Applied Soil Ecology 73 (2014) 19–25

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Applied Soil Ecology

journal homepage: www.elsevier.com/locate/apsoil

Effects of wastewater sludge, urea and charcoal on greenhouse gas

emissions in pots planted with wheat
Martín Díaz-Rojas a , Ángel Aguilar-Chávez a , María del Rosario Cárdenas-Aquino a ,
Víctor Manuel Ruíz-Valdiviezo a , Eduardo Hernández-Valdez a , Marco Luna-Guido a,∗ ,
Víctor Olalde-Portugal b , Luc Dendooven a
a
Laboratory of Soil Ecology, ABACUS, Cinvestav, Av. I.P.N. 2508, C.P. 07360 México, D.F., Mexico
b
Laboratory of Ecological Biochemistry, Department Biotechnology and Biochemistry, Cinvestav Campus Guanajuato, Km 9.6 Libramiento Norte Carretera
Irapuato-León, Apdo. Postal 629, C.P. 36500 Irapuato, Guanajuato, Mexico

a r t i c l e i n f o a b s t r a c t

Article history: Agriculture is an important source of greenhouse gases (GHG), mostly carbon dioxide (CO2 ), nitrous oxide
Received 3 April 2013 (N2 O) and methane (CH4 ). Application of charcoal to agricultural soils is known to reduce GHG emissions
Received in revised form 13 July 2013 while application of fertilizer or wastewater sludge increases them. Therefore, the objective of this work
Accepted 4 August 2013
was to study the effect of charcoal application on GHG emissions from soil planted with wheat (Triticum
aestivum L.) and amended with wastewater sludge or urea, or left unamended. Wheat fertilized with
Keywords:
urea or wastewater sludge, at a rate of 150 kg N ha−1 , was cultivated in soil amended with or without
Greenhouse gases
2% (w/w) charcoal, a biochar used mostly for heating, in a greenhouse. Emission of CO2 , CH4 and N2 O,
Charcoal
Wastewater sludge
soil characteristics and plant development were monitored. Charcoal had no significant effect on the
Plant development emission of CO2 , CH4 and N2 O in wastewater sludge or urea-amended soil. The wheat development and
Soil characteristics yields, and soil pH and electrolytic conductivity were also not affected by charcoal application. It was
found that charcoal did not affect the emissions of the monitored GHG, wheat or soil characteristics.
© 2013 Elsevier B.V. All rights reserved.

1. Introduction organic material also increases the concentration of NH4 + in soil,

Nitrous oxide (N2 O), methane (CH4 ) and carbon dioxide (CO2 )
are important greenhouse gases (GHG) (IPCC, 2007) and agricul-
tural activities are an important source of anthropogenic GHG,
contributing up to 20% of the annual atmospheric increase (Lemke
et al., 2007). For instance, application of inorganic N fertilizers, such
as urea, to an N depleted soil can increase emissions of CO2 as micro-
bial activity is stimulated. Additionally, hydrolysis of urea releases
CO2 . (Fertilizing plants with inorganic N fertilizer, such as urea,
increases generally the emissions of N2 O). Abiotic or biotic hydroly-
sis of urea generates ammonium (NH4 + ) in soil, which is oxidized to
nitrite (NO2 − ) and then to nitrate (NO3 − ). During this oxidation pro-
cess, i.e. nitrification process, N2 O is formed. The produced NO3 −
is then reduced to N2 O under anaerobic conditions, i.e. denitrifica-
tion, further contributing to GHG emissions. Application of organic
fertilizer, such as wastewater sludge, also increases emissions of
CO2 , N2 O and CH4 (Chiaradia et al., 2009). The organic material
in the organic fertilizer is mineralized increasing emissions of CO2
and CH4 when anaerobic microsites are formed. Mineralization of
∗ Corresponding author. Tel.: +52 55 5747 3319; fax: +52 55 5747 3313.
E-mail address: mluna@cinvestav.mx (M. Luna-Guido).
which, as mentioned earlier, increases emissions of N2 O.
Charcoal, a biochar primarily used for fuel, is made by burn-
ing wood in a kiln with the least possible flame (Gómez-Luna et al.,
2009). The process lasts from 12 to 14 days (Vázquez-Marrufo et al.,
2003). The organic material is combusted above 200 ◦ C, but struc-
tural changes occur in the organic material when the temperature
is higher than 300 ◦ C, e.g. decarboxylation of the humic and ful-
vic acids and an increase in aromatic components, making it far
more resistant to microbial degradation (Knicker et al., 2005). Char-
coal is still a commonly used fuel in many rural communities of
Latin America, Africa and the Middle East (Nepstad et al., 2006;
Ndayambaje and Mohren, 2011). For instance, charcoal is still pro-
duced in the Santa Rosa forest in the central highlands of México
(Guanajuato).
It has been reported that application of charcoal to a soil can raise
pH, water holding capacity and sometimes decreases N mineral-
ization (Streubel et al., 2011). The charcoal is resistant to microbial
degradation so C is sequestered in soil (Sohi et al., 2010; Foereid
et al., 2011). Charcoal has been shown to improve plant develop-
ment. For instance, Kammann et al. (2011) showed that charcoal
increased growth, drought tolerance and leaf-N and water use effi-
ciency of the pseudo-cereal Chenopodium quinoa Willd. despite a
larger plant leaf area. The effect of charcoal on emissions of N2 O,

