Hyperpolarized NMR Metabolomics at Natural ¹³C Abundance

Abstract

Metabolomics plays a pivotal role in systems biology, and NMR is a central tool with high precision and exceptional resolution of chemical information. Most NMR metabolomic studies are based on 1 H 1D spectroscopy, severely limited by peak overlap. 13 C NMR benefits from a larger signal dispersion but is barely used in metabolomics due to ca. 6000-fold lower sensitivity. We introduce a new approach, based on hyperpolarized 13 C NMR at natural abundance, that circumvents this limitation. A new untargeted NMR-based metabolomic workflow based on dissolution dynamic nuclear polarization (d-DNP) for the first time enabled hyperpolarized natural abundance 13 C metabolomics. Statistical analysis of resulting hyperpolarized 13 C data distinguishes two groups of plant (tomato) extracts and highlights biomarkers, in full agreement with previous results on the same biological model. We also optimize parameters of the semiautomated d-DNP system suitable for high-throughput studies.

Reply on RC1

“…A volume of 600 L of the hyperpolarized liquid was directly injected into an NMR tube that was treated with strongly oxidizing rinsing solutions (Helmanex III) beforehand to reduce the likelihood of gas inclusions forming upon injection into the NMR tube and disrupting NMR detection. (Dey, Charrier et al 2020) In the liquid state, 1 H single pulse acquisitions were repeated at one-second intervals, using a flip angle of 10 degrees. The spectral width was 30 ppm at a carrier frequency centered at 5 ppm.…”

A novel sample handling system for dissolution dynamic nuclear polarization experiments

“…A volume of 600 µL of the hyperpolarized liquid was directly injected into an NMR tube that was treated with strongly oxidizing rinsing solutions (Helmanex III) beforehand to reduce the likelihood of gas inclusions forming upon injection into the NMR tube and disrupting NMR detection (Dey et al, 2020) In the liquid state, 1 H single pulse acquisitions were repeated at 1 s intervals, using a flip angle of 10 • . The spectral width was 30 ppm at a carrier frequency centered at 5 ppm.…”

“…Dissolution dynamic nuclear polarization (DDNP) (Ardenkjaer-Larsen et al, 2003;Kovtunov et al, 2018;Jannin et al, 2019) is a method used for hyperpolarizing nuclear spins at cryogenic temperatures (Abragam and Goldman, 1978) close to 1 K -typically attained in a liquid-helium-cooled cryostat at low pressures -coupled to a subsequent temperature jump and detection at ambient conditions in a conventional liquid-state nuclear magnetic resonance (NMR) spectrometer. Spin hyperpolarization is herein understood as a strong increase in the population difference between the populations of two eigenstates in a magnetic field B 0 .…”

Trendbericht: Kernmagnetische Resonanz

“…33) Von Interesse dürfte die Entwicklung von 13 C-basierten Metabolomics mit hoher, Dissolution-DNP-gesteigerter Empfindlichkeit sein (siehe dazu Abschnitt Hyperpolarisation). 34) Weitere Entwicklungen mit paramagnetischen Gruppen sind etwa die ultraempfindliche aminosäurespezifische Detektion durch Gadolinium-Markierungen, 35) die 300-fache Empfindlichkeitssteigerung bei der Detektion der Iminoprotonen in RNA und DNA durch hyperpolarisiertes Wasser 36) sowie die Detektion von Lösungsmittelzugänglichkeit und damit vorübergehend existierender Strukturelemente in intrinsisch ungeordneten Proteinen (IDPs) durch Gadolinium-dotiertes Wasser. 37 41) Kürzlich ist es gelungen, die Struktur der transmembranen Domäne und einer Wirkstoffbindungsstelle des Sars-Cov-2-Virus-Hüllproteins E mit 2,1 Å Genauigkeit zu bestimmen.…”