SEPARATION AND IDENTIFICATION OF ISOFLAVONES WITH ONLINE LIQUID-CHROMATOGRAPHY ELECTROCHEMISTRY-THERMOSPRAY MASS-SPECTROMETRY

The on-line high-performance liquid chromatography-electrochemistry-thermospray mass spectrometry (LCEC-LC/MS) for separation and identification of the phenolic isoflavones genistein, formononetin, and Biochanin A is described. The isoflavones are separated on a C-18 reversed phase column with methanol: 0.1 M ammonium acetate pH 5 (70:30 v/v) as eluent, which is compatible for both LCEC and LC/MS detection. Low ng detection is achieved with a conventional glassy carbon thin-layer electrochemical cell at 1.3 V vs. Ag/AgCl. Selected ion monitoring (SIM) mode LC/MS results in detection levels at the 20-50 ng range, and the compounds are positively identified. In the on-line detection, the electrochemical cell is connected to the thermospray and the column effluent flows from the working electrode into the heated thermospray capillary tube. The results show that on-line coupling between a thin-layer LCEC and a thermospray-LC/MS can be used to characterize analytical systems.