0929-1393/$ – see front matter © 2013 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.apsoil.2013.08.001
20 M. Díaz-Rojas et al. / Applied Soil Ecology 73 (2014) 19–25
CO2 and CH4 is not always unequivocal. Increases in emissions of
CO2 have been reported, decreases or no effect at all depending
on charcoal and soil characteristics (Spokas and Reicosky, 2009;
Zimmerman et al., 2011). Although charcoal often decreases emis-
sions of N2 O (Zhang et al., 2010; Rogovska et al., 2011), Cheng et al.
(2012) reported that charcoal of wheat straw did not affect the
emissions of N2 O.
Organic wastes, such as wastewater sludge, are often applied
to soil (Bastida et al., 2012). Wastewater sludge is rich in plant
nutrients and organic material (Guo et al., 2012). The organic mat-
ter content increases when wastewater sludge is applied to soil,
which improves soil structure and increases soil fertility (Jindo
et al., 2011). Consequently, plant development is often better when
wastewater sludge is used to fertilize crops than urea (Fernandez-
Luqueño et al., 2010). However, application of organic wastes is also
known to increase emissions of CO2 (Paramasivam et al., 2008), N2 O
(Jager et al., 2013) and CH4 (Sistani et al., 2010).
Wheat is one of the most important crops in the world
with a total production of 704.1 million metric tonnes culti-
vated on 220,385,284 ha in 2011 (http://faostat3.fao.org/home/).
Wheat was cultivated on 662,221 ha in Mexico with a total
production of 3.6 million metric tonnes in the same year
(http://faostat3.fao.org/home/). Consequently, soil cultivated with
wheat plays an important role in the contribution of farmland to
global warming (Lenka and Lal, 2013). As such, whatever change
occurs in agricultural practices, such as changes in fertilizer use
or application of charcoal, might have a large impact on climate
change.
In the study reported here, the effect of charcoal on plant growth,
soil characteristics, and the emission of GHG was studied in a
soil cultivated with wheat (Triticum aestivum L.) and amended
with an inorganic fertilizer (urea) or an organic fertilizer (waste-
water sludge) at 150 kg N ha−1 . The objectives of the study were
to investigate how the application of charcoal to soil cultivated
with wheat and fertilized with wastewater sludge or urea affected
plant growth, soil characteristics and emissions of GHG. It was
hypothesized that charcoal would reduce emissions of GHG, while
wastewater sludge would increase it.
2. Materials and methods
2.1. Area description and soil sampling
The sampling site is located in Otumba in the State of México
(19◦ 42 N 98◦ 49 W) and the soil was classified as Typic Frag-
iudepts. The soil is mainly cultivated with maize (Zea mays L.) for
>20 years, receiving a minimum amount of mineral fertilizer, i.e.
<50 kg N ha−1 , without being irrigated. Soil was sampled randomly
by augering the 0–15 cm top-layer of three plots of approximately
0.5 ha spatially separated. At each plot 30 soil samples were taken,
pooled so that three soil samples were obtained and characterized
separately. The sandy loam soil with pH 8.9 and electrolytic con-
ductivity (EC) 1.23 dS m−1 , had a total C content of 29.1 g C kg−1 soil
and a total N content of 2.41 g N kg−1 and water holding capacity
(WHC) of 567 g kg−1 dry soil. The field based replication (n = 3) was
maintained in the greenhouse experiment.
2.2. Wastewater sludge characteristics
Sludge was obtained from Reciclagua (Sistema Ecológico de
Regeneración de Aguas Residuales Ind., S. A. de C. V.) in Lerma,
State of México (México). Reciclagua treats wastewater derived
from a number of industries (90%), mostly textile, and the rest
