Isotope Dilution Analysis of Combined Nitrogen in Natural Waters. III. Nitrate and nitrite

Abstract

The measurement of nitrate uptake is of major importance to our investigations of primary production and nitrogen regeneration in the photic zone. Endogenous nitrate production (nitrification), by diluting the enrichment of added 15N-nitrate, can lead to underestimation of nitrate uptake. Although nitrification is assumed to be minimal in the photic zone, this needs to be confirmed by experiment. We have developed a specific isotope dilution method based on the analysis of a volatile derivative of the azo dye, 1-phenylazo-2-naphthol (Sudan-1). Sea water is spiked to approximately 10% of the nitrate pool with a known quantity of 15N-nitrate. An aliquot is filtered immediately. A second aliquot is incubated in situ for a fixed period and is then filtered. Nitrate concentration is estimated by isotope dilution analysis of the first filtrate. Nitrification rate is estimated from the change in 15N enrichment between the first and second filtrate. Nitrate from a 10 mL aliquot is first reduced to nitrite. Sudan-1 is formed by diazotization with aniline and coupling to 2-naphthol. It is extracted by solid phase extraction using C-18 cartridges, with ethyl acetate as eluant. 4-Phenylazophenol is added as internal standard and the samples are dried. They are analysed as tert-butyldimethylsilyl derivatives, using gas chromatography/mass spectrometry in selected ion recording mode. The intense M-57 fragment ion formed under electron impact ionization is used for 15N analysis. The procedure is precise, sensitive and, unlike most previous methods, it is free from interference by other forms of nitrogen. This method compliments our existing isotope dilution methods for the analysis of ammonium and dissolved free amino acids. Here, the new method has been used to confirm that our estimates of nitrate uptake in the photic zone of the Southern Ocean are not compromised by high nitrification rates. © 1998 John Wiley & Sons, Ltd.