Adaptation of micro-diffusion method for the analysis of 15N natural abundance of ammonium in samples with small volume

RATIONALE: There are several preparation methods for the measurement of the nitrogen (N) isotopic composition of ammonium (NH4+) in different types of samples (freshwater, saltwater and soil extracts). The diffusion method is the most popular and it involves NH4+ in solutions being released under alkaline conditions and then immediately trapped by an acidified filter. However, the traditional preparation is designed for samples with large volume and relatively high N concentrations. The performance of diffusion for small-volume samples (e.g., a few milliliters) remains unknown. METHODS: We examined the overall performance of micro-diffusion on 5 mL samples on varying the incubation time, temperature and initial NH4+ concentration. The trapped ammonia was chemically converted into nitrous oxide (N2O) with hypobromite and hydroxylamine in sequence. The produced N2O was analyzed by a commercially available purge and cryogenic trap system coupled to an isotope ratio mass spectrometer. RESULTS: We found that diffusion can be complete with no more than 7 days of treatment at 37 degrees C. Increasing the temperature to 50 degrees C and the incubation time to 11 days did not improve the overall performance. There were no significant differences in the overall performance during diffusion with NH4+ concentrations from 15 to 60 mu M. The blank size was relatively large, and the N contamination might come from the reagents especially KCl salts. CONCLUSIONS: The method presented here combines micro-diffusion and hypobromite oxidation and hydroxylamine reduction. It is suitable for samples with small volume and low NH4+ concentrations. Our study demonstrates that the NH4+ concentrations in samples can be as low as 15 mu M, and a volume of 5 mL is sufficient for this method. We suggest that this method can be used for the routine determination of N-15/N-14 for either natural abundance or N-15-enriched NH4+. Copyright (C) 2015 John Wiley & Sons, Ltd.