High-precision measurement of phenylalanine [delta]15N values for environmental samples: A new approach coupling high-pressure liquid chromatography purification and elemental analyzer isotope ratio mass spectrometry

• Published in: Rapid communications in mass spectrometry 2013-11, Vol.27 (21), p.2327
• Main Authors: Broek, Taylor A B, Walker, Brett D, Andreasen, Dyke H, McCarthy, Matthew D
• Format: Article
• Language: English
• ISSN: 0951-4198; 1097-0231
• DOI: 10.1002/rcm.6695

RATIONALE Compound-specific isotope analysis of individual amino acids (CSI-AA) is a powerful new tool for tracing nitrogen (N) source and transformation in biogeochemical cycles. Specifically, the [delta]15N value of phenylalanine ([delta]15NPhe) represents an increasingly used proxy for source [delta]15N signatures, with particular promise for paleoceanographic applications. However, current derivatization/gas chromatography methods require expensive and relatively uncommon instrumentation, and have relatively low precision, making many potential applications impractical. METHODS A new offline approach has been developed for high-precision [delta]15N measurements of amino acids ([delta]15NAA), optimized for [delta]15NPhe values. Amino acids (AAs) are first purified via high-pressure liquid chromatography (HPLC), using a mixed-phase column and automated fraction collection. The [delta]15N values are determined via offline elemental analyzer-isotope ratio mass spectrometry (EA-IRMS). RESULTS The combined HPLC/EA-IRMS method separated most protein AAs with sufficient resolution to obtain accurate [delta]15N values, despite significant intra-peak isotopic fractionation. For [delta]15NPhe values, the precision was ±0.16[per thousand] for standards, 4× better than gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS; ±0.64[per thousand]). We also compared a [delta]15NPhe paleo-record from a deep-sea bamboo coral from Monterey Bay, CA, USA, using our method versus GC/C/IRMS. The two methods produced equivalent [delta]15NPhe values within error; however, the [delta]15NPhe values from HPLC/EA-IRMS had approximately twice the precision of GC/C/IRMS (average stdev of 0.27[per thousand]±0.14[per thousand] vs 0.60[per thousand]±0.20[per thousand], respectively). CONCLUSIONS These results demonstrate that offline HPLC represents a viable alternative to traditional GC/C/IMRS for [delta]15NAA measurement. HPLC/EA-IRMS is more precise and widely available, and therefore useful in applications requiring increased precision for data interpretation (e.g. [delta]15N paleoproxies). Copyright © 2013 John Wiley & Sons, Ltd. [PUBLICATION ABSTRACT]