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Influence of biochar on drought tolerance of Chenopodium quinoa Willd and

on soil-plant relations

Article in Plant and Soil · August 2011

DOI: 10.1007/s11104-011-0771-5

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Plant Soil (2011) 345:195–210
DOI 10.1007/s11104-011-0771-5

REGULAR ARTICLE

Influence of biochar on drought tolerance

of Chenopodium quinoa Willd and on soil–plant relations
Claudia Irene Kammann & Sebastian Linsel &
Johannes W. Gößling & Hans-Werner Koyro

Received: 28 December 2010 / Accepted: 10 March 2011 / Published online: 5 April 2011
# Springer Science+Business Media B.V. 2011

Abstract The application of pyrogenic carbon, bio- and chlorophyll-concentrations. In this regard, plant
char, to agricultural soils is currently discussed as a win- responses to biochar closely resembled those to
win strategy to sequester carbon in soil, thus improving elevated CO2. However, neither soil- nor plant–soil-
soil fertility and mitigate global warming. Our aim was respiration was higher in the larger plants, indicating
to investigate if biochar may improve plant eco- less respiratory C losses per unit of biomass produced.
physiological responses under sufficient water supply Soil-N2O emissions were significantly reduced with
as well as moderate drought stress. A fully randomized biochar. The large application rate of 200 t ha−1
greenhouse study was conducted with the pseudo-cereal biochar did not improve plant growth compared to
Chenopodium quinoa Willd, using three levels of 100 t ha−1; hence an upper beneficial level exists. For
biochar addition (0, 100 and 200 t ha−1) to a sandy quinoa grown in a sandy soil, biochar application
soil and two water treatments (60% and 20% of the might hence provide a win-win strategy for increased
water holding capacity of the control), investigating crop production, GHG emission mitigation and soil C
growth, water use efficiency, eco-physiological param- sequestration.
eters and greenhouse gas (GHG) fluxes. Biochar
application increased growth, drought tolerance and Keywords CO2 gas exchange . Halophyte crop .
leaf-N- and water-use efficiency of quinoa despite Biochar . Water use efficiency . Nitrogen use
larger plant–leaf areas. The plants growing in biochar- efficiency . N2O emission . Quinoa
amended soil accumulated exactly the same amount of
nitrogen in their larger leaf biomass than the control Abbreviations
plants, causing significantly decreased leaf N-, proline- BC Biochar
WUE Water use efficiency
Responsible Editor: Johannes Lehmann. NUE Nitrogen use efficiency
C. I. Kammann (*) : S. Linsel : J. W. Gößling :
WHC Water holding capacity
H.-W. Koyro SOC Soil organic carbon
Department of Plant Ecology,
Justus-Liebig-University Gießen,
Gießen 35392, Germany
e-mail: [email protected]
Introduction
C. I. Kammann
School of Biology and Environmental Sciences,
University College Dublin, Atmospheric CO2 concentrations have already in-
Dublin, Ireland creased from 275 ppm in preindustrial times to
196 Plant Soil (2011) 345:195–210

387 ppm today which is higher than at any time soils (Blackwell et al. 2009; Major et al. 2010), (c) it
during the past 20 million years, resulting in global reduces nutrient leaching (Chan et al. 2007, 2008;
warming (IPCC 2007b). At the same time, arable land Laird et al. 2009; Steiner et al. 2008). Additionally it
areas worldwide decline by soil erosion, drought, could be shown (d) that the cation exchange capacity
salinization, loss of soil organic carbon (SOC) (CEC) of soils increases with BC addition (Liang et
contents (Lal 2004; IPCC 2007a; Kimetu et al. 2009) al. 2006), in particular over time as the functional
or other forms of degradation. Global warming and a groups are oxidized (Cheng et al. 2006).
fast-growing world population intensify the need to Although using BC seems to be promising, and
develop solutions for our future food and energy despite the fact that several international projects have
needs (Mathews 2008; Hansen et al. 2008; Lal 2009). been initiated, there is still a considerable lack of
In recent years, application of black, charred knowledge on its effects and their causes (Blackwell
carbon (in the following termed ‘biochar’) has been et al. 2009). In particular, the plant physiological
increasingly discussed as a mitigation strategy for response (other than crop yield) to BC in soils
sequestering recalcitrant carbon into agricultural soils, remains poorly understood and has, to our knowl-
which can, at the same time, improve soil fertility edge, seldom been investigated (Elad et al. 2010;
(Glaser et al. 2002; Marris 2006; Lehmann 2006, Graber et al. 2010).
2007a, b). The idea originates in Amazonian dark Before BC can be applied large scale in agricultural
earth or Terra preta research (Glaser et al. 2001; practice, possible counterproductive effects must be
Marris 2006). Terra preta (TP) soils enable several investigated. Negative effects could theoretically lead to
harvests per year without extra fertilization, or the increased greenhouse gas (GHG) fluxes of CO2, CH4 or
need to move and cut new forest after a few years N2O, (Wardle et al. 2008; Clough et al. 2010) or
(Glaser 2007; Steiner et al. 2008). TP soils have reduced plant stress resistances, e.g. drought tolerance
distinct bacterial communities with a significantly with improved water supply. However, first lines of
greater species richness (Kim et al. 2007), exhibit evidence suggest that N2O emissions may decline
significantly larger cation exchange capacities (Glaser rather than increase with BC addition (Lehmann
et al. 2001; Steiner et al. 2008), contain significantly 2007a; Spokas et al. 2009; van Zwieten et al. 2010).
higher phosphorus amounts, and have larger stocks of Increased soil CO2 effluxes may result from soil
soil organic matter besides the black carbon than near- organic carbon (SOC) decomposition via "priming"
by Ferralsols (Glaser et al. 2001), suggesting that of old soil carbon (Kolb et al. 2009; Wardle et al.
additional C sequestration in soil organic matter has 2008). In addition, suboptimal large application rates
occurred. of BC may lead to other negative effects, or less
Modern pyrolysis techniques, which are currently positive responses (compare Rondon et al. 2007), e.g.
undergoing a rapid technical development (Laird et al. by nutrient immobilization at BC surfaces or pH
2009) allow energy production from syngas (mainly changes (Chan and Xu 2009; Laird et al. 2009).
CO, H2 and CH4 and other hydrocarbons) and/or Beside the investigation of possible productive BC
liquid–fuel production while simultaneously generat- effects on arable land, in particular BC effects on less
ing different types of biochar (abbreviated BC in the productive soils must be investigated because there is
following; Gaunt and Lehmann 2008; McHenry also need to extend arable land into less suitable
2009). The resulting biochars can greatly differ in areas, or to avoid further desertification. Drought is a
their material properties (CHO-concentrations, aro- worldwide problem, seriously constraining global
maticity, cation exchange capacity, pH, nutrient crop production and quality. It is well known that
contents, porosity, energy density etc.), depending soil characteristics influence plant communities
on feedstock and pyrolysis conditions (Amonette and through water relations (Sperry and Hacke 2002).
Joseph 2009, Downie et al. 2009). Experimental The addition of BC to sandy soil changes soil
evidence so far shows that (a) BC is quite stable and characteristics such as its texture and porosity.
hence principally suitable for C sequestration (Cheng Hypothetically, finer textured soils (after BC addition)
et al. 2008; Kuzyakov et al. 2009; Major et al. 2010), in arid climates should be associated with more
(b) BC addition often promotes plant growth, in negative plant and soil water potentials during
particular combined with N-fertilizer addition in poor drought, inducing a greater resistance of xylem to
Plant Soil (2011) 345:195–210 197

