Skip to main content
OSTI.GOVU.S. Department of Energy Office of Scientific and Technical Information
U.S. Department of Energy
Office of Scientific and Technical Information
 

Position-specific carbon isotope analysis of serine by gas chromatography/Orbitrap mass spectrometry, and an application to plant metabolism

Journal Article · · Rapid Communications in Mass Spectrometry
DOI:https://doi.org/10.1002/rcm.9347· OSTI ID:1976395
 [1];  [2];  [2];  [2];  [3];  [4];  [2]
  1. California Institute of Technology (CalTech), Pasadena, CA (United States); OSTI
  2. California Institute of Technology (CalTech), Pasadena, CA (United States)
  3. University of Louisiana, Lafayette, LA (United States)
  4. University of Oslo (Norway)
+ Show Author Affiliations
Position-specific 13C/12C ratios within amino acids remain largely unexplored in environmental samples due to methodological limitations. We hypothesized that natural-abundance isotope patterns in serine may serve as a proxy for plant metabolic fluxes including photorespiration. Here we describe an Orbitrap method optimized for the position-specific carbon isotope analysis of serine to test our hypothesis and discuss the generalizability of this method to other amino acids. Position-specific carbon isotope ratios of serine were measured using a Thermo Scientific™ Q Exactive™ GC Orbitrap™. Amino acids were hydrolyzed from Arabidopsis biomass, purified from potential matrix interferences, and derivatized alongside standards. Derivatized serine (N,O-bis(trifluoroacetyl)methyl ester) was isolated using gas chromatography, trapped in a reservoir, and purged into the electron ionization source over tens of minutes, producing fragment ions containing different combinations of atoms from the serine-derivative molecule. The 13C/12C ratios of fragments with monoisotopic masses of 110.0217, 138.0166, and 165.0037 Da were monitored in the mass analyzer and used to calculate position-specific δ13C values relative to a working standard. This methodology constrains position-specific δ13C values for nanomole amounts of serine isolated from chemically complex mixtures. The δ13C values of fragment ions of serine were characterized with ≤1‰ precisions, leading to propagated standard errors of 0.7–5‰ for each carbon position. Position-specific δ13C values differed by up to ca 28 ± 5‰ between serine molecules hydrolyzed from plants grown under contrasting pCO2, selected to promote different fluxes through photosynthesis and photorespiration. The method was validated using pure serine standards characterized offline. Here this study presents the first Orbitrap-based measurements of natural-abundance, position-specific carbon isotope variation in an amino acid isolated from a biological matrix. We present a method for the precise characterization of isotope ratios in serine and propose applications probing metabolism in plants. We discuss the potential for extending these approaches to other amino acids, paving the way for novel applications.
Research Organization:
California Institute of Technology (CalTech), Pasadena, CA (United States)
Sponsoring Organization:
Agouron Institute; National Aeronautics and Space Administration (NASA); National Science Foundation (NSF); Research Council of Norway; Simons Foundation; USDOE; USDOE Office of Science (SC)
Grant/Contract Number:
SC0016561
OSTI ID:
1976395
Alternate ID(s):
OSTI ID: 1877313
Journal Information:
Rapid Communications in Mass Spectrometry, Journal Name: Rapid Communications in Mass Spectrometry Journal Issue: 18 Vol. 36; ISSN 0951-4198
Publisher:
WileyCopyright Statement
Country of Publication:
United States
Language:
English

References (57)

