Directory UMM :Data Elmu:jurnal:S:Soil Biology And Chemistry:Vol32.Issue11-12.oct2000:
Soil Biology & Biochemistry 32 (2000) 1529±1536
www.elsevier.com/locate/soilbio
Ion exchange resin±soil mixtures as a tool in net nitrogen
mineralisation studies
JuÈrgen K. Friedel a,*, Anke Herrmann b, Markus Kleber c
b
a
University of Agricultural Sciences, Institute of Organic Farming, Gregor-Mendel-Street 33, A-1180 Vienna, Austria
Department of Soil Sciences, Division of Soil Fertility and Plant Nutrition, Swedish University of Agricultural Sciences (SLU), S-75007 Uppsala,
Sweden
c
Martin-Luther University Halle-Wittenberg, Institute of Soil Science and Plant Nutrition, Weidenplan 14, D-06108 Halle, Germany
Accepted 29 February 2000
Abstract
Mixed-bed ion exchange resins (IER) were mixed with intact soil aggregates and incubated at 60% water ®lled pore space in
closed polyethylene bags for 12 weeks. To test IER eects on N losses, nitri®cation and net N mineralisation, an arable soil and
a grassland soil, diering in organic matter content, were chosen and two crop residues (wheat straw, sugar-beet leaves) with
dierent C-to-N ratios were added to the arable soil. It was proposed that IER might exert an in¯uence on N cycling similar to
that of plant roots. Nitri®cation was inhibited by adsorption of NH4 in the +IER treatments. Net N mineralisation was greater
in the grassland soil than in the arable soil which had less soil organic matter. Without incorporation of additional organic
substrates, net N mineralisation was not aected by IER in both soils. Straw addition to the arable soil caused immediate N
immobilisation in the ÿIER treatment, whereas N mineralisation continued in the +IER treatment. Incorporation of sugar-beet
leaves into the arable soil highly increased net N mineralisation and microbial biomass N in the ÿIER treatment. In the +IER
treatment, the enhancement of both N mineralisation and microbial biomass N was less pronounced. Thus, IER mixed into soil
samples can exert either a stimulating (wheat straw) or dampening (sugar-beet leaves) eect on N mineralisation. Soil±IER
mixtures can prevent losses and re-immobilisation of mineralised N and mimic nutrient exchange properties of plant roots. It is
concluded that in incubation experiments they can better re¯ect conditions in the vicinity of roots than incubations without IER
or with incorporation of IER in con®ned resin bags as long as water and aeration conditions are not largely changed. Soil±IER
mixtures may also be a useful tool for studying root-induced changes in net N mineralisation. 7 2000 Elsevier Science Ltd. All
rights reserved.
Keywords: Ion exchange resins; Soil±resin mixtures; Soil nitrogen cycling; Nitrogen mineralisation; Nitri®cation
1. Introduction
Quanti®cation of the relationships between soil N
mineralisation and other ecosystem processes has been
restricted by the lack of suitable methods to measure
N mineralisation under ®eld conditions. Methods used
to measure or estimate patterns of N mineralisation
under ®eld conditions include: (i) exposure of dis-
* Corresponding author. Fax: +43-1-47654-3792.
E-mail address: friedel@edv1.boku.ac.at (J.K. Friedel).
turbed soil in plastic bags buried in the ®eld, (ii) exposure of relatively undisturbed soil columns under
®eld conditions, and (iii) measurement of mineral N
collected by ion exchange resins placed in the ®eld for
extended periods (Raison et al., 1987).
All methods of containment developed to date alter
the soil environment through (i) cessation of the carbon (C) input from decomposing litter and from ®ne
root turnover, (ii) increased C inputs from severed
roots, (iii) modi®cation of the moisture and temperature regimes relative to bulk soil, and (iv) accumulation of inorganic N (Adams et al., 1989). An
0038-0717/00/$ - see front matter 7 2000 Elsevier Science Ltd. All rights reserved.
PII: S 0 0 3 8 - 0 7 1 7 ( 0 0 ) 0 0 0 6 4 - X
5.9
5.2
0
0
8.7
10.2
0.15
0.46
1.3
4.7
45.0
59.5
a
b
Horizon designators according to A.G. Boden (1994).
Textural classes according to USDA (1996).
1.4
1.0
0
0
Silt loam
Clay loam
Ap
Ah
Arable
Grassland
0-30
0-10
Ccarbonate (%)
C/N
Nt (%)
TOC (%)
Total pore space (vol%)
Bulk density (g cmÿ3)
Rock fragments (vol%)
Textureb
Depth (cm)
Horizona
Soil
additional source of error is the inability to prevent
mineralised N (Nmin) from being either re-immobilised
into microbial biomass or denitri®ed within the containment period. As assay conditions alter the C and
N availability within the containers, none of the
methods may be considered to measure mineralisation
rates accurately.
The use of intact soil cores (Nordmeyer and Richter,
1985; Raison et al., 1987) may reduce the sources of
error due to disturbance of the soil and modi®cations
of the moisture and temperature regimes. The central
problem however, the absence of living root functions,
remains.
In undisturbed soil inhabited by living plants, root
N uptake reduces or even inhibits denitri®cation or reimmobilisation of mineralised N and limits microbial
N availability. Simultaneously occurring rhizodeposition leads to an increase in the amount of substrate
available to soil micro-organisms, enabling them to
mineralise ``surplus'' amounts of organic N. Consecutive predation by soil fauna may release part of this
extra N as NH4 (Clarholm, 1985).
Any attempt to advance the methodology for studying ¯uxes of soil mineral-N in situ must therefore seek
to ®nd a means which takes root functions into
account. Such a procedure should immobilise inorganic N throughout the incubation, thus preventing it
from being nitri®ed, denitri®ed or re-immobilised into
microbial biomass.
If this process occurs with the same magnitude as is
performed by living roots, it should eliminate the most
prominent source of error.
IER were shown to adsorb NH4 and NO3 eectively
in soils (Binkley, 1984). Only few attempts (e.g. HuÈbner et al., 1991; Binkley et al., 1986; DiStefano and
Gholz, 1986) to establish the use of IER as a procedure to measure net N mineralisation were known
when growing concern about the reliability of the buried bag method (Eno, 1960) led Zeller et al. (1997) and
Bhogal et al. (1999) to investigate further into the subject. In all previous attempts to employ IER as a tool
in net nitrogen mineralisation studies, these were
applied in ``resin bags'' to the soil. Such design may
prevent Nmin from being leached out of cores as well
as from being washed into them. Due to the low mobility of NH4 in soil, however, nitri®cation and re-immobilisation of NH4 cannot be prevented. Thus, with
resin bags of traditional design, Nmin may continue to
accumulate in soil samples during the incubation.
We assumed that by mixing IER thoroughly into
soil samples, the accumulation of microbially-available
mineralised N in soil samples can be prevented and
one of the most important eects of living roots on
soil N transformations (i.e. the removal of Nmin) may
be simulated. Conditions in soil samples containing
IER can therefore be expected to be closer to con-
pH, CaCl2
J.K. Friedel et al. / Soil Biology & Biochemistry 32 (2000) 1529±1536
Table 1
Selected soil properties
1530
J.K. Friedel et al. / Soil Biology & Biochemistry 32 (2000) 1529±1536
ditions in the vicinity of roots than conditions in
samples without IER. For this reason, N mineralisation rates measured in soil samples containing IER
should be a more realistic measure for N mineralisation rates under ®eld conditions.
To achieve this, we tested (i) if mixing IER into soil
samples can reduce losses of Nmin either by denitri®cation or immobilisation, (ii) whether nitri®cation can be
reduced or even prevented by IER in soil samples, and
(iii) if net N mineralisation rates are enhanced in the
presence of both IER and microbially-available substrate.
