INTRODUCTION: Nuclear magnetic resonance (NMR) measurement of 1 JCC coupling by two-dimensional (2D) INADEQUATE (incredible natural abundance double quantum transfer experiment), which is a special case of double-quantum (DQ) spectroscopy that offers unambiguous determination of 13 C-13 C spin-spin connectivities through the DQ transitions of the spin system, is especially suited to solving structures rich in quaternary carbons and poor in hydrogen content (Crews rule). OBJECTIVE: To review published literature on the application of NMR methods to determine structure in the liquid-state, which specifically considers the interaction of a pair of carbon-13 (13 C) nuclei adjacent to one another, to allow direct tracing out of contiguous carbon connectivity using 2D INADEQUATE. METHODOLOGY: A comprehensive literature search was implemented with various databases: Web of Knowledge, PubMed and SciFinder, and other relevant published materials including published monographs. The keywords used, in various combinations, with INADEQUATE being present in all combinations, in the search were 2D NMR, 1 JCC coupling, natural product, structure elucidation, 13 C-13 C connectivity, cryoprobe and CASE (computer-assisted structure elucidation)/PANACEA (protons and nitrogen and carbon et alia). RESULTS: The 2D INADEQUATE continues to solve "intractable" problems in natural product chemistry, and using milligram quantities with cryoprobe techniques combined with CASE/PANACEA experiments can increase machine time efficiency. The 13 C-13 C-based structural elucidation by dissolution single-scan dynamic nuclear polarisation NMR can overcome disadvantages of 13 C insensitivity at natural abundance. Selected examples have demonstrated the trajectory of INADEQUATE spectroscopy from structural determination to clarification of metabolomics analysis and use of DFT (density functional theory) and coupling constants to clarify the connectivity, hybridisation and stereochemistry within natural products. CONCLUSIONS: Somewhat neglected over the years because of perceived lack of sensitivity, the 2D INADEQUATE NMR technique has re-emerged as a useful tool for solving natural products structures, which are rich in quaternary carbons and poor in hydrogen content.

Centre for Natural Products Discovery School of Pharmacy and Biomolecular Sciences Liverpool John Moores University Byrom Street Liverpool Merseyside L3 3AF UK

Laboratory of Growth Regulators Institute of Experimental Botany ASCR and Palacký University Olomouc Czech Republic

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