American Journal of Climate Change, 2012, 1, 205-209 Published Online December 2012 (
Wet Soil Redox Chemistry as Affected by Organic
Matter and Nitrate
Duane T. Gardiner, Stephanie James
Texas A&M University-Kingsville, Kingsville, USA
Received September 12, 2012; revised October 13, 2012; accepted October 23, 2012
Wet soil microcosms were established to determine effects of organic matter and nitrate additions on microbial respira-
tion and redox potentials. Organic matter (1%) and nitrate (100 ppm and 200 ppm) treatments were applied in factorial
combination. Soil pH, redox potential, and CO2 emissions were measured. Data were analyzed by ANOVA for repeated
measures and separately by sampling day. Addition of organic matter significantly (P < 0.05) and consistently increased
CO2 emissions and decreased redox potentials. On Day 42 nitrate significantly (P < 0.05) increased redox values. This
study indicates a tendency for organic matter to decrease soil redox potential both in absolute terms and relative to the
suboxic-anoxic boundary. Our findings portend that additions of organic matter may quickly and markedly decrease soil
redox potentials and increase CO2 emissions in wetlands, whereas additions of nitrate may have complex and sporadic
effects on redox potentials.
Keywords: Oxidation; Reduction; Wetlands; Carbon Dioxide Emission
1. Introduction
Microbial respiration in the soil, accomplished by linking
oxidation and reduction half-reactions, directly impacts
the redox potential (Eh) of soil-water systems. Soil redox
potential affects, and is affected by, many attributes of
the ecosystem. In soil the primary electron donor for oxi-
dation half-reactions is organic matter. In aerobic soil the
prevailing electron acceptor for reduction half-reactions
is molecular oxygen (O2). Unlike higher organisms, mi-
crobial populations can use electron acceptors less sus-
ceptible to reduction than O2, especially in environments
where O2 is absent. Where microbes and organic matter
are abundant, the absence of O2 (i.e., an anaerobic condi-
tion) leads to the reduction of other substances, creating a
low redox potential as expressed in units of millivolts or
by the term pe implying the negative logarithm of con-
centration of electrons in search of an electron acceptor.
Electron acceptors are generally reduced in the fol-
lowing order as redox potentials decline: O2, 3
Mn4+, Fe3+, [1]. Oxic soils (also termed normal,
oxidized, and aerobic) are those soils with high redox
potentials indicating that O2 is present. Anoxic soils (also
termed reduced, and water-logged) are soils in which O2
has been reduced and is absent, as are sulfate
and other less preferable electron acceptors. Suboxic
soils (also termed wet but not water-logged) are those
having intermediate redox potentials in which O2 is ab-
sent but sulfate has not been completely reduced. A con-
cept used to delineate anoxic and suboxic soils is the
pe-pH boundary between oxidized and reduced sulfur, as
described by Baas Becking et al. [2] and modified by
Sposito [1] and Essington [3]. This boundary between
suboxic and anoxic conditions is described by the equa-
tion: pe = 9 pH.
is a particularly effective electron
accepter in the absence of O2, and can presumably miti-
gate the reduction of sulfate and chemicals less prefer-
able as electron acceptors, preventing very low redox
potentials. Whitmire and Hamilton [4] demonstrate an
increase in sulfate coinciding with denitrification, sug-
gesting that the presence of nitrate for an electron accep-
tor may alleviate sulfate reduction or encourage sulfur
oxidation. A decrease in wet soil redox potential has
been linked to the loss of nitrate [5]. The phenomenon of
denitrification occurs when nitrate, functioning as an
electron acceptor, is reduced to gaseous nitrogen. The
extent to which denitrification occurs depends upon
many environmental factors, including the quality of the
carbon in the system [6], the relation between organic N
mineralization and the presence of inorganic electron ac-
ceptors [7], and competing fates of nitrate [8]. The ef-
fects of nitrate on soil redox potential are intrinsically
complex. The presence of three soil factors, organic mat-
ter, O2, and nitrate, could strongly influence microbial
respiration and soil redox potential.
opyright © 2012 SciRes. AJCC
Wetlands, either natural or constructed, are used for
nitrogen removal from water catchments. The effective-
ness of wetlands for nitrate attenuation has been assessed
in lab [9], mesocosm [10], and field studies [11], with
differing results.