2. Materials and methods
2.1. Soil sampling
Soil samples were taken from the A horizons of an
arable soil (Stagnic Luvisol; ``Arable soil'') and a
humus-rich grassland soil (Cumulic Anthrosol; ``Grassland soil''). Basic soil properties are given in Table 1.
In both cases, a surface area of 1 m2 was cleared of
plant materials and the soil subsequently sampled
down to the horizon boundary. The soil was then
freed from root residues and homogenised. This was
done with maximum care to preserve crumbs and
aggregates. As the material was stone-free, no further
sieving was necessary until preparation of individual
incubates.
2.2. Ion exchange resin (IER)
When both cations and anions are included as target
ions, the various species can be accumulated by the
resin if a mixture of cation- and anion-resins are
placed into the medium (Skogley and Dobermann,
1996). Consequently, a mixed-bed resin (Amberlite1
MB-3) from CWG GmbH, Mannheim, Germany, was
used. Amberlite1 MB-3 is a mixture of the strong cation exchange resin Amberlite1 IR-120 and the strong
anion exchange resin Amberlite1 IRA-420 Type 1.
Grain size of the moist resin was determined to be in
the range 0.3±1.2 mm, with most of the grains within
0.5 and 1.0 mm. The speci®c weight was 0.7 g cmÿ3.
Prior to the experiment, IER was washed once with
deionised water. By this treatment, the N content
extractable with 2 M KCl (soil : extractant ratio=1 :
20) was reduced to about 50% (data not shown).
2.3. Incubation experiment
To
rates
ganic
poor
test the in¯uence of IER on N mineralisation
under dierent conditions, soils diering in ormatter content and an addition of N-rich and Nsubstrates were chosen. The treatments were:
1531
``Arable soil ÿ IER'' (low organic matter), ``Arable
soil + IER'', ``Grassland soil ÿ IER'' (high organic
matter), ``Grassland soil + IER'', ``Arable soil +
wheat straw ÿ IER'', ``Arable soil + wheat straw +
IER'', ``Arable soil + sugar-beet leaves ÿ IER'',
``Arable soil + sugar-beet leaves + IER''.
2.4. Preparation of organic additives
Wheat straw and sugar-beet leaves were dried at
508C for 72 h and ground to pass a 1 mm sieve. After
this, wheat straw had a water content of 7.7% and
sugar-beet leaves of 16.2%. Wheat straw had a C content of 409 mg gÿ1 and an N content of 7 mg gÿ1,
yielding a C-to-N ratio of 58. C and N contents in
sugar-beet leaves were 397 and 28.3 mg gÿ1; which
resulted in a C-to-N ratio of 14. Organic additives
were added to soil at a 1:30 ratio by weight.
2.5. Preparation of individual incubates
Polyethylene freezer bags manufactured by Fa.
Haaf, 85716 Lohhof, Germany, were used as containers for the soil. Bags had an average standard thickness of 50 mm. Polyethylene ®lm is only slightly
permeable to water vapour, while its O2 and CO2
transmission rates are relatively high. Soil structure in
the samples was not disturbed, i.e. complete aggregates
were incubated. Soil sample size was 300 g. The same
amount of resin was added in the treatments with IER
addition. As the density of quartz is about double the
density of the resin selected, 600 g of quartz sand of a
grain size similar to that of the resin were added to
treatments without IER in order to maintain identical
grain size distribution and soil physical conditions.
For each treatment, 21 replicate samples were prepared and incubated at 158C and at 60% water ®lled
pore space. Three samples per treatment were selected
the next day (t0 = 16 h) at random and analysed to
determine starting values. Every 2 weeks, another
three samples per treatment were selected at random
from the incubator and analysed in the same manner
to represent a total of 7 (t0 . . .t6) data points spanning
incubation periods from 0 to 12 weeks in 2-week increments tn1 ÿ tn 2 weeks). Microbial biomass N
(Nmic) was determined at t0 and t6 (in the beginning of
the experiment and after 12 weeks).
2.6. Determination of inorganic N and microbial N
Inorganic N was extracted by 2 M KCl solution
(extractant : soil ratio = 40:1) and measured by means
of a Skalar autoanalyser. NH4 was determined colorimetrically by a modi®ed Berthelot reaction according
to Krom (1980) and NO3 after reduction to NO2 by
the Griess reaction as a-naphthylamine-para-diazoben-
1532
J.K. Friedel et al. / Soil Biology & Biochemistry 32 (2000) 1529±1536
zene-parasulphuric acid at 540 nm in an automated
analyser (Houba et al., 1987).
Microbial biomass N was measured with the slightly
modi®ed standard fumigation-extraction method
(Brookes et al., 1985). Brie¯y, soil samples (20 g for
IER±soil incubates and 30 g for quartz±soil incubates,
equivalent to 10 g of wet soil each) were fumigated
with CHCl3 for 24 h at 258C. For analysis at t6, 250 ml
liquid CHCl3 was added to each portion of the soil
samples before fumigation, according to the method
recommended by Mueller et al. (1992) for soil samples
with a high moisture content. After removal of the
CHCl3, soluble N was extracted from fumigated and
unfumigated samples with 2 M KCl (extractant:soil
ratio = 40:1) for 2 h. N in ®ltrated solution was determined by the Griess reaction colorimetrically after an
automatic UV peroxide digestion to NO3 and reduction to NO2 by means of a Skalar autoanalyser
(Houba et al., 1987). Nmic was calculated using a kEN
factor of 0.54.
Fig. 2. Amount of NO3±N in the soil samples during the incubation
experiment. Bars indicate one standard deviation. r = Treatments
without IER, R = treatments with soil±IER mixtures. n = 3 for
each treatment. : treatments dier (Mann and Whitney U-test, P <
0.05).
3.2. IER eects on net N-mineralisation
3. Results
3.1. IER eects on nitri®cation
In ÿIER samples, mineralised N was readily nitri®ed (Figs. 1 and 2). At the end of the incubation (12
weeks) all Nmin was nitri®ed to NO3 (Table 2). In
+IER samples, NO3 concentrations found at t0
remained constant throughout the incubation (Fig. 2,
Table 2). Mineralisation occurring during the incubation merely increased the NH4 pool.
In the ``Arable soil'' without substrate addition,
Nmin accumulated gradually at a low rate during the
incubation (Fig. 3). No signi®cant dierences in Nmin
Table 2
Inorganic N and soil microbial biomass N (mg gÿ1) in the dierent
soil treatments with or without IER (standard deviation in parentheses) after 16 h (t0) and 12 weeks (t6)
NO3 ±N
NH+
4 ±N
Inorganic N
Microbial N
t0
t0
t0
t0
t6
t6
t6
Arable soil
ÿIER
8.5
38.6
0.4
0
8.9 38.6
67
(0.7) (7.1) (0.6) (0)
(0.4) (7.1) (13)
+ IER 7.9
9.7a
3.3a 24.3a 11.1 34.0
±b
(1.2) (2.5) (2.1) (3.1) (1.8) (5.5)
Arable soil + wheat straw
ÿIER
2.8
0
3.7
0
6.5
0
123
(0.7) (0)
(3.8) (0)
(3.1) (0)
(2)
+ IER 9.5a 11.4a 14.4a 35.3a 23.9a 46.6a ±b
(0.5) (3.7) (1.6) (4.7) (1.3) (7.0)
Arable soil + sugar-beet leaves
ÿIER
1.8 528.0 28.6
0
30.3
528 280
(1.4) (14.9) (11.7) (0) (10.5) (14.9) (46)
+ IER 29.6a 27.9a 42.3 232.0a 71.9a 260a ±b
(5.9) (10.2) (8.3) (44.6) (5.6) (34.8)
Grassland soil
ÿIER 28.4 124.0 15.1
0
43.5
124 166
(2.5) (30.1) (2.8) (0)
(5.1) (30.1) (12)
+ IER 22.8
23.7a 23.3a 74.1a 46.1 97.8
±b
(1.6) (1.6) (2.6) (0.9) (1.5) (1.9)
Fig. 1. Amount of NH4 ±N in the soil samples during the incubation
experiment. Bars indicate one standard deviation. r = Treatments
without IER; R = treatments with soil±IER mixtures. n = 3 for
each treatment; : treatments dier (Mann and Whitney U-test, P <
0.05).
t6
113
(18)
70a
(17)
73
(24)
40a
(5)
654
(69)
167a
(90)
133
(15)
141
(9)
a
Diers signi®cantly from the respective treatment without IER
(Mann and Whitney U-test, P < 0.05).
b
± data missing due to equipment failure.
n = 3 for each treatment.
