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Gas chromatography/combustion/isotope ratio mass spectrometry (GC-C-IRMS) is a highly sensitive approach which allows the analysis of the 13C/12C and 15N/14N isotope composition of amino acids in the range of natural abundance or in slightly 13C- and 15N-enriched samples. However, the accuracy of measurements remains a permanent challenge. Here we show the effect of the presence of slightly 15N-enriched compounds in physiological samples on the accuracy and reproducibility of 15N-abundances of amino acids within or between analytical runs. We spiked several individual amino acids with the respective 15N-labelled isotopomer and measured the 15N/14N ratios of other amino acids in the same sample or in the following analytical runs. Intra- and inter-run memory effects can be observed in 15N/14N ratios of amino acids. Sample throughput is reduced when cleaning runs using standard mixtures are required to restore initial conditions after runs of samples with 15N-enriched analytes. Possible reasons for the observed phenomenon and its implications for work in the lower 15N-enrichment range (<0.5 APE) are discussed and include different aspects of gas chromatography, derivatisation, and hot catalytic metal surface effects. Results need to be interpreted with caution if complex physiological samples contain 15N-enriched amino acids beyond 500‰ δ15N (~0.18 APE). Copyright © 2011 John Wiley & Sons, Ltd.