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

Authors

  • Marc Staal,

    Corresponding author
    1. Department of Marine Microbiology, NIOO-Center for Marine and Coastal Ecology, PO Box 140, 4400 AC Yerseke, The Netherlands.
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  • Sacco te Lintel-Hekkert,

    1. Life Science Trace Gas Facility, Department of Molecular and Laser Physics, University of Nijmegen, Toernooiveld, 6525 ED Nijmegen, The Netherlands.
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  • Frans Harren,

    1. Life Science Trace Gas Facility, Department of Molecular and Laser Physics, University of Nijmegen, Toernooiveld, 6525 ED Nijmegen, The Netherlands.
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  • Lucas Stal

    1. Department of Marine Microbiology, NIOO-Center for Marine and Coastal Ecology, PO Box 140, 4400 AC Yerseke, The Netherlands.
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*For correspondence. E-mail staal@cemo.nioo.knaw.nl; Tel. (+31) 113 577 479; Fax (+31) 113 573 616.

Abstract

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).

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