natural concentrations (Oberemm et al. 1999). 

Cells of C. reinhardtii were exposed to cell-free filtrates from A. flos-aquae, pure microcystin-LR or anatoxin-a, or combinations of the toxins. Both the position of the cells and the chlorophyll-a concentration of the cultures were observed for 12 days. Exposure to crude extracts as well as to combinations of the toxins significantly decreased chlorophyll levels in the cultures. Furthermore, these compounds were all capable of paralyzing the algae and thus promoted the settlement of C. reinhardtii cells. One intriguing aspect of this dynamic interaction is the separate finding that C. reinhardtii may actually induce toxin synthesis in A. flos-aquae (Kearns and Hunter 2000), essentially signaling its own demise. 

Komarek J, Kling H (2003) Filamentous cyanobacteria. In Wehr JD, Sheath RG (eds) Freshwater algae of North America ecology and classification. Academic, San Diego, CA, pp 117-196 Kotak BG, Zurawell RW, Prepas EE, Holmes CFB (1996) Microcystin-LR concentration in aquatic food web compartments from lakes of varying trophic status. Can J Fish Aquatic Sci 53 1974-1985... 

An international intercomparison exercise in the determination of microcystin, carried out by using the most common methods (LC/DAD, ELISA and LC/MS) indicated that LC/DAD is affected by lower precision [234], while the coupling of the LC technique with ELISA permit the achievement of high sensitivity and specificity in the determination of microcystins and nodularin [235] without the need of pre-concentration the method meets the World Health Organization guidelines (1 pg L ). The combination of ELISA characterization and LC analysis with fluorescence, UV, and tandem MS detections, allowed the first identification of cylindrospermopsin, an algal toxin that caused the poisoning of up to 148 persons in Australia [236],... 

Furthermore, the structure of microcystin includes an electrophilic carbon atom, (Fig. 7.26), which is part of the Mdha amino acid. If microcystin is ingested from contaminated water, for example, it is taken up into the liver by an organic anion transporter (OAT) system and therefore is concentrated in the liver. The structure of the microcystins means they are able to associate with the enzymes protein phosphatases, such as PP-1, PP-2A, and PP-2B via hydrophobic and ionic interactions. 

In order to demonstrate the viability of the approach, protein phosphatase inhibition was first performed with the enzyme in solution and detected by colorimetric methods. Two microcystin variants, microcystin-LR and microcystin-RR, were used. Both enzymes were inhibited by these toxins, although to a different extent. The 50% inhibition coefficients (IC50) towards microcystin-LR were 0.50 and 1.40 pgL 1 (concentrations in the microtitre well) for the Upstate and the GTP enzymes, respectively. Hence, the Upstate enzyme was more sensitive. The IC50 towards microcystin-RR were 0.95 and 2.15 pgL-1 for the Upstate and the GTP enzymes, respectively. As expected, microcystin-LR was demonstrated to be a more potent inhibitor. 

Speed VAC concentrator (Organomation Associates Inc., USA) under nitrogen for solvent evaporation in the microcystin extraction process. 

Since at the beginning the microcystin content in the real samples is unknown, pure and 100-fold diluted solutions are analysed in order to obtain, at least for one of the two dilutions, inhibition values into the linear range. Concentration values (pg L-1) are then converted to toxin contents (pgg-1 dry weight). 

Saxitoxin has been labeled with fluorescamine, o-phthaldialdehyde (OPA) and dansyl chloride and detection limits as low as 0.1 attomole were reported for the OPA derivative of saxitoxin (26). Labeling, separation, and analysis of saxitoxin was best accomplished using fluorescamine, which produces ionic derivatives that can be separated from other fluorescently labeled marine toxins, such as tetrodotoxin and microcystin. However, the precolumn labeling methods required xM concentrations of analyte, limiting the utility of the technique for trace analysis. 

Toxins such as microcystin LR and associated substances can be very difficult to analyse at low concentrations in water. Therefore, it is preferable to control blue-green algae by preventing algal blooms in source waters. There are treatment options for microcystin LR and related substances, but these require careful assessment for example, it is particularly important to ensure that algal cells are removed. 

In Japan, the first detection of AN and its degradation product, epoxy-AN, have been reported (Park et al. 1993). This was also the first study to show that Microcystis could produce both AN and microcystins. The predominant species were Anabanea and Planktothrix with toxin concentrations in the range 0.4-16 pg AN/g. In addition, AN, HMAN, and a new compound, hydroxy-HMAN, were isolated from Raphidiopsis mediterranea in Japan (Namikoshi et al. 2003 Watanabe et al. 2003). AN was also found in four of 26 samples from Korean lakes, collected during 1992-1995 (Park etal. 1998). 

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