Cooperating scientist: Rene Diocares , Griffith University, Brisbane Qld, Australia
Dedication: This website is dedicated to
the memory of the late Mexican mangrove researcher
Dr. Gina Holguin who
participated in the development of this technique.
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A team from the group of Environmental Microbiology, the Northwestern Center for Biological Research, Mexico evaluates nitrogen fixation of mangroves in the field |
Why do we need this technique? Heavy nitrogen gas can be purchased in a pressurized cylinder,
or produced in the laboratory with specialized equipment. However
, both methods of obtaining the gas are expensive, and for some laboratories,
particularly those in developing countries, this represents an insurmountable
obstacle to their research. These methods of obtaining the gas also introduce
a complication, because a typical nitrogen fixation experiment requires samples
of gas at atmospheric pressure. The pressure must be reduced with a pressure
regulator in the case of gas in a pressurized cylinder, and traditional laboratory
methods produce heavy nitrogen at pressures well below atmospheric that are
not generally suitable for direct use in nitrogen fixation experiments. This is because
there is no easy way in which to equalize the gas to atmospheric pressure for
subsequent injection into a sample bottle.
Previously, a simple technique was developed in Australia by Wood and
Kennedy (2001) to produce 15N2 gas at a research laboratory involved in biological
nitrogen fixation research (not a specialized chemical production facility).
This technique, and the modification to it described here, are both limited by
the volume of available plastic syringes, but the procedure here does not require
the special apparatus described by Wood and Kennedy (2001). The technique
produces small quantities of heavy nitrogen at atmospheric pressure with
readily available analytical chemicals, and inexpensive laboratory supplies and
equipment.
Chemical principle The advantages of the technique are:
For additional reading
The heavy nitrogen non-radioactive isotope (15N2) is often used as a label for
nitrogen fixation experiments, in which it is desirable to monitor directly the
rate of nitrogen fixation and the destination of the resulting metabolites in an
organism or ecosystem. This is one of the most acceptable and reliable techniques
for this purpose, and it has been used for decades.
The technique produces heavy nitrogen through the reaction of 15N-labeled
ammonium sulfate [(15NH4)2SO4] with lithium hypobromite (LiOBr), and the
gas subsequently is sparged first with acidified potassium permanganate
(KMnO4) and then with acidified water to remove contaminating oxides that
otherwise would be taken up by the organisms and thus produce errors in
nitrogen fixation experiments.
This website contains technical specifications and construction of the assembly
and operational procedure, two technical diagrams, a PowerPoint presentation
of the procedure for rapidly producing the gas, and the original scientific report
(as PDF).