Also, wetlands are often constructed in locations where
they did not formerly exist to compensate for the loss of
wetlands elsewhere. As wetlands come into existence,
one might well ask whether or not wetland functions
have also come into existence, or has the process only
created lands that are wet [12]. Answers to such ques-
tions hinge on the chemistry, biology, and hydrology of a
site. Moreover, legally recognized diagnostic characteris-
tics for wetlands in the United States [13] include the
presence of hydric soils, i.e., soils with anaerobic condi-
tions. Yet some soils, natural or otherwise, can be inun-
dated for long periods of time while maintaining high
redox potentials, indicating a lack of anaerobic condi-
tions. Soil redox potential influences many processes of
practical importance, such as the degradation of pesti-
cides [14]. Therefore understanding the nature of both
constructed and natural wetlands requires an understand-
ing of the factors associated with aerobic and anaerobic
conditions as indicated by measurements of soil redox
The primary objective of this study was to determine if,
and to what extent, organic matter addition would in-
crease respiration in wet soils therefore consuming O2
and decreasing redox potentials. If redox potentials were
reduced by organic matter, the secondary objective
would be to determine whether or not nitrate would mi-
tigate those effects, either in an absolute sense or in rela-
tion to the wet-waterlogged (suboxic-anoxic) boundary.
2. Materials and Methods
Experimental units were 18 wet soil microcosms con-
sisting of 1-liter opaque bottles, each containing 1.0 kg of
soil collected from a seasonally wet site of the Aransas
National Wildlife Refuge in southern coastal Texas, USA.
The soil was excavated from the upper 30 cm of a prob-
able Mustang soil (Mixed, hyperthermic Typic Psamm-
aquents) mapped as the Galveston-Mustang Association
[15]. The soil contained 89.5% sand, 3.8% clay, 6.2% silt,
0.6% organic matter, 2.6 g kg–1 total nitrogen, and 31 mg
kg–1 nitrate.
Experimental treatments were applied in a 2 × 3 facto-
rial combination of 0 and 1% organic matter additions (0
and 10 g kg–1); and 0, 100, and 200 ppm nitrate additions
(0, 100, and 200 mg kg–1), replicated three times. The
organic matter added was dried Burmudagrass containing
1.33 mg kg–1 nitrogen. Nitrate was added as NaNO3. The
treatment combination that included neither organic mat-
ter nor nitrate constituted the control. Each microcosm
was randomly assigned to a treatment. Soils were main-
tained in a wet condition with a few mm of water cover-
ing the surface, and incubated in a growth chamber held
a 30˚C with 11 hours of simulated sunlight.
Electrode measurements of pH and redox potential
were taken at 14-day intervals for a period of 112 days.
Redox values were determined using a calomel electrode,
then adjusted to standard Eh values relative to a hydro-
gen electrode by adding 245 mV to the calomel-electrode
values [16]. Redox values relative to the sloping line
separating suboxic and anoxic soils were calculated by
determining the difference between the measured redox
value and the threshold redox value corresponding to the
soil pH, given that pe = 9 pH, and that Eh expressed in
millivolts = 59.16 pe.
Carbon dioxide emissions were determined by meas-
uring CO2 concentrations in the headspace above each
soil [17] on day 1, then again on days 8, 15, 22, 29, and
70. Tops of the containers were open during incubation,
but sealed for 10 min for CO2 sampling, at which time
100 mL samples were collected from the 415 mL head-
space using a Sensidyne gas detection pump (Sensidyne
Inc., Clearwater, FL) connected in series to an RAE gas
detection tube (RAE Systems Inc., Sunnyvale, CA).
Effects of organic matter and nitrate on dependent va-
riables CO2 and redox potential were analyzed by ANOVA
for a repeated measures design. Also, separate data ana-
lyses for each individual sampling day were conducted
by ANOVA for a factorial design [18].
