Isolation, purification, structural characterization, detection and microbial effects of a polyether toxic compound produced by the chain-forming dinoflagellate, Alexandrium monilatum

The chain-forming dinoflagellate Alexandrium monilatum has been reported to be associated with widespread discolored water and massive fish mortality in the Mississippi Sound and off the eastern and western coasts of Florida. Previous studies over the last 70 years have determined that A. monilatum produces a harmful, lipophilic substance(s) that is predominantly contained in the cell mass and is unlike that of saxitoxin-like toxins. After 70 years of research, no molecular structure of the toxic extract(s) has been characterized. Using sophisticated chemical, chromatographic, and analytical techniques, we have successfully purified and identified the molecular structure of the toxin produced by A. monilatum. Utilizing a 500 MHz NMR to carry out a number of experiments (i.e. 1H, 13C, COSY, HSQC, HMBC) and ESI-MS and MALDI-TOF-MS mass spectral techniques, we unambiguously determined the molecular structure of the toxic extract(s). This extract(s) was representative of a polyether macrolide with an empirical formula of C43H60O12. This toxic compound was shown to be identical to a Japanese tidepool toxin identified as Goniodomin A, which is produced by another Alexandrium species.;With structure and mass spectrometry data in hand, a detection mechanism using an LC-MS was developed to monitor toxin production in culture. Additionally, toxin production and algal growth was monitored in antibiotic-treated cultures. There was no change in algal growth in antibiotic-treated cultures, and slightly higher cellular toxin content in the cells of the untreated, control cultures. It appeared that microbial presence influences toxin production.;This is the first report of structural characterization and development of a LC-MS detection method for monitoring of the toxic extract produced by the red-tide associated dinoflagellate, A. monilatum. Additionally, we have reported toxin production in culture over the algal growth with and without antibiotic addition, as well as monitored microbial community changes in each treatment.