Sample preservation methods for nitrous oxide concentration and isotope ratio measurements in aquatic environments

The nitrogen (N) and oxygen (O) stable isotope analysis of dissolved nitrous oxide (N 2 O) can provide important constraints on the sources and cycling of N 2 O in aquatic environments. The isotopic composition of aqueous N 2 O, both in field (natural abundance) or experimental ( 15 N‐labeling) samples, however, may be altered by abiotic reactions involving nitrite () or hydroxylamine (NH 2 OH) and microbial activity during sample storage, if samples are not adequately preserved. Here we tested five different preservatives, mercuric chloride (HgCl 2 ), copper sulfate (CuSO 4 ), zinc chloride (ZnCl 2 ), hydrochloric acid (HCl) mixed with sulfamic acid (SFA), and sodium hydroxide (NaOH), for fixing natural water samples from an estuary and a lake with different concentrations over a range of different storage times for N 2 O analyses. ZnCl 2 and CuSO 4 decreased the pH, and led to abiotic N 2 O production from , shifting the N 2 O isotopic composition significantly. Removal of with a mixture of SFA and HCl did not always prevent the alteration of the original N and O isotope composition of N 2 O, confirming the requirement for complete removal, and underscoring the biasing effects of at very low pH, even at trace levels. At low background, HgCl 2 preservation led to robust and accurate results. However, in light of its toxicity and bioaccumulation potential in food webs, this fixative should be avoided. The addition of NaOH reduced abiotic side reactions involving , and yielded the most reliable and reproducible results for N 2 O concentration and isotope ratio measurements across tested settings and storage times.