J.K. Friedel et al. / Soil Biology & Biochemistry 32 (2000) 1529±1536
1533
concentrations occurred between the two treatments
(+IER or ÿIER).
In samples of the ``Grassland soil'', Nmin concentrations and N mineralisation rate were greater compared with ``Arable soil''. Signi®cant dierences in
Nmin concentrations occurred between the two treatments (ÿIER or +IER) only at 4 and 8 weeks of incubation.
In ``Arable soil + wheat straw'', the amount of
Nmin also was generally small in both +IER and
ÿIER treatments. Nmin values were signi®cantly higher
in +IER compared with ÿIER. In this treatment,
values decreased to zero during the incubation.
``Arable soil + sugar-beet leaves'' showed high mineralisation activity in both +IER and ÿIER treatments. The Nmin values were signi®cantly lower from 4
weeks until the end of the incubation in the +IER
compared with the ÿIER treatment.
incubation (12 weeks), Nmic contents were increased in
``Arable soil ÿ IER'' and slightly decreased in ``Grassland soil ÿ IER''. Addition of wheat straw and sugarbeet leaves immediately (16 h) increased Nmic contents
in ÿIER samples by a factor of approximately 2 and
4, respectively. Whereas in ``Arable soil + wheat straw
ÿ IER'', Nmic contents at 12 weeks had decreased to
the original concentrations found in ``Arable soil ÿ
IER'' at 16 h, in ``Arable soil + sugar-beet leaves
ÿIER'' values had further increased.
At the end of the incubation, in ``Arable soil +
IER'' and ``Grassland soil + IER'', Nmic values were
similar to the initial values of the soils (ÿIER) at 16 h.
In both, ``Arable soil + wheat straw + IER'' and
``Arable soil + sugar-beet leaves + IER'', Nmic contents were signi®cantly lower than in the respective ÿ
IER treatments.
3.3. IER eects on soil microbial biomass N
4. Discussion
Nmic contents were unrealistically low for +IER
samples at t0, although in a preliminary experiment
results of the chloroform fumigation extraction
method were not aected by mixing of IER into soil
samples (data not shown). Therefore, at 12 weeks 250
ml CHCl3 was added to each portion of soil sample
before fumigation to ensure complete fumigation.
Results of the chloroform fumigation method still
showed a high variability in both the soil + IER and
the soil + quartz mixtures and must be considered as
estimates and not exact values for Nmic contents.
Nmic contents at the beginning of the incubation (16
h) were smaller in ``Arable soil ÿ IER'' than in
``Grassland soil ÿ IER'' (Table 2). At the end of the
4.1. IER eect on losses of mineralised N and
nitri®cation
Fig. 3. Amount of Nmin in the soil samples during the incubation experiment. Bars indicate one standard deviation. r = Treatments
without IER; R = treatments with soil±IER mixtures. n = 3 for
each treatment; : treatments dier (Mann and Whitney U-test, P <
0.05).
A prerequisite for preventing losses of mineralised N
either due to denitri®cation or immobilisation during
the incubation of con®ned soil samples is to prevent
NH4 and NO3 from being transformed by soil microorganisms. No indication of removal or transformation of NH4 was found in pot incubation experiments
by Binkley (1984), who investigated NH4 adsorbed to
IER in resin bags at dierent rates of cellulose addition. NO3 adsorbed to IER contained in resin bags
was not transformed during 4 weeks exposure in the
®eld (HuÈbner et al., 1991). Over long deployment
times in the ®eld (up to 44 weeks), NO3 but not NH4,
was desorbed from preloaded IER (Giblin et al.,
1994).
In our experimental approach, IER and soil were
not separated before analysis. Thus, it cannot be determined directly, if NH4 or NO3 once adsorbed to IER
was removed or transformed during the further incubation period. However, the lack of nitri®cation in all
+IER treatments (Fig. 2) clearly indicates, that in
these treatments, in contrast to the ÿIER variants,
NH4 was not accessible to nitri®ers. The amounts of
NH4 in the +IER treatments either increased or
remained constant throughout the incubation in contrast to decreases in NH4 concentrations occurring in
the ÿIER variants (Fig. 1). This also indicates that no
transformation of NH4 occurred once it was adsorbed
to the IER. NO3 concentrations in the +IER samples
present at the beginning of the incubation were also
conserved until the end of the incubation (Table 2).
Therefore, we conclude that the mixture of strongly
acidic cation exchange resins and strongly basic anion
1534
J.K. Friedel et al. / Soil Biology & Biochemistry 32 (2000) 1529±1536
exchange resins mixed into the soil samples in our experiment can eectively adsorb mineralised N and prevent it from being further transformed by either
immobilisation, nitri®cation or denitri®cation. By separating IER from soil samples, e.g. by density fractionation (Thien and Myers, 1991), amounts of Nmin
adsorbed to the IER may be determined separately.
This separation procedure might also reduce the variability in determination of Nmic contents by the chloroform fumigation extraction method.
4.2. IER eects on net N mineralisation rates and
microbial biomass N
To test the eect of IER on net N mineralisation
rates, two soils diering in organic matter contents
and two organic substrates diering in C-to-N ratio
were chosen. Higher values of mineralised N in the
``Grassland soil'' than in the ``Arable soil'' (Fig. 3) can
be explained by higher organic matter contents of the
grassland topsoil (Table 1). Without substrate addition, IER had no eect on N mineralisation in both
soils (Fig. 3). Nmic contents slightly decreased in the
``Grassland soil'' (both ÿIER and +IER) during the
incubation (Table 2), indicating no eect of IER on N
in soil micro-organisms. The increase in Nmic contents
from 16 h to 12 weeks in the ``Arable soil ÿ IER''
treatment cannot be explained since no substrate containing easily available organic matter was added here.
We assume that this value may be overestimated due
to the heterogeneity of the samples. Nmic contents
remained unchanged in the ``Arable soil +IER'' treatment at 12 weeks compared with ``Arable soil ÿ IER''
at 16 h, also indicating a non-existing IER eect on N
in soil microorganisms.
IER eects on N mineralisation or Nmic contents
reported in most of the other studies (e.g. Binkley et
al., 1986; Zeller et al., 1997; Bhogal et al., 1999) are
not comparable with our experiments, because in such
investigations IER were used in resin bags and not
mixed into the soil. Only Saeed et al. (1994) used mixtures of soil with K+ saturated IER in anaerobic incubation experiments with lowland rice soils. Nitrogen
mineralisation was higher in IER±soil mixtures than in
control soil samples and the eect was greater in
strongly NH4-®xing soils. The authors concluded that
N mineralisation is underestimated by standard anaerobic incubations due to ®xation of mineralised
NH4. The contrasting eects of IER (without any substrate addition) on N mineralisation in our study is
probably due to the dierent experimental conditions
(NH4-®xing capacity of the soils, anaerobic versus
aerobic incubations).
