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Development of methods for Measuring Total Microcystins in Fish Tissue using the 2-methoxy-3-methyl-4-phenylbutyric acid (MMPB) procedure.
Citation:
Lazorchak, Jim, T. Sanan, D. Sundaravadivelu, J. Kickish, J. Jones, AND R. Venkatapathy. Development of methods for Measuring Total Microcystins in Fish Tissue using the 2-methoxy-3-methyl-4-phenylbutyric acid (MMPB) procedure. 2018 SETAC Europe, Rome, ITALY, May 13 - 17, 2018.
Impact/Purpose:
This presentation will present findings of our 4.01D research on development of methods to detect algal toxins in tissues. This is the first phase looking at extraction method efficiencies.
Description:
There are limited methods for the analyses of multiple algal toxins in aquatic food webs, phytoplankton, zooplankton, periphyton, macroinvertebrates, forage fish, bottom feeders and top carnivore fish. Algal toxins in freshwater systems do not necessarily occur as single contaminants; mixtures of toxins may be produced from Cyanobacteria, Prymnesium parvum (Prymnesins), and Euglena sanguinea, including microcystins, saxitoxins, cylindrospermopsin, anatoxin-a., prymnesins and euglenophycin. The objective of the first phase of this research was to spike existing fillet and whole fish homogenates with 3 congeners of microcystins (LR, LA and RR) individually and as mixtures, and to develop a method for their recovery and measurement using the MMPB derivatization method. The second phase of the project is to field-test this method on fish collected from water bodies experiencing algal blooms and compare results with individual congener measurements. Extraction methods and analytical methods being developed for this research will be a starting point for developing extraction procedures for plankton, periphyton, and macorinvertebrates. Ten and 100 mg of fish homogenates from fish containing 1, 4 and 14% lipids were spiked with 4 and 40 ng of each of the microcystin congeners, LR, LA and RR. Various extraction techniques and conditions were tested to optimize recovery and simplify the procedure. Overall toxin recoveries were found to range from 30 to 50%. The lipid content was found to not interfere with generation of MMPB; however, it did impact the workup/extraction procedure in ways which were accountable through the use of a surrogate standard. The MMPB technique can be reliably employed for microcystin quantification in fish tissue. Detections in non-spiked samples (10-20 ug/kg) are comparable to literature precedent. For tissue quantification the MMPB method provides considerable improvements over extraction of individual toxin congeners and is consistent even with very polar or hydrophobic MCs.
