Selective extraction of microcystins and nodularins with immobilized polyclonal antibodies for LC-MS analysis.

Microcystins (MCs) and nodularins (NODs) are groups of toxins produced by cyanobacteria, and occur in surface and drinking water, recreational waters, dietary supplements and seafood. The presence of these toxins in drinking water and food may cause poisoning in humans and animals. More than 300 MCs and 10 NODs have been reported, and this large number of variants, combined with complex sample matrices, makes their analysis challenging for both LC-MS and ELISA. We developed immunoaffinity chromatography (IAC) columns using antibodies with broad specificity for MCs and NODs originally produced for ELISA analysis. The IAC columns were tested on a range of sample matrices including cyanobacterial blooms, dietary supplements, and cultures. Good recoveries were obtained, with remarkable clean-up of the samples, removing contaminants and thereby reducing matrix interferences in subsequent LC-MS analysis. This approach can improve analytical capacity in laboratories lacking the most expensive LC-MS equipment. It also opens up possibilities for simplifying omics studies of various types of samples containing the toxins and their metabolites, including identification of conjugated and metabolized forms of microcystins and nodularins. The IAC columns were reusable at least five times, with a capacity of ca. 500 ng MC-LR, with recoveries over 90% for tested MCs and NOD-R. Several conjugated derivatives of [D-Asp3]MC-RR from a Planktothrix prolifica culture were captured, concentrated and released, illustrating the potential of combining LC-HRMS and IAC to identify heavily modified or metabolized MCs.