On-line high-performance liquid chromatography-electrospray ionization mass spectrometry for the determination of...

On-line high-performance liquid chromatography-electrospray ionization mass spectrometry (HPLC-ESMS) has been successfully applied to the separation and identification of brevetoxins associated with "red tide" algae. Brevetoxins are toxic polyethers produced by the marine dinoflagellate Gymnodinium breve. They are responsible for fish kills, and they pose certain health risks to humans. The LC-MS method employs reversed-phase microbore HPLC on a C18 column with a mobile phase consisting of 85:15 methanol/water, a flow rate of 8 microL/min, and a postcolumn split ratio of 3:1 (UV absorbance detector/mass spectrometer). A brevetoxin culture sample was found to contain at least six components, including two well-separated peaks corresponding to the brevetoxins PbTx-2 and PbTx-1, as well as several unknown compounds, including one with a molecular mass of 899 Da (possibly an isomer of PbTx-9). The brevetoxin molecules exhibited a high tendency to bind to alkali cations in positive ion ESMS. For standard PbTx-9, PbTx-2, and PbTx-1 brevetoxins analyzed on our LC-MS system, the detection limits (employing mass spectrometer scans of 100 m/z units) were determined to be less than 600 fmol, 1 pmol, and 50 fmol, respectively (S/N = 3); the total analysis time was about 35 min.