Addition of wheat straw with a wide C-to-N ratio of
58 at a high rate (33 mg gÿ1) led to immobilisation of
inorganic N at the beginning of the incubation in the
``Arable soil + wheat straw ÿ IER'' treatment (Fig. 3,
Table 2). Throughout the incubation period, no remineralisation of N occurred. Nmic values at t0 16 h
were almost doubled compared with ``Arable soil ÿ
IER'' without addition of organic substrate (Table 2),
also indicating immediate N immobilisation. Addition
of straw to soil samples also resulted in N immobilisation for several weeks in incubation experiments of
Aoyama and Nozawa (1993) and Jensen (1997).
In the ``Arable Soil + wheat straw + IER'' treatment, however, IER prevented microbial transformation of mineralised N due to immobilisation (Fig. 3).
Throughout the incubation period, in ``Arable soil'' N
mineralisation after wheat straw addition was slightly
greater than in the treatment without substrate addition (Fig. 3, Table 2). Thus, in this soil sample, N
mineralisation from the soil organic matter was
obviously little aected by straw addition when N immobilisation was excluded by IER. This ®nding is in
accordance with the concept of dierent microbial
groups, living independently on dierent substrates in
soil (Cochran et al., 1988). Results also indicate that,
in the ``Arable soil + wheat straw ÿ IER'' treatment,
gross N mineralisation continued, but was overcompensated by a greater N immobilisation, leading to net
N immobilisation. In accordance with this assumption,
N mineralisation and N immobilisation have been
shown to occur simultaneously in soil by gross N mineralisation measurements (Sparling et al., 1995).
In ``Arable soil + sugar-beet leaves ÿ IER'', addition of sugar-beet leaves with a narrow C-to-N ratio
of 14 at a high rate (33 mg gÿ1) strongly enhanced N
mineralisation rates during the ®rst 6 weeks (Fig. 3)
and increased Nmic contents at 16 h and 12 weeks
(Table 2). In other studies, increases in N mineralisation (Aoyama and Nozawa, 1993; Aulakh et al.,
1995) and Nmic contents (Aoyana and Nozawa, 1993),
mostly after a short lag period, have been described
after addition of crop residues with a narrow C-to-N
ratio.
In the ``Arable soil + sugar-beet leaves + IER''
treatment, N mineralisation rates were dampened in
the presence of IER from week 4 until the end of the
incubation (Fig. 3). Signi®cantly reduced Nmic contents
at the end of the incubation (Table 2) indicate that
IER mixed into the soil prevented or strongly reduced
microbial N immobilisation after incorporation of
sugar-beet leaves, as in the variants with straw addition. The eect of IER on Nmin concentrations
(Table 2) and N mineralisation, however, was negative
in this case, in contrast to the positive eect observed
after wheat straw addition. We assume that IER
strongly reduced the N availability in the ``Arable soil
+ sugar-beet leaves + IER'' treatment after an initial
phase of the incubation, when N was probably beginning to be released from the micro-organisms and was
J.K. Friedel et al. / Soil Biology & Biochemistry 32 (2000) 1529±1536
re-used by the micro-¯ora in the ``Arable soil +
sugar-beet leaves ÿ IER'' treatment. In the +IER
treatment, this reduced N availability obviously limited
microbial growth (Nmic contents) and activity (N mineralisation rates). This eect of IER on N release, to
our knowledge, has not been described before.
Due to this, we have to modify our initial hypothesis
that net N mineralisation rates would be enhanced in
the presence of both IER and microbially-available
substrate. This increasing eect of IER in the presence
of available substrate was only found for substrate
with a wide C-to-N ratio (wheat straw, N-limited conditions), in accordance with the positive eect of growing roots (through NH4 uptake and deposition of Crich exudates) on net N mineralisation described by
Clarholm (1985). IER in the presence of substrate with
a low C-to-N ratio (sugar-beet leaves, conditions of
high N availability) diminished net N mineralisation
rates due to their limiting eect on Nmin availability
and consecutive microbial growth and activity.
4.3. Use of IER to approximate conditions in the
rhizosphere
Mixing IER into soil may alter soil physical conditions with respect to micro-sites of diering water
and aeration potential and change net N mineralisation rates in this way. First, conditions and N mineralisation rates in the disturbed samples may dier
from undisturbed conditions and N mineralisation
rates in situ. We tried to reduce this disturbing eect
by preserving natural soil aggregation. Secondly, we
assume that we could reduce artefacts due to mixing
IER into the soil to a minimum by mixing sand of a
grain size equivalent to that of the moistened resin
into the control soil samples (see Section 2.5).
IER mixed into the soil accumulate nutrients that
diuse through water ®lled pore spaces. Hence, they
may be used to assess plant-available nutrient quantity
as a function of diusion (Yang et al., 1991). IER also
mimic exchange properties of plant roots (Yavitt and
Wright, 1996). Consequently, we conclude that adsorption of Nmin to IER with a suciently high sorption
capacity can be regarded as a functional analogy to
the N uptake by roots.
IER used in our experiment had a speci®c surface
area in the range of 30±60 cm2cmÿ3 (calculated from a
mean diameter of 0.5±1.0 mm, assuming a spherical
shape, see Section 2.2). The speci®c surface area of
IER in the IER±soil mixture (1:1 by weight) therefore
is about 18±36 cm2 cmÿ3 (assuming a bulk density of
0.7 g cmÿ3 for IER, see Section 2.2 and 1.0 g cmÿ3 for
the soil). This is many times higher than the active
root area of herbaceous crop plants in soil (11 cm2
cmÿ3; Larcher, 1994, 21). Regarding the speci®c surface area and the sorption capacity of IER, they were
1535
added in surplus compared with the conditions in the
rhizosphere. We regard this useful because in soil±IER
mixtures there is no water ¯ow due to transpiration directed to the sink as in the rhizosphere. No depletion
of the sorption capacity of the IER was visible in our
experiment.
Our results indicate, depending on microbial N
availability, either positive (straw addition) or negative
(sugar-beet leaves addition) IER eects on net N mineralisation. Assuming a functional analogy of IER and
plant roots, this suggests that, by the combined eects
of N uptake and rhizodeposition, plant roots may
either enhance (under N-limited conditions) or diminish (under conditions of high N availability) net N
mineralisation.
4.4. Conclusions
In IER±soil mixtures, NH4 mineralised from organic
compounds is readily adsorbed by the IER due to its
intimate contact with the soil and short diusion ways.
Nitri®cation, accumulation and subsequent losses of
Nmin are prevented. Therefore, we suggest net N mineralisation rates may be quanti®ed more precisely by
IER±soil mixtures than by traditional methods (buried
bags, soil cores, resin cores).
IER may mimic sink functions of plant roots, and
nutrient sorption can be regarded as a functional analogy to nutrient uptake by roots. From this, we assume
that net N mineralisation rates of soil±IER mixtures
approximate conditions in the rhizosphere better than
results of traditional approaches, as long as soil water
and aeration conditions are not largely changed.
Due to this assumed functional analogy, soil±IER
mixtures may also be a useful model system to study
root-induced N mineralisation eects.
Acknowledgements
We would like to thank Corinna Kuûmaul, MartinLuther-University Halle-Wittenberg, and Irma Schumacher, University of Hohenheim, for their technical
assistance. We are indebted to Martin Kaupenjohann,
University of Hohenheim, for his contributions to the
experimental set-up and the interpretation of the
results, and Karl Stahr, University of Hohenheim, for
his readiness to ®nancially support the experiment.
