Is a Molecule More Than the Sum of its Parts: Disassembling the 1 H NMR Spectra of Steviol Glycosides

From a simplistic perspective, molecules are collections of atoms bound to each other in a 3D space. Readily NMR-observable nuclei such as ¹H, ¹³C, and ¹⁵N yield characteristic resonance patterns that reflect these atom connectivities and spatial arrangements. In the narrow ¹H chemical shift window, complex resonance patterns due to frequent signal overlap pose a challenge to structure determination/verification by NMR. This raises the question of whether the NMR parameters of well-characterized molecular motifs can be fully or partially transferred to other molecules to facilitate structural analysis. In this report, we introduce a “Lego-like” approach to the complete 1D ¹H NMR analysis of complex, congeneric natural products, exemplified with eight steviol glycosides from Stevia rebaudiana. The strategy combines a detailed interpretation of 1D/2D NMR datasets with the development of digital NMR profiles using ¹H iterative Full Spin Analysis (HiFSA). The common aglycone of the eight glycosides, steviol, was used as starting point for this analysis. Characteristic NMR profiles of steviol and individual sugar units were used as building blocks to recreate the 1D ¹H NMR spectra of increasingly complex glycosylated derivatives. As a result, comprehensive NMR fingerprints of the eight steviol glycosides were generated. The potential impact of this methodology on important aspects of natural product research such as chemical dereplication, configurational analysis, and structural elucidation of new congeners will be discussed.