from households. The wastewater from the different companies
complies with the following guidelines: biological oxygen demand
(BOD) less than 1000 mg l−1 , lipids less than 150 mg l−1 and phe-
nol less than 1 mg l−1 . The wastewater is digested aerobically in a
reactor and the sludge obtained after the addition of a flocculant is
passed trough a belt filter. Thirty kilogram of the wastewater sludge
that had passed the belt filter was sampled aseptically in plas-
tic bags, characterized and used in the experiment. The pH of the
sludge was 8.7 and with a water content of 820 g kg−1 . The organic
C content was 451 g kg−1 and total N content 13.4 g kg−1 , while
extractable P was 0.67 g kg−1 , total P 2.33 g kg−1 and concentration
of NH4 + 2.94 g N kg−1 expressed on a dry weight base.
2.3. Charcoal production
Charcoal is still used commonly as fuel in rural communities
of central México. Details of the production process and charac-
teristics are described in Gómez-Luna et al. (2009, 2012). Briefly,
charcoal is made by burning wood in a kiln with the least possi-
ble flame for 12–14 days. The organic material is combusted above
200 ◦ C and structural changes occur in the organic material when
the temperature is higher than 300 ◦ C, e.g. decarboxylation of the
humic and fulvic acids and an increase in aromatic components,
making it far more resistant to microbial degradation.
2.4. Cultivation of wheat in the greenhouse
The experimental design was a completely randomized 4
factorial with nine replications (maintained from the field site
replications). The arable soil was cultivated with wheat and four
treatments were applied: (1) plants fertilized with urea (0.646 g
urea was applied per soil column or an equivalent of 150 kg N ha−1 );
(2) soil mixed with charcoal at 2% (w/w) applied to the upper 10 cm
layer and plants fertilized with urea (0.646 g urea was applied per
column or an equivalent of 150 kg N ha−1 ); (3) soil mixed with
wastewater sludge (235 g fresh sludge per column) so that plants
were fertilized with 150 kg N ha−1 considering the mineral N in the
sludge and that 40% of the organic N in the sludge was mineralized
within the experimental period and (4) soil mixed with charcoal
at 2% (w/w)) applied at the upper 10 cm layer and sewage sludge
(235 g fresh sludge per column) so that plants were fertilized with
150 kg N ha−1 .
Nine samples from each of the three soil samples taken in the
field were used for each of the four treatments. As such, a total
of 108 sub-samples of 6 kg soil were added to PVC tubes (length
50 cm and diameter 16 cm) filled at the bottom with 7 cm of gravel
topped up with 3 cm sand (Bellini et al., 1996). Three soil columns
from each of the three soil samples and each of the four treatments
were sampled after 45 days and six columns at harvest.
Seeds of wheat cultivar temporalera M87 were provided by the
‘Research Centre for Agricultural and Forestry of the State of México,
Campo Experimental Valle de México’ (INIFAP) (Texcoco, México).
Six wheat seeds were planted in each of the 108 PVC tubes. Seeds
were placed at 3 cm depth. The PVC tubes were placed at random
in the greenhouse until grain maturity. The experiment mentioned
above was done three times to account for possible effects of cli-
matic conditions. During the first experiment that started the 9th
of November 2010, 500 ml water was added to each column every
seven days. In the second experiment that started on the 7th of
February 2011 and the third on the 18th of April 2011, the same
amount of water was added every seven days. The temperature,
relative humidity and light intensity in the greenhouse were moni-
