Soil Biology & Biochemistry 35 (2003) 1289–1298

www.elsevier.com/locate/soilbio

Natural 15N abundances of inorganic nitrogen in soil treated with fertilizer

and compost under changing soil moisture regimes
Woo-Jung Choia, Hee-Myong Roa,*, Sang-Mo Leeb
a
Soil Science Laboratory, School of Agricultural Biotechnology, College of Agriculture and Life Sciences, Seoul National University,
Suwon 441-744, South Korea
b
National Instrumentation Center for Environmental Management, College of Agriculture and Life Sciences, Seoul National University,
Suwon 441-744, South Korea
Received 8 July 2002; received in revised form 26 February 2003; accepted 13 May 2003

Abstract
This study was conducted to examine whether the applications of N-inputs (compost and fertilizer) having different N isotopic
compositions (d 15N) produce isotopically different inorganic-N and to investigate the effect of soil moisture regimes on the temporal
variations in the d 15N of inorganic-N in soils. To do so, the temporal variations in the concentrations and the d 15N of NHþ 2
4 and NO3 in
soils treated with two levels (0 and 150 mg N kg21) of ammonium sulfate (d 15N ¼ 2 2.3‰) and compost (þ 13.9‰) during a 10-week
incubation were compared by changing soil moisture regime after 6 weeks either from saturated to unsaturated conditions or vice versa.
Another incubation study using 15N-labeled ammonium sulfate (3.05 15N atom%) was conducted to estimate the rates of nitrification
and denitrification with a numerical model FLUAZ. The d 15N values of NHþ 2
4 and NO3 were greatly affected by the availability of
substrate for each of the nitrification and denitrification processes and the soil moisture status that affects the relative predominance
between the two processes. Under saturated conditions for 6 weeks, the d 15N of NHþ 4 in soils treated with fertilizer progressively
increased from þ2.9‰ at 0.5 week to þ18.9‰ at 6 weeks due to nitrification. During the same period, NO2 3 concentrations were
consistently low and the corresponding d 15N increased from þ16.3 to þ39.2‰ through denitrification. Under subsequent water-
unsaturated conditions, the NO2 15 2
3 concentrations increased through nitrification, which resulted in the decrease in the d N of NO3 . In
15 þ
soils, which were unsaturated for the first 6-weeks incubation, the d N of NH4 increased sharply at 0.5 week due to fast nitrification.
On the other hand, the d 15N of NO2 3 showed the lowest value at 0.5 week due to incomplete nitrification, but after a subsequence
increase, they remained stable while nitrification and denitrification were negligible between 1 and 6 weeks. Changing to saturated
conditions after the initial 6-weeks incubation, however, increased the d 15N of NO2 3 progressively with a concurrent decrease in NO3
2
15 2
concentration through denitrification. The differences in d N of NO3 between compost and fertilizer treatments were consistent
throughout the incubation period. The d 15N of NO2 3 increased with the addition of compost (range: þ 13.0 to þ35.4‰), but decreased
with the addition of fertilizer (2 10.8 to þ11.4‰), thus resulting in intermediate values in soils receiving both fertilizer and compost
(23.5 to þ 20.3‰). Therefore, such differences in d 15N of NO2 15
3 observed in this study suggest a possibility that the d N of upland-
2
grown plants receiving compost would be higher than those treated with fertilizer because NO3 is the most abundant N for plant uptake
in upland soils.
q 2003 Elsevier Ltd. All rights reserved.
Keywords: Compost; d 15N; Denitrification; Fertilizer; Isotopic fractionation; Nitrification

1. Introduction (Kirchmann and Thorvaldsson, 2000). With increased

Recently, organic farming has been widely adopted as an
alternative agricultural practice to sustain economically
viable crop production with minimal environmental impacts
* Corresponding author. Tel.: þ 82-312902403; fax: þ 82-31293-8608.
E-mail address: hmro@snu.ac.kr (H.-M. Ro).
0038-0717/03/$ - see front matter q 2003 Elsevier Ltd. All rights reserved.
doi:10.1016/S0038-0717(03)00199-8
economic rewards of organic farming, consumer concerns
have grown about whether organically-labeled produce is
truly grown with organic inputs.
The use of composted manure as an organic input has
increased in areas where the use of chemical fertilizer is
prohibited or organic farming is encouraged (Hodges, 1991;
Choi et al., 2001a). In general, the 15N contents (d 15N) of
1290 W.-J. Choi et al. / Soil Biology & Biochemistry 35 (2003) 1289–1298