cavitation, and shallower root systems than coarse and 0 (control), 100 or 200 t ha−1 BC application rates
soils. (n=24 pots in total). The unusually high rate of 200 t
Therefore, the aim of this study was the investiga- ha−1 (compare Chan et al. 2007, 2008) was chosen to
tion of BC application effects on plant responses such investigate if there is an upper limit of BC addition,
as water relations, C-, N-content or gas exchange and which has negative effects on plant growth. Temper-
plant–soil interactions including greenhouse gas ature, relative humidity (RH) and light regimes were
(GHG) fluxes of CO2 and N2O under high and low set to 22±2°C, 60±5% RH and >10.000 lux at a 16
water availability. and 8 h day-night cycle, respectively.
In recent years there has been a growing interest in To obtain a poor sandy soil medium, 1/4 (v/v) of a
introducing alternative crops in Europe able to resist sandy loam brown earth (obtained from Kieswerk
conditions of nutrient-poor soil, drought, salinization or Gießen, Germany) was mixed with 3/4 of pure
other forms of degradation. One such crop is quinoa washed sand (<1.4 mm particle size). Each of the 24
(Chenopodium quinoa Willd) originating from the pots (inner diameter 10.2 cm, height 20 cm) was filled
South American highlands and therefore considered with 2 kg of air-dried sandy soil (=1.872±1.2 g dry
as a hardy plant with good drought tolerance (Galwey soil). For BC treatments, 81.7 and 163.4 g dry BC
1989; Jacobsen and Stølen 1993; Jensen et al. 2000). It were added per pot and thoroughly mixed with the
was chosen as a suitable candidate for this study sandy soil, equivalent to 100 and 200 t BC ha−1 and
because it exhibits the attributes of drought tolerance, 20 cm ploughing depth, respectively. Thus, the soil
sustainability in the context of global changes and a surface in the biochar-amended pots was 2–5 cm
high economic potential. Until now, few investigations higher than in the control pots. The pots were made
have been performed to study the drought tolerance of from commercial polyethylene pipes capped at the
quinoa under controlled conditions. bottom with five draining holes per cap. The biochar
We studied the validity of the following hypothe- (particle size <2 mm) had been purchased from
ses: (1) BC addition will promote comparatively more EPRIDA, Athens, USA, where it had been produced
plant growth under limited water supply than under from peanut hull residues at 498°C and 26269 Pa
good water supply. (2) BC amendment will not alter (0.263 bar), with a biomass feed rate of 16.3 kg hr−1,
basic plant parameters such as the leaf N or C content, a steam temperature of 550.7°C and a steam flow rate
the relative chlorophyll content, transpiration or of 10.2 kg hr−1 (Fig. 1a). The total C and N contents
respiration. (3) Soil-derived CO2 effluxes and N2O were 71.6% and 1.84%, determined following ISO
emissions (based on denitrification) may be stimulat- 10694 and 13878, respectively. The pHCaCl2 and
ed at first, due to priming (Wardle et al. 2008) or to pHH2O were 7.6 and 8.1, respectively. The macro
initial oxidation of labile C fractions on the BC nutrient contents (in g kg−1 biochar) were: K 18.7, Ca
surfaces (Cheng et al. 2006). Later (4), N 2 O 5.41, Mg 2.83, P 2.13, S 0.83 (all: analysis according
emissions might be reduced, but CO2 effluxes of to ISO 11885) and CaCO3 <10 (ISO 10693); Gaskin
the plant–soil systems might continue to increase, et al. 2010 report quite similar values for a peanut hull
when BC-grown plants (with an unchanged respira- biochar also produced by Eprida at 400°C. The micro-
tion per leaf area) become larger than controls. (5) A nutrients or heavy metal contents (in mg kg−1
very large BC application dose will have negative biochar) were as follows: Al 2900, As 0.545, Cd
effects, e.g. via N immobilization (Chan and Xu 0.05, Cu 21.6, Fe 1190, Hg <0.01; Ni 3.75, Pb 1.58
2009). and Zn 42.1 (all: analysis according to ISO 11885).
Thus, with 100 t ha−1 biochar, 1,840 kg of N ha−1 and
1,870 kg K ha−1 were applied. Gaskin et al. (2010)
Material and methods conclude from their results that not much of the
applied biochar-N must have become plant-available
Experimental setup and growth conditions via mineralization, despite a rather low C/N ratio of
38 (here: 39) for a plant-residue based biochar.
The greenhouse study was designed as a completely Potassium, on the other hand, had in the first year
randomized experiment with n = 4 replicates per contributed to higher tissue-K+ contents but not in the
treatment, with sufficient or reduced water supply second year (Gaskin et al. 2010). Hence the peanut
198 Plant Soil (2011) 345:195–210

