Optimization of heteronuclear ultrafast 2D NMR for the study of complex mixtures hyperpolarized by dynamic nuclear polarization

Hyperpolarized 13C NMR at natural abundance, based on dissolution dynamic nuclear polarization (d-DNP), provides rich, sensitive and repeatable 13C NMR fingerprints of complex mixtures. However, the sensitivity enhancement is associated with challenges such as peak overlap and the difficulty to assign hyperpolarized 13C signals. Ultrafast (UF) 2D NMR spectroscopy makes it possible to record heteronuclear 2D maps of d-DNP hyperpolarized samples. Heteronuclear UF 2D NMR can provide correlation peaks that link quaternary carbons and protons through long-range scalar couplings. Here, we report the analytical assessment of an optimized UF long-range HETCOR pulse sequence, applied to the detection of metabolic mixtures at natural abundance and hyperpolarized by d-DNP, based on repeatability and sensitivity considerations. We show that metabolite-dependent limits of quantification in the range of 1-50 mM (in the sample before dissolution) can be achieved, with a repeatability close to 10% and a very good linearity. We provide a detailed comparison of such analytical performance in two different dissolution solvents, D2O and MeOD. The reported pulse sequence appears as an useful analytical tool to facilitate the assignment and integration of metabolite signals in hyperpolarized complex mixtures. Ultrafast 2D NMR spectroscopy combined with dissolution dynamic nuclear polarization provides rich, sensitive and repeatable NMR fingerprints of complex mixtures.

Automated summary

Hyperpolarized 13C NMR based on dissolution dynamic nuclear polarization technology can provide rich, sensitive and repeatable NMR fingerprints of complex mixtures at natural abundance. Ultrafast 2D NMR spectroscopy allows the recording of heteronuclear 2D maps of d-DNP hyperpolarized samples and provides correlation peaks between carbon and proton through long-range scalar couplings. The optimized UF long-range HETCOR pulse sequence shows good repeatability, linearity and limits of quantification for metabolic mixtures in two different dissolution solvents. It is a useful analytical tool for assigning and integrating metabolite signals in hyperpolarized complex mixtures.