tored during each of the experiments (No data shown). The mean air
temperature in the greenhouse was 36.3 ◦ C, the relative humidity
16.9 (%) and the light intensity 1620 lux.
Forty-five days after planting, three PVC tubes from each plot
(n = 9) and at harvest six PVC tubes were selected from each
 M. Díaz-Rojas et al. / Applied Soil Ecology 73 (2014) 19–25
treatment. The entire soil column was removed from the PVC tube
and the 0–15 cm and a 15–30 cm layer sampled taken care not
to damage the root structure. The roots were separated from the
shoots and the fresh weight determined and the root and shoot
length measured. The roots and shoots were dried at 45 ◦ C in an
oven for a week and weighed. At grain maturity, i.e. 129 days in
experiment 1, 90 days in experiment 2 and 92 days in experiment
3, the six remaining columns were sampled and root and shoot
characteristics measured as described above. In addition, the
weight of grains dried at room temperature and number of grains
per plant were determined.
2.5. Emissions of CO2 , CH4 and N2 O
From the onset of the experiment until day 45, a cylindrical PVC
chamber (length 50 cm and Ø 16 cm) fitted with two sampling ports
was placed on the PVC tube and air-tied sealed with Teflon tape
three times a week. Zero, 15 and 30 min after the chamber was
sealed, 15 cm3 air was injected into the PVC chamber headspace.
The gas in the headspace was mixed by flushing the air inside
the chamber five times. A 15 cm3 sample was then taken from the
headspace for analysis. The 15 cm3 air sample was injected into 15-
ml evacuated vials closed with a butyl rubber stopper and sealed
with an aluminium cap pending analysis.
The headspace of the vials was analyzed on an Agilent Tech-
nology 4890D gas chromatograph (GC) fitted with an electron
capture detector (ECD) for the determination of CO2 and N2 O and
a flame ionization detector (FID) for CH4 . Details of the settings
of the GC can be found in Serrano-Silva et al. (2011). Concentra-
tions of CO2 , N2 O and CH4 were calculated by comparing peak areas
against a standard curve prepared from known concentrations, i.e.
1 and 10 ppm N2 O in N2 , 5 ppm CH4 in N2 and 2500, 20,000 and
40,000 ppm CO2 in N2 , every time samples were analyzed.
2.6. Soil analysis
The techniques used to measure pH in a 1:2.5 soil-H2 O sus-
pension, EC in a 1:5 soil-H2 O suspension, the organic C, total N,
WHC, particle size distribution and the NH4 + , NO2 − and NO3 − in
the K2 SO4 extracts can be found in Serrano-Silva et al. (2011).
2.7. Statistical analysis
Emission of CO2 , CH4 and N2 O was regressed on elapsed time, i.e.
after 0, 15 and 30 min, using a linear model forced to pass through
the origin, but allowing different slopes (production rates). The
sample at time 0 accounted for the atmospheric CO2 , CH4 and N2 O
and was subtracted from the measured values.
Significant differences among plant characteristics as a result of
the treatments were determined by analyses of variance (ANOVA)
and based on the least significant difference using the general lin-
ear model procedure (PROC GLM, SAS Institute, 1989). Effect of
layer, time of sampling and treatment and their interactions on soil
characteristics was determined in the same way. Significant dif-
ferences between treatments for production of CO2 , CH4 and N2 O
were determined using PROC MIXED considering repeated mea-
surements (SAS Institute, 1989).
Plant characteristics were separately explored with a princi-
pal component analysis (PCA) using PROC FACTOR (SAS Institute,
1989). The matrix of nine columns (plant variables: shoot and
root length, dry and fresh root and shoot weight, chlorophyll
content, number of seeds and seed weight) and 54 lines (in the
