NMR spectroscopy is a central method for the analysis of complex mixtures, and this is particularly true in metabolomics. However, such analysis mostly relies on 1D 1H spectroscopy, which provides an acceptable sensitivity but suffers from ubiquitous overlap between complex analyte patterns. While NMR offers a wide range of multi-nuclear and multi-dimensional techniques for analyzing complex samples, these tools are underutilized in metabolomics. In the past few years, we showed how fast 2D NMR methods could be systematically incorporated into metabolomics workflows, providing improved sample classification and/or biomarker identification.1–3 These methods mainly homonuclear 2D experiments for sensitivity reasons provide a first stage of dispersion improvement. 13C NMR spectroscopy would be even more advantageous as it provides narrow singlets spread over a broad spectral range. In fact, 13C NMR would be ideal for metabolomics, were it not for the fact that its low sensitivity is not compatible with the detection of low-concentrated analytes at natural abundance.
In this context, we recently showed that Dissolution Dynamic Nuclear Polarization (d-DNP) could provide a unique way to detect 13C NMR metabolomics spectral signatures with a sensitivity enhanced by several orders of magnitude.4 After a systematic optimization of a prototype d-DNP equipment, we showed that 13C NMR at natural abundance could be applied to plant extracts and incorporated in a full metabolomics workflow. More recently, we reported the first dDNP-enhanced 13C NMR analysis of a biofluid -urine- at natural abundance, offering unprecedented resolution and sensitivity for this challenging type of sample.5 We also showed that accurate quantitative information on multiple targeted metabolites could be retrieved through a standard addition procedure. Finally, we incorporated this approach into a first clinical metabolomics study.6 The analysis of urine samples from patients with different stages of chronic kidney disease (CKD) was performed using 13C d-DNP NMR and conventional 1H NMR metabolomics to explore the complementarity between the two methods. Results from 13C d-DNP NMR highlighted several biomarkers known to be biologically relevant, but also showed interesting complementarity with conventional 1H NMR, while raising exciting challenges associated with the analysis of spectral fingerprints stemming from this new approach.