below). The solution contained 8% N including

3.7% NH4+, 2.3% NO3− and 2% carbamid-N, 8%
P2O5, 6% K2O, 0.01% B, 0.007% Cu, 0.001% Mo
and 0.005% Zn.
The water holding capacity (WHC) of the soil-
biochar mixtures was determined by submerging the
entire pots for 24 h, and subsequent draining for another
24 h, and weighing (n=8 per BC treatment) before the
experiment started. Five seeds of Chenopodium quinoa
Willd were sown into each pot and seedlings were
counted per pot four times during germination. After
9 days (defined as day zero), when seedlings had about
four leaves and were approximately 5 cm high, all but
the largest seedling per pot were weeded. A WHC of
60% of the control was chosen as “sufficient water
supply” treatment, i.e. pots were watered regularly to a
target weight set to 60% of the WHC of the control
soils without BC. For a “low water supply” treatment,
pots were allowed to further dry out during seedling
growth in the first 3 weeks of the culture. The low-
water supply pots first dried down to 15% WHC.
However, since plants showed severe wilting symp-
toms, the “low water” treatment was adjusted to 20%
of the WHC of the control (also in case of the BC
treatments) from day 27 on. Afterwards, the pots were
weighed every 1 to 2 days, the respective water loss
was recorded, and water was applied to achieve the
desired target WHC"s of 60% and 20% of the control.
Since the WHC of the BC-amended soils was
significantly larger this treatment ensured that a
potential benefit from simply “more water per pot in
the BC-amended soils” was excluded.
On day 28 and 29, two plants were accidentally
broken during respiration measurements: The respec-
Fig. 1 a Scanning electron microscope (SEM) picture of the tive pots 17 and 22, a BC-200-wet and a BC-200-dry
structural characteristics of the peanut hull biochar with narrow treatment were therefore not included in subsequent
pores and gaps; b Sampling for greenhouse gas (GHG) flux
measurements, figure is partially sliced. FC: foam coating; HS:
analyses.
headspace; PE: grey PE capping; PP: plant pot; RS: rubber
septum; S: syringe for gas sampling (details not to scale) Measurements performed during plant growth

hull biochar was expected to improve cation nutrition Greenhouse gas (GHG: CO2, N2O, CH4) flux mea-
but not to contribute significant labile nitrogen surements on the entire plant–soil system as well as
amounts over the course of this study. To avoid side gas exchange measurements on the green plant parts
effects due to either nutrient limitations, or probably were performed during the study. For the GHG fluxes,
due to an improved micro-nutrient supply with BC pots were covered tightly by grey PE chambers
addition, a compound-fertilizer solution including constructed from pipes, closed with a lid, and fitted
micro-nutrients was applied in three doses (50, 25 with a rubber septum to allow gas sampling via
and 25 kg N ha−1) with the regular watering on days syringe (Fig. 1b). Chambers or chamber extensions of
1, 25 and 29, respectively (day numbering: see 10–20 cm length were fitted gas-tight to the pots via
Plant Soil (2011) 345:195–210 199

