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Soil Biology & Biochemistry 32 (2000) 449±456
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Elevated atmospheric carbon dioxide concentration: e€ects of
increased carbon input in a Lolium perenne soil on
microorganisms and decomposition
J.H. van Ginkel, A. Gorissen*, D. Polci
Plant Research International, P.O. Box 16, 6700 AA Wageningen, Netherlands
Accepted 28 May 1999

Abstract
E€ects of ambient and elevated atmospheric CO2 concentrations (350 and 700 ml lÿ1) on net carbon input into soil, the
production of root-derived material and the subsequent microbial transformation were investigated. Perennial ryegrass plants (L.
perenne L.) were labelled in a continuously labelled 14C-CO2 atmosphere to follow carbon ¯ow through the plant and all soil
compartments. After 115 days, root biomass was 41% greater at elevated CO2 than at ambient CO2 and this root biomass
seemed to be the driving force for the increase of 14C-labelled carbon in all compartments examined, i.e. carbon in the soil
solution, soil microbial biomass and soil residue. After incubation for 230 days at 148C, roots grown at elevated CO2
decomposed slower (14%) than roots grown at ambient CO2. Increasing the incubation temperature of the roots grown at
elevated CO2 by 28C could not compensate for this delay in decomposition. In addition, `elevated CO2' root-derived material
(14C-labelled soil microorganisms plus 14C-labelled soil residue) decomposed signi®cantly slower (29%) than `ambient CO2 ' rootderived material. At the end of the incubation experiment, the ratio between 14C-labelled microorganisms and total 14CO2
evolved showed no di€erence among root incubation and incubation of root-derived material. Thus, the substrate use eciency

of microorganisms, involved with decomposition of roots and root-derived material, seems not to be a€ected by an increase in
atmospheric CO2 concentrations. Therefore, the lower decomposition rate at elevated CO2 is not due to a change in the internal
metabolism of microorganisms. 7 2000 Published by Elsevier Science Ltd. All rights reserved.

1. Introduction
Atmospheric CO2 levels are increasing steadily
(Keeling et al., 1995), mainly due to combustion of
fossil fuels and deforestation. Increasing CO2 levels are
expected to have numerous direct and indirect e€ects
on terrestrial ecosystems (Bazzaz, 1990). Among those
e€ects, changes in above-ground primary production
and litter fall are relatively easy to assess, but changes
in carbon allocation below-ground are more dicult to
determine. Yet, this process is thought to be of great
importance both for the functioning of terrestrial eco-

* Corresponding author. Tel.: +31-317-475-846; fax: +31-317-423110.
E-mail address: a.gorissen@plant.wag-ur.nl (A. Gorissen).

systems and for carbon sequestering. Most intriguing

is the impact of root growth on soil microorganisms
and associated transformation processes. After all,
microorganisms play an essential role in the cycling of
nutrients associated with primary production (Paul
and Clark, 1989; Killham, 1994) and this largely determines the overall ecosystem response. To date, the response of the soil microorganisms to elevated
atmospheric CO2 concentrations is poorly understood
(O'Neill, 1994). Newton et al. (1995) and Ross et al.
(1995) found no change in microbial biomass in soils
with plants grown elevated CO2. Rillig et al. (1997)
measured di€erent carbon-substrate utilisation of rhizosphere extracts applied to Biolog microplates with
fewer polymers oxidised at elevated CO2. Because only
a minority of the microbial community is active, while
the majority is dormant and forms a high background

0038-0717/00/$ - see front matter 7 2000 Published by Elsevier Science Ltd. All rights reserved.
PII: S 0 0 3 8 - 0 7 1 7 ( 9 9 ) 0 0 0 9 7 - 8