3. Results
Tables 1 and 2 show P values indicating significance of
treatment effects. We considered any P value less than
0.05 to be significant. Results from the analysis of vari-
ance for repeated measures of the dependent variables
CO2, redox potential, and relative redox potential are
presented in Table 1. For CO2, organic matter main ef-
fects and the organic matter interaction with time were
significant. Nitrate did not significantly affect CO2 levels.
For redox values and relative redox values, interactions
between time and organic matter, and between nitrate
and time, were very highly significant (P < 0.001). Also
for both redox and relative redox values, the main effects
of organic matter were very highly significant (P < 0.001)
but main effects of nitrate were not significant (P > 0.05).
Because of the interactions with time (i.e., sampling day)
in a temporally repeated measures design, data were also
analyzed for each individual sampling day. Table 2 in-
dicates results of analysis of variance on redox values
and relative redox values performed for individual sam-
pling days.
Microbial respiration as measured by CO2 evolution
(Figure 1) increased significantly in response to addi-
tions of organic matter as measured on Days 8, 15, 22, and
Copyright © 2012 SciRes. AJCC
Table 1. P values for temporally repeated measures of car-
bon dioxide emissions (CO2), soil redox potential (redox),
and soil redox potential relative to the suboxic-anoxic boun-
dary (relative redox). Variables were nitrate (N), organic
matter (O) and time (T).
Redox Relative Redox
Source dfP valuedfP valuedfP value
N 20.281 20.143 2 0.094
O 1<0.0011<0.001 1 <0.001
N × O 20.374 20.384 2 0.355
T 50.011 8<0.001 8 <0.001
N × T 100.769 16<0.001 16<0.001
O × T 50.028 8<0.001 8 <0.001
N × O × T 100.839 160.103 160.120
Table 2. P values for effects of nitrate (N) and organic mat-
ter (O) on soil redox potential (redox) and soil redox poten-
tial relative to the suboxic-anoxic boundary (relative redox)
as determined by analysis of variance for a factorial design.
On no sampling day was the N × O interaction significant.
Redox Relative Redox
Sampling Day N O N O
1 0.940 0.063 0.543 0.046
14 0.098 <0.001 0.016 0.004
28 0.215 <0.001 0.393 <0.001
42 0.042 <0.001 0.010 0.001
56 0.093 <0.001 0.245 0.003
70 0.151 <0.001 0.226 <0.001
84 0.075 <0.001 0.076 <0.001
98 0.137 0.004 0.191 0.006
112 0.274 <0.001 0.216 <0.001
29. Because neither nitrate main effects nor nitrate × or-
ganic matter interactions were significant on any sam-
pling day, data from nitrate treatments were pooled such
that organic matter effects presented in Table 1 include
averages of all treatments with the organic matter addi-
tion vs all treatments without it.
Organic matter additions consistently produced a strong
negative effect on soil redox values, with highly signifi-
cant results observed on all days after Day 1 (Table 2;
Figure 2). Nitrate did not significantly affect soil redox
values except on Day 42, when redox values were higher
for the 100 ppm nitrate treatment than for the control. No
significant interactions between nitrate and organic mat-
ter were observed.
4. Discussion
The data strongly support the hypothesis that adding soil
organic matter will decrease soil redox potentials. Furth-
Figure 1. Carbon dioxide emitted from 1.0-kg wet soil mi-
crocosms. Values are means of all treatments either with or
without a 1% addition of organic matter. Vertical bars in-
dicate standard errors of the means.
Figure 2. Mean soil redox potentials as affected by organic
matter and nitrate. Treatments levels were 100 ppm nitrate
(N1), 200 ppm nitrate (N2), and 1% organic matter (OM).
ermore, the control treatment having native levels of or-
ganic matter and nitrate resulted in intermediate redox
potentials through Day 42, after which it produced the
highest redox values of all treatments. This further sup-
ports the hypothesis that the negative impact on redox
values was caused by the organic matter addition, not by
the native organic matter in the soil.
