Abstract
The periodical occurrence of harmful algal blooms (HABs) in freshwater lakes requires the determination of potential cyanobacterial toxins, especially microcystins (MCs). On demand of an adequate risk assessment, the high diversity of these hepatotoxic cyclic heptapeptides implicates the need of an unambiguous detection of their specific structural variants. Therefore, LC-MS and LC-MS/MS methods are the approaches of choice for determination of MCs. In contrast, even tandem mass spectromic fragmentation patterns are not even sufficient in any kind of structural determination requirements, whereas NMR methods require very high amounts of MCs. In this study, we present a novel method for chromatographic separation of desmethylated microcystins (dm-MCs). Based on the isolation of the specific structural variants using semi-preparative HPLC, a method was developed for the structure elucidation of cyclic peptides with special appliance for the determination of dm-MCs via analysis of the specific amino acid composition after peptide hydrolysis followed by stereospecific detection of the amino acids and resulting keto acids. On the basis of this method it is demonstrated that dm-MC-RR with the structure [Dha(7)]MC-RR represented the major compound in the microcystin pattern of Microcystis aeruginosa bloom events in 2005 and 2006 in Lake Senftenberg, Germany.