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J.H. van Ginkel et al. / Soil Biology & Biochemistry 32 (2000) 449±456

noise when measuring soil microbial biomass, responses of soil microorganisms to elevated CO2 are
hard to measure. Therefore, the use of carbon tracers
such as 14C or 13C is a good tool to distinguish
between already present native-soil organic carbon and
incoming plant-derived carbon in the soil and to
measure soil microorganisms actively involved in the
transformation of either of the carbon sources.
To study the metabolic behaviour of soil microorganisms, we calculated the ratio between the 14Clabelled soil microbial biomass and total 14CO2
evolved at the end of an incubation with roots and
root-derived material originated from ambient and elevated CO2. What consequences a temperature
increase accompanying the raise of atmospheric CO2
will have on microbial behaviour and subsequently on
decomposition of root material grown at elevated CO2
is not clear yet. Gorissen et al. (1995) found that slow
decomposition of grass roots grown at elevated CO2
could be compensated by incubation at a 68C higher
temperature. However, the projected 28C increase with
atmospheric CO2 levels rising from 350 to 700 ml lÿ1
(IPPC, 1995) may make soil a sink for atmospheric

CO2.
In this study, grass plants (L. perenne ) were grown
and homogeneously 14C-labelled at two di€erent 14CCO2concentrations (350 and 700 ml lÿ1). Our objectives
were to determine the e€ects of elevated atmospheric
CO2 concentrations on: (i) possible changes in behaviour of the soil microbial biomass due to altered
below-ground 14C-carbon input and (ii) the in¯uence
of such changes on decomposition of 14C-roots, incubated at two di€erent temperatures.

2. Materials and methods

mosphere (speci®c activity 0.70 kBq mgÿ1 C) with
either CO2 concentrations of 350 or 700 ml lÿ1. The
plants were grown under the following conditions:
light 16h dayÿ1; PAR (Photosynthetically Active Radiation) 400 mmol mÿ2 sÿ1; temperature 18/148C (day/
night); relative humidity 65/70% (day/night) and wind
speed 0.1 m sÿ1. All environmental variables were
checked with a third independent meter to ensure identical conditions in the growth chambers. During the
experiment, soil water was kept at about 14% w/w
(60±70% of ®eld capacity) by adjusting to a predetermined weight adjusted for the growth of the plants.
The mineral N content (KCl extractable NH+

4 and
NOÿ
3 ) of the soil at the start of the experiment was
10.5 mg N gÿ1 dry soil. In addition, at the start of the
experiment all soil containers received phosphorus and
potassium at a concentration of 29 and 78 mg gÿ1, respectively. The shoots were cut after 49, 66, 83 and
115 days.
On day 115, the plants were removed from the
growth chambers and the shoots and any stubble were
cut o€ at ground level. The roots and soil were separated by gently shaking the soil-root core and the
remaining roots were removed from the soil by handpicking. Subsequently, the root material was rinsed
with tap water to remove adhering soil particles.
Shoots, roots and soil were dried (708C) and ground (1
mm) and dry weight, total carbon and 14C-carbon
were determined. Roots were also analysed for total
nitrogen.
Plant weight at the time of transfer to ESPAS and
the start of 14C-labelling (day 32) was less than 1% of
plant weight at the end of the experiment (data not
shown) and was therefore assumed negligible. Before

and after analysis, the soils of the harvested plants
were kept at 48C to minimise microbial activity, pending the incubation experiment.

2.1. Plant and soil
2.2. Incubation experiment
We used L. perenne L. cv. `Barlet' and fresh soil
from the top layer (20 cm) of an arable loamy sand
soil. Soil properties were: particle size distribution 3%
< 2 mm, 12% 2±50 mm and 85% > 50 mm; organic
carbon 1.7%; total nitrogen 0.10%; pH 6.4.
Grass seeds were germinated on moist paper for two
days in the dark and transferred to 50 ml pots containing 70 g of soil. Half of the seedlings were grown for
30 days in a greenhouse at 350 ml lÿ1 CO2 and the
other half at 700 ml lÿ1 CO2. Subsequently, eight plants
were transferred to 1150 ml polythene soil containers
(one plant per container) ®lled with 1745 g of moistened soil with a bulk density of 1.3 g cmÿ3. (dry
weight basis). These containers were transferred to the
ESPAS (Experimental Soil Plant Atmosphere System;
Gorissen et al., 1996) growth chambers and the four
plants were grown in a continuously labelled 14CO2 at-

