NMR Spectroscopy in Complex Mixture Analysis and Structure Elucidation of Natural Products: Rethinking the Need for Separations

Qualitative and quantitative analysis of complex mixtures and structure elucidation is generally impeded by the intrinsic complexity of the NMR spectra and the extensive signal overlap. The conventional approach to characterizing individual metabolites from complex crude extracts of natural products relies on multistep separation workflows employing diverse liquid chromatographic approaches and/or hyphenated techniques, which combine online integration of NMR with separation methods and other forms of spectroscopy. In recent decades, considerable efforts have been devoted to NMR applications in crude extracts without previous separation and isolation of the individual analytes. We present herein a critical overview of several NMR applications using chemical shift ranges of common organic functional groups, which can provide significant resolution advantages under specific experimental conditions. Particular emphasis is placed on: (i) characteristic chemical shift regions of strongly deshielded phenol OH groups, aldehyde CHO groups, hydroperoxide C-O-O-H groups and olefinic protons in conjugated double bonds; (ii) the advantages of using 13C chemical shift ranges through 2D 1H-13C HSQC and HMBC experiments of strongly deshielded phenol OH groups, aldehyde CHO groups, hydroperoxide groups, conjugated double bonds, and deshielded aliphatic CH groups; (iii) selective 1D NMR-spin chromatography techniques (1D TOCSY, 1D NOE); (iv) multiple suppression of strong resonances for minor analyte identification and (v) band-selective excitation techniques for minor analyte identification and quantification. The complementary contributions of statistical heterospectroscopy and computational chemical shift prediction are also considered, together with a brief assessment of the NMR experimental parameters and performance characteristics.