Determination of FVIIa-sTF Inhibitors in Toxic Microcystis Cyanobacteria by LC-MS Technique

Mar Drugs. 2015 Dec 30;14(1):7. doi: 10.3390/md14010007.


The blood coagulation cascade involves the human coagulation factors thrombin and an activated factor VII (fVIIa). Thrombin and fVIIa are vitamin-K-dependent clotting factors associated with bleeding, bleeding complications and disorders. Thrombin and fVIIa cause excessive bleeding when treated with vitamin-K antagonists. In this research, we explored different strains of toxic Microcystis aeruginosa and cyanobacteria blooms for the probable fVIIa-soluble Tissue Factor (fVIIa-sTF) inhibitors. The algal cells were subjected to acidification, and reverse phase (ODS) chromatography-solid phase extraction eluted by water to 100% MeOH with 20%-MeOH increments except for M. aeruginosa NIES-89, from the National Institute for Environmental Studies (NIES), which was eluted with 5%-MeOH increments as an isolation procedure to separate aeruginosins 89A and B from co-eluting microcystins. The 40%-80% MeOH fractions of the cyanobacterial extract are active against fVIIa-sTF. The fVIIa-sTF active fractions from cultured cyanobacteria and cyanobacteria blooms were subjected to liquid chromatography-mass spectrometry (LC-MS). The 60% MeOH fraction of M. aeruginosa K139 exhibited an m/z 603 [M + H]⁺ attributed to aeruginosin K139, and the 40% MeOH fraction of M. aeruginosa NIES-89 displayed ions with m/z 617 [M - SO3 + H]⁺ and m/z [M + H]⁺ 717, which attributed to aeruginosin 89. Aeruginosins 102A/B and 298A/B were also observed from other toxic strains of M. aeruginosa with positive fVIIa-sTF inhibitory activity. The active fractions contained cyanobacterial peptides of the aeruginosin class as fVIIa-sTF inhibitors detected by LC-MS.

Keywords: aeruginosins; anticoagulant; blood coagulation cascade; cyanobacteria; fVIIa-sTF inhibitors; peptides; toxic Microcystis.

MeSH terms

  • Anticoagulants / pharmacology*
  • Chromatography, Liquid
  • Cyanobacteria / chemistry*
  • Factor VIIa / antagonists & inhibitors*
  • Factor VIIa / drug effects
  • Fresh Water
  • Humans
  • Japan
  • Mass Spectrometry
  • Structure-Activity Relationship
  • Thromboplastin / antagonists & inhibitors*
  • Thromboplastin / drug effects


  • Anticoagulants
  • Thromboplastin
  • Factor VIIa