The new face of isotopic NMR at natural abundance: The new face of isotopic NMR at natural abundance journal December 2016
Effects of hydrolysis on the?13C values of individual amino acids derived from polypeptides and proteins journal January 2003
Natural intramolecular isotope measurements in physiology: elements of the case for an effort toward high-precision position-specific isotope analysis journal January 2001
Optimisation of derivatisation procedures for the determination ofδ13C values of amino acids by gas chromatography/combustion/isotope ratio mass spectrometry journal January 2007
Isolation and desalting with cation-exchange chromatography for compound-specific nitrogen isotope analysis of amino acids: application to biogeochemical samples: Compound-specific nitrogen isotope analysis of amino acids journal July 2010
Observation of ion coalescence in Orbitrap Fourier transform mass spectrometry: Ion coalescence in Orbitrap FTMS journal June 2012
Large-scale plant growth chamber design for elevated p CO 2 and δ 13 C studies : Growth chamber design for journal January 2015
Position‐specific 13 C/ 12 C analysis of amino acid carboxyl groups – automated flow‐injection analysis based on reaction with ninhydrin journal May 2018
Plant growth chamber design for subambient p CO 2 and δ 13 C studies journal June 2018
Sulfur isotope analysis of cysteine and methionine via preparatory liquid chromatography and elemental analyzer isotope ratio mass spectrometry journal January 2021
13C Gas Chromatography–Combustion Isotope Ratio Mass Spectrometry Analysis of N-Pivaloyl Amino Acid Esters of Tissue and Plasma Samples journal February 2000
Patterns of intramolecular carbon isotopic heterogeneity within amino acids of autotrophs and heterotrophs journal April 2004
Molecular biology of the plastidic phosphorylated serine biosynthetic pathway in Arabidopsis thaliana journal April 2001
Intramolecular distribution of 13C/12C isotopes in amino acids of diverse origins journal June 2020
Isotope effect in gas—liquid chromatography of labelled compounds journal December 1991
Gasometric Determination of Carboxyl Groups in free Amino Acids journal November 1941
Insights into the role of methionine synthase in the universal 13 C depletion in O - and N -methyl groups of natural products journal December 2017
Incorporating the effects of photorespiration into terrestrial paleoclimate reconstruction journal February 2018
The effect of atmospheric CO2 concentration on carbon isotope fractionation in C3 land plants journal November 2012
Carbon isotope evidence for the substrates and mechanisms of prebiotic synthesis in the early solar system journal January 2021
Differences in carbon isotope discrimination between angiosperm and gymnosperm woody plants, and their geological significance journal May 2021
Analysis of molecular isotopic structures at high precision and accuracy by Orbitrap mass spectrometry journal November 2017
Scanning the isotopic structure of molecules by tandem mass spectrometry journal November 2018
Using Orbitrap mass spectrometry to assess the isotopic compositions of individual compounds in mixtures journal November 2020
Exploring frontiers of orbitrap performance for long transients journal August 2021
Methods and limitations of stable isotope measurements via direct elution of chromatographic peaks using gas chromotography-Orbitrap mass spectrometry journal July 2022
Seasonal and within-plant gradients in the intramolecular carbon isotopic composition of amino acids of Spartina alterniflora journal September 2004
Ab initio calculation of the carbon isotope signatures of amino acids journal June 2009
Practical considerations for amino acid isotope analysis journal February 2022
Serine in plants: biosynthesis, metabolism, and functions journal September 2014
On-line high-precision carbon position-specific stable isotope analysis: A review journal February 2016
Electrostatic Axially Harmonic Orbital Trapping:  A High-Performance Technique of Mass Analysis journal March 2000
Simultaneous, High-Precision Measurements of δ 2 H and δ 13 C in Nanomole Quantities of Acetate Using Electrospray Ionization-Quadrupole-Orbitrap Mass Spectrometry journal December 2021
Stable Isotope Analysis of Intact Oxyanions Using Electrospray Quadrupole-Orbitrap Mass Spectrometry journal January 2020
Reaction Energetics and 13 C Fractionation of Alanine Transamination in the Aqueous and Gas Phases journal January 2020
Improved Uranium Isotope Ratio Analysis in Liquid Sampling–Atmospheric Pressure Glow Discharge/Orbitrap FTMS Coupling through the Use of an External Data Acquisition System journal April 2021
Derivatization of organic compounds prior to gas chromatographic–combustion–isotope ratio mass spectrometric analysis: identification of isotope fractionation processes journal January 1994
Viewpoint: How large is the carbon isotope fractionation of the photorespiratory enzyme glycine decarboxylase? journal January 2006
On the Relationship Between Carbon Isotope Discrimination and the Intercellular Carbon Dioxide Concentration in Leaves journal January 1982
Carbon Isotope Fractionation In Formation Of Amino Acids By Photosynthetic Organisms journal May 1961
Fourier Transform Mass Spectrometry journal July 2011
An Examination of the Carbon Isotope Effects Associated with Amino Acid Biosynthesis journal December 2006
Investigating the Regulation of One-carbon Metabolism in Arabidopsis thaliana journal March 2003
Carbon Isotope Fractionation during Photorespiration and Carboxylation in Senecio journal October 2008
Evidence for a Nonstatistical Carbon Isotope Distribution in Natural Glucose journal June 1991
Carbon isotope fractionation of amino acids in fish muscle reflects biosynthesis and isotopic routing from dietary protein: Carbon isotope fractionation of fish muscle amino acids journal July 2010
Estimating the rate of photorespiration in leaves journal May 1988
Current methods for estimating the rate of photorespiration in leaves journal November 2012
Photorespiration in the context of Rubisco biochemistry, CO 2 diffusion and metabolism journal January 2020
The Organic Isotopologue Frontier journal May 2021
Tracing Carbon Sources through Aquatic and Terrestrial Food Webs Using Amino Acid Stable Isotope Fingerprinting journal September 2013
Stable carbon isotope diagnostics of mammalian metabolism, a high-resolution isotomics approach using amino acid carboxyl groups journal October 2019
Stable isotope fingerprinting: a novel method for identifying plant, fungal, or bacterial origins of amino acids journal December 2009
Fractionation of Carbon and Hydrogen Isotopes in Biosynthetic Processes journal January 2001
Insights Into the Known 13C Depletion of Methane—Contribution of the Kinetic Isotope Effects on the Serine Hydroxymethyltransferase Reaction journal January 2022
The Glycerate and Phosphorylated Pathways of Serine Synthesis in Plants: The Branches of Plant Glycolysis Linking Carbon and Nitrogen Metabolism journal March 2018
A Guide to Using Compound-Specific Stable Isotope Analysis to Study the Fates of Molecules in Organisms and Ecosystems journal January 2019

Similar Records

Position-specific carbon isotopes of Murchison amino acids elucidate extraterrestrial abiotic organic synthesis networks
Journal Article · Tue Jun 13 20:00:00 EDT 2023 · Geochimica et Cosmochimica Acta · OSTI ID:2424906
Practical considerations for amino acid isotope analysis
Journal Article · Mon Dec 27 19:00:00 EST 2021 · Organic Geochemistry · OSTI ID:1977525

Related Subjects

59 BASIC BIOLOGICAL SCIENCES