Soil from the pots of the harvested plants was incubated: (1) 60 g of root-free 14C-labelled soil and, (2) as
(1) but mixed with the dried and ground 14C-labelled
roots in the same root/soil ratio as was present at harvest. This resulted in four treatments: root-free 14Clabelled soil (includes 14C-labelled soil microbial biomass plus 14C-labelled microbial products) from plants
grown at 350 and 700 ml lÿ1 CO2 (later referred to as
350S and 700S) and 14C-labelled soil amended with
14
C-labelled roots originating from 350 and 700 ml lÿ1
CO2 (referred to as 350R and 700R). Each beaker was
placed in a hermetically closed 1.5 l glass jar together
with a beaker containing NaOH solution and subsequently subjected to decomposition at 148C for 230
days. One additional treatment of 700R was prepared
in the same way but incubated at a 28C higher tem-

J.H. van Ginkel et al. / Soil Biology & Biochemistry 32 (2000) 449±456

perature (700R-HT). During incubation, soil water
was about 14% w/w (60±70% of ®eld capacity) and as
the containers were hermetically closed, no water was
lost. After 32 and 128 days, the containers were

¯ushed with CO2-free air to prevent lack of oxygen.
NaOH solutions were refreshed and analysed after 1,
2, 4, 8, 16, 32, 64, 128 and 230 days.
2.3. Analyses

451

the 14C-CO2 respiration determined by liquid scintillation counting (see Analyses).
2.5. Statistics
Four plants were exposed to 350 ml lÿ1 CO2 and
four plants to 700 ml lÿ1 CO2. The data of the incubation experiment were analysed based on a completely randomised design with ®ve treatments in four
replicates. All pots were incubated in duplicate and the

Total carbon and 14C-carbon content of shoots,
roots and soil were determined using a modi®ed wet
combustion method (Dalal, 1979). Dried and ground
plant material (30 mg) and soil (1 g) were digested in 5
ml of a 10% (w/v) solution of K2Cr2O7, dissolved in a
mixture of concentrated H2SO4 and H3PO4 (3:2 v/v),
at 1608C for 2 h. The CO2 evolved was trapped in 10

ml of a 0.5 N NaOH solution. Total CO2 (12CO2 and
14
CO2) adsorbed in the NaOH solution was determined by titrating the remaining NaOH with HCl
2ÿ
species by excess
after precipitation of HCOÿ
3 /CO3
BaCl2. Liquid scintillation counting was used to determine 14C-carbon: 0.5 ml of the 0.5 N NaOH solution
was mixed with 3 ml of Ultima Gold (Packard) and
counted on a liquid scintillation counter (Tri-Carb
2100TR; Packard). Total nitrogen in the roots was
determined by acid digestion and determination of
nitrogen with Nessler's reagent by continuous ¯owanalysis (Van Ginkel and Sinnaeve, 1980). The 14Clabelled soil microbial biomass (14C-smb) was determined using the fumigation±centrifugation technique:
soil solutions were obtained by centrifugation of nonfumigated and chloroform-fumigated (20 h) soils and
in an aliquot of the soil solution 14C was subsequently
determined (Van Ginkel et al., 1994). The proportionality factor Kcc relating the ¯ush size obtained by centrifugation with the microbial biomass of the soil was
determined by in situ labelling of the microbial biomass with D(U-14C)glucose (Van Veen et al., 1985),
assuming that after 3 days the 14C-label is totally incorporated into the microbial biomass. The proportionality factor so obtained for the FC-method (Kcc)
was 0.152.
2.4. Calculations

