Structure elucidation of beta-carotene isomers by HPLC-NMR coupling using a C-30 bonded phase

The separation of cis/trans isomers of beta-carotene has been performed with a C-30 stationary phase employing H-1 NMR spectroscopy as an on-line detection technique. 1D as well as 2D NMR spectra have been recorded in the stopped-flow mode for the predominant chromatographic peaks. Structural assignment of the five identified isomers was performed via comparison of simulated 1D H-1 NMR spectra on the basis of the structures of beta-carotene cis/trans isomers with the experimental data, and also by the analysis of the proton-proton connectivities in the 2D NMR spectra of three isomers with the highest concentration. The chromatographic retention behaviour of the isomers agreed well with previously reported data, The advantage of the applied hyphenated coupling technique compared to conventional off-line techniques lies in the fact that chromatographic separation and NMR detection are performed in a closed system, so that reisomerization of the separated compounds is inhibited.