Practical considerations for improved reliability and precision during determination of δ15 N values in amino acids using a single combined oxidation-reduction reactor.

Abstract

There has been increased interest in the measurement of δ15 N values in amino acids (AAs) to gain simultaneous insight into both trophic relationships and the composition of biogeochemical sources used by producers at the base of the food web. A new combustion reactor design in gas chromatography/combustion isotope ratio mass spectrometry (GC/C-irMS) equipment has brought to light variable outcomes in performance, highlighting the need for better information about best practices for new systems. Precision for δ15 N values in amino acids using the single combined oxidation-reduction reactor is improved across a sequence of analyses if the reactor is oxidized for a substantial period (2 h) and subsequently maintained throughout the sequence with 12-17 s seed oxidation before each run during GC/C-irMS. A five-point calibration curve using amino acids with a range of δ15 N values from -2.4‰ to +61.5‰ was used in combination with a 13-15 amino acid mixture to consistently normalize measurements to internationally calibrated reference materials. Combining this oxidation method with normalization techniques using both internal and external standards provided a reliable throughput of ~25 samples per week. It allowed for a reproducible level of precision of <±0.5‰, n = 10 within a derivatized standard mixture across each sequence and an average sample precision of ±0.27‰ n = 3, which is lower than the analytical precision typically associated with δ15 N values for amino acid analysis (<±1‰). A few practical considerations regarding oxidation and conditioning of the combustion reactor allow for increased sequence capacity with the single combined oxidation-reduction reactor. These considerations combined with normalization techniques result in a higher throughput and reduced analytical error during the measurement of δ15 N values in amino acids.