Rapid Quantitation of Anatoxins in Benthic Cyanobacterial Mats Using Direct Analysis in Real-Time-High-Resolution Tandem Mass Spectrometry

Toxic benthic cyanobacterial mats are increasingly 80 reported worldwide as being responsible for animal mortalities due to their production of the potent neurotoxin anatoxin-a (ATX) and its analogues. Improved analytical methods for anatoxins are needed to address public health and watershed management challenges arising from extremely high spatial and temporal variability within impacted systems. We present the development, validation, and application of a direct analysis in real-time-high-resolution tandem mass spectrometry (DART-HRMS/MS) method for analysis of anatoxins in cyanobacterial field samples, including a simplified sample preparation approach. The method showed excellent sensitivity and selectivity for ATX, homoanatoxina, and dihydroanatoxin-a. Isotopically labeled ATX was used as an internal standard for all three analogues and successfully corrected for the matrix effects observed (86 +/- 16% suppression). The limit of detection and recovery for ATX was estimated as 5 ng/g and 88%, respectively, using spiked samples. The total analysis time was similar to 2 min, and excellent agreement was observed with results from a liquid chromatography-HRMS reference method. Finally, the DART-HRMS/MS method was applied to a set of 45 Microcoleus-dominated benthic cyanobacterial mat samples from the Wolastoq near Fredericton, Canada, demonstrating its power and applicability in enabling broad-scale field studies of ATX distribution.

Automated summary

Toxic benthic cyanobacterial mats, which produce potent neurotoxins called anatoxin-a (ATX) and its analogues, have been increasingly reported worldwide as responsible for animal mortalities. This study presents a new method for analyzing anatoxins in cyanobacterial field samples using direct analysis in real-time-high-resolution tandem mass spectrometry (DART-HRMS/MS).