composted manures (. þ 8‰) are higher than those of
fertilizer (2 3 to þ 2‰) due to faster ammonia volatilization
of the lighter N isotope (14N) than 15N during the
composting process (Wassenaar, 1995; Kerley and Jarvis,
1996). This d 15N difference between N inputs suggests a
probability of different N isotope signatures of plants and/or
soils between compost and fertilizer inputs. Recently, Choi
et al. (2001b, 2002) observed that maize treated with
compost was significantly enriched more in 15N than that
receiving fertilizer, suggesting a possible usage of d 15N as a
marker in labeling organic produce.
Because inorganic-N is directly available for plant
uptake, the differences in d 15N of the inorganic-N in
soils treated with compost and fertilizer would provide
affecting the d 15N of inorganic-N through N isotopic
fractionation.
2. Materials and methods
2.1. Soil, compost, and fertilizer
For the incubation study, soil was collected from an
experimental field of Seoul National University, Korea, air-
dried, and passed through a 2 mm sieve. The soil was coarse
loamy, Typic Dystrudepts, and had 13.8 g C kg21, 1.3 g
N kg21, and a pH (H2O) of 5.8. Initial d 15N values of total-,
NHþ 2
4 -, NO3 -, and organic-N of the soil were þ 8.9, þ 12.4,