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www.elsevier.com/locate/soilbio
Ion exchange resin±soil mixtures as a tool in net nitrogen
mineralisation studies
JuÈrgen K. Friedel a,*, Anke Herrmann b, Markus Kleber c
b
a
University of Agricultural Sciences, Institute of Organic Farming, Gregor-Mendel-Street 33, A-1180 Vienna, Austria
Department of Soil Sciences, Division of Soil Fertility and Plant Nutrition, Swedish University of Agricultural Sciences (SLU), S-75007 Uppsala,
Sweden
c
Martin-Luther University Halle-Wittenberg, Institute of Soil Science and Plant Nutrition, Weidenplan 14, D-06108 Halle, Germany
Accepted 29 February 2000
Abstract
Mixed-bed ion exchange resins (IER) were mixed with intact soil aggregates and incubated at 60% water ®lled pore space in
closed polyethylene bags for 12 weeks. To test IER eects on N losses, nitri®cation and net N mineralisation, an arable soil and
a grassland soil, diering in organic matter content, were chosen and two crop residues (wheat straw, sugar-beet leaves) with
dierent C-to-N ratios were added to the arable soil. It was proposed that IER might exert an in¯uence on N cycling similar to
that of plant roots. Nitri®cation was inhibited by adsorption of NH4 in the +IER treatments. Net N mineralisation was greater
in the grassland soil than in the arable soil which had less soil organic matter. Without incorporation of additional organic
substrates, net N mineralisation was not aected by IER in both soils. Straw addition to the arable soil caused immediate N
immobilisation in the ÿIER treatment, whereas N mineralisation continued in the +IER treatment. Incorporation of sugar-beet
leaves into the arable soil highly increased net N mineralisation and microbial biomass N in the ÿIER treatment. In the +IER
treatment, the enhancement of both N mineralisation and microbial biomass N was less pronounced. Thus, IER mixed into soil
samples can exert either a stimulating (wheat straw) or dampening (sugar-beet leaves) eect on N mineralisation. Soil±IER
mixtures can prevent losses and re-immobilisation of mineralised N and mimic nutrient exchange properties of plant roots. It is
concluded that in incubation experiments they can better re¯ect conditions in the vicinity of roots than incubations without IER
or with incorporation of IER in con®ned resin bags as long as water and aeration conditions are not largely changed. Soil±IER
mixtures may also be a useful tool for studying root-induced changes in net N mineralisation. 7 2000 Elsevier Science Ltd. All
rights reserved.
Keywords: Ion exchange resins; Soil±resin mixtures; Soil nitrogen cycling; Nitrogen mineralisation; Nitri®cation
1. Introduction
Quanti®cation of the relationships between soil N
mineralisation and other ecosystem processes has been
restricted by the lack of suitable methods to measure
N mineralisation under ®eld conditions. Methods used
to measure or estimate patterns of N mineralisation
under ®eld conditions include: (i) exposure of dis-
* Corresponding author. Fax: +43-1-47654-3792.
E-mail address: friedel@edv1.boku.ac.at (J.K. Friedel).
turbed soil in plastic bags buried in the ®eld, (ii) exposure of relatively undisturbed soil columns under
®eld conditions, and (iii) measurement of mineral N
collected by ion exchange resins placed in the ®eld for
extended periods (Raison et al., 1987).
All methods of containment developed to date alter
the soil environment through (i) cessation of the carbon (C) input from decomposing litter and from ®ne
root turnover, (ii) increased C inputs from severed
roots, (iii) modi®cation of the moisture and temperature regimes relative to bulk soil, and (iv) accumulation of inorganic N (Adams et al., 1989). An
0038-0717/00/$ - see front matter 7 2000 Elsevier Science Ltd. All rights reserved.
PII: S 0 0 3 8 - 0 7 1 7 ( 0 0 ) 0 0 0 6 4 - X
5.9
5.2
0
0
8.7
10.2
0.15
0.46
1.3
4.7
45.0
59.5
a
b
Horizon designators according to A.G. Boden (1994).
Textural classes according to USDA (1996).
1.4
1.0
0
0
Silt loam
Clay loam
Ap
Ah
Arable
Grassland
0-30
0-10
Ccarbonate (%)
C/N
Nt (%)
TOC (%)
Total pore space (vol%)
Bulk density (g cmÿ3)
Rock fragments (vol%)
Textureb
Depth (cm)
Horizona
Soil
additional source of error is the inability to prevent
mineralised N (Nmin) from being either re-immobilised
into microbial biomass or denitri®ed within the containment period. As assay conditions alter the C and
N availability within the containers, none of the
methods may be considered to measure mineralisation
rates accurately.
The use of intact soil cores (Nordmeyer and Richter,
1985; Raison et al., 1987) may reduce the sources of
error due to disturbance of the soil and modi®cations
of the moisture and temperature regimes. The central
problem however, the absence of living root functions,
remains.
In undisturbed soil inhabited by living plants, root
N uptake reduces or even inhibits denitri®cation or reimmobilisation of mineralised N and limits microbial
N availability. Simultaneously occurring rhizodeposition leads to an increase in the amount of substrate
available to soil micro-organisms, enabling them to
mineralise ``surplus'' amounts of organic N. Consecutive predation by soil fauna may release part of this
extra N as NH4 (Clarholm, 1985).
Any attempt to advance the methodology for studying ¯uxes of soil mineral-N in situ must therefore seek
to ®nd a means which takes root functions into
account. Such a procedure should immobilise inorganic N throughout the incubation, thus preventing it
from being nitri®ed, denitri®ed or re-immobilised into
microbial biomass.
If this process occurs with the same magnitude as is
performed by living roots, it should eliminate the most
prominent source of error.
IER were shown to adsorb NH4 and NO3 eectively
in soils (Binkley, 1984). Only few attempts (e.g. HuÈbner et al., 1991; Binkley et al., 1986; DiStefano and
Gholz, 1986) to establish the use of IER as a procedure to measure net N mineralisation were known
when growing concern about the reliability of the buried bag method (Eno, 1960) led Zeller et al. (1997) and
Bhogal et al. (1999) to investigate further into the subject. In all previous attempts to employ IER as a tool
in net nitrogen mineralisation studies, these were
applied in ``resin bags'' to the soil. Such design may
prevent Nmin from being leached out of cores as well
as from being washed into them. Due to the low mobility of NH4 in soil, however, nitri®cation and re-immobilisation of NH4 cannot be prevented. Thus, with
resin bags of traditional design, Nmin may continue to
accumulate in soil samples during the incubation.
We assumed that by mixing IER thoroughly into
soil samples, the accumulation of microbially-available
mineralised N in soil samples can be prevented and
one of the most important eects of living roots on
soil N transformations (i.e. the removal of Nmin) may
be simulated. Conditions in soil samples containing
IER can therefore be expected to be closer to con-
pH, CaCl2
J.K. Friedel et al. / Soil Biology & Biochemistry 32 (2000) 1529±1536
Table 1
Selected soil properties
1530
J.K. Friedel et al. / Soil Biology & Biochemistry 32 (2000) 1529±1536
ditions in the vicinity of roots than conditions in
samples without IER. For this reason, N mineralisation rates measured in soil samples containing IER
should be a more realistic measure for N mineralisation rates under ®eld conditions.
To achieve this, we tested (i) if mixing IER into soil
samples can reduce losses of Nmin either by denitri®cation or immobilisation, (ii) whether nitri®cation can be
reduced or even prevented by IER in soil samples, and
(iii) if net N mineralisation rates are enhanced in the
presence of both IER and microbially-available substrate.