elastic rubber seals or a rubber gasket. During Plant H2O/CO2 gas exchange measurements
chamber closure, pots were set onto soft, water- at the leaf level
proof sealing foam to close the pot bottom. Gas
samples (50 ml) were withdrawn with 60 ml PE Plant gas exchange characteristics were measured
syringes (Pastipak®, Medica, Germany) three times exemplarily on one plant leaf per treatment. Fully
after 0, 2 and 4 h of closure; temperature and air expanded upper leaves (10–15 days old) were inserted
pressure were recorded. Samples were analyzed into a hermetically sealed compact minicuvette
within 24 h on a gas chromatograph fitted with an system (Waltz, Effeltrich, Germany) with a 500 ml
electron capture detector (ECD) and a flame ioniza- cuvette, type GK-022. Leaf surfaces were positioned
tion detector (FID: Shimadzu GC-14B; analytical horizontally towards a polychromatic lamp, type LA-
scheme after Mosier and Mack 1980) and 4, within a distance of 0.05 m. Photosynthetically
an automated sample-injection unit for syringes active photon flux density (PPFD) on the lamina was
(Loftfield et al. 1997). The exact chamber headspace stepwise increased by light filters NG3 (28 μmol m−2
was calculated from the volume of the chamber itself, s−1), NG4 (162 μmol m−2 s−1), NG5 (555 μmol m−2
the respective extension tube, and the pot space s−1), NG11 (1,031 μmol m−2 s−1) up to full light
between pot brim and soil surface (V=π * r² * h; (2,050 μmol m−2 s−1). The cuvette was covered with a
h determined as the mean of five measurements per non-transparent black rag for dark conditions. CO2
pot surface). Fluxes were calculated according to concentration in the cuvette was held constant at 380
Hutchinson and Mosier (1981) by linear regression of ±5 μl l−1 by purifying ambient air via potassium
the concentration increase inside the chambers and permanganate and afterwards adding CO2. CO2 and
related to the total soil weight (respective the soil H2O concentrations were detected by infrared absorp-
surface) of the pots. Methane uptake rates were near- tion using a dual comparison analyzer (Binos 100,
zero at the detection limit and hence not reported. and CO2 analyzer, type Binos4P, Rosemount, Hanau,
To measure the total aboveground plant respiration, a Germany). Data acquisition was carried out at the
LI-8100 soil CO2 efflux chamber (LICOR, Nebraska, steady state H2O concentration. Apparent photosyn-
USA; with the small chamber for 10.2 cm diameter soil thesis efficiency (A), stomatal conductance (cond.)
frames) was employed 1 day prior to harvest. and transpiration rate (transp.) were calculated (based
Immediately before measurement the soil surface on Fick´s law of diffusion) using data of CO2/H2O
around the Quinoa shoot was covered by a flexible concentration variation, ambient air pressure, leaf
diffusion-tight foam disc with a fitting slit. Next, an temperature and leaf area. Leaf water use efficiency
extension tube was fitted air-tight with a rubber gasket was calculated as the ratio of transpiration rate to
to the pot and the LI-8100 10-cm diameter survey apparent photosynthesis efficiency. Calculation of the
chamber was set on top above the plant. The offset leaf respiration rate (Rleaf) and maximum photosyn-
(height) of the entire pot-extension-tube system was thesis efficiency (Amax) were done by curve fitting
entered into the LI-8100-driving software for correct (SigmaPlot, Sysstat Software Inc., Richmond, USA)
flux calculations. Measurement time and observation using the asymptotic saturation function described by
delay were set to 120 s and 40 s, respectively, to allow Schulte and Brooks (2003).
sufficient chamber-volume mixing time. CO2-concen-
tration increase always showed a linear slope with R² Quantification of LSU-RuBisCO
>0.9. The respiration rate was expressed per m² (of pot
area, automatic function of the LI-8100), but also per g Juvenile leaves (one sample per treatment) were ground
of aboveground dry biomass, alternatively per m² of in liquid nitrogen. A micro spoon of polyvinyl-
total leaf area, i.e. based on values that were obtained polypyrrolidon (PVPP) was added. Subsequently, pro-
1 day later at the final harvest. tein digestion was performed according to Granier
The relative chlorophyll content was measured (1988) in cold (0°C) extraction buffer (100 mM
four times on day 23, 31, 37 and 49, respectively, Imidazol and 1.25 mM EDTA; pH 7.8). Next, 75 μg
with five replicated measurements on three leaves Bovine serum albumin (BSA) was added as an internal
(youngest fully expanded leaf) per plant using a standard. Low range proteins (14.3 to 220 kDa) were
SPAD 502 device (Minolta, USA). separated using sodium dodecylsulfate polyacrylamide
200 Plant Soil (2011) 345:195–210