some information on the variations in d 15N of plants as þ 9.8, and þ 8.8‰, respectively (Table 1). The water
affected by N applications (Choi et al., 2002). However, holding capacity (WHC) at saturation of this soil measured
measurements of the differences in d 15N of inorganic-N by saturating a known amount of soil (Gill et al., 2001) was
in soils under field conditions are very difficult due to the 0.40 kg kg21.
complexity of N isotopic fractionation during N trans- Compost was made by mixing pig manure and sawdust
formations and the disturbance of the inorganic-N pool using a pilot scale enclosed reactor in the livestock farming
by plant uptake (Högberg, 1997). Therefore, a laboratory station at Seoul National University, Korea. Detailed
incubation under controlled conditions is a prerequisite to description of the composting reaction has been provided
by Choi et al. (2002). Compost was crushed to pass through a
study the d 15N differences of inorganic soil-N between
2 mm sieve. Chemical and physical properties of compost are
compost and fertilizer treatments.
summarized in Table 1. The d 15N values of total-, NHþ 4 -,
The high biological activity of inorganic soil-N (NHþ 4
NO2 3 -, and organic-N of the compost were þ 13.9, þ 25.9,
and NO2 3 ) means that levels of inorganic N change
þ 14.9, and þ 13.3‰, respectively.
quickly in response to various N transformation processes
For the incubation experiments, two ammonium sulfate
including nitrification and denitrification. Because many
solutions of different N-isotope composition were prepared
processes of the N cycle fractionate against the heavy
using either 15N-unlabeled (2 2.3‰ d 15N, solution A) or
isotope (15N), the d 15N patterns of inorganic soil-N are
-labeled (3.05 atom% 15N, solution B) compound. The
also sensitive to N transformation processes (Högberg,
concentration of N in each ammonium sulfate solution was
1997). For examples, nitrification is the dominant N
6000 mg l21.
process responsible for 15N enrichment of the remaining
NHþ 4 in aerobic soil, while denitrification is mainly Table 1
responsible for 15N enrichment of the remaining NO2 3 in
Chemical and physical properties of compost and soil
anaerobic soil (Choi et al., 2001c; Choi and Ro, 2003). Parameter Compost Soil
Since the predominance between nitrification and deni-
trification varies depending on soil water status, altera- pH (1:5) 8.1 5.8
tions in soil water regime can influence the d 15N of soil Total organic C (g kg21) 376 13.8
inorganic-N by modifying the two processes (Ostrom Total N
Content (g kg21) 20.6 1.3
et al., 1998). Therefore, the information on variations in d 15N (‰) þ13.9 þ 8.9
d 15N of soil inorganic-N under changing soil moisture 2 M KCl-extractable NHþ 4 -N
conditions will improve understandings of the d 15N of Content (mg kg21) 1135 0.2
soils and plants receiving compost or fertilizer. d 15N (‰) þ25.9 þ 12.4
2 M KCl-extractable NO2 3 -N
In this study, we measured temporal variations in the
Content (mg kg21) 380 20.3
d 15N of inorganic-N in soils treated with fertilizer in d 15N (‰) þ14.9 þ 9.8
combination with compost under two different soil Organic Na
moisture treatment scenarios. Our objectives were to Content (g kg21) 19.1 1.3
examine whether the applications of N-inputs having d 15N (‰) þ13.3 þ 8.8
C/N ratio 18.2 10.6
different N isotopic compositions produce isotopically Texture NAb Sandy loamc
different inorganic-N and to investigate the effect of soil
a
moisture regimes on the temporal variations in the d 15N Content of organic N is the difference in N content between total N and
inorganic N, and the corresponding d 15N value was calculated using the
of inorganic-N in soils. Additionally, we conducted isotope mass balance (Karamanos and Rennie, 1981).
another incubation using 15N-labeled chemical fertilizer b
Not applicable.
c
to estimate the rates of nitrification and denitrification USDA classification.
 W.-J. Choi et al. / Soil Biology & Biochemistry 35 (2003) 1289–1298
2.2. Incubation experiments
Two sets of incubations (15N-unlabeled and -labeled)
were conducted with soils pre-incubated at 27 8C for 10
days. During pre-incubation, soil moisture contents were
adjusted to 40 and 90% WHC for water-unsaturated and
-saturated conditions, respectively. To examine the tem-
poral variations in the d 15N of inorganic-N of soil under
changing soil moisture regimes, we laid out both drying
(saturated to unsaturated, S/U) and wetting (unsaturated to
saturated, U/S) scenarios. Drying span denotes the treat-
ments in which initial soil moisture content was adjusted
from saturated to unsaturated after 6 weeks of incubation,
while wetting is the reverse (Table 2). Soil moisture
contents were maintained at 100% WHC for saturated and
50% WHC for unsaturated conditions throughout the
