Method for High Frequency Underway N₂ Fixation Measurements: Flow-Through Incubation Acetylene Reduction Assays by...

Because of the difficulty in resolving the large variability of N₂ fixation with current methods which rely on discrete sampling, the development of new methods for high-resolution measurements is highly desirable. We present a new method for high-frequency measurements of aquatic N₂ fixation by continuous flow-through incubations and spectral monitoring of the acetylene (C₂H_2, a substrate analog for N₂) reduction to ethylene (C₂H₄). In this method, named Flow-through Incubation Acetylene Reduction Assays by Cavity Ring-Down Laser Absorption Spectroscopy (FARACAS), dissolved C₂H₂ is continuously admixed with seawater upstream of a continuous-flow stirred-tank reactor (CFSR) in which C₂H₂ reduction takes place. Downstream of the flow-through incubator, the C₂H₄ gas is stripped using a bubble column contactor and circulated with a diaphragm pump into a wavelength-scanned cavity ring down laser absorption spectrometer (CRDS). Our method provides high-resolution and precise mapping of aquatic N₂ fixation, its diel cycle, and its response to environmental gradients, and can be adapted to measure other biological processes. The short-duration of the flow-through incubations without preconcentration of cells minimizes potential artifacts such as bottle containment effects while providing near real-time estimates for adaptive sampling. We expect that our new method will improve the characterization of the biogeography and kinetics of aquatic N₂ fixation rates.