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 consists of a gas-flow cell connected to an electronic gas-mixing system and an automatic sample loop in the gas chromatograph. Alternatively, ethylene can be determined by using laser-based trace gas detection. The laser-based trace gas detection technique achieves a detection limit that is three orders of magnitude better than gas chromatography. We have applied the on-line method to the measurement of nitrogen fixation in a culture of the heterocystous cyanobacterium Nodularia spumigena and compared it with conventional batch-type incubations. Incubation of N. spumigena in the gas-flow cell resulted in very short response times with a steady-state flux of ethylene obtained within 2 min. Nitrogenase was shown to respond immediately to changes in light and oxygen. Monitoring of nitrogenase activity could be continued for several hours without having a negative impact on nitrogen fixation rates in N. spumigena. This was not the case in batch incubations, in which changes in nitrogenase activities were recorded during incubations, probably as a result of varying oxygen concentrations. It was therefore concluded that the on-line method is superior to batch incubations when rates of nitrogenase activity are to be measured. The method is suitable for natural samples (water or sediment).
Staal
M
M
Department of Marine Microbiology, NIOO-Center for Marine and Coastal Ecology, PO Box 140, 4400 AC Yerseke, The Netherlands. staal@cemo.nioo.knaw.nl
Lintel-Hekkert
S T
ST
Harren
F
F
Stal
L
L
D000114:Acetylene / Q000378:metabolism*; D001395:Azotobacter / Q000201:enzymology; D018533:Biomass; D002734:Chlorophyll / Q000378:metabolism; D000077194:Chlorophyll A; D002849:Chromatography, Gas / Q000295:instrumentation / Q000379:methods*; D000458:Cyanobacteria / Q000201:enzymology* / Q000378:metabolism; D005030:Ethylenes / Q000378:metabolism; D007700:Kinetics; D008027:Light; D009586:Nitrogen Fixation; D009591:Nitrogenase / Q000378:metabolism*; D010084:Oxidation-Reduction; D010100:Oxygen / Q000378:metabolism; D013816:Thermodynamics
D005030:Ethylenes; D002734:Chlorophyll; C036216:ethylene; D009591:Nitrogenase; D000114:Acetylene; D010100:Oxygen; D000077194:Chlorophyll A