incubation by daily addition of distilled water as necessary
to adjust their respective initial soil moisture content.
In Experiment 1, for the measurement of the temporal
variations in d 15N of inorganic-N of soils treated with
fertilizer and/or compost, 40 g of each soil sample (dry
basis) was weighed into a 100 ml polyethylene bottle
(3.5 cm in diameter and 5.5 cm deep) and mixed with
solution A (F) and/or compost (C) at rates of 0 and 0 (F0C0-
U/S), 150 and 0 (F1C0-U/S), 0 and 150 (F0C1-U/S), and
150 and 150 (F1C1-U/S) mg N kg21 for the wetting
treatments. For drying treatments, on the other hand, only
the first two treatments (F0C0-S/U and F1C0-S/U) without
compost were compared. In Experiment 2, for the
estimation of the rates of nitrification and denitrification,
solution B was used and three treatments (F1C0-U/S, F1C1-
U/S, and F1C0-S/U) were established. The bottles were
individually sealed with perforated cap to ensure gas
exchange and then incubated at 27 ^ 2 8C in the dark.
Three bottles from each treatment were sampled period-
ically throughout the 10-week incubation period. Details of
N application rates and changing soil-water regimes are
described in Table 2.
Table 2
Details of experiment settings for experiment 1a
Treatment N application rate Water conditions
(mg N kg21soil)
Incubation period (week)
b
AS Compost 0.5 1 2
R Q
F0C0-S/U 0 0 Saturated
F1C0-S/U 150 0 (100% WHC)c
F0C0-U/S 0 0 Unsaturated
F1C0-U/S 150 0 (50% WHC)
F0C1-U/S 0 150 Unsaturated
F1C1-U/S 150 150 (50% WHC)
a
For experiment 2 with 15N-labeled ammonium sulfate, only three treatments (F1C0-S/U, F1C0-U/S, F1C1-U/S) were included.
b
Ammonium sulfate.
c
Water holding capacity at saturation is 0.40 kg kg21.
1291
2.3. Analytical procedures
Each 35 g (dry basis) of moist soil sample was extracted
with 2 M KCl at a 5:1 extractant-to-soil ratio, and each
30 ml of the extracts were steam-distilled using MgO and
Devarda’s alloy for the N concentration of NHþ 4 and NO3
2
(Keeny and Nelson, 1982). Residual extracts were used for
d 15N analysis. The NHþ 2
4 and NO3 in the extracts were
steam-distilled, collected, and trapped in H2SO4, instead of
H3BO3. To prevent cross-contamination, acetic acid and
ethanol were used to clean the steam distillation apparatus
between each distillation (Hauck, 1982). After adjustment
of the NHþ 4 solution into pH 2 –3 using 0.1N H2SO4 or
0.1N NaOH, the solution was evaporated to dryness under
an infrared lamp (Feast and Dennis, 1996; Choi et al.,
2001c).
For the determination of N isotope compositions of the
samples, two types of stable isotope ratio mass spec-
trometers (dual-inlet and continuous-flow) were used. For
the samples from Experiment 1 using naturally abundant N,
the isotope composition of N2 gas, which was prepared
through alkaline LiOBr oxidation under vacuum (Hauck,
1982), was determined using a dual-inlet mass spectrometer
(VG OPTIMA, Micromas, UK). Fine powder (ammonium
sulfate) of the dried samples from Experiment 2 using 15N-
labeled ammonium sulfate was analyzed for atom% 15N
through a combustion unit (Feast and Dennis, 1996; Choi
et al., 2001c) linked to a continuous-flow mass spectrometer
(Isoprime-EA, Micromass, UK). The d 15N of samples
þ
containing NO2 3 or NH4 below 4 mg kg
21
soil was not
determined since the amount of N was too low to meet the N
requirement for reproducible analysis. Although the steam
distillation method may cause N isotope fractionation
during isolation of N (Robinson, 2001), the accuracy and
reproducibility of the analytical procedure checked with
reference materials (RM 8548: IAEA-N2 and RM 8549:
IAEA-N3) from the International Atomic Energy Agency
were better than 0.4 and 0.2‰, respectively.
3 4 5 6 7 8 9 10
R Q
Unsaturated
(50% WHC)
Saturated
(100% WHC)
Saturated
(100% WHC)
1292 W.-J. Choi et al. / Soil Biology & Biochemistry 35 (2003) 1289–1298
2.4. Calculations and statistical analysis
Nitrogen isotopic composition (d 15N) was expressed in
parts per thousand deviations from the atmospheric N2 as
defined by the following equation:
d15 Nð‰Þ ¼ ½ðRsample =Rstandard Þ 2 1 £ 1000 ð1Þ
where, Rsample and Rstandard are the 15N/(14N þ 15N) ratios of
the sample and standard, respectively.
To estimate gross rates of nitrification and denitrification,
which cause great isotopic alteration, N fluxes were
calculated by a numerical model FLUAZ (Mary et al.,
1998; Sørensen, 2001) using the five variables (NHþ 4 -N and
15
N, NO23 -N and
15
N, and organic-15N) collected from the
single 15NHþ 4 labeling experiment (Experiment 2). The
model combines a numerical integration of the differential
equations describing the changes in four N pools
(ammonium, nitrate, biomass and organic N) and their 15N
excess over time, and a nonlinear fitting program.