gel electrophoresis (SDS-PAGE) according to Laemmli sured, the total leaf number per plant was counted, and
(1970), containing a 6% acrylamide stacking gel and the total leaf area per plant was determined with a
12.5% acrylamide separation gel. An internal SDS- planimeter (type LI-3000A, Licor, Nebraska, USA).
PAGE molecular weight standard (BIO-RAD Labora- The dry leaf biomass was ground to powder with a
tories GmbH, Munich, GER) was used for calibration. Retsch type MM ball mill (Retsch, Düsseldorf,
Protein staining was done with Coomassie brilliant Germany). For the quantification of carbon and nitro-
blue R-250 (B2025-1EA Sigma-Aldrich, St. Louis, gen, approximately 50 mg dry powder was analyzed by
USA) for 45 min. Gels were de-stained in 10% acetic combustion with a macro CNS analyzer (type Vario
acid and digitalized with a flat bed scanner after 1 day. MAX, Elementar Analysensysteme GmbH, Hanau,
After detection of the molecular weight quantification Germany). Leaves that were removed for subsequent
took place by integrating the signal strength of all analyses of RuBisCO, proline and osmotic potential
impulses of a band with the image processing and were weighed, their area was measured, and the weights
analysis program ImageJ (National Institute of Health, added to the final harvest results. The leaf nitrogen use
Maryland, USA). The large subunit of Ribulose-1,5- efficiency of productivity NUEProd, i.e. the net primary
bisphosphate-carboxylase/-oxygenase (LSU) was iden- production (NPP) per unit of nitrogen absorbed
tified as a band in the range of 53 kDa (Ishida et al. (Golluscio 2007) was calculated as aboveground dry
1997). The calculated total amount of LSU-RuBisCO matter produced per mg leaf-N.
was corrected with the BSA recovery rate (internal Osmotic active substances were measured in the
standard, s. a.). press-sap of approx. 1 g fresh leaf material with a
cryo-osmometer (Osmomat 030, Gonotec GmbH,
Quantification of proline Berlin, Germany). Subsequently, cationic macronu-
trients such as K, Ca, Mg, and Na were measured in
Proline content was determined according to Bates et al. diluted solutions (1:10 or 1:100v/v) with a flame
(1973) 1 day after harvest. Proline was extracted from atomic absorption spectrophotometer (PE2100, Perkin
approximately 50 mg (exact weight noted) of leaf dry Elmer, USA). Anionic macronutrients (such as nitrate,
mass by homogenizing in 10 ml of 3% (v/v) phosphate and sulfate) were determined by ion-
sulfosalicylic acid using a liquid nitrogen-chilled exchange chromatography (Metrohm 690 ion chro-
mortar and pestle. After filtration 1 ml of filtrate was matograph, Metrohm, SUI).
added to 1 ml glacial acetic acid and 1 ml reaction
dilution (0.63 g Ninhydrin dissolved in 15 ml glacial Statistical analysis
acetic acid, 10 ml 6 M phosphoric acid). After agitation
and incubation in a water bath at 100°C, the reaction Effects of the two water supply treatments and three
was terminated in ice. The formed colour complex was levels of BC addition on all fully replicated measure-
extracted with 2 ml of toluene. After vortexing ments were tested via two-way analysis of variance
absorbance of the toluene extract was recorded at (ANOVA). Significance of differences among treatment
546 nm (Beckman photometer, Beckman Coulter inc., groups was determined with the Tukey test. Data were
Fullerton, USA) and final proline concentration was log- or root-transformed to normality if necessary.
calculated on basis of a standard curve (proline Differences at the P<0.05 level are reported as
buffered and solved in 3% (v/v) sulfosalicylic acid). significant and P<0.1 results are reported as trends.
All statistical tests were carried out with SigmaPlot
Harvest 11.0 (Systat Software, Inc., Richmond, USA).

Total aboveground plant biomass was harvested on day

50 post germination. Plants had not yet reached full Results
maturity, where they become senescent and dry, but had
already begun to produce seeds. Fresh weight and dry Plant and soil variables at the harvest date
weight (at 105°C to weight constancy) were determined
of each plant separated into leaves (including stem), The water holding capacities were 0.223, 0.276 and
shoots, tap roots and seeds. Stem heights were mea- 0.304 g H2O g−1 soil (mixture) dry weight (dwt) in 0,
Plant Soil (2011) 345:195–210 201

100 and 200 t ha−1 soil-BC mixtures, respectively. The highest absolute WUEProd value was reached at
The BC application significantly increased the WHC 100 t ha−1 BC application and reduced water supply
by 23.9% and 36.1% compared to the control (one- (Fig. 2a: 300% of the well-watered control). Lower
way ANOVA, P<0.001, n=8). After the target value water supply reduced leaf nitrogen use efficiency
of WHC had been reached (see methods), the total or (NUEProd, aboveground dry matter produced per mg
average daily water consumption of the plants over leaf-N) compared to high water supply (F=51.43, P<
the experimental duration was significantly lower in 0.001), however, BC application significantly in-
all 20% WHC compared to the 60% WHC treatments. creased NUEProd in both water treatments (F=28.02,
In addition, there was a small, but non-significant P<0.001), with a significant water×BC interaction
reduction in water consumption with BC application (F=5.415, P=0.016).
(Table 1). Leaf N concentration, leaf proline concentration
As expected, the reduced water availability (WHC (Fig. 3a, b) and the relative chlorophyll content in
20%) significantly affected nearly all measured leaves (Table 2) were all significantly reduced with
variables (Table 1; Figs. 2a, b, 3a), with the exception BC application. However, the total amount of N that
of the leaf mass to area ratio (LMA, g m−2, Table 1) taken up into the leaves was nearly identical in all
and the leaf proline concentration (Fig. 3b). treatments; it was neither effected by BC- nor by
Biochar application significantly increased the total water supply-treatments (BC: F=1.051, P=0.372;
leaf area and leaf biomass per plant in both water WS: F = 0.179, P = 0.678; over-all mean: 48.6 ±
treatments (Table 1), while the total number of leaves 6.2 mg leaf-N plant−1). Biochar hence significantly
per plant tended to increase with BC application (P= widened C to N ratios in plant leaves (P<0.001),
0.090, Table 1). Although the mean area per leaf was mirroring the reduction of leaf N concentration; the
larger with BC application, the increase was not high leaf C concentration was unchanged (not shown).
enough to become significant. With reduced water
supply the average area per leaf tended to be reduced Treatment effects on the plant gas exchange
(P=0.062, Table 1). The biomass of all plant parts
and also of the total biomass (Table 1) were Plant gas exchange measurements at the leaf level
significantly increased by BC application (P values were performed with one plant per treatment (Table 2),
between 0.022 and <0.001). Tap root biomass also i.e. are unreplicated. However, each given variable is
increased significantly (compared to the respective logically connected to one or more repeated variables
controls) and, in contrast to other parameters, linearly (e.g. transpiration with water consumption and leaf
(P<0.001) with increasing BC application rates under area, Amax with N concentration and SPAD etc.). The
both water treatments, i.e. by 88% and 191%, or by stomatal conductance (cond.) was high at good soil
63% and 133% at WHC’s of 60% or 20%, respec- water availability (Table 2). Low soil water availabil-
tively. Plant height was significantly lower with ity (20% WHC) generally decreased the stomatal
reduced water supply, but unchanged due to BC conductance, resulting in a decrease of transpiration
treatment, although the leaf area per plant was rates, increased the maximum apparent photosynthe-
significantly larger with BC application (Table 1). sis (Amax) at light saturation, and decreased the leaf-
H2O content in leaves tended to be higher at 20% level respiration (RLeaf) compared to the higher water
WHC, and tended to be lower with BC application, in availablility. Within both soil water availability levels,
particular at 60% WHC. Water use efficiency of BC seemed to decrease transpiration and also Amax
productivity (WUEProd), i.e. the total amount of compared to the un-amended control, while RLeaf was
aboveground biomass (without tap roots) produced visibly reduced only with the highest BC application,
per H2O consumed, generally increased with reduced respectively. Without BC addition the RuBisCo
water supply (F=228.09, P<0.001; Fig. 2a). Biochar concentration (per g fresh weight) was higher at
application further significantly increased (in compar- 20% than 60% WHC, respectively. The relative
ison to the respective controls) WUEProd (F=63.72, change of RuBisCo concentration between zero-BC-
P < 0.001) by +54% and +62%, and by +65% and highest BC-treatment was much higher than the
and +52% in well-watered and reduced-watered BC- change of Amax suggesting an increased RuBisCo
100 and BC-200 treatments, respectively (Fig. 2a). content.
Table 1 Plant variables measured at the harvest of Chenopodium quinoa plants grown at sufficient and reduced water supply (i.e. 60% and 20% WHC of the control without BC,
202