2. Materials and methods
2.1. Soil sampling
Soil samples were taken from the A horizons of an
arable soil (Stagnic Luvisol; ``Arable soil'') and a
humus-rich grassland soil (Cumulic Anthrosol; ``Grassland soil''). Basic soil properties are given in Table 1.
In both cases, a surface area of 1 m2 was cleared of
plant materials and the soil subsequently sampled
down to the horizon boundary. The soil was then
freed from root residues and homogenised. This was
done with maximum care to preserve crumbs and
aggregates. As the material was stone-free, no further
sieving was necessary until preparation of individual
incubates.
2.2. Ion exchange resin (IER)
When both cations and anions are included as target
ions, the various species can be accumulated by the
resin if a mixture of cation- and anion-resins are
placed into the medium (Skogley and Dobermann,
1996). Consequently, a mixed-bed resin (Amberlite1
MB-3) from CWG GmbH, Mannheim, Germany, was
used. Amberlite1 MB-3 is a mixture of the strong cation exchange resin Amberlite1 IR-120 and the strong
anion exchange resin Amberlite1 IRA-420 Type 1.
Grain size of the moist resin was determined to be in
the range 0.3±1.2 mm, with most of the grains within
0.5 and 1.0 mm. The speci®c weight was 0.7 g cmÿ3.
Prior to the experiment, IER was washed once with
deionised water. By this treatment, the N content
extractable with 2 M KCl (soil : extractant ratio=1 :
20) was reduced to about 50% (data not shown).
2.3. Incubation experiment
To
rates
ganic
poor
test the in¯uence of IER on N mineralisation
under dierent conditions, soils diering in ormatter content and an addition of N-rich and Nsubstrates were chosen. The treatments were:
1531
``Arable soil ÿ IER'' (low organic matter), ``Arable
soil + IER'', ``Grassland soil ÿ IER'' (high organic
matter), ``Grassland soil + IER'', ``Arable soil +
wheat straw ÿ IER'', ``Arable soil + wheat straw +
IER'', ``Arable soil + sugar-beet leaves ÿ IER'',
``Arable soil + sugar-beet leaves + IER''.
2.4. Preparation of organic additives
Wheat straw and sugar-beet leaves were dried at
508C for 72 h and ground to pass a 1 mm sieve. After
this, wheat straw had a water content of 7.7% and
sugar-beet leaves of 16.2%. Wheat straw had a C content of 409 mg gÿ1 and an N content of 7 mg gÿ1,
yielding a C-to-N ratio of 58. C and N contents in
sugar-beet leaves were 397 and 28.3 mg gÿ1; which
resulted in a C-to-N ratio of 14. Organic additives
were added to soil at a 1:30 ratio by weight.
2.5. Preparation of individual incubates
Polyethylene freezer bags manufactured by Fa.
Haaf, 85716 Lohhof, Germany, were used as containers for the soil. Bags had an average standard thickness of 50 mm. Polyethylene ®lm is only slightly
permeable to water vapour, while its O2 and CO2
transmission rates are relatively high. Soil structure in
the samples was not disturbed, i.e. complete aggregates
were incubated. Soil sample size was 300 g. The same
amount of resin was added in the treatments with IER
addition. As the density of quartz is about double the
density of the resin selected, 600 g of quartz sand of a
grain size similar to that of the resin were added to
treatments without IER in order to maintain identical
grain size distribution and soil physical conditions.
For each treatment, 21 replicate samples were prepared and incubated at 158C and at 60% water ®lled
pore space. Three samples per treatment were selected
the next day (t0 = 16 h) at random and analysed to
determine starting values. Every 2 weeks, another
three samples per treatment were selected at random
from the incubator and analysed in the same manner
to represent a total of 7 (t0 . . .t6) data points spanning
incubation periods from 0 to 12 weeks in 2-week increments tn1 ÿ tn 2 weeks). Microbial biomass N
(Nmic) was determined at t0 and t6 (in the beginning of
the experiment and after 12 weeks).
2.6. Determination of inorganic N and microbial N
Inorganic N was extracted by 2 M KCl solution
(extractant : soil ratio = 40:1) and measured by means
of a Skalar autoanalyser. NH4 was determined colorimetrically by a modi®ed Berthelot reaction according
to Krom (1980) and NO3 after reduction to NO2 by
the Griess reaction as a-naphthylamine-para-diazoben-
1532
J.K. Friedel et al. / Soil Biology & Biochemistry 32 (2000) 1529±1536
zene-parasulphuric acid at 540 nm in an automated
analyser (Houba et al., 1987).
Microbial biomass N was measured with the slightly
modi®ed standard fumigation-extraction method
(Brookes et al., 1985). Brie¯y, soil samples (20 g for
IER±soil incubates and 30 g for quartz±soil incubates,
equivalent to 10 g of wet soil each) were fumigated
with CHCl3 for 24 h at 258C. For analysis at t6, 250 ml
liquid CHCl3 was added to each portion of the soil
samples before fumigation, according to the method
recommended by Mueller et al. (1992) for soil samples
with a high moisture content. After removal of the
CHCl3, soluble N was extracted from fumigated and
unfumigated samples with 2 M KCl (extractant:soil
ratio = 40:1) for 2 h. N in ®ltrated solution was determined by the Griess reaction colorimetrically after an
automatic UV peroxide digestion to NO3 and reduction to NO2 by means of a Skalar autoanalyser
(Houba et al., 1987). Nmic was calculated using a kEN
factor of 0.54.
Fig. 2. Amount of NO3±N in the soil samples during the incubation
experiment. Bars indicate one standard deviation. r = Treatments
without IER, R = treatments with soil±IER mixtures. n = 3 for
each treatment. : treatments dier (Mann and Whitney U-test, P <
0.05).
3.2. IER eects on net N-mineralisation
3. Results
3.1. IER eects on nitri®cation
In ÿIER samples, mineralised N was readily nitri®ed (Figs. 1 and 2). At the end of the incubation (12
weeks) all Nmin was nitri®ed to NO3 (Table 2). In
+IER samples, NO3 concentrations found at t0
remained constant throughout the incubation (Fig. 2,
Table 2). Mineralisation occurring during the incubation merely increased the NH4 pool.
In the ``Arable soil'' without substrate addition,
Nmin accumulated gradually at a low rate during the
incubation (Fig. 3). No signi®cant dierences in Nmin
Table 2
Inorganic N and soil microbial biomass N (mg gÿ1) in the dierent
soil treatments with or without IER (standard deviation in parentheses) after 16 h (t0) and 12 weeks (t6)
NO3 ±N
NH+
4 ±N
Inorganic N
Microbial N
t0
t0
t0
t0
t6
t6
t6
Arable soil
ÿIER
8.5
38.6
0.4
0
8.9 38.6
67
(0.7) (7.1) (0.6) (0)
(0.4) (7.1) (13)
+ IER 7.9
9.7a
3.3a 24.3a 11.1 34.0
±b
(1.2) (2.5) (2.1) (3.1) (1.8) (5.5)
Arable soil + wheat straw
ÿIER
2.8
0
3.7
0
6.5
0
123
(0.7) (0)
(3.8) (0)
(3.1) (0)
(2)
+ IER 9.5a 11.4a 14.4a 35.3a 23.9a 46.6a ±b
(0.5) (3.7) (1.6) (4.7) (1.3) (7.0)
Arable soil + sugar-beet leaves
ÿIER
1.8 528.0 28.6
0
30.3
528 280
(1.4) (14.9) (11.7) (0) (10.5) (14.9) (46)
+ IER 29.6a 27.9a 42.3 232.0a 71.9a 260a ±b
(5.9) (10.2) (8.3) (44.6) (5.6) (34.8)
Grassland soil
ÿIER 28.4 124.0 15.1
0
43.5
124 166
(2.5) (30.1) (2.8) (0)
(5.1) (30.1) (12)
+ IER 22.8
23.7a 23.3a 74.1a 46.1 97.8
±b
(1.6) (1.6) (2.6) (0.9) (1.5) (1.9)
Fig. 1. Amount of NH4 ±N in the soil samples during the incubation
experiment. Bars indicate one standard deviation. r = Treatments
without IER; R = treatments with soil±IER mixtures. n = 3 for
each treatment; : treatments dier (Mann and Whitney U-test, P <
0.05).
t6
113
(18)
70a
(17)
73
(24)
40a
(5)
654
(69)
167a
(90)
133
(15)
141
(9)
a
Diers signi®cantly from the respective treatment without IER
(Mann and Whitney U-test, P < 0.05).
b
± data missing due to equipment failure.
n = 3 for each treatment.