Assimilation of organic N by microorganisms, ammonia
volatilization and N humification rates were fixed at 0.
Nitrification and denitrification were assumed to be a first
order process in the model, and it was assumed that the ratio
between immobilized NHþ 2
4 -N and NO3 -N was 0.95/0.05
(Mary et al., 1998).
Data were statistically analyzed through a two-factorial
analysis (treatments £ time intervals) of variance
(ANOVA) using Generalized Linear Models procedures
(SAS Institute, 1989). Least significant differences (LSD)
are given to indicate significant variations between means
for selected subsets of data.
3. Results
3.1. Temporal variations in the inorganic N content
Although fertilizer addition increased the NHþ 4 concen-
tration in soils (Fig. 1b), in soils which were unsaturated to
begin with (F1C0-U/S and F1C1-U/S), the initial increase
disappeared rapidly such that values were as low as those of
treatments receiving no fertilizer within 1 or 2 weeks. On
the other hand, in the initially saturated soil (F1C0-S/U),
NHþ 4 concentrations decreased gradually to the level of
treatments without fertilizer (Fig. 1a). Changing the soil
moisture content after 6-week incubation did not affect the
overall NHþ 4 concentration in any treatments.
In soils which were unsaturated to begin with (F1C0-U/S
and F1C1-U/S), the addition of fertilizer significantly
increased the NO2 3 level during the first 0.5 week, but the
increase was small in the initially saturated (F1C0-S/U)
soils (Fig. 1b). In particular, the NO2 3 concentrations of
F1C0-U/S and F1C1-U/S soils receiving fertilizer rose to
the maximum value (. 150 mg N kg21) within the first
week and this value was maintained until 6 weeks (the point
of soil moisture change), while those of the remaining soils
were maintained below 50 mg kg21 throughout the incu-
bation. When the soil moisture status was changed from
unsaturated to saturated conditions after 6 weeks, the NO2 3
concentrations began to decrease considerably in soils with
compost (F0C1-U/S and F1C1-U/S) and slightly without
compost (F0C0-U/S and F1C0-U/S). In contrast, the NO2 3
concentrations of F0C0-S/U and F1C0-S/U soils began to
increase slightly after changing the soil moisture conditions
from saturated to unsaturated.
3.2. Temporal variations in the d15N of inorganic N
At 0.5-week incubation of unsaturated soils, the d 15N of
NHþ 4 increased with increasing N-application rates, irre-
spective of compost or fertilizer treatment (Fig. 2a). For
soils mixed at a given compost level, the d 15N of NHþ 4
increased from þ 8.1‰ for F0C0-U/S to þ 21.1‰ for F1C0-
U/S and from þ 18.1‰ for F0C1-U/S to þ 27.3‰ for F1C1-
U/S with the addition of fertilizer (Fig. 2a). Similarly, for
soils mixed at a given fertilizer level, the d 15N of NHþ 4
increased from þ 8.1‰ for F0C0-U/S to þ 18.1‰ for F0C1-
U/S and from þ 21.1‰ for F1C0-U/S to þ 27.3‰ for F1C1-
U/S with the addition of compost. In contrast, the addition of
fertilizer to soils under saturated conditions (F1C0-S/U)
significantly ðP , 0:05Þ lowered the d 15N of NHþ 4,
compared to soils (F0C0-S/U) receiving no N input. During
further incubation until 6 weeks, the d 15N of NHþ 4 of
saturated soils increased from þ 11.5 to þ 26.9‰ for F0C0-
S/U and from þ 2.9 to þ 18.9‰ for F1C0-S/U, while those
of unsaturated soils decreased and some were not measur-
able due to low N concentration below 4 mg kg21. After 6
weeks, d 15N data were not collected in all treatments
because of low N concentrations.
In soils under unsaturated conditions, the d 15N of NO2 3
was significantly ðP , 0:05Þ lowered by the addition of
fertilizer, while increased by compost (Fig. 2b). In
particular, the d 15N values of NO2 3 of F1C1-U/S and
F1C0-U/S soils receiving fertilizer were negative (2 3.5 and
2 10.8‰, respectively) at 0.5 week of incubation, while
those of the remaining soils were positive. During further
incubation until 6 weeks, irrespective of fertilizer and
compost treatments, the d 15N of NO2 3 in unsaturated soils
remained stable (about þ 3.0‰ for F1C0-U/S, þ 10.0‰ for
F1C1-U/S, þ 15.0‰ for F0C0-U/S, þ 19.0‰ for F0C1-U/
S). However, changing the soil moisture regime to saturated
conditions after 6 weeks resulted in significant increases in
d 15N; the values were þ 11.4‰ for F1C0-U/S, þ 20.3‰ for
F1C1-U/S, þ 29.2‰ for F0C0-U/S, þ 35.4‰ for F0C1-U/S
at 8 weeks of incubation.
In contrast, the addition of fertilizer significantly
increased the d 15N of NO2 3 in the initially saturated soils
(F1C0-S/U). In general, the d 15N values of NO2 3 rose from
þ 6.9 to þ 22.5‰ for F0C0-S/U soils and from þ 16.3 to
þ 38.7‰ for F1C0-S/U soils during the first 3 weeks of
incubation and formed stable plateaus for another 3 weeks.
 W.-J. Choi et al. / Soil Biology & Biochemistry 35 (2003) 1289–1298 1293