Total carbon (mg C) recovered in the di€erent
plant-soil compartments was calculated by dividing the
14
C-recovered (kBq) in all compartments by the mean
speci®c activity (kBq mgÿ1 C) of the ESPAS atmosphere. The percentage respired 14C-labelled material
from the soil was calculated as 14CO2 respired (mg C)
divided by the initial 14C-labelled soil content (mg C)
times 100. The respiration of the native-soil organic
carbon was calculated as the total respiration determined by acid digestion (12C-CO2 plus 14C-CO2) minus

Fig. 1. (A) Relation between the amount of 14C-labelled carbon in
the roots (mg C) and the amount of 14C-labelled carbon in the soil
solution (mg C gÿ1 soil) after 115 days of L. perenne growth at 350
and 700 ml Lÿ1 CO2 (four replicates). (B) The same relation between
the amount of 14C-labelled carbon in the soil solution (mg C gÿ1 soil)
and the amount of 14C-labelled carbon in the soil microbial biomass
(mg C gÿ1 soil). (C) The same relation between the amount of 14Clabelled carbon in the soil microbial biomass (mg C gÿ1 soil) and the
amount of 14C-labelled carbon in the soil residue (mg C gÿ1 soil).

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J.H. van Ginkel et al. / Soil Biology & Biochemistry 32 (2000) 449±456

HT) to 39%, which was not signi®cantly di€erent
from the 700R treatment, but still signi®cantly lower
than the decomposition in the 350R treatment. The decomposition of the root-derived material (consisting of
14
C-labelled soil microbial biomass plus 14C-labelled
microbial products) in the `elevated' soil (700S) and in
the `ambient' soil (350S) was 27 and 38% ( p < 0.001),
respectively.
The 14C-labelled soil microbial biomass (14C-smb) in
the 700R soil was 28% larger ( p < 0.01) than that in
the 350R soil (Table 2). The 14C-smb in the 700R-HT
soil was 20.5 mg C gÿ1 dry soil, which was not di€erent
from that in the 350R soil but signi®cantly lower
(41%; p < 0.01) than that in the 700R soil. The 14Csmb of 350S was 32% lower than that of 700S, but
this di€erence was not signi®cant.
No di€erence was observed between the decomposition of native-soil organic matter (SOM) of the 350R
and 700R treatment (290 mg C gÿ1 dry soil). The SOM
decomposition of the 700R-HT treatment was 422 mg
C gÿ1, which was signi®cantly higher than the SOM
decomposition at the lower temperature. There was no
di€erence in SOM decomposition between 350S and
700S treatment (248 mg C gÿ1), but it was signi®cantly
lower ( p < 0.01) than the SOM decomposition of the
350R and the 700R.
A ratio between 14C-labelled soil microbial biomass
(mg C) and total 14CO2 evolved (mg C) at the end of
the incubation experiment was calculated. There was
no di€erence in this ratio between the 350R and 700R
treatment or between the 350S and 700S treatment
(Fig. 1). This ratio was signi®cantly lower in the 700RHT treatment than in the 700R treatment.

analysis was performed based on the averaged values.
The signi®cance of di€erences and LSD test were
assessed by ANOVA (Genstat 5; release 3.1). Treatment e€ects are considered signi®cant when p values
were lower than 0.05, unless stated otherwise.

