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Practical Aspects of NMR-Based Metabolomics

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Part of the Handbook of Experimental Pharmacology book series (HEP,volume 277)


While NMR-based metabolomics is only about 20 years old, NMR has been a key part of metabolic and metabolism studies for >40 years. Historically, metabolic researchers used NMR because of its high level of reproducibility, superb instrument stability, facile sample preparation protocols, inherently quantitative character, non-destructive nature, and amenability to automation. In this chapter, we provide a short history of NMR-based metabolomics. We then provide a detailed description of some of the practical aspects of performing NMR-based metabolomics studies including sample preparation, pulse sequence selection, and spectral acquisition and processing. The two different approaches to metabolomics data analysis, targeted vs. untargeted, are briefly outlined. We also describe several software packages to help users process NMR spectra obtained via these two different approaches. We then give several examples of useful or interesting applications of NMR-based metabolomics, ranging from applications to drug toxicology, to identifying inborn errors of metabolism to analyzing the contents of biofluids from dairy cattle. Throughout this chapter, we will highlight the strengths and limitations of NMR-based metabolomics. Additionally, we will conclude with descriptions of recent advances in NMR hardware, methodology, and software and speculate about where NMR-based metabolomics is going in the next 5–10 years.


  • Applications
  • Experimental methods
  • NMR spectroscopy
  • Targeted metabolomics
  • Untargeted metabolomics

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The authors wish to thank Genome Alberta (a division of Genome Canada), The Canadian Institutes of Health Research (CIHR), Western Economic Diversification (WED), and Alberta Innovates Health Solutions (AIHS) for financial support.


Baseline correction:

A spectral processing technique that yields a more pleasant looking NMR spectrum where signal-free regions appear as completely flat lines with zero intensity


Band-Selective Excitation Transient


A branch of information science that uses mathematical and statistical methods to identify patterns and extract information from large data sets collected on analytical instruments, such as UV, IR, and NMR spectrometers



CPMG experiment:

A pulse that can filter out the signals arising from large molecules, such as proteins or lipoproteins, from the spectrum (without the need for ultrafiltration or solvent extraction)


COrrelated SpectroscopY


4,4-dimethyl-4-silapentane-1-sulfonic acid, a chemical shift reference compound


Electronic REference To access In vivo Concentrations, an electronic reference signal


Heteronuclear Single Quantum Coherence Spectroscopy


Inborn error of metabolism. They are rare genetic disorders characterized by significant changes (several-fold increase or absence) in the concentration of specific metabolites that result from disturbances in normal metabolism


Incredible Natural Abundance DoublE QUAntum Transfer Experiment


The International Union of Pure and Applied Chemistry


The International Union of Biochemistry and Molecular Biology


A branch of analytical chemistry that comprehensively characterizes the molecules in various biofluids and tissues


The chemical constituents of the metabolome


The complete collection of all chemicals or metabolites found within cells, biofluids, organs, or organisms

Metnoesy experiment:

A simple 1D NOESY pulse sequence that provides solvent suppression before the experiment and during the mixing time without the use of gradients


Nuclear Overhauser Effect Spectroscopy


An NMR spectral adjustment process that is designed to maximize the absorptive character of NMR peaks over all regions of an NMR spectrum

Relaxation delay:

The sum of the acquisition time and acquisition delay prior to the next scan


Signal Amplification by Reversible Exchange in SHield Enables Alignment Transfer to Heteronuclei


Band-Selective Optimized Flip Angle Short Transient


Statistical total correlation spectroscopy


Longitudinal relaxation time


Transverse relaxation time

Targeted metabolomics:

A metabolomics technique that uses spectral deconvolution software to identify and quantify fluid-specific or targeted metabolites in individual spectra


Trimethylsilylpropanoic acid, a chemical shift reference compound

Untargeted metabolomics:

A metabolomics technique that uses spectral alignment, spectral binning, and multivariate statistical analysis to identify spectral features of interest

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Correspondence to David S. Wishart .

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Wishart, D.S., Rout, M., Lee, B.L., Berjanskii, M., LeVatte, M., Lipfert, M. (2022). Practical Aspects of NMR-Based Metabolomics. In: Ghini, V., Stringer, K.A., Luchinat, C. (eds) Metabolomics and Its Impact on Health and Diseases. Handbook of Experimental Pharmacology, vol 277. Springer, Cham.

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