Environmental Engineering Reference
In-Depth Information
We have included a third method, water column analysis for inorganic N, which
represents a very different approach. Nitrogen removal from the water column
can be calculated by comparing the inorganic N level in the input water from the N
level in the output water. This is an indirect approach; it does not directly assess
denitrification and there are other pathways for inorganic N removal (i.e., macro-
phyte assimilation). Furthermore, total N loss can only be determined if a hydro-
logic budget is created. However, relative differences in water quality services
capacity between wetlands can be characterized by this approach.
7.5.5.2 Measurement of Denitrification in Intact Soil Cores
using an Acetylene Block
This method is based on Bohlen and Gathumbi (
2007
), modified from Horwath et al.
(
1998
) and Mosier and Klemedtsson (
1994
). A number of 2-cm-diameter soil cores
are taken with a hammer corer fitted with cylindrical plastic inserts to the desired
depth, the number of cores dictated by the resources available and the expected
experimental variability. The plastic insert should be only partially filled, leaving
room for sampling headspace gasses. The bottom end of the plastic insert is immedi-
ately sealed with a rubber stopper, and the cores kept at 4
C until analyzed (
24 h).
The cores are allowed to equilibrate briefly at ambient or some standard temperature.
The top of the plastic cylinder is sealed with a stopper fitted with a rubber septum and
C
2
H
2
(See comments) is added to a concentration of ~10 kPa using a syringe. The
C
2
H
2
is distributed throughout the cylinder by repeated pumping of the headspace
gasses with a large (~60-ml) syringe. Samples of headspace gasses are taken at
intervals (e.g., after 2 and 6 h; see comments) and placed in evacuated vials fitted
with rubber septa. Gas samples, including blanks and standards, are analyzed by gas
chromatography using a Poropak Q (or equivalent) packed column in combination
with an electron capture detector. For examples of specific chromatographic run
conditions, refer to Hunt et al. (
2007
), Jordan et al. (
2007
), and Sirivedhin and Gray
(
2006
). The amount of N
2
O produced between selected sampling times is used to
estimate denitrification.
<
7.5.5.3 Denitrification Enzyme Activity (DEA)
The following method is taken from Jordan et al. (
2007
) as modified by Tiedje
et al. (
1989
). Aweighed amount of soil (~25 g) is placed in a 125-ml flask containing
25 ml of a solution of 1 mM glucose, 1 mM KNO3, and 1 g/L chloramphenicol.
The chloramphenicol is used to prevent
de novo
synthesis of denitrification
enzymes. The flask is flushed with N
2
or another inert gas such as Ar and closed
with a rubber stopper fitted with a gas-sampling septum. The flask is injected with
11 ml of C
2
H
2
and incubated for a standard period of time (e.g., 2 h; see comments)
at ambient or some standard temperature. Samples of headspace gasses are then
taken by syringe and transferred to evacuated vials fitted with rubber septa. Gas
samples, including blanks and standards, are analyzed as described for intact cores.