experiment wheat was cultivated three times × three different soil
samples were taken in the field and used separately in each exper-
iment × plants were taken from six columns at harvest) was used
21
for principal component analysis. Further details of the PCA analysis
can be found in López-Valdez et al. (2011).
3. Results
3.1. Emissions of CH4 , CO2 and N2 O
The emission of CO2 fluctuated over time, but no clear pattern
emerged (Fig. 1a). The emission of CO2 was significantly greater
in the sludge-amended soil than in the urea and urea + charcoal
amended soil (P < 0.05) (Table 1).
The emission of CH4 was highly variable and no clear pattern
emerged (Fig. 1b). Oxidation of CH4 (negative production) occurred
only occasionally throughout the experiment. The emission of CH4
was not affected by treatment (Table 1).
The emission of N2 O showed large variations over time espe-
cially in the beginning of the experiment (Fig. 1c). The largest
emissions in the sludge and charcoal + sludge-amended soil were
found at day 0 and day 3 and in the urea and charcoal + urea-
amended soil on day 7. Thereafter, emission of N2 O remained below
<60 mg N ha−1 day−1 in all treatments. The emissions of N2 O were
not affected by treatment in the first 10 days or when the emissions
were considered between days 11 to 45 (Table 1).
3.2. Plant and soil characteristics
After 45 days, the application of wastewater sludge increased
root fresh weight and root length significantly compared to the
urea-amended soil, but not the other plant characteristics (P < 0.05)
(Table 2). After 90 days, plant chlorophyll and shoot length
were significantly affected by treatment, but not the other plant
characteristics (P < 0.05). The PCA analysis indicated that plant
development was more affected by the type of fertilizer applied
than by the application of charcoal, i.e. on the scatter plot the treat-
ments were more separated due to fertilizer application than by
charcoal application (Fig. 2).
None of the soil characteristics was significantly different
between the treatments in the 0–15 or 15–30 cm soil layer and
after 45 or 90 days (No data shown). The pH, EC and concentra-
tion of NH4 + and NO3 − were significantly different between day 45
and 90. The soil layer had a significant effect on the concentration
of NH4 + and NO3 − and treatment on EC. The interaction between
layer and day, and layer and time had a significant effect on the
concentration of NO3 − .
4. Discussion
4.1. Emissions of CO2 , CH4 and N2 O
Application of charcoal had no effect on the CO2 emission rate in
the soil amended with wastewater sludge or urea. Different results
have been reported in literature (Zimmerman et al., 2011). Some
researchers found an increase, a decrease or no effect at all (Spokas
and Reicosky, 2009; Smith et al., 2010; Liu et al., 2011). Differ-
ences have been attributed to the type of charcoal used, charcoal
application rate, an ageing effect or soil type (Singh et al., 2010).
The application of charcoal to a soil might affect the emission of
CO2 in various ways. First, Smith et al. (2010) reported that miner-
alization of the charcoal contributed to emissions of CO2 , although
most of the organic material in the charcoal is recalcitrant with a
mean residence time >100 years (Jha et al., 2010). The application
of charcoal had no effect on the emission of CO2 in this study, so it
can be speculated that no mineralization of the charcoal occurred. It
can be speculated that the production process, i.e. burning wood in
a kiln for 12 to 14 days, reduced the easily decomposable organic
22 M. Díaz-Rojas et al. / Applied Soil Ecology 73 (2014) 19–25

