EGU24-7124, updated on 08 Mar 2024
EGU General Assembly 2024
© Author(s) 2024. This work is distributed under
the Creative Commons Attribution 4.0 License.

Development of a comprehensive and ultrasensitive isotope calibration method for soil amino acid profiles using Orbitrap mass spectrometry

Tao Li1,2, Yuhua Li1,2, Ye Tian1,2, Erika Salas1,2, Xiaofei Liu1,2,3, and Wolfgang Wanek1
Tao Li et al.
  • 1Division of Terrestrial Ecosystem Research, Centre for Microbiology and Environmental Systems Science, University of Vienna, Vienna, Austria
  • 2Doctoral School in Microbiology and Environmental Science, University of Vienna, Vienna, Austria
  • 3State Key Laboratory for Subtropical Mountain Ecology of the Ministry of Science and Technology and Fujian Province, Fujian Normal University, Fuzhou, China

Bound amino acids constitute a significant portion of soil organic nitrogen, which represents an essential source of nitrogen for plant and microbial nutrition. The analysis of the content and isotope enrichment of bound amino acids still represents a significant challenge due to the degradation of certain amino acids following the conventional acid hydrolysis method, due to the low isotope enrichment levels reached under near-native soil conditions and due to the lack of isotopically labelled standards for some key amino acids. In this study, we used both a 13C-labeled and unlabeled 16 algal amino acid mixture to establish standard calibration curves for various amino acids, using the 6-Aminoquinolyl-N-hydroxysccinimidyl carbamate (AQC) derivatization method and the ultra-high-performance liquid chromatography with high-resolution Orbitrap mass spectrometry (UPLC-Orbitrap MS) platform. Molecular ions of AQC-derivatives for all amino acids were identified at the expected m/z values of the respective isotopologues, and the isotope calibration curves exhibited excellent linearity for those amino acids where we had isotope standards at hand (polynomial R2 > 0.9896). However, the polynomial fitting terms differed between single amino acids. Subsequently, we developed equations to relate the calibrated regression terms to physicochemical properties of the respective amino acids. First, we conducted a linear regression using the Orbitrap-derived 13C atom % of unlabeled standards against the carbon atom number of the specific amino acid-AQC derivative molecules, demonstrating great linearity (R2 = 0.9728). This linear regression curve allowed us to predict the natural 13C abundance of amino acids unavailable as isotopically labelled standards (e.g. hydroxyproline, meso-diaminopimelic acid). Consequently, based on further regressions, we could ultimately develop isotope calibration curves for those amino acids unavailable as 13C labelled standards based on the integrated isotope calibration functions. This general predictive model can be applied to comprehensively and highly sensitively (13C enrichment ~0.01 at %) quantify isotope enrichments of the whole soil amino acids profile, providing valuable insights for a better understanding of the overall fate of different amino acids in soils.

How to cite: Li, T., Li, Y., Tian, Y., Salas, E., Liu, X., and Wanek, W.: Development of a comprehensive and ultrasensitive isotope calibration method for soil amino acid profiles using Orbitrap mass spectrometry, EGU General Assembly 2024, Vienna, Austria, 14–19 Apr 2024, EGU24-7124,, 2024.