Application of on-line capillary high-performance liquid chromatography-nuclear magnetic resonance spectrometry coupling for the analysis of vitamin A derivatives

The direct on-line coupling between capillary high-performance liquid chromatography (capillary HPLC) and proton high-field nuclear magnetic resonance (NMR) spectrometry has been used to derive structural information about constituents of a mixture of vitamin A derivatives. ¹H NMR spectra were recorded in the stopped-flow and continuous-flow mode within a 180 μm I.D. capillary column mounted in a micro probe on a 600 MHz NMR spectrometer. The resolution of the ¹H NMR spectra obtained in capillary HPLC-NMR coupling experiments is sufficient to determine coupling constants in the order of 1.5 Hz. The detection limit is in the lower nanogram range. A stopped-flow 2D-TOCSY experiment of a 1% solution of vitamin A acetate acquired within 4 h reveals that the acquisition of 2D NMR spectra is possible in the nanoliter detection scale without any loss of structural information.