200 a) CO2 emission rate

160

(g CO2 ha-1 day-1)

120

80

40

0
0 5 10 15 20 25 30 35 40 45

0.15 b) CH4 emission rate

0.10

(g CH4 ha-1 day-1)

0.05

0.00

-0.05
0 5 10 15 20 25 30 35 40 45

0.75 c) N2O emission rate

Charcoal+sludge Sludge
0.60 Charcoal+Urea Urea
(g N2O ha-1 day-1)

0.45

0.30

0.15

0.00
0 5 10 15 20 25 30 35 40 45
Time (days)

Fig. 1. (a) The CO2 , (b) CH4 rates and (c) N2 O emission rates from soil cultivated with wheat (Triticum aestivum L.) amended with urea (䊉), with charcoal plus urea (),
wastewater sludge () and wastewater sludge plus charcoal (). Bars are ± one standard error of the estimates of three different soil samples and three experiments (n = 9).

material sharply, so little or no organic material in the charcoal
was mineralized. Second, reduction in emissions of CO2 has been
explained by the fact that charcoal might inhibit soil organic carbon
(SOC) mineralization as organic material get fixed on the charcoal
rendering it unavailable as C substrate for the soil microorganisms
(Zimmerman et al., 2011). In this study, the fluxes of CO2 were not
affected by the application of the charcoal so it appears that none
of the added soil organic material (i.e. the wastewater sludge) or
soil organic matter got fixed on the charcoal. Third, application of
charcoal might induce a priming effect, but that will depend again
on the type of charcoal applied and the easily decomposable soil
organic matter fraction (Keith et al., 2011). If the charcoal still con-
tains easily decomposable organic material, then a priming effect
might occur (Luo et al., 2011; Zavalloni et al., 2011; Zimmerman
et al., 2011). No increase in emission of CO2 was found in our study
so no priming effect occurred. The alkaline condition of our soil (pH
8.9) might have had an inhibitory effect on the labile organic matter
degradation of the charcoal.
Application of wastewater sludge increased emission of CO2 ,
which is a well-described phenomenon. In a laboratory study, 40%

Table 1
CO2 , CH4 and N2 O emission rate from soil cultivated with wheat (Triticum aestivum L.) in three different experiments and fertilized with urea, urea and amended with
charcoal, amended with wastewater sludge or amended with wastewater sludge and charcoal measured for 45 days.

Treatment CO2 (g CO2 ha−1 day−1 ) CH4 (g CH4 ha−1 day−1 ) N2 O (g N2 O ha−1 day−1 )

0–45 days 0–45 days 0–10 days 11–45 days

Urea 51.5 C † , § 0.025 0.110 0.022

Urea + charcoal 56.7 BC 0.023 0.113 0.022
Wastewater sludge 71.1 A 0.026 0.164 0.026
Sludge + charcoal 62.5 AB 0.024 0.195 0.023
SEE ‡ 4.6 0.058 0.036 0.002
F-value 6.60 0.16 2.58 1.87
P-value 0.025 0.917 0.149 0.236
†
Values with the same letter are not significantly different between the treatments, i.e. the columns (P < 0.05).
§
Mean of n = 9.
‡
SEE: standard error of the estimate (P < 0.05).
 M. Díaz-Rojas et al. / Applied Soil Ecology 73 (2014) 19–25 23

Table 2
Characteristics of wheat (Triticum aestivum L.) in three different experiments fertilized with urea, fertilized with urea and amended with charcoal, amended with wastewater
sludge or amended with wastewater sludge and charcoal.

Plant characteristics Urea Urea + charcoal Sludge Sludge + charcoal MSD † F value P value

Plant cultivated for 45 days

Chlorophyl content 43.2 § , ‡ 43.9 43.1 43.8 1.7 0.89 0.451
Shoot length (cm) 23.3 20.8 21.8 21.6 7.3 0.28 0.832
Shoot fresh weight (g) 16.5 15.9 21.1 19.9 5.5 2.25 0.087
Shoot dry weight (g) 3.1 3.0 3.8 4.1 1.6 1.52 0.215
Root length (cm) 38.5 b 45.5 ab 49.8 a 46.8 ab 9.0 3.85 0.012
Root fresh weight (g) 4.7 b 5.8 b 8.2 a 6.5 ab 2.3 5.42 0.002
Root dry weight (g) 0.70 0.74 0.92 0.96 0.33 2.08 0.107
Plant sampled at harvest
Chlorophyl content 39.9 c 43.9 a 42.6 ab 41.2 bc 2.3 7.51 <0.001
Shoot length (cm) 67.0 a 66.1 ab 65.1 b 65.2 b 1.5 4.62 0.004
Shoot fresh weight (g) 16.9 15.9 16.5 16.2 3.8 0.17 0.920
Shoot dry weight (g) 7.7 7.5 7.6 7.6 1.4 0.04 0.991
Root length (cm) 37.9 37.9 39.2 38.3 5.8 0.15 0.928
Root fresh weight (g) 4.7 4.7 5.5 4.3 1.7 1.22 0.303
Root dry weight (g) 1.05 1.04 1.11 1.13 0.35 0.19 0.901
Number of seeds 5.4 5.5 5.2 4.9 0.8 1.31 0.272
Dry weight seeds 147 148 136 130 21 2.22 0.087
†
MSD: least significant difference (P < 0.05).
§
Values within the row with the same letter are not significantly different (P < 0.05).
‡
Mean of n = 9.