respectively), and with application rates of 0, 100 and 200 t/ha biochar in each water supply treatment. Statistical results: Two-way ANOVA"s, post-hoc Tukey test. A result of P<0.05 is
considered significant (different uppercase letters after treatment means), and a result of P<0.1 is reported as a trend (not reflected in different uppercase letters). dm = dry matter

Measured variable per pot (units) Treatment means Treatment factor, interaction

WHC 60% WHC 20% Water supply BC applicat. Water x BC

BC-0 BC-100 BC-200 BC-0 BC-100 BC-200 F P F P F P

∑ water consumpt.X (ml) 1604a 1484a 1555a 677b 508b 491b 249.8 <0.001 2.18 0.146 0.39 0.686
a a a b b
plant height (cm) 65.0 72.6 70.5 50.7 49.2 47.5b 83.35 <0.001 0.72 0.503 1.95 0.174
Y
∑ biomass (g) 3.43b 4.95a 5.45a 2.66b 3,21b 2.94b 48.44 <0.001 9.61 0.002 4,22 0,034
biomass leaves (g dm) 1.57b 2.30a 2.67a 1.27d 1.59c 1.58c 36.73 <0.001 13.41 <0.001 3.69 0.048
b a a d c c
biomass shoots (g dm) 1.76 2.41 2.58 1.26 1.48 1.26 48.40 <0.001 4.92 0.022 3.14 0.071
biomass seeds (g dm) 0.10b 0.25a 0.20a 0.13b 0.14b 0.10b 13.21 0.002 8.96 0.002 7.96 0.004
biomass tap roots (g dm) 0.33c 0.63b 0.97a 0.25f 0.41e 0.58d 21.86 <0.001 32.30 <0.001 3.20 0.065
−1 b a a d c
∑ leaf area (cm² plant ) 563 666 728 412 501 494c 44.23 <0.001 7.52 0.005 0.80 0.468
leaf number (plant−1) 63.0a 70.5a 73.0a 48.8b 57.3b 52.3b 26.93 <0.001 2.81 0.090 0.51 0.607
−1 a a a a a a
mean leaf area (cm² leaf ) 7.17 8.28 8.54 6.50 7.32 7.36 4.01 0.062 2.32 0.131 0.095 0.910
leaf mass area (g m−2) 28.0a 34.3a 36.8a 31.3a 31.5a 31.7a 0.902 0,356 2,81 0.09 2.313 0.131
Z
H2O in biomass (g H2O g−1 dm) 5.10a 4.78a 4.12a 4.21a 4.28a 4.37a 2.50 0.134 0.935 0.413 1.802 0.197
X
Sum of water (ml) consumed from day 27 on (after target WHC had been established, see methods)
Y
Aboveground biomass: sum of shoots, leaves and seeds, in g dry matter
Z
Water content of the total biomass of leaves, shoots and seeds; water content of the leaves was not significantly different between treatments
Plant Soil (2011) 345:195–210
Plant Soil (2011) 345:195–210 203

Fig. 2 a Mean water use efficiency of productivity (above- under high and low water availability, and with biochar
ground yield per H2O consumed) and b leaf-nitrogen use application rates of zero, 100 and 200 t biochar ha−1,
efficiency (aboveground dry matter produced per mg leaf-N) respectively; different letters indicate significant differences
plus one standard deviation of the mean of quinoa plants grown between treatments

