Nitrogenase activity in cyanobacteria measured by the acetylene reduction assay: a comparison between batch incubation and on-line monitoring

A new on-line method for measuring acetylene reduction is described. It con sists of a gas-flow cell connected to an electronic gas-mixing system and a n automatic sample loop in the gas chromatograph. Alternatively, ethylene c an be determined by using laser-based trace gas detection. The laser-based trace gas detection technique achieves a detection limit that is three orde rs of magnitude better than gas chromatography. We have applied the on-line method to the measurement of nitrogen fixation in a culture of the heteroc ystous cyanobacterium Nodularia spumigena and compared it with conventional batch-type incubations. Incubation of N. spumigena in the gas-flow cell re sulted in very short response times with a steady-state flux of ethylene ob tained within 2 min. Nitrogenase was shown to respond immediately to change s in light and oxygen. Monitoring of nitrogenase activity could be continue d for several hours without having a negative impact on nitrogen fixation r ates in N. spumigena. This was not the case in batch incubations, in which changes in nitrogenase activities were recorded during incubations, probabl y as a result of varying oxygen concentrations. It was therefore concluded that the on-line method is superior to batch incubations when rates of nitr ogenase activity are to be measured. The method is suitable for natural sam ples (water or sediment).