of the organic material in wastewater sludge from the same treat-
ment plant was mineralized in an aerobic incubation within 7 days
increasing the emission of CO2 substantially (Rodriguez et al., 2011)
although lower values were also found, i.e. 28% within 42 days
(Franco-Hernández et al., 2003).
In this study, emissions of N2 O were not affected by application
of charcoal in both the urea and wastewater sludge amended soil.
It has been reported that application of charcoal to soil decreases
emissions of N2 O, has no effect at all or increases it (Singh et al.,
2010; Bruun et al., 2011; Scheer et al., 2011; Wu et al., 2013). The
differences can be attributed to an ageing effect and the initial N
content of the soil. Scheer et al. (2011) reported no significant differ-
ences in GHG emissions between the control and biochar amended
subtropical pasture soil, but that was three years after the biochar
was applied.
Application of charcoal can affect emissions of N2 O in different
ways. First, biochar can increase the emission of N2 O when it is
mineralized (Bruun et al., 2011). Mineralization of the biochar will
increase the concentration of NH4 + , which is then oxidized to NO3 − ,
and nitrification is known to contribute to the emission of N2 O. As
mentioned before, the charcoal was not mineralized as no increase
in the emission of CO2 was found. Consequently, no N mineraliza-
tion occurred and no increase in the emission of N2 O as a result of
nitrification. Second, charcoal can reduce the emission of N2 O when
organic material is fixed on it and mineralization is reduced (Singh
et al., 2010; Bruun et al., 2011; Rogovska et al., 2011). Reduction in
soil organic matter mineralization will reduce the concentration of
NH4 + and thus nitrification. As mentioned before, there was no evi-
dence that organic material was fixed on the charcoal and became
unavailable for degradation, so no reduction in the emission of
N2 O occurred. Third, charcoal can improve soil aeration thereby
decreasing denitrification and thus emission of N2 O (Singh et al.,
2010). As we did not have a reduction in emission of N2 O when
charcoal was applied, charcoal did not increase soil aeration.
The application of wastewater sludge in this study strongly
increased the emission rate of N2 O in the first 10 days compared
to the N2 O fluxes in rest of the experiment. A number of pro-
cesses might contribute to an increase in the emission of N2 O. The
NH4 + content of the wastewater sludge was high (2.94 g N kg−1 ) so
its oxidation increased emission of N2 O. The organic material in

Number of seeds Seed weight

0.8

0.6

0.4 Shoot length

Root dry weight
0.2 Charcoal+Urea
Shoot dry weight
Chlorophyl Urea
0
Sludge Root fresh weight
-0.2 Factor 1: 0.39
Shoot fresh weight
Root length
-0.4 Charcoal+Sludge