Leaf osmotic active compounds effect, F = 27.68, P < 0.001), but BC treatment
resulted in no significant effects (F= 2.488, P=
Low water supply significantly increased the os- 0.133). Sulfate concentrations also followed the
motic value (Fig. 4a; WS: F=62.22, P<0.001), same increasing trend as sucrose (WS: P<0.001,
which further increased when BC was present, in BC: P=0.064). Most other osmotic active substances
particular at reduced water supply (BC: F=11.48, were in tendency or significantly increased by the
P=0.002; WS×BC: P=0.067). Improved osmotic lower water supply, but significantly decreased by
values in leaves with BC application were mainly BC application (Cl−: P=0.007; Na+: P<0.001; Ca2+:
caused by higher potassium concentrations (Fig. 4b, P<0.001; Mg2+: P<0.003). Phosphate tends to be
F=65.35, P<0.001) and sucrose. Here, the all-over reduced at low-water treatment (WS × BC: P =
trends were the same as for K+ (significant WS 0.081). Nitrate was unchanged.

Fig. 3 a Mean leaf N concentration, and b mean leaf proline and with biochar application rates of zero, 100 and 200 t
concentration per g leaf dry matter plus one standard deviation biochar ha−1 respectively; different letters indicate significant
of quinoa plants grown under high and low water availability, differences between treatments
204 Plant Soil (2011) 345:195–210

Table 2 Fully replicated (chlorophyll) and un-replicated 11.0); Amax (= maximum photosynthesis at light saturation
measurements of plant gas exchange parameters (i.e. one plant (2,200 μmol m-2 s−1), transp. = transpiration at light saturation;
per treatment). Un-replicated measurements are only reported if RuBisCo = ribulose-1,5-bis-phosphate-carboxylase/oxygenase
they correspond to related, replicated parameters (e.g. to (fwt = fresh weight); WUEp = water use efficiency of
RuBisCo to leaf N and chlorophyll concentration). WS = water photosynthesis, measured at the leaf level (μmol m−2 s−1
supply; BC = biochar application rate (see Table 1); parameters CO2/mmol m−2 s−1 H2O); Cond. = stomatal conductivity; Rleaf =
Amax and Rleaf were calculated from light curve fits (SigmaPlot leaf dark respiration

WS BC Chlorophyll Amax RuBisCo Transp. WUEp Cond. Rleaf

WHC% (t/ha) (relative) μmol m−2 s−1 mg * g fwt−1 mmol m−2 s−1 (Amax/transp.) mmol m−2 s−1 μmol m−2 s−1

60 0 43.1c 8.789 1.59 2.224 3.95 111.73 −0.287

60 100 34.8 d
7.910 1.65 1.781 4.44 111.18 −0.316
60 200 34.1d 8.033 1.28 1.958 4.10 113.61 −0.237
20 0 46.7a 9.139 2.53 1.746 5.23 92.37 −0.298
20 100 41.8b 8.692 1.00 1.543 5.63 79.88 −0.262
20 200 40.3b 8.225 0.27 1.357 6.06 61.66 −0.143
X
Mean relative chlorophyll content (WST: P<0.001; BC: P<0.001; WST x BC: P=0.081); the relative chlorophyll content was
measured on the 1st, 9th, 15th and 27th of July; all dates showed the same significance (WS and BC P<0.001)

Treatment effects on plant–soil CO2 efflux and soil respectively. This revealed that biomass-based Rplant
N2O emissions was significantly reduced in the BC-grown plants
compared to respective controls (Table 3). Based on
At the beginning of the experiment (day 15), BC total leaf area, Rplant was significantly reduced with BC
significantly increased plant–soil CO2 efflux (Rplant+soil, application in the 20% WHC treatment but not in the
P=0.013), however this was due to the100-kg BC 60% WHC treatment, as indicated by a significant
treatment at 60% WHC which had significantly larger water×BC interaction (Table 3).
CO2 efflux rates than all other treatments (Table 3). N2O emissions were significantly lower (and often
After application of the fertilizer, the effect of BC near the detection limit) with reduced water supply
vanished and remained non-significant in later measure- (Table 4). In the beginning of the study and directly
ments (Table 3). Rather, the BC treatments tended to after the first N-fertilizer application, BC-application
have lower soil respiration rates (Rsoil, P=0.077) did not reduce N2O emissions. However, BC reduced
expressed on a soil weight basis after harvest. After N2O emissions significantly during the later part of
the targeted low water supply treatment of 20% WHC the study (repeated measures ANOVA, Table 4).
of the control had been fully established, Rplant+soil or
Rsoil efflux rates at 20% WHC were significantly lower
(P<0.001) compared to the 60% WHC treatments. The Discussion
effect of the water supply (20% vs. 60% WHC) on the
aboveground plant respiration was significant, based on The plant response patterns and growth-increasing
the ground area (larger plants). It was much less mechanisms observed in this study with BC amend-
pronounced or absent, when based on the total plant– ments were surprising and did not match all initial
leaf area or the aboveground biomass, respectively hypotheses. As intended, the reduced water supply
(Table 3). BC application significantly increased above- treatment significantly impacted several measured
ground plant respiration with sufficient water supply, parameters without generating severe water stress.
but not with limiting water supply (related to the ground This is indicated by the lack of a significant change of
area, Table 3). Respiration was also calculated based on the proline concentrations (Ibarra-Caballer et al.
the aboveground biomass1 or on the total leaf area1, 1988) but increases in the osmotic value.
1 In accordance with hypothesis #1 (Blackwell et al.
Calculated with the total dry matter and leaf area at the
harvest date, but without the few leaves that were harvested 2009), application of BC did increase aboveground
earlier for proline- Rubisco- and osmotic potential analyses. biomass of quinoa by 10–61%. However, the largest
Plant Soil (2011) 345:195–210 205