3. Results
3.1. Elevated CO and
plant growth

14

C-allocation after 115 days

At the end of the growth period, elevated CO2 had
signi®cantly increased the accumulated carbon content
of the shoots by 27% and of roots by 41% (Table 1).
Root mass showed a positive correlation …r ˆ 0:75, p ˆ
0:03† with the amount of soluble 14C-carbon in the soil
solution of the nonfumigated soil (Fig. 1A). Subsequently, the amount of soluble 14C-labelled carbon
in the soil solution showed a positive correlation
…r ˆ 0:79, p ˆ 0:02† with the amount of 14C-labelled
soil microbial biomass (Fig. 1B). The 14C-labelled soil
microbial biomass was 46% greater at elevated CO2
than that at ambient CO2 (Table 1) and showed a
strong positive correlation …r ˆ 0:95, p < 0.001) with
the amount of 14C-labelled soil residue (Fig. 1C).
Expressing the 14C-labelled soil residue per unit 14Clabelled soil microbial biomass showed that at elevated
CO2 this ratio was signi®cantly higher (100%, p <
0.001). The amount of 14C-labelled soil residue was
about three times greater at elevated CO2 than at
ambient CO2 (Table 1). Increasing the atmospheric
CO2 concentration from 350 to 700 ml lÿ1 caused total
14
C-labelled carbon in the soil (14C-labelled roots, 14Clabelled soil microbial biomass and 14C-labelled soil
residue) to increase by 53% (Table 1).

4. Discussion
14

C-allocation after 115 days of plant growth

3.2. Incubation of roots and root-derived material

4.1.

The mean decomposition of the `elevated CO2' roots
after 230 days (700R) was 36%, which was less ( p <
0.01) than the 42% of the `ambient CO2' roots (350R,
Table 2). Raising the incubation temperature by 28C
increased the decomposition of the `700' roots (700R-

The 27% increase in shoot yield at elevated CO2
agrees well with the observations reviewed by Kimball
(1983). The total plant-derived below-ground carbon
input was 53% higher at elevated CO2 than at ambient
CO2 and agrees with studies by Newton et al. (1994;

Table 1
Total amounts of 14C-labelled C recovered in shoots, roots, soluble C in the soil solution, soil microbial biomass, soil residue and total belowground of L. perenne grown at 350 and 700 ml lÿ1 for 115 days. Each value represents four replicates
CO2a treatment

Shoot (mg C)

Root (mg C)

Soluble (mg C gÿ1)

smbb (mg C gÿ1)

Residue (mg C gÿ1)

Total

350
700

264
335

182
257

0.126
0.163

14.8
21.6

10.9
31.4

147
225

LSD0.05

36

59

0.057

4.6

8.6

50

a

350=atmosphere 350 ml lÿ1 CO2; 700=atmosphere 700 ml lÿ1 CO2.
C-labelled soil microbial biomass.

b 14

14

C-soil content (mg C gÿ1)

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J.H. van Ginkel et al. / Soil Biology & Biochemistry 32 (2000) 449±456

Table 2
Initial 14C-labelled carbon content, 14C-labelled soluble carbon in the soil solution, 14C-labelled soil microbial biomass, respired native-soil organic matter (som) and respired 14C-carbon after incubation (four replicates) of L. perenne roots and/or root derived material at 148C and 168C
(HT) for 230 days
CO2/temperature treatment

Initial 14C-soil content
(mg C gÿ1)

Soluble carbon
(mg C gÿ1)

smba
(mg C gÿ1)

Respired som
(mg C gÿ1)

Respired
(%)

350Rc
700R
700R-HT
350S
700S
LSD0.05

147
225
225
26
53

0.148
0.181
0.146
0.051
0.066
0.048

22.0
28.9
20.5
7.6
10.0
5.0

294
286
422
245
251
25

42.2
36.2
38.6
38.3
27.1
3.7

14

Cb

a 14

C-labelled soil microbial biomass.
Expressed as a percentage of the initial total 14C-labelled soil carbon content.
c
350R=soil from atmosphere 350 ml lÿ1 CO2 with 14C-labelled roots, 14C-labelled soil microbial biomass and 14C-labelled microbial products;
700R=same as 350R but from a 700 ml lÿ1 CO2 atmosphere; 700R-HT=same as 700R but with a 28C higher incubation temperature;
350S=soil from atmosphere 350 ml lÿ1 CO2 with 14C-labelled soil microbial biomass and 14C-labelled microbial products; 700S=same as 350S
but from a 700 ml lÿ1 CO2 atmosphere.
b

