Expanding CyanoHAB Monitoring: New Micropeptins and Generalizable MS/MS Workflows for the Annotation of Cyanopeptide...

Area of Science:

• Environmental Science
• Marine Biology
• Toxicology

Background:

• Cyanobacterial harmful algal blooms (cyanoHABs) pose ecological and public health risks.
• Current monitoring primarily focuses on a limited number of regulated toxins like microcystins, cylindrospermopsins, anatoxins, and saxitoxins.
• The full spectrum of bioactive metabolites produced by cyanoHABs remains largely uncharacterized.

Purpose of the Study:

• To isolate and characterize novel bioactive metabolites from a cyanobacterial bloom.
• To develop and apply a new analytical pipeline for comprehensive cyanopeptide identification and annotation.
• To assess the ecological and toxicological relevance of identified cyanopeptides.

Main Methods:

• Isolation and structural elucidation of compounds using chromatography, mass spectrometry, and NMR spectroscopy.
• Analysis of LC-MS/MS data using the GNPS2 Analysis Hub for product ion searching and cyanopeptide class annotation.
• Biochemical assays to determine the bioactivity of isolated compounds, specifically neutrophil elastase inhibition.
• Analysis of field samples from Lake Erie blooms.

Main Results:

• Two new micropeptins (1 and 2) and ferintoic acid C (3) were isolated and structurally confirmed.
• A novel LC-MS/MS analysis pipeline enabled rapid annotation of cyanopeptide classes and modifications across multiple samples.
• Micropeptin 1 demonstrated moderate inhibition of neutrophil elastase.
• Field samples from Lake Erie blooms contained recurring micropeptins but lacked detectable microcystins.

Conclusions:

• Routine monitoring of cyanoHABs often overlooks a wide array of under characterized cyanopeptides.
• The developed LC-MS/MS analysis pipeline provides a practical approach for rapid, class-level annotation of cyanopeptides.
• Expanded monitoring of non-microcystin cyanopeptides is crucial for accurate assessment of cyanoHAB hazards.