Fig. 1. Temporal changes in (a) NHþ 2

4 -N and (b) NO3 -N concentrations. Symbols: X, F0C0-S/U; A, F1C0-S/U; K, F0C0-U/S; £ , F1C0-U/S; þ , F0C1-U/S; W,
F1C1-U/S. Values are the means of triplicates. Vertical bars represent LSDs ðP ¼ 0:05Þ between treatments at each time. LSDs ðP ¼ 0:05Þ between time
intervals for F0C0-S/U, F1C0-S/U, F0C0-U/S, F1C0-U/S, F0C1-U/S, and F1C1-U/S are 1.6, 2.5, 2.0, 1.4, 1.1, and 1.2 mg kg21 for NHþ 4 -N and 2.2, 8.5, 3.3,
12.1, 8.4, and 7.3 mg kg21 for NO23 -N, respectively. The values for fertilizer-treated soils are the averaged concentrations obtained through both Experiments 1
and 2 since the concentrations from Experiment 2 were in good agreement with those of Experiment 1. Details of treatments are shown in Table 2. Soil water
regimes were changed after 6 weeks of incubation.

However, after the soil moisture regime was switched to
unsaturated conditions, the d 15N of NO2 3 decreased; the
values were þ 5.7‰ for F0C0-S/U and þ 12.6‰ for F1C0-
S/U at the end of incubation. The missing data points of the
d 15N values of NO2 3 were due to low N concentration at
sampling times.
3.3. Gross rates of nitrification and denitrification
The time-course patterns of nitrification and
denitrification estimated using the FLUAZ model with
the respective 15N dataset were greatly different between
soil moisture treatments (Fig. 3). The rate of nitrification
of soils (F1C0-U/S and F1C1-U/S) under unsaturated
conditions was significantly greater than that of soils
(F1C0-S/U) under saturated conditions during the first
week of incubation, but thereafter sharply decreased,
showing that most of nitrification occurred during this
period (Fig. 3a). On the other hand, meaningful nitrifica-
tion was predicted in the initially saturated soil (F1C0-S/
U) for a relatively long time compared to the initially
unsaturated soils.
1294 W.-J. Choi et al. / Soil Biology & Biochemistry 35 (2003) 1289–1298

Fig. 2. Temporal changes in d 15N values of (a) NHþ 2

4 and (b) NO3 . Symbols: X, F0C0-S/U; A, F1C0-S/U; K, F0C0-U/S; £ , F1C0-U/S; þ , F0C1-U/S; W,
F1C1-U/S. Values are the means of triplicates. Vertical bars represent LSDs ðP ¼ 0:05Þ between treatments at each time. For d 15N of NO2
3 in Fig. 2 (b), LSDs
ðP ¼ 0:05Þ between time intervals for F0C0-S/U, F1C0-S/U, F0C0-U/S, F1C0-U/S, F0C1-U/S, and F1C1-U/S are 1.8, 2.6, 3.3, 2.1, 2.1 and 2.6‰, respectively.
Some data were not determined due to the low N concentrations. Details of treatments are shown in Table 2. Soil water regimes were changed after 6 weeks of
incubation as indicated in Fig. 1.

In F1C0-S/U soil, denitrification rate rapidly increased 4. Discussion

during the first 2 weeks of incubation under saturated
conditions, but gradually decreased thereafter (Fig. 3b).
However, virtually no denitrification was estimated in
F1C0-U/S and F1C1-U/S soils under unsaturated conditions
during the first 6 weeks, but under subsequent saturated
conditions, the rate of denitrification gradually increased for
F1C1-U/S soils, while denitrifiaction began to occur during
the last 2 weeks of incubation for F1C0-U/S soil.
4.1. Variations in the d15N of NHþ
4
The effects of kinetic isotope fractionation associated
with nitrification, which preferentially incorporates 14N
rather than 15N into NO2 3 (Mariotti et al., 1981), on the
increase in d 15N of NHþ4 persisted for a relatively long time
in the initially saturated F0C0-S/U and F1C0-S/U soils
 W.-J. Choi et al. / Soil Biology & Biochemistry 35 (2003) 1289–1298 1295

Fig. 3. Rates of (a) nitrification and (b) denitrification in F1C0-S/U, F1C0-U/S, and F1C1-U/S soils estimated with the FLUAZ model. Vertical bars represent
standard deviation of the means ðn ¼ 3Þ: Details of treatments are shown in Table 2.