1995), i.e. +64% for pasture turves, by Cotrufo and
Gorissen (1997), i.e. +26% for L. perenne, Agrostis
capillaris and Festuca ovina and by Van Ginkel et al.
(1996; 1997), i.e. +45 and +57%, respectively, for L.
perenne. It emphasises the relatively large impact of
increased CO2 concentrations in the atmosphere on
below-ground processes.
However, it remains extremely dicult to assess
whether roots release also more carbon per unit of
root weight into the soil under elevated CO2 since
carbon ¯ows through the di€erent below-ground compartments can hardly be quanti®ed (Cardon, 1996).
We applied continuous 14C-labelling during the entire
plant growth period and found a positive correlation
between carbon ¯ows through the subsequent belowground compartments (roots 4 carbon in the soil solution 4 soil microbial biomass 4 soil residue; Fig. 1).
This seems to support the conclusion that the root biomass is the driving parameter for all subsequent
below-ground processes in our plant-soil system.
The positive correlation between roots and soil microbial biomass is in contrast with the results of Kampichler et al. (1998) who found no e€ect of a plant
community (Cardamine hirsuta, Poa annua, Senico vulgaris and Spergula arvensis ) grown in an Ecotron at elevated CO2 on soil microbial biomass. In their ®rst
experiment, root biomass at elevated CO2 was 31%
lower than that at ambient CO2, whereas in their second experiment root biomass at elevated CO2 was
69% higher. We consider it unlikely that 69% more
root biomass would not stimulate the size or activity
of the microbial biomass in the soil. In our experiment,
41% more root biomass signi®cantly increased the size
of soil microbial biomass. Other experiments (Cotrufo
and Gorissen, 1997; Van Ginkel and Gorissen, 1998)
also showed a positive correlation between the weight
of the root biomass of L. perenne, A. capillaris and F.

ovina and the size the soil microbial biomass. Kampichler et al. (1998) suggested that a detectable response of soil microorganisms cannot be found in
experiments on ecosystem level as compared with experiments at a single species level. However, we feel
that techniques used by Kampichler et al. (1998) are
not sensitive enough to accurately measure small
changes in the size of the soil microbial biomass and
their suggestion seems rather premature.
4.2. Microbial behaviour in soil
Ladd et al. (1995) pointed out that faster turnover
rates would result in less biomass-14C accumulated per
unit 14CO2 evolved. Comparing this ratio in the 350R
soil with that in the 700R soil (Fig. 2) revealed no
di€erence, indicating that the turnover of plant-derived

Fig. 2. Ratio between 14C-labelled soil microbial biomass (mg C gÿ1
soil) and total 14C-CO2 (mg C gÿ1 soil) evolved. Incubations at 148C
of roots plus root-derived materials originating from plant growth
(L. perenne ) at 350 and 700 ml lÿ1 CO2 (350R and 700R), incubations at 168C of roots plus root-derived materials originating from
plant growth (L. perenne ) at 700 ml lÿ1 CO2 (700R-HT) and incubations at 148C of root-derived materials originating from plant
growth (L. perenne ) at 350 and 700 ml lÿ1 CO2 (350S and 700S).
Each treatment represents four replicates. Values indicated by di€erent letters are signi®cantly di€erent ( p < 0.05).

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J.H. van Ginkel et al. / Soil Biology & Biochemistry 32 (2000) 449±456

