Intramolecular distribution of 13C/12C isotopes in amino acids of diverse origins

Abstract

Carbon stable isotope analysis can provide information about the origin and synthetic pathways that produce organic molecules, with applications in chemical, medical and (bio)geochemical sciences. The 13C/12C isotope ratios of organics such as amino acids are most commonly obtained as whole molecule averages. In this study, we apply proton nuclear magnetic resonance spectroscopy to conduct position-specific carbon isotope analyses of l-/d-alanine, l-threonine and l-histidine from different sources, in addition to molecule average stable isotope analyses obtained via mass spectrometry. Our results demonstrate that carbon isotope ratios can vary significantly between the individual carbon positions within an amino acid. For example, the β- and γ- carbons of l-threonine can differ in 13C/12C ratio by > 20 ‰. Comparisons of the position-specific and whole molecule average stable isotope abundances show that whole molecule analyses can mask the intramolecular isotope variation. These results provide the first experimentally measured position-specific isotope ratios for alpha and side chain carbons of alanine, threonine and histidine. Comparison with previous ab initio calculations of intramolecular equilibrium fractionation shows that the carbon isotope distributions are not at equilibrium, thus kinetic isotope effects play a significant role in amino acid synthesis. We hypothesize that position-specific 13C/12C isotope ratios provide an “isotopic fingerprint” that can give insight into the origin or synthesis pathway that formed an amino acid, and that this emerging analytical field will be a valuable addition to traditional stable isotope analysis.

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Abbreviations

(1H) NMR:

(Proton) Nuclear magnetic resonance spectroscopy

IRMS:

Isotope ratio mass spectrometry

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Acknowledgements

We thank Steven Sorey of the University of Texas at Austin NMR facility for his exceptional support and helpful advice during the NMR analyses and methods development. This research was supported by a Special Research Grant from the Office of the Vice President for Research at the University of Texas at Austin, a CNS Catalyst grant from the University of Texas at Austin, and a Petroleum Research Fund New Directions grant (PRF # 61080-ND2). This is UTIG contribution no. 3660. This is CPSH contribution no. 0007.

Funding

Petroleum Research Fund New Directions grant (PRF # 61080-ND2). College of Natural Sciences Catalyst grant, University of Texas at Austin.

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Correspondence to Cornelia Rasmussen.

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Rasmussen, C., Hoffman, D.W. Intramolecular distribution of 13C/12C isotopes in amino acids of diverse origins. Amino Acids 52, 955–964 (2020). https://doi.org/10.1007/s00726-020-02863-y

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Keywords

  • 1H NMR
  • Position specific
  • Isotope analysis
  • Amino acids