J.K. Friedel et al. / Soil Biology & Biochemistry 32 (2000) 1529±1536
1533
concentrations occurred between the two treatments
(+IER or ÿIER).
In samples of the ``Grassland soil'', Nmin concentrations and N mineralisation rate were greater compared with ``Arable soil''. Signi®cant dierences in
Nmin concentrations occurred between the two treatments (ÿIER or +IER) only at 4 and 8 weeks of incubation.
In ``Arable soil + wheat straw'', the amount of
Nmin also was generally small in both +IER and
ÿIER treatments. Nmin values were signi®cantly higher
in +IER compared with ÿIER. In this treatment,
values decreased to zero during the incubation.
``Arable soil + sugar-beet leaves'' showed high mineralisation activity in both +IER and ÿIER treatments. The Nmin values were signi®cantly lower from 4
weeks until the end of the incubation in the +IER
compared with the ÿIER treatment.
incubation (12 weeks), Nmic contents were increased in
``Arable soil ÿ IER'' and slightly decreased in ``Grassland soil ÿ IER''. Addition of wheat straw and sugarbeet leaves immediately (16 h) increased Nmic contents
in ÿIER samples by a factor of approximately 2 and
4, respectively. Whereas in ``Arable soil + wheat straw
ÿ IER'', Nmic contents at 12 weeks had decreased to
the original concentrations found in ``Arable soil ÿ
IER'' at 16 h, in ``Arable soil + sugar-beet leaves
ÿIER'' values had further increased.
At the end of the incubation, in ``Arable soil +
IER'' and ``Grassland soil + IER'', Nmic values were
similar to the initial values of the soils (ÿIER) at 16 h.
In both, ``Arable soil + wheat straw + IER'' and
``Arable soil + sugar-beet leaves + IER'', Nmic contents were signi®cantly lower than in the respective ÿ
IER treatments.
3.3. IER eects on soil microbial biomass N
4. Discussion
Nmic contents were unrealistically low for +IER
samples at t0, although in a preliminary experiment
results of the chloroform fumigation extraction
method were not aected by mixing of IER into soil
samples (data not shown). Therefore, at 12 weeks 250
ml CHCl3 was added to each portion of soil sample
before fumigation to ensure complete fumigation.
Results of the chloroform fumigation method still
showed a high variability in both the soil + IER and
the soil + quartz mixtures and must be considered as
estimates and not exact values for Nmic contents.
Nmic contents at the beginning of the incubation (16
h) were smaller in ``Arable soil ÿ IER'' than in
``Grassland soil ÿ IER'' (Table 2). At the end of the
4.1. IER eect on losses of mineralised N and
nitri®cation
Fig. 3. Amount of Nmin in the soil samples during the incubation experiment. Bars indicate one standard deviation. r = Treatments
without IER; R = treatments with soil±IER mixtures. n = 3 for
each treatment; : treatments dier (Mann and Whitney U-test, P <
0.05).
A prerequisite for preventing losses of mineralised N
either due to denitri®cation or immobilisation during
the incubation of con®ned soil samples is to prevent
NH4 and NO3 from being transformed by soil microorganisms. No indication of removal or transformation of NH4 was found in pot incubation experiments
by Binkley (1984), who investigated NH4 adsorbed to
IER in resin bags at dierent rates of cellulose addition. NO3 adsorbed to IER contained in resin bags
was not transformed during 4 weeks exposure in the
®eld (HuÈbner et al., 1991). Over long deployment
times in the ®eld (up to 44 weeks), NO3 but not NH4,
was desorbed from preloaded IER (Giblin et al.,
1994).
In our experimental approach, IER and soil were
not separated before analysis. Thus, it cannot be determined directly, if NH4 or NO3 once adsorbed to IER
was removed or transformed during the further incubation period. However, the lack of nitri®cation in all
+IER treatments (Fig. 2) clearly indicates, that in
these treatments, in contrast to the ÿIER variants,
NH4 was not accessible to nitri®ers. The amounts of
NH4 in the +IER treatments either increased or
remained constant throughout the incubation in contrast to decreases in NH4 concentrations occurring in
the ÿIER variants (Fig. 1). This also indicates that no
transformation of NH4 occurred once it was adsorbed
to the IER. NO3 concentrations in the +IER samples
present at the beginning of the incubation were also
conserved until the end of the incubation (Table 2).
Therefore, we conclude that the mixture of strongly
acidic cation exchange resins and strongly basic anion
1534
J.K. Friedel et al. / Soil Biology & Biochemistry 32 (2000) 1529±1536
exchange resins mixed into the soil samples in our experiment can eectively adsorb mineralised N and prevent it from being further transformed by either
immobilisation, nitri®cation or denitri®cation. By separating IER from soil samples, e.g. by density fractionation (Thien and Myers, 1991), amounts of Nmin
adsorbed to the IER may be determined separately.
This separation procedure might also reduce the variability in determination of Nmic contents by the chloroform fumigation extraction method.
4.2. IER eects on net N mineralisation rates and
microbial biomass N
To test the eect of IER on net N mineralisation
rates, two soils diering in organic matter contents
and two organic substrates diering in C-to-N ratio
were chosen. Higher values of mineralised N in the
``Grassland soil'' than in the ``Arable soil'' (Fig. 3) can
be explained by higher organic matter contents of the
grassland topsoil (Table 1). Without substrate addition, IER had no eect on N mineralisation in both
soils (Fig. 3). Nmic contents slightly decreased in the
``Grassland soil'' (both ÿIER and +IER) during the
incubation (Table 2), indicating no eect of IER on N
in soil micro-organisms. The increase in Nmic contents
from 16 h to 12 weeks in the ``Arable soil ÿ IER''
treatment cannot be explained since no substrate containing easily available organic matter was added here.
We assume that this value may be overestimated due
to the heterogeneity of the samples. Nmic contents
remained unchanged in the ``Arable soil +IER'' treatment at 12 weeks compared with ``Arable soil ÿ IER''
at 16 h, also indicating a non-existing IER eect on N
in soil microorganisms.
IER eects on N mineralisation or Nmic contents
reported in most of the other studies (e.g. Binkley et
al., 1986; Zeller et al., 1997; Bhogal et al., 1999) are
not comparable with our experiments, because in such
investigations IER were used in resin bags and not
mixed into the soil. Only Saeed et al. (1994) used mixtures of soil with K+ saturated IER in anaerobic incubation experiments with lowland rice soils. Nitrogen
mineralisation was higher in IER±soil mixtures than in
control soil samples and the eect was greater in
strongly NH4-®xing soils. The authors concluded that
N mineralisation is underestimated by standard anaerobic incubations due to ®xation of mineralised
NH4. The contrasting eects of IER (without any substrate addition) on N mineralisation in our study is
probably due to the dierent experimental conditions
(NH4-®xing capacity of the soils, anaerobic versus
aerobic incubations).