(Fig. 2a) due to slow nitrification under saturated conditions conditions, a negative correlation between the N concen-
(Fig. 3a). However, 15N-enrichment of NHþ 4 during trations and the d 15N of NHþ 4 was observed (Fig. 4),
nitrification decreased with decreasing concentrations of indicating 15N-enrichment of the remaining NHþ 4 increased
the substrate (NHþ 4 ). In the F1C0-S/U soils under saturated as nitrification progressed (Choi and Ro, 2003). For F0C0-S/U
1296 W.-J. Choi et al. / Soil Biology & Biochemistry 35 (2003) 1289–1298
Fig. 4. Relationship between concentrations and d 15N of the remaining
NHþ 4 in F1C0-S/U soil treated with fertilizer under saturated conditions
during the first 6 weeks. Vertical and horizontal bars represent standard
deviation of the means ðn ¼ 3Þ of the d 15N and concentrations of the NHþ
4,
respectively.
soil, however, the negative correlation was not observed
because of low NHþ 4 concentration. The consistently lower
d 15N of NHþ 4 in soil with fertilizer (F1C0-S/U) than that
without fertilizer (F0C0-S/U) suggested that the effect of
isotopic composition of N source on the d 15N of NHþ 4 was
apparent in the initially saturated soil with low nitrifying
capacity.
In contrast, in the initially unsaturated soils, the d 15N
of NHþ 4 at 0.5 week increased with increasing N-
application rates (F1C1-U/S . F1C0-U/S . F0C1-
U/S . F0C0-U/S) as seen in Fig. 1a, and this suggested
that 15N composition of NHþ 4 in soils with high nitrifying
capacity was affected more by the availability of
substrate for nitrification than by the d 15N of N inputs.
Since the N isotopic fractionation associated with
nitrification is most marked when NHþ 4 is abundant in
soils, it becomes greater when NHþ 4 is abundant in soils
(Feigen et al., 1974; Choi et al., 2002). In particular, in
F1C1-U/S soil, the decreased d 15N of NHþ 4 from þ 27.3
at 0.5 week to þ 10.1‰ at 1 week showed the
contribution of mineralized NHþ 4 from organic-N pool
to lowering d 15N of NHþ 4 (Ostrom et al., 1998), since the
d 15N of NHþ 4 is the result of both nitrification resulting
in 15N-enriched NHþ 4 and ammonification producing
15
N-
þ þ
depleted NH4 compared to the original NH4 pool
(Nadelhoffer and Fry, 1994).
At 0.5 week, the higher d 15N of NHþ 4 in soils receiving
fertilizer alone under unsaturated conditions (F1C0-U/S,
þ 21.1‰) compared to that under saturated conditions
(F1C0-S/U, þ 2.9‰) again showed the greater effect of
nitrification on the 15N-enrichment of the remaining NHþ 4 in
unsaturated soils than that in saturated soils. The gross rate
of nitrification during the first 0.5 week of incubation,
estimated using the FLUAZ model, was also significantly
greater in soils under unsaturated (13.7 mg N kg21 d21)
conditions than that under saturated (9.1 mg N kg21 d21)
conditions (Fig. 3a).
4.2. Variations in the d15N of NO2
3
The d 15N of NO2 3 was controlled chiefly by denitrifica-
tion under saturated, but by nitrification under unsaturated
conditions (Fig. 2b). Extremely low or high d 15N values of
NO3 were due to 15N-discrimination during nitrification and
denitrification. However, these extreme values are not likely
to occur under field conditions because the amount of N is
too small to detect and is diluted considerably by the same
compounds form other N sources (Högberg, 1997).
In the initially saturated soils (F0C0-S/U and F1C0-
S/U), the d 15N of NO2 3 increased during the first 3
weeks of incubation, indicating the isotopic segregation
during denitrification (Mariotti et al., 1981; Choi et al.,
2001c). However, during the subsequent 3 weeks, the
increment of d 15N was reduced with decreasing
availability of the substrate as denitrification progressed
(Fig. 3b), and this observation was consistent with the
pattern of temporal variations in d 15N of NHþ 4 . Changing
the soil moisture to unsaturated conditions after 6 weeks
gradually decreased the d 15N of NO2 3 (Fig. 2b) as the
NO2 3 concentration gradually increased (Fig. 1b) through
nitrification. The higher d 15N of NO2 3 in soil treated
with fertilizer (F1C0-S/U) than that without fertilizer
(F0C0-S/U) could also be attributed to higher availability
of the substrate for denitrification in F1C0-S/U soil, as
mentioned previously.
In contrast, for the initially unsaturated soils (F0C0-U/S,
F1C0-U/S, F0C1-U/S, and F1C1-U/S), the maximum 15N-
depletion of the NO2 3 pool was observed at the first 0.5 week
(Fig. 2b) due to the isotope effect caused by incomplete
nitrification of NHþ 4 (Karamanos and Rennie, 1981; Choi
and Ro, 2003). In particular, the much lower d 15N of NO2 3
(2 10.8‰) in F1C0-U/S soil than that of applied-NHþ 4
(2 2.3‰) indicated that considerable isotopic discrimi-
nation occurred during nitrification. However, the initial
drop in the d 15N of NO2 3 was soon followed by an increase
in 15N-enrichment as nitrification proceeded to completion
(Fig. 3b). As reported by Högberg (1997) and Choi and Ro
(2003), the initial 15N-depletion followed by 15N-enrich-
ment is a typical pattern of the variations in d 15N during a
single step unidirectional reaction in a closed system.
During the subsequent incubation up to 6 weeks, the
stable NO2 3 concentrations and the corresponding d N
15
values (Figs. 1b and 2b) could be explained by decreased
nitrification due to very low concentrations of NHþ 4 (Figs.
1a and 3a), which serves as the substrate for nitrification.
After changing the soil moisture status to saturated
conditions, a greater increase in d 15N of NO2 3 in soils
receiving fertilizer and compost (F1C1-U/S) than that
receiving fertilizer alone (F1C0-U/S) probably resulted
from the enhanced denitrification (Fig. 3b) due to high
organic-C content of the compost-treated soils. Aulakh and
Rennie (1987) have suggested that decomposition of
organic materials and mineralizaiton of organic-C would
 W.-J. Choi et al. / Soil Biology & Biochemistry 35 (2003) 1289–1298
increase microbial demand for O2 and thereby create
anaerobic conditions facilitating enhanced denitrifcation.
Although the d 15N of inorganic soil-N is the result of the
interactions between N sources and N isotopic fractiona-
tions (Handley and Scrimgeour, 1996; Robinson, 2001),
the significant differences in d 15N of NO2 3 between the four
treatments (F0C0-U/S, F1C0-U/S, F0C1-U/S, and F1C1-U/
S) were virtually uniform throughout the incubation (Fig.
2b). Overall, the d 15N of NO2 3 increased with the addition
of compost, but decreased with fertilizer, thus resulting in
intermediate values in soils receiving both fertilizer and
compost due to the mixing of two N pools having different
isotopic compositions. These data corroborate the findings
of Yoneyama et al. (1990) and Choi et al. (2001b, 2002) that
plants grown in soils receiving compost had more 15N
contents than those grown with fertilizer under upland
conditions.
