Microcystins, a class of hepatotoxins produced by a variety of cyanobacteria species, hold the potential to significantly impact the health of both animal and human populations utilizing surface waters for drinking water, daily living, and recreation worldwide. Cyanobacterial blooms are predicted to become more severe due to rising surface water temperatures and increasing eutrophication of water bodies. In such conditions cyanobacteria often predominate, often creating toxic blooms. Anoxic conditions caused by these blooms can also result in fish kills, the hepatotoxic characteristics of the toxin may cause mammalian mortalities, and falling debris from blooms can have profound impacts on the invertebrates in the sediment below. Analytical techniques must allow for accurate quantification of microcystins in biological and environmental specimens; yet, this task is challenging as microcystins can become covalently bound to proteins. Thus, methods solely based on solvent extraction may lead to false negative results as they are unable to liberate bound microcystin residues. Extraction methods must be modified to subsequently determine the concentrations of bound and free microcystins.<br/><br/>Here we describe an optimization approach for the detection of free and covalently-bound microcystins. The method utilizes initial sample cleanup with methyl tert-butyl ether extraction, which allows removal of the lipid layer from the sample to eliminate competition with the oxidizing agent. Microcystins are then oxidized via the Lemieux reaction causing cleavage of the Adda amino acid moiety and releasing the compound 3-methyl-2-methoxy-phenylbutric acid (MMPB). Resulting MMPB are subsequently recovered and further cleaned up with solid phase extraction and the final sample is quantified with gas chromatography mass spectrometry. The developed method is highly robust and will be further applied to evaluate bioaccumulation and biomagnification in sediment, clam, and water samples from the Sacramento-San Joaquin Delta. Microcystin quantification of this environment is crucial the Delta feeds water into one of the largest estuaries on the west coast of North America, supplying water to upwards of 25 million people and home to a number of threatened or endangered aquatic organisms. Results of microcystin analysis in environmental samples will be further discussed.

Optimization of free and protein bound microcystin detection in sediments and aquatic organisms

255th National Meeting (2018)

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