Abstract
The increasing use of the stable isotope 15N-NO3− for the quantification of ecological processes requires analytical approaches able to distinguish between labelled and unlabeled N forms. We present a method coupling anoxic sediment slurries and membrane inlet mass spectrometry to quantify dissolved 15N-NO3− and 14N-NO3−. The approach is based on the microbial reduction of 14N-NO3− and 15N-NO3− mixed pool, the determination of the produced 29N2 and 30N2, and the calculation of the original 15N-NO3− and 14N-NO3− concentrations. The reduction is carried out in 12 mL exetainers containing 2 mL of sediment and 10 mL of water, under anoxia. To validate this approach, we prepared multiple standard solutions containing 15N-NO3− alone or in combinations with 14N-NO3−, with final concentrations varying from 0.5 to 3000 µM. We recovered nearly 90% of the initial 14N-NO3− or 15N-NO3−, over a wide range of concentrations and isotope ratios in the standards. We applied this method to a 15N-NO3− dilution experiment targeting the measurement of nitrification in sediments with and without the burrower Sparganophilus tamesis. The oligochaete did not stimulate nitrification, likely due to limited ventilation and unfavorable conditions for nitrifiers growth. The proposed method is reliable, fast, and could be applied to multiple ecological studies.
Highlights
Nitrogen (N) pollution and increasing eutrophication of aquatic environments have increased attention concerning the regulation of the N cycle by microbial and primary producer communities [1]
We propose a relatively simple, inexpensive and quick method to measure a wide range of concentrations of N-NO3 − and N-NO3 − combining anoxic sediment slurries and MIMS analysis
As the assumptions for normality were met, two-way analysis of variance (ANOVA) was employed to test for differences between the rates measured in the two treatments and between the methods utilized for the calculations
Summary
Nitrogen (N) pollution and increasing eutrophication of aquatic environments have increased attention concerning the regulation of the N cycle by microbial and primary producer communities [1]. 15 N-NO3 − determination, in combination with 15 N-NH4 + analysis, may allow the precise quantification of relevant processes of the N cycle as the dissimilative reduction of nitrate to ammonium (DNRA) or the nitrification rate [8,9,10]. The latter are much less studied processes as compared to denitrification [6]. We present here results from multiple laboratory tests aiming at the precise calibration of the method and of the specific performances of the sediment we employed (e.g., denitrification capacity, co-occurrence of anammox) under different experimental conditions (e.g., slurry concentrations, incubation time, variable isotopic ratios). Such production leads to decreasing 15 N-NO3 − to 14 N-NO3 − ratios in the water column, which are proportional to nitrification rates and can be accurately quantified
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