High-precision measurement of phenylalanine and glutamic acid δ 15 N by coupling ion-exchange chromatography and purge-and-trap continuous-flow isotope ratio mass spectrometry

• Published in: Rapid communications in mass spectrometry 2021-07, Vol.35 (13), p.e9085
• Main Authors: Zhang, Lin, Lee, Wing-Man Charlotte, Kreider-Mueller, Ava, Kuhnel, Evelyn, Baca, Jesus, Ji, Chongxiao, Altabet, Mark
• Format: Article
• Language: English
• ISSN: 0951-4198; 1097-0231
• DOI: 10.1002/rcm.9085

Nitrogen isotopic compositions (δ N) of source and trophic amino acids (AAs) are crucial tracers of N sources and trophic enrichments in diverse fields including archeology, astrobiochemistry, ecology, oceanography, and paleo-sciences. The current analytical technique using gas chromatography-combustion-isotope ratio mass spectrometry (GC/C/IRMS) requires derivatization which is not compatible with some key AAs.. Another approach using the high-performance liquid chromatography-elemental analyzer-IRMS (HPLC/EA/IRMS) may experience co-elution issues with other compounds in certain types of samples and the highly sensitive Nano-EA/IRMS instrumentations are not widely available. We present a method for high-precision δ N measurements of amino acids (δ N-AA) optimized for canonical source AA-phenylalanine (Phe) and trophic AA-glutamic acid (Glu). This offline approach entails a purification and separation step via high-pressure ion-exchange chromatography (IC) with automated fraction collection, followed by sequential chemical conversion of AA to nitrite and then to nitrous oxide (N O), and final determination of δ N of the produced N O via purge-and-trap continuous-flow isotope ratio mass spectrometry (PT/CF/IRMS). The cross-plots of δ N of Glu and Phe standards (four different natural abundance levels) generated by this method and their accepted values have a linear regression slope of 1 and small intercepts demonstrating high accuracy. The precisions were 0.36~0.67‰ for Phe standards, and 0.27~0.35‰ for Glu standards. Our method and the GC/C/IRMS approach produced equivalent δ N values for two lab standards (McCarthy Lab AA mixture and cyanobacteria) within error. We further tested our method on a wide range of natural sample matrices and obtained reasonable results. Our method provides a reliable alternative to the current methods for δ N-AA measurement as IC or HPLC-based techniques that can collect underivatized AAs are widely available. Our chemical approach that converts AA to N O can be easily implemented into laboratories currently analyzing δ N of N O using PT/CF/IRMS. This method will help promote the use of δ N-AA in important studies of N cycling and trophic ecology in a wide range of research areas.