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Liquid State NMR for Metabolomics

Nuclear magnetic resonance (NMR) is a powerful analytical technique yielding quantitative, structural information. Liquid state NMR studies molecules dissolved in solution. NMR can study mixtures or pure compounds, returning spectra rich in chemical information. NMR experiments can be targeted or untargeted, and a high degree of automation lends the technique to high-throughput studies, where established statistical methods can pull out key features to discriminate between sample groups.

Nuclear magnetic resonance
EMSL's nuclear magnetic resonance supports the Cell Signaling and Communications, Rhizosphere Function, and Biomolecular Pathways Integrated Research Platforms.

NMR is routinely applied in metabolomics studies and provides highly complementary data to other prominent methods like mass spectrometry. EMSL routinely employs NMR in metabolomics studies of bacteria, fungi, and plants. Key features include:

  • High resolution and high sensitivity that provide one-dimensional and multidimensional data for biological samples. Commonly observed nuclei include the biologically relevant proton (1H), carbon (13C), nitrogen (15N), and phosphorus (31P). NMR spectroscopy can provide unequivocal identification of isomers and can facilitate identification of unknowns.
  • Chenomx software to provide extensive metabolite identification and reliable quantification by fitting one-dimensional proton NMR spectra using the calibrated Chenomx spectral database. Additional metabolite identifications via the Human Metabolome Database and custom metabolite standards additions.
  • Measure and trace carbon and nitrogen through metabolic pathways by incorporating feedstocks enriched with stable NMR active isotopes (e.g., 13C and 15N).
  • An EMSL-developed spectral database that enables simultaneous querying of multiple orthogonal data sources, including NMR and mass spectrometry, to expand and confirm metabolite identifications in the complex metabolite mixtures characteristic of environmental (eco-metabolomics) and other biological samples.
  • Automation for high sample throughput.

Research applications

Available instruments

  • 800 MHz Bruker Avance Neo (Tava), TCI H&F/C/N-D-05 Z XT, and QCI H-P/C/N-D-05 Z ET extended temperature range CryoProbes
  • 750 MHz Bruker Avance III (Bokan), TCI H/C-N-D-05 Z CryoProbe, QXI H-P-C-N-05, BBO, BBI
  • 750 MHz Varian VNMRS (Rainier): 5 mm HCN Z-gradient probe with variable temperature (VT) control of −20 to 80 °C and 5 mm Broad-Band Observe (31P to 15N) with VT control of −80 to +150 °C 
  • 600 MHz Agilent VNMRS (Hood): 5 mm HCN salt tolerant Z-gradient cold probe with VT control of 0 to 80 °C 
  • 600 MHz Agilent DD2 (Baker): 5 mm HCN Z-gradient cold probe with VT control of 0 to 50 °C 

Tips for success

  • Solution-state NMR is non-destructive and only requires small volumes of sample (200 μl for 3 mm tubes, 600 μl for 5 mm tubes). It is possible and routine to use a 3 mm sample tube in a 5 mm probe when samples are limited.
  • NMR is not as sensitive as mass spectrometry, so providing more sample will be advantageous.
  • NMR is sensitive to salt—higher salts reduce sensitivity, so limiting salts below 100 mM is ideal. Desalting methods (i.e., SPE) are an option but will bias the later-detected compounds.
  • CryoProbe or Cold Probe equipped instruments have higher sensitivity but are more affected by salt levels.
  • 1H NMR is the most sensitive nuclei, with approximate limits-of-detection (in practical instrument time on our best instruments) for small molecules around 1–10 μM. Our most sensitive instrument for 1H experiments is the 800 MHz (Tava).

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