In conclusion, despite lack of information supporting the
comparative effect of fertilizer with or without compost on
the concentration and the d 15N of NHþ 2
4 and NO3 under soil
drying scenario, this study showed that the temporal
variations in d 15N of inorganic N were affected greatly by
the predominance between the nitrification and denitrifica-
tion associated with soil moisture status. In water-saturated
soils, slow nitrification gradually increased the d 15N of
NHþ 4 , and fast denitrification resulted in considerable
15
N-
enrichment of NO3 . In unsaturated soils, the d N of NHþ
2 15
4
increased sharply due to fast nitrification, but the effect of
denitrification on the d 15N of NO2 3 was not observed due to
aerobic conditions. However, the effects of these two
processes on the d 15N variations decreased with decreasing
availability of the substrates. On the other hand, the isotopic
compositions of N inputs apparently affected the d 15N of
NHþ 4 in saturated sols with low nitrifying capacity, and that
of NO2 3 in unsaturated soils with low denitrifying capacity.
In particular, the results showing the higher d 15N of NO2 3 in
soils receiving compost than fertilizer under unsaturated
conditions could explain that the d 15N values of plants
grown with compost were higher than those with fertilizer
(Yoneyama et al., 1990; Choi et al., 2001b, 2002), since
NO2 3 is the most abundant N for plant uptake. However, it
should be noted that the significant difference in d 15N of
soil NO2 3 between compost and fertilizer treatments in
upland fields would be apparent only in early plant-growth
period following N fertilization, since plant uptake of NO2 3 natural abundance stable isotope discrimination. Plant and Soil 178,
and increased microbial activity stimulated by plant
exudates may disturb the NO2 3 pool.
Acknowledgements
Authors thank NICEM, SNU for allowing the use of
stable isotope ratio mass spectrometers, and Dr. Bruno Mary
for sharing the FLUAZ program in estimating the rates of
nitrification and denitrification.
1297
References
Aulakh, M.S., Rennie, D.A., 1987. Effect of wheat straw incorporation on
denitrification under anaerobic and aerobic conditions. Canadian
Journal of Soil Science 67, 825–834.
Choi, W.J., Jin, S.A., Lee, S.M., Ro, H.M., Yoo, S.H., 2001a. Corn uptake
and microbial immobilization of 15N-labeled urea-N in soil as affected
by composted pig manure. Plant and Soil 235, 1–9.
Choi, W.J., Lee, S.M., Kim, K.C., Kim, P.G., Yoo, J.H., Yoo, S.H., 2001b.
Natural d 15N abundances of corn treated with urea or composted pig
manure in a pot experiment. Korean Journal of Soil Science and
Fertilizer 34, 292–301.
Choi, W.J., Lee, S.M., Ro, H.M., Kim, K.C., Yoo, S.H., 2002. Natural 15N
abundances of maize and soil amended with urea and composted pig
manure. Plant and Soil 245, 223–232.
Choi, W.J., Lee, S.M., Yoo, S.H., 2001c. Increase in d 15N of nitrate
through kinetic isotope fractionation associated with denitrificaiton in
soil. Agricultural Chemistry and Biotechnology 44, 135–139.
Choi, W.J., Ro, H.M., 2003. Differences in isotopic fractionation of
nitrogen in water-saturated and unsaturated soils. Soil Biology &
Biochemistry 35, 483–486.
Feast, N.A., Dennis, P.F., 1996. A comparison of methods for nitrogen
isotope analysis of groundwater. Chemical Geology 129, 167 –171.
Feigen, A., Kohl, D.H., Shearer, G., Commoner, B., 1974. Variation in the
natural nitrogen-15 abundance in nitrate mineralized during incubation
of several Illinois soils. Soil Science Society of America Proceeding 38,
90– 95.
Gill, J.S., Sivasithamparam, K., Smettem, K.R.J., 2001. Soil moisture
affects disease severity and colonization of wheat roots by Rhizoctonia
solani AG-8. Soil Biology & Biochemistry 33, 1363–1370.
Handley, L.L., Scrimgeour, C.M., 1996. Terrestrial plant ecology and 15N
natural abundance: the present limits to interpretation for uncultivated
systems with original data from a Scottish old field. Advances in
Ecological Research 27, 133 –212.
Hauck, R.D., 1982. Nitrogen-isotope ratio analysis. In: Page, A.L., (Ed.),
Methods of Soil Analysis. Part 2: Chemical and Micorbiological
Properties, ASA and SSSA, Madison and Wisconsin, pp. 735–779.
Hodges, R.D., 1991. Soil organic matter: its central position in organic
farming. In: Wilson, W.S., (Ed.), Advances in Soil Organic Matter
Research: The Impact on Agriculture and the Environment, The Royal
Society of Chemistry, Redwood Press, Wiltshire, pp. 355–364.
Högberg, P., 1997. 15N natural abundance in soil-plant systems. New
Phytologist 137, 179 –203.
Karamanos, R.E., Rennie, D.A., 1981. Changes and significance in natural
15
N abundance in residual nitrogen fertilizer studies. Canadian Journal
of Soil Science 61, 553 –559.
Keeney, D.R., Nelson, D.W., 1982. Nitrogen-inorganic forms. In: Page,
A.L., (Ed.), Methods of Soil Analysis. Part 2: Chemical and
Micorbiological Properties, ASA and SSSA, Madison and Wisconsin,
pp. 643–698.
Kerley, S.J., Jarvis, S.C., 1996. Preliminary studies of the impact of
excreted N on cycling and uptake of N in pasture systems using
287– 294.
Kirchmann, Thorvaldsson, 2000. Challenging targets for future agriculture.
European Journal of Agronomy 12, 145–161.
Mariotti, A., Germon, J.C., Hubert, P., Kaiser, P., Letolle, R., Tardieux, A.,
Tardieux, P., 1981. Experimental determination of nitrogen kinetic
isotope fractionation: some principles; illustration for the denitrification
and nitrification processes. Plant and Soil 62, 423 –430.
Mary, B., Recous, S., Robin, D., 1998. A model for calculating nitrogen
fluxes in soil using 15N tracing. Soil Biology & Biochemistry 30,
1963–1963.
Nadelhoffer, K.J., Fry, B., 1994. Nitrogen isotope studies in forest
ecosystems. In: Lajtha, K., Michener, R.H. (Eds.), Stable Isotopes in
1298 W.-J. Choi et al. / Soil Biology & Biochemistry 35 (2003) 1289–1298
Ecology and Environment Science, Blackwell, London, pp.
643– 698.
Ostrom, N.E., Knoke, K.E., Hedin, L.O., Robertson, G.P., Smucker,
A.J.M., 1998. Temporal trends in nitrogen isotope values of nitrate
leaching from an agricultural soil. Chemical Geology 146, 219–227.
Robinson, D., 2001. d 15N as an integrator of the nitrogen cycle. Trends in
Ecology and Evolution 16, 153–162.
SAS Institute, 1989. SAS/STAT user’s guide. Version 6.4th ed, vol. 2. SAS
Inst, North Carolina.
Sørensen, P., 2001. Short-term nitrogen transformations in soil amended
with animal manure. Soil Biology & Biochemistry 33, 1211–1216.
Wassenaar, L.I., 1995. Evaluation of the origin and fate of nitrate in the
Abbotsford Aquifer using the isotopes of 15N and 18O in NO2 3 . Applied
Geochemistry 10, 391 –404.
Yoneyama, T., Kouno, K., Yazaki, J., 1990. Variations of natural 15N
abundance of crops and soils in Japan with special reference to the
effects of soil conditions and fertilizer application. Soil Science and
Plant Nutrition 36, 667–675.