A Bacterial Method for the Nitrogen Isotopic Analysis of Nitrate in Seawater and Freshwater

TLDR

The study introduces a new natural‑abundance nitrate isotopic analysis method for seawater and freshwater, detailing the protocol and nitrogen isotope results in the first of two companion papers. The method employs denitrifying bacteria lacking N2O‑reductase to convert nitrate to nitrous oxide, enabling measurement of both nitrogen and oxygen isotopic compositions. The technique achieves <0.2‰ precision at nitrate levels as low as 1 µM, accurately reproduces nitrogen isotope differences among standards and samples, and maintains a blank below 10 % of the signal for concentrations ≥1 µM.

Abstract

We report a new method for measurement of the isotopic composition of nitrate (NO3-) at the natural-abundance level in both seawater and freshwater. The method is based on the isotopic analysis of nitrous oxide (N2O) generated from nitrate by denitrifying bacteria that lack N2O-reductase activity. The isotopic composition of both nitrogen and oxygen from nitrate are accessible in this way. In this first of two companion manuscripts, we describe the basic protocol and results for the nitrogen isotopes. The precision of the method is better than 0.2‰ (1 SD) at concentrations of nitrate down to 1 μM, and the nitrogen isotopic differences among various standards and samples are accurately reproduced. For samples with 1 μM nitrate or more, the blank of the method is less than 10% of the signal size, and various approaches may reduce it further.