14

C-carbon in soil was not a€ected by elevated CO2.
Substrate use eciency seems unaltered and soil microorganisms seem to transform root material originating
from both elevated and ambient CO2 metabolically in
the same way. It remains unclear why relatively less
root material in the 700R soil had been decomposed
as compared with roots in the 350R soil. In absolute
amounts, more carbon was transformed by microorganisms in the 700R soil, but signi®cantly more carbon
remained in this soil than in the 350R soil due to a signi®cantly higher carbon input at elevated CO2. The
lowest ratio between biomass-14C and 14CO2 evolved
was found in the incubation of the `700 roots' at a
higher temperature, indicating that this 14C-material
had the highest turnover rate. The relative amount of
decomposed 14C-material in the 700R-HT soil showed
no signi®cant di€erence with the 700R soil; the higher
turnover rate can therefore only have been caused by
faster turnover of the soil microbial biomass itself as
can be derived from the decrease in soil microbial biomass. No di€erence was found in the ratio between
14
C-labelled microorganisms and total 14CO2 evolved
in the 350S soil and in the 700S soil, again indicating
that turnover of microorganisms and their metabolites
occurs in a similar way. Microbial transformation processes involved in the turnover of roots and root-derived material in soil ecosystems do not seem to be
a€ected by an increase in atmospheric CO2 concentrations. Therefore, a lower decomposition rate at elevated CO2 is not due to a change in the internal
metabolism of microorganisms.
One could argue that available mineral nitrogen
could cause the reduced carbon mineralisation at elevated CO2. Soil mineral nitrogen concentration at the
start of the incubation experiment was 2.4 and 3.2 mg
gÿ1 dry soil at ambient and elevated CO2, respectively.
The amount of nitrogen mineralised in the soil during
the incubation experiment can be calculated from the
respiration of the native-soil organic carbon (Table 2).
Assuming a substrate assimilation eciency of the soil
microbial biomass for recalcitrant soil organic carbon
of about 20% (Van Veen et al., 1984) for both treatments, 80% of this material has been evolved as
12
CO2. Division of the total amount of carbon mineralised from soil organic matter (carbon incorporated in
the microbial biomass plus CO2 evolved) by the C-toN ratio of the soil (17.0) results in an estimated nitrogen mineralisation of 20 mg gÿ1 dry soil. This amount
makes it unlikely that mineral nitrogen was a limiting
factor during the incubation experiment.
4.3. Elevated CO2 and decomposition of roots and root
residues
Although the C-to-N ratio of the grass roots (62)
was not a€ected, the decomposition of grass roots

grown at elevated CO2 (700R) was signi®cantly lower
(14%) than that of roots grown at ambient CO2
(350R) after incubation for 230 days. The lower decomposition is in agreement with other studies on
grass roots (Gorissen et al., 1995; Van Ginkel et al.,
1996; Van Ginkel and Gorissen, 1998). Norby and
Cotrufo (1998) argued that litter decomposition is not
a€ected by elevated CO2, but they mainly refer to studies with above-ground plant materials. However, the
importance of below-ground decomposition processes
on the carbon cycle is considerable since 40±60% of
all assimilated photosynthates is transported belowground in grasses (Van Ginkel and Gorissen, 1998).
Although both increased carbon input and a change in
decomposability play a role in soil carbon storage in
grasslands (Van Ginkel et al., 1999), their relative contributions are still unclear. Several studies have shown
a decreased decomposition of roots grown under elevated CO2 without a consistent correlation with the Cto-N ratio (Cotrufo and Ineson, 1995; Van Ginkel et
al., 1996; Van Ginkel and Gorissen, 1998). Most likely,
the C-to-N ratio is not the appropriate indicator for
`CO2-induced' changes in `quality' and subsequent
changes in decomposition rates. Norby and Cotrufo
(1998) derived their conclusions from studies with litterbags. Unfortunately, using litterbags to measure the
overall e€ect of elevated CO2 on decomposition processes, the amount and fate of (labile) carbon leaching
into the soil matrix are neglected. Litterbags containing 14C-labelled plant material could provide a powerful tool to examine this artefact.
Raising the incubation temperature by 28C, the predicted increase in temperature when atmospheric CO2
rises from 350 to 700 ml lÿ1 (IPPC, 1995), increased
the decomposition of the 14C-labelled `700' roots from
36.2 to 38.6%. Still, this is lower than the decomposition of roots in the 350R soil (42.2%) indicating that
the retarded decomposition of 700R could not fully be
compensated by a 28C higher temperature. In a shortterm experiment, we observed that a slower decomposition of grass roots grown at elevated CO2 (ÿ24%
compared with ambient CO2) even counteracted the
increased decomposition at a 68C higher temperature
(Gorissen et al., 1995).
Ball (1996) found decreased respiration levels of L.
perenne plant litter after incubation for 30 days and
argued that this e€ect was due to a decreased C-to-N
ratio of the plant material grown at elevated CO2.
These results are in contrast with those of Ross et al.
(1995) who found no di€erences in decomposition of
roots originating from ryegrass/white clover turves
grown at ambient and elevated CO2, but they incubated their roots for only 20 days. As both worked
with nonlabelled material in a mineral soil and subtracted the background respiration of nontreated soil,
side e€ects cannot be precluded due to priming or con-

