Dataset: Nitrifier C fixation release

DIC fixation was measured via the incorporation of [14C]-bicarbonate as previously described (Herndl et al. 2005) with modifications. [14C]-bicarbonate (specific activity 56 mCi mmol-1/2.072 x 109 Bq mmol-1, Perkin Elmer) was added to 5 mL of culture (between 6 and 65 µCi were added depending on the activity of the culture). For every culture condition, at least three replicate live samples and one formaldehyde-fixed blank (3% v/v) were incubated in temperature-controlled incubators in the dark. Parallel incubations without [14C]-tracer additions were used to determine cell abundance and nitrite concentration. Incubations were terminated by adding formaldehyde (3% v/v) to 5 mL of sample. After 30-60 min, every sample was individually filtered onto 25 mm, 0.2 µm pore size polycarbonate filters (Millipore) and rinsed with 0.5 mL of artificial seawater using a glass filtration set (Millipore). The individual filtrates (5.5 mL per sample) were collected and transferred to scintillation vials to determine the fraction of [14C]-dissolved organic carbon ([14C]-DOC). Excess [14C]-bicarbonate from the filters was removed by exposing them to fumes of concentrated HCl (37 %) for 24 h. The filters were transferred to scintillation vials and 10 mL of scintillation cocktail (Ultima Gold, Perkin Elmer) was added. The filtrates were acidified to pH ∼2 with HCl (25 %) as previously described (Marañón et al. 2004), and filtrates were kept for 24 h in open scintillation vials placed on an orbital shaker before 10 mL scintillation cocktail was added to each vial. Samples were shaken for ca. 30 sec and incubated in the dark for at least 24 hours prior to counting the disintegrations per minute (DPM) in a scintillation counter (Beckman Coulter LS6500) for 15 min. Total radioactivity measurements were performed to verify added [14C]-bicarbonate concentrations by pipetting 100µl of sample into scintillation vials containing 400µl beta-phenylethylamine (to prevent outgassing of 14CO2). Scintillation cocktail was added, vials were shaken for ca. 30 sec and immediately measured in the scintillation counter.  

A detailed description of materials and methods can be found in Bayer et al. 2022.