A simple, high-precision, high-sensitivity tracer assay for N2 fixation

Abstract

We describe a simple, precise, and sensitive experimental protocol for direct measurement of N fixation rising the conversion of 15N2 to organic N. Our protocol greatly reduces the limit of detection for N2 fixation by taking advantage of the high sensitivity of a modern, multiple-collector isotope ratio mass spectrometer. This instrument allowed measurement of N2 fixation by natural assemblages of plankton in incubations lasting several hours in the presence of relatively low-level (ca. 10 atom%) tracer additions of 15N2 to the ambient pool of N2. The sensitivity and precision of this tracer method are comparable to or better than those associated with the C2H2 reduction assay. Data obtained in a series of experiments in the Gotland Basin of the Baltic Sea showed excellent agreement between 15N2 tracer and C2H2 reduction measurements, with the largest discrepancies between the methods occurring at very low fixation rates. The ratio of C2H2 reduced to N2 fixed was 4.68 ± 0.11 (mean ± standard error, n = 39). In these experiments, the rate of C2H2 reduction was relatively insensitive to assay volume. Our results, the first for planktonic diazotroph populations of the Baltic, confirm the validity of the C2H2 reduction method as a quantitative measure of N2 fixation in this system. Our 15N2 protocols are comparable to standard C2H2 reduction procedures, which should promote use of direct 15N2 fixation measurements in other systems.