Simultaneous de Novo Identification of Molecules in Chemical Mixtures by Doubly Indirect Covariance NMR Spectroscopy

The detailed characterization of complex molecular mixtures plays a key role in many areas of modern Chemistry. Here we report a novel NMR spectroscopic method that deconvolutes a complex mixture of organic molecules simultaneously into individual components and depicts their chemical structure without requiring physical separation of the components. Doubly indirect covariance spectroscopy is introduced and applied to 2D C-13-H-1 HSQC and 2D H-1-H-1 COSY spectra, which results in a C-13-C-13 2D spectrum with unprecedented high resolution. This reconstituted spectrum is indeed a carbon-connectivity map that can be directly analyzed with basic graph theory to obtain the skeletal structures of individual mixture components or their fragments. The method is demonstrated for a model mixture and a natural product mixture extracted from cancer cells. Its suitability for automation makes this approach attractive for the analysis of a broad range of mixtures of natural or synthetic products.