High-performance liquid chromatography on-line coupled to high-field NMR and mass spectrometry for structure elucidation of constituents of Hypericumperforatum L.

The on-line separation and structure elucidation of naphthodianthrones, fla vonoids, and other constituents of an extract from Hypericum perforatum L, using high performance liquid chromatography (HPLC) coupled on-line with ul traviolet-visible, nuclear magnetic resonance (NMR), and mass spectrometry (MS) is described. A conventional reversed-phase HPLC system using ammonium acetate as the buffer substance in the eluent tvas used, and proton NMR sp ectra were obtained on a 500 MHz NMR instrument. The MS and MS/MS analyses were performed using negative electrospray ionization, In the present study , all of the major known constituents in extracts from Hypericum perforatum L, were identified, and two new substances which had not previously been r eported as constituents of extracts of Hypericum perforatum L. were identif ied and their structures elucidated.