Topic 13.2 Measurement of Nitrogen Fixation

Topic 13.2 Measurement of Nitrogen Fixation

Besides N2, the nitrogenase enzyme can reduce other substrates such as cyanide, azide and acetylene. Direct measurement of N2 fixation (reduction) requires a mass spectrometer, instrumentation that is not readily available, but the reduction of acetylene to ethylene can easily be measured by gas chromatography (Dilworth 1966). Acetylene reduction involves two electrons, C2H2 + 2e + 2 H+ → C2H4, as opposed to the eight electrons required for the reduction of N2 and 2 H+:

Therefore, reduction of four ethylene molecules correspond to the reduction of one N2 molecule. The acetylene method has limitations, however: Acetylene blocks gas diffusion into nodules, and rhizosphere microorganisms produce ethylene. For these reasons, the acetylene method is suitable for comparative studies (Dennison et al. 1992), while mass spectroscopy is used for direct measurements of N2 reduction, which is required for precise quantification of nitrogen fixation.