Compound-Specific and Intramolecular δ¹⁵N of Histidine in Marine Particles and Organisms: A New Tool for Investigating Nitrogen (N) Metabolism and Cycling

Histidine (HIS) exhibits minimal ¹⁵N fractionation during trophic transfer, potentially preserving the δ¹⁵N baseline of the food web. With its three N atoms incorporated into the HIS molecule and catabolized via distinct enzymatic pathways, site-specific δ¹⁵N values of HIS can provide insights into N metabolism and associated isotopic fluxes. While molecular average δ¹⁵N_HIS (δ¹⁵N_HIS-Total) is rarely reported and no intramolecular δ¹⁵N_HIS data for natural samples exist due to technical challenges with conventional analytical methods. We developed a new approach using ion-exchange chromatography (IC) to analyze δ¹⁵N of underivatized HIS. δ¹⁵N_HIS-Total is determined by converting all N to nitrate through UV-persulfate oxidation. Site-specific δ¹⁵N values of the α-N and sidechain N (δ¹⁵N_HIS-α and δ¹⁵N_HIS-s, respectively) are calculated by isotope mass balance. δ¹⁵N_HIS-Total matched corresponding EA/IRMS δ¹⁵N values within ±1‰ for in-house isotopic standards, and uncertainty ≤ ±0.6‰. In both commercial HIS standards and biological samples, including cyanobacteria, zooplankton, and fish, the α-N was found to be significantly enriched in ¹⁵N relative to the side-N (Δδ¹⁵N_α-s = +4 – 26‰), suggesting greater catabolic processing of α-N compared to side-N. This study establishes a new framework for combining chromatographic purification, chemical oxidation, and isotope analysis to obtain both molecular average and intramolecular δ¹⁵N values for poly-N amino acids, offering a valuable tool for investigating N metabolic pathways.

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Primary Presenter: Charlotte Wing Man Lee, Texas A&M University - Corpus Christi (wlee4@islander.tamucc.edu)

Authors:

Charlotte Wing Man Lee, Texas A&M University - Corpus Christi  (wlee4@islander.tamucc.edu)

Mark Altabet, University of Massachusetts Dartmouth  (maltabet@umassd.edu)

Jesus Baca, Texas A&M University - Corpus Christi  (jbaca2@islander.tamucc.edu)

Jason Barrera, Texas A&M University - Corpus Christi  (jbarrera35@islander.tamucc.edu)

Lin Zhang, Texas A&M University - Corpus Christi  (lin.zhang@tamucc.edu)