-0.6

-0.8
Factor 2: 0.19
-1
-1 -0.8 -0.6 -0.4 -0.2 0 0.2 0.4 0.6 0.8 1

Fig. 2. Principal component analysis (PCA) performed on plant characteristics (shoot and root length, dry and fresh root and shoot weight, chlorophyll content, number of
seeds and seed weight) after a varimax rotation. The two axes explained 49% of the variation.
24 M. Díaz-Rojas et al. / Applied Soil Ecology 73 (2014) 19–25
the wastewater sludge (total N content 13.4 g kg−1 ) was mineral-
ized readily, which further increased the NH4 + content in soil and
thus nitrification. Additionally, the large microbial activity induced
by the mineralization of the organic material increased anaerobic
microsites inducing denitrification and thus emissions of N2 O.
Application of charcoal had no significant effect on emissions
of CH4 in this study. Similar results have been found by Scheer
et al. (2011) when charcoal was applied to a pasture soil in New
South Wales (Australia) in the field, but three years after the appli-
cation of the biochar (ageing effect). Castaldi et al. (2011) found
no effect of biochar application on emissions of CH4 in a field of
a Mediterranean wheat crop immediately after biochar applica-
tion (no ageing effect) and Wu et al. (2013) also found no effect
of biochar on CH4 emission from an agricultural soil (no ageing), in
a laboratory study after 100 days. However, application of charcoal
to paddy soils reduced emissions of CH4 in a laboratory study of 49
days (Zhang et al., 2010; Liu et al., 2011), to paddy and forest soils,
but depending on the soil water content in a 84 days incubation
experiment (Yu et al., 2013) and when added at very high concen-
trations (>20%, w/w) to an agricultural soil in a 100 day laboratory
experiment (Spokas et al., 2009).
Fluxes of CH4 from arable soil are normally low and mostly oxi-
dation of CH4 is larger than production of CH4 so arable soils are a
sink for CH4 (Smith et al., 2011). Application of organic wastes to
soil might stimulate emission of CH4 , but not always even when
applied to poorly drained soils (Thangarajan et al., 2013). Sistani
et al. (2010) applied swine effluent to soil and found an increase in
emission of CH4 . However, Contin et al. (2012) found that methane
oxidation rates from an arable and a grassland soil that received
7.5 mg ha−1 y−1 aerobically treated sewage sludge for ten years
were similar to those found in an unamended control soil. In this
study, application of 39 g wastewater sludge kg−1 soil did not
change the CH4 emission.
4.2. Soil and plant characteristics
Charcoal had no significant effect on soil characteristics in this
study, although various effects have been reported (Yuan et al.,
2011). An increase in soil pH has been reported after charcoal was
applied to soil, but no such effect was found in our study (Van
Zwieten et al., 2010; Bell and Worrall, 2011). It has been reported
that charcoal can affect nutrient availability in soil (Kookana et al.,
2011), but the mineral N content was not affected by charcoal
application in this study. On the one hand, mineralization of char-
coal with a high C-to-N ratio will induce N immobilization, but if
the C-to-N ratio is low then N mineralization will occur increasing
mineral N content in soil (Novak et al., 2010).
It has been reported that charcoal stimulated plant growth and
can increase crop yields (Kammann et al., 2011). For instance, Jha
et al. (2010) reported increases with 45% to 250%, Uzoma et al.
(2011) reported a 150% and 98% increase in maize yield when soil
was mixed with 15 and 20 t charcoal ha−1 in a greenhouse exper-
iment after 85 days, while Vaccari et al. (2011) reported increases
up to 30% in biomass production and yields of durum wheat in
two consecutive years when soil was applied with 30 and 60 t
charcoal ha−1 . However, no such effect was found in this study
even though the cases mentioned above were done in similar
conditions (no ageing). Application of charcoal might affect plant
growth in several ways. First, mineralization of the charcoal might
increase the nutrient content in soil thereby stimulating plant
growth. Second, charcoal might improve soil structure and aera-
tion thereby facilitating plant growth and increasing plant yield.
As mentioned before, the charcoal did not affect the soil nutri-
ent content and its effect on soil structure did not stimulate crop
growth.
Application of organic material can improve physical, chemical
and biological soil ‘fertility’ (Thangarajan et al., 2013). Soil struc-
ture improves, soil nutrient content and soil microbial activity
increases when organic material is applied to soil. Consequently,
plant growth is faster and crop yields increase. In the study reported
here, however, application of wastewater sludge did not affect
wheat growth compared to plants amended with urea. It is clear
that the characteristics of the wastewater sludge, i.e. nutrient,
micro-elements and humic acid content, might change over time so
that a possible effect on plant development changes. Additionally, it
might be that a positive effect is determined by the cultivated crop.
For instance, growth and yield of common bean (Phaseolus vulgaris
L.) increased when fertilized with wastewater sludge compared to
plants fertilized with urea (Fernandez-Luqueño et al., 2010), but not
with sunflower (Helianthus annuus L.) (López-Valdez et al., 2011).
5. Conclusions
Application of charcoal had no significant effect on the emission
of the GHG in a soil cultivated with wheat and fertilized with urea or
wastewater sludge, but application of wastewater sludge increased
the GHG. Wheat development and yields, and soil characteristics
(pH and electrolytic conductivity) were not affected by charcoal
application.
Acknowledgements
The research was funded by Cinvestav (México). M. D.-R. and
V.M. R.V. received grant-aided support from ‘Consejo Nacional de
Ciencia y Tecnología (CONACyT)’ while M.R. C.-A., E. H.V. from ‘Sis-
tema Nacional de Investigadores (SNI)’ México.
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