This study uniquely measured redox potential relative
to the pH-dependent boundary line separating suboxic
and anoxic soils (Figure 3). Effects of organic matter
were significant on every sampling day, whereas effects
of nitrate were significant on Days 14 and 42 only (Ta-
ble 2). Interactions between nitrate and organic matter
were not significant on any sampling day. Redox values
relative to the suboxic-anoxic boundary can be more
telling than absolute redox values because of their rela-
tion to actual events or reactions. In suboxic soil sulfate
and perhaps other species ranging between O2 and sulfate
in redox susceptibility remain in their oxidized state;
whereas in anoxic soil sulfate is absent, having been re-
duced to sulfide or other forms of reduced sulfur. As one
might expect, many data points in Figure 3 are in close
proximity to the boundary line, suggesting that for these
soils sulfate may be reducing but not completely reduced.
With the disappearance of sulfate, soil redox potentials
Copyright © 2012 SciRes. AJCC
Figure 3. Soil redox potential relative to the boundar y sepa-
rating suboxic and anoxic soils. The boundary is a function
of pH. Treatments levels were 100 ppm nitrate (N1), 200
ppm nitrate (N2), and 1% organic matter (OM).
can fall appreciably below the boundary line, as was ob-
served in most measurements here.
The findings raise the question of why redox potentials
relative to the suboxic-anoxic boundary were somewhat
more strongly influenced by nitrate than were absolute
redox values. Denitrification rates have long been known
to be inhibited by acidic soil; therefore denitrification
may have occurred to a lesser extent where pH values
were lower. However, we postulate that a second expla-
nation may also apply, i.e., that denitrification may have
produced the mildly alkaline reaction product bicarbon-
ate as suggested Equation (1) [19].
 
Figure 3 indicates that the combination of N plus or-
ganic matter generally tended to increase pH; and illus-
trates that as pH increases the boundary line slopes
downward. With all other factors remaining constant, a
treatment causing a slight elevation in pH would increase
redox value relative to the boundary line, and would tend
to shift equilibria between reduced and oxidized sulfur
toward the oxidized state.
Only for Days 14 and 42 did the results support the
hypothesis that nitrate would mitigate the impact of or-
ganic matter. Because denitrification is one of many pos-
sible fates of nitrate, this result was not unexpected.
Studies of N attenuation in wetlands reveals a wide range
of results, including 100% nitrate and 69% total N at-
tenuation [11], 13% to 73% nitrate attenuation, [9], and
51% total N attenuation [20]. Investigators [10,21] show
generally positive correlations between nitrate reduction
and soil organic matter. Matheson et al. [8] found that in
unplanted wetland microcosms 49% of nitrate nitrogen
was reduced to ammonium, 29% was denitrified, and
22% was immobilized. Adding to the complexities of
quantifying denitrification in wetlands is its tendency to
occur simultaneously or sequentially with nitrogen recy-
cling [22]. Whitmire and Hamilton [4] observed nitrate
depletion within 5 to 20 hours, leading to the speculation
for the present study that added nitrate might have al-
ready been reduced to another form when the first redox
reading was taken, perhaps to cycle back as nitrate later
in the study, such as on day 42 when nitrate treatment
effects were surprisingly significant.
In conclusion, this study of wet soil microcosms de-
scribed the effects of organic matter and nitrate on respi-
ration, redox potentials, and relative redox potentials (i.e.,
relative to the suboxic-anoxic boundary). Relative redox
potentials are of particular relevance because they pro-
vide an indication of the effect of a factor on electron
acceptors having lesser susceptibility to reduction than
O2 but greater than sulfate. We found that an organic
matter addition of 1% markedly decreased the redox po-
tential of wet soil both in the absolute sense and relative
to the suboxic-anoxic boundary. On two sampling days
nitrate raised redox potentials relative to the suboxic-
anoxic boundary. Our findings suggest that adding orga-
nic matter to a wetland may quickly and markedly de-
crease soil redox potential and increase CO2 emission.
Our findings offer little information from which a res-
ponse to nitrate could be predicted, but rather serve to
corroborate previous descriptions of the complex nature
of nitrate in wetlands.
5. Acknowledgements
We thank the USDA Natural Resources Conservation
Service for funding this project.
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