LC-MS and GC-MS analytical conditions have been developed to detect the cis- and trans-epimers (relative configuration of the carbon bearing the acetyl or propionyl group) of dihydroanatoxin-a and dihydrohomoanatoxin-a, in biological samples. These compounds epimerize under acidic conditions, yielding a major species that was tentatively assigned as the cis-epimer. Cylindrospermum stagnale PCC 7417 was definitively shown to produce dihydroanatoxin-a (1.2 mg/g dried cells). Oscillatoria sp. PCC 9107, Oscillatoria sp. PCC 6506, and C. stagnale PCC 7417, which produce anatoxin-a, homoanatoxin-a, and dihydroanatoxin-a, respectively, were cultivated in the presence of isotopically labeled proline, and the toxins were extracted. Interpretation of the GC-MS electron ionization mass spectra of these labeled anatoxins showed that they are all biosynthesized from proline and that the positions of the labels in these molecules are identical. These data and the fact that the ana cluster of genes is conserved in these cyanobacteria suggest that dihydroanatoxin-a is formed by the reduction of either anatoxin-a or its precursor in a specific step involving AnaK, an F420-dependent oxido-reductase whose gene is found in the ana gene cluster in C. stagnale PCC 7417. This is the first report of a cyanobacterium producing dihydroanatoxin-a, suggesting that other producers are present in the environment.

In the last decades, an apparent increase in the frequency of benthic cyanobacterial blooms has occurred in coral reefs and tropical lagoons, possibly in part because of global change and anthropogenic activities. In the frame of the survey of marine benthic cyanobacteria proliferating in the lagoon of Moorea Island (French Polynesia), 15 blooms were collected, mainly involving three species—Anabaena sp.1, Lyngbya majuscula and Hydrocoleum majus-B. Their chemical fingerprints, obtained through high performance liquid chromatography combined with UV detection and mass spectrometry (HPLC-UV-MS) analyses, revealed a high extent of species-specificity. The chemical profile of Anabaena sp.1 was characterized by three major cyclic lipopeptides of the laxaphycin family, whereas the one of L. majuscula was characterized by a complex mixture including tiahuramides, trungapeptins and serinol-derived malyngamides. Toxicity screening analyses conducted on these cyanobacterial samples using Artemia salina and mouse neuroblastoma cell-based (CBA-N2a) cytotoxic assays failed to show any toxicity to a degree that would merit risk assessment with regard to public health. However, the apparently increasing presence of blooms of Lyngbya, Hydrocoleum, Anabaena or other benthic cyanobacteria on coral reefs in French Polynesia encourages the implementation of ad hoc monitoring programs for the surveillance of their proliferation and potential assessment of associated hazards.