Addition of wheat straw with a wide C-to-N ratio of
58 at a high rate (33 mg gÿ1) led to immobilisation of
inorganic N at the beginning of the incubation in the
``Arable soil + wheat straw ÿ IER'' treatment (Fig. 3,
Table 2). Throughout the incubation period, no remineralisation of N occurred. Nmic values at t0 16 h
were almost doubled compared with ``Arable soil ÿ
IER'' without addition of organic substrate (Table 2),
also indicating immediate N immobilisation. Addition
of straw to soil samples also resulted in N immobilisation for several weeks in incubation experiments of
Aoyama and Nozawa (1993) and Jensen (1997).
In the ``Arable Soil + wheat straw + IER'' treatment, however, IER prevented microbial transformation of mineralised N due to immobilisation (Fig. 3).
Throughout the incubation period, in ``Arable soil'' N
mineralisation after wheat straw addition was slightly
greater than in the treatment without substrate addition (Fig. 3, Table 2). Thus, in this soil sample, N
mineralisation from the soil organic matter was
obviously little aected by straw addition when N immobilisation was excluded by IER. This ®nding is in
accordance with the concept of dierent microbial
groups, living independently on dierent substrates in
soil (Cochran et al., 1988). Results also indicate that,
in the ``Arable soil + wheat straw ÿ IER'' treatment,
gross N mineralisation continued, but was overcompensated by a greater N immobilisation, leading to net
N immobilisation. In accordance with this assumption,
N mineralisation and N immobilisation have been
shown to occur simultaneously in soil by gross N mineralisation measurements (Sparling et al., 1995).
In ``Arable soil + sugar-beet leaves ÿ IER'', addition of sugar-beet leaves with a narrow C-to-N ratio
of 14 at a high rate (33 mg gÿ1) strongly enhanced N
mineralisation rates during the ®rst 6 weeks (Fig. 3)
and increased Nmic contents at 16 h and 12 weeks
(Table 2). In other studies, increases in N mineralisation (Aoyama and Nozawa, 1993; Aulakh et al.,
1995) and Nmic contents (Aoyana and Nozawa, 1993),
mostly after a short lag period, have been described
after addition of crop residues with a narrow C-to-N
ratio.
In the ``Arable soil + sugar-beet leaves + IER''
treatment, N mineralisation rates were dampened in
the presence of IER from week 4 until the end of the
incubation (Fig. 3). Signi®cantly reduced Nmic contents
at the end of the incubation (Table 2) indicate that
IER mixed into the soil prevented or strongly reduced
microbial N immobilisation after incorporation of
sugar-beet leaves, as in the variants with straw addition. The eect of IER on Nmin concentrations
(Table 2) and N mineralisation, however, was negative
in this case, in contrast to the positive eect observed
after wheat straw addition. We assume that IER
strongly reduced the N availability in the ``Arable soil
+ sugar-beet leaves + IER'' treatment after an initial
phase of the incubation, when N was probably beginning to be released from the micro-organisms and was
J.K. Friedel et al. / Soil Biology & Biochemistry 32 (2000) 1529±1536
re-used by the micro-¯ora in the ``Arable soil +
sugar-beet leaves ÿ IER'' treatment. In the +IER
treatment, this reduced N availability obviously limited
microbial growth (Nmic contents) and activity (N mineralisation rates). This eect of IER on N release, to
our knowledge, has not been described before.
Due to this, we have to modify our initial hypothesis
that net N mineralisation rates would be enhanced in
the presence of both IER and microbially-available
substrate. This increasing eect of IER in the presence
of available substrate was only found for substrate
with a wide C-to-N ratio (wheat straw, N-limited conditions), in accordance with the positive eect of growing roots (through NH4 uptake and deposition of Crich exudates) on net N mineralisation described by
Clarholm (1985). IER in the presence of substrate with
a low C-to-N ratio (sugar-beet leaves, conditions of
high N availability) diminished net N mineralisation
rates due to their limiting eect on Nmin availability
and consecutive microbial growth and activity.
4.3. Use of IER to approximate conditions in the
rhizosphere
Mixing IER into soil may alter soil physical conditions with respect to micro-sites of diering water
and aeration potential and change net N mineralisation rates in this way. First, conditions and N mineralisation rates in the disturbed samples may dier
from undisturbed conditions and N mineralisation
rates in situ. We tried to reduce this disturbing eect
by preserving natural soil aggregation. Secondly, we
assume that we could reduce artefacts due to mixing
IER into the soil to a minimum by mixing sand of a
grain size equivalent to that of the moistened resin
into the control soil samples (see Section 2.5).
IER mixed into the soil accumulate nutrients that
diuse through water ®lled pore spaces. Hence, they
may be used to assess plant-available nutrient quantity
as a function of diusion (Yang et al., 1991). IER also
mimic exchange properties of plant roots (Yavitt and
Wright, 1996). Consequently, we conclude that adsorption of Nmin to IER with a suciently high sorption
capacity can be regarded as a functional analogy to
the N uptake by roots.
IER used in our experiment had a speci®c surface
area in the range of 30±60 cm2cmÿ3 (calculated from a
mean diameter of 0.5±1.0 mm, assuming a spherical
shape, see Section 2.2). The speci®c surface area of
IER in the IER±soil mixture (1:1 by weight) therefore
is about 18±36 cm2 cmÿ3 (assuming a bulk density of
0.7 g cmÿ3 for IER, see Section 2.2 and 1.0 g cmÿ3 for
the soil). This is many times higher than the active
root area of herbaceous crop plants in soil (11 cm2
cmÿ3; Larcher, 1994, 21). Regarding the speci®c surface area and the sorption capacity of IER, they were
1535
added in surplus compared with the conditions in the
rhizosphere. We regard this useful because in soil±IER
mixtures there is no water ¯ow due to transpiration directed to the sink as in the rhizosphere. No depletion
of the sorption capacity of the IER was visible in our
experiment.
Our results indicate, depending on microbial N
availability, either positive (straw addition) or negative
(sugar-beet leaves addition) IER eects on net N mineralisation. Assuming a functional analogy of IER and
plant roots, this suggests that, by the combined eects
of N uptake and rhizodeposition, plant roots may
either enhance (under N-limited conditions) or diminish (under conditions of high N availability) net N
mineralisation.
4.4. Conclusions
In IER±soil mixtures, NH4 mineralised from organic
compounds is readily adsorbed by the IER due to its
intimate contact with the soil and short diusion ways.
Nitri®cation, accumulation and subsequent losses of
Nmin are prevented. Therefore, we suggest net N mineralisation rates may be quanti®ed more precisely by
IER±soil mixtures than by traditional methods (buried
bags, soil cores, resin cores).
IER may mimic sink functions of plant roots, and
nutrient sorption can be regarded as a functional analogy to nutrient uptake by roots. From this, we assume
that net N mineralisation rates of soil±IER mixtures
approximate conditions in the rhizosphere better than
results of traditional approaches, as long as soil water
and aeration conditions are not largely changed.
Due to this assumed functional analogy, soil±IER
mixtures may also be a useful model system to study
root-induced N mineralisation eects.
Acknowledgements
We would like to thank Corinna Kuûmaul, MartinLuther-University Halle-Wittenberg, and Irma Schumacher, University of Hohenheim, for their technical
assistance. We are indebted to Martin Kaupenjohann,
University of Hohenheim, for his contributions to the
experimental set-up and the interpretation of the
results, and Karl Stahr, University of Hohenheim, for
his readiness to ®nancially support the experiment.
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