J.H. van Ginkel et al. / Soil Biology & Biochemistry 32 (2000) 449±456

serving e€ect on soil organic matter decomposition.
Poorter et al. (1997) pointed out that leaves of 27 C3species, grown at elevated CO2, contained about 50%
more nonstructural carbohydrates, while Wong (1990)
found double the amount of nonstructural carbohydrates in `elevated CO2' roots of cotton plants. It is
conceivable that this energy-rich plant-derived carbon
may have a conserving e€ect on native-soil organic
carbon decomposition, since it has been hypothesised
that microorganisms may prefer the more energy-rich
carbon to native-soil organic carbon (Lekkerkerk et
al., 1990; Van Veen et al., 1993). This preferential substrate use (and thus less native-soil organic carbon respiration) can incorrectly be considered as retarded
respiration of plant material grown at elevated CO2.
On the other hand, energy-rich plant-derived carbon
incubated in a rather nutrient-poor soil may have a
priming e€ect on soil organic carbon decomposition to
satisfy the nutrient demand of microorganisms, resulting in extra CO2 release. Summarising, carbon balances in such studies may be inconclusive with respect
to decomposition of plant litter. Therefore, it seems
inevitable to work with homogeneously 14C-labeled
plant material for studies concerning biodegradability
in soils; this also allows comparative monitoring of
native-soil organic carbon decomposition.

5. Conclusions
The present experiment supports other studies showing that decomposition of grass roots might be
retarded when grown under elevated CO2 (Gorissen et
al., 1995; Jongen et al., 1995; Van Ginkel et al., 1996;
Van Ginkel and Gorissen, 1998). The total response of
our root/soil system of L. perenne towards elevated
CO2 (more carbon input, no altered microbial metabolic behaviour and retarded decomposition of roots
and root-derived material) is most interesting in the
debate whether soils may act as a sink for atmospheric
CO2. Whatever the reason for altered below-ground
processes, these processes will always be driven by
plant carbon inputs and mediated by soil microorganisms. Until now, the question whether soil microorganism behaviour is a€ected by elevated CO2 has been
underexposed. This is why we have tried to elucidate
their role in carbon decomposition in a changing climate. After all, microorganisms are the most important precursors in any altered soil organic carbon
build-up.
Tans et al. (1990) suggested that ecosystems (e.g.
temperate grasslands) in the Northern Hemisphere
could possibly function as a sink for missing carbon in
the global carbon budget. Models describing the response of the soil carbon content of ecosystems to elevated CO2 (Parton et al., 1995; Thornley and

455

Cannell, 1997) often su€er from a lack of experimental
data of elevated CO2-induced below-ground changes
like increased carbon input (Newton et al., 1994; Newton et al., 1995; Van Ginkel et al., 1997; Cotrufo and
Gorissen, 1997; Schapendonk et al., 1997) and
retarded decomposition of roots (Gorissen et al., 1995;
Van Ginkel et al., 1996). Thornley and Cannell (1997)
argued that experiments should try to lessen uncertainty about processes within models rather than try to
predict ecosystem responses directly. This point of
view seems realistic and a good guideline for future experimental research.

Acknowledgements
The authors thank J.A. van Veen for critically reading the manuscript and useful suggestions.

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