Fig. 4 a mean osmotic value in the leaf press-sap and b under high and low water availability, and with biochar
concentrations of the main osmotic substances potassium (left, application rates of zero, 100 and 200 t biochar ha−1,
open bars) and sucrose (right, hatched bars) per leaf fresh respectively; different letters (a–d; or x–z) indicate significant
weight plus one standard deviation of quinoa plants grown differences between treatments

relative increase occurred at 60%- rather than at 20%- N-fertilizer use efficiency. Rondon et al. (2007)
WHC, contradicting hypothesis #1. In two green- reported increased N2 fixation, legume-biomass and
house studies with radish grown in a hard-setting bean yields on BC-amended soils. Lower water
Chromosol, Chan et al. (2007, 2008) observed consumption and larger biomass resulted in 160%
biomass improvements and a significantly improved up to 300% greater water use efficiencies with BC

Table 3 Respiration of the entire pots (plant and soil) at three area), of the aboveground plant biomass, or of the plants' leaf
dates during the study; respiration of the soil with remaining area. Expressing Rplant+soil or Rsoil on an area rather than a kg-
roots after the harvest (Rsoil); and respiration of the above- soil basis does not change the statistical results on any
ground plant biomass 1 day before the harvest, expressed as a measurement day, except for the BC-effect trend (P=0.077)
function of the entire system (m² pot-s.(surface) = m² ground on Rsoil which becomes non-significant

Respiration Mean respiration rate Treatment factor, interaction

variable, units
WHC 60% WHC 20% Water supply BC Water x BC
applicat.

BC-0 BC-100 BC-200 BC-0 BC-100 BC-200 F P F P F P

Rplant+soil, day 15 μg CO2 329b 685a 508b 378b 529b 652b 1.28 0.274 5.64 0.013 4.82 0.022
kg−1 h−1
Rplant+soil#, day 18 μg CO2 843a 1381a 964a 911a 768a 973a 2.04 0.170 0.84 0.450 3.03 0.074
kg−1 h−1
Rplant+soil, day 28 μg CO2 1695a 2107a 1699a 1313b 1057b 1111b 23.96 <0.001 0.56 0.583 2.06 0.156
kg−1 h−1
Rsoil, day 51 μg CO2 370a 326a 300a 201b 139b 110b 66.40 <0.001 3.03 0.077 0.90 0.428
kg−1 h−1
Rplant (per m² pot-s) μmol CO2 11.9b 16.0a 16.6a 9.5b 8.3b 7.3b 55.65 <0.001 1.16 0.339 5.70 0.014
m−2 s−1
Rplant (per g dwt) nmol CO2 28.37a 26.71b 24.81b 28.86a 21.44b 20.92b 2.35 0.145 3.53 0.054 0.93 0.42
g−1 s−1
Rplant (per m² leaf) μmol CO2 2.16a 2.24a
X
2.19a 2.50a 1.63b 1.60b 6.01 0.026 3.94 0.041 5.60 0.014
m−2 s−1

# One day after N fertilizer application; pot-s = pot surface (ground area)
X
relating Rplant to m² leaf area is not entirely correct since the entire aboveground plant material respired; however, the calculation is
provided to include the significantly larger leaf areas of biochar-grown plants (see Table 1)
206 Plant Soil (2011) 345:195–210

Table 4 Mean N2O flux rates (ng N2O-N kg−1 soil dwt h−1) day” was always significant (P<0.001). Post-hoc Tukey test:
measured during the study. Statistical results, single dates: Two- different uppercase letters following mean N2O flux rates
way ANOVA; several measurement dates (bottom): Repeated indicate significant differences at the P<0.05 level
measurement ANOVA; the within-subject factor “measurement

Day of flux measure-ment N2O flux rate, treatment means Treatment factor, interaction

WHC-60% WHC-20% water supply BC applicat. water x BC

BC-0 BC-100 BC-200 BC-0 BC-100 BC-200 F P F P F P

Day 15 24.7a 7.0a 9.8a 12.7a 7.4a 10.8a 0.003 0.955 1.38 0.277 1.12 0.349
a a a a a
Day 18 13.3 2.5 5.5 6.5 2.7 10.6a 0.276 0.605 0.570 0.575 0.218 0.806
Day 25# 58.7a 69.1a 51.0a 43.4b 48.9b 50.2b 6.08 0.024 1.25 0.310 1.41 0.271
a a b c c
Day 28 88.1 87.0 63.3 42.7 33.1 43.9c 57.30 <0.001 1.70 0.211 3.97 0.037
Day 51* 18.4a 3.9a 4.4a 2.8b 2.8b 2.7b 9.82 0.006 3.57 0.049 2.920 0.080
X a a b b b b
all 5 days 40.6 33.9 26.8 21.6 19.0 23.7 7.89 0.012 3.40 0.056 2.14 0.146
last 2 daysY 53.2a 45.4b 33.8b 22.7c 17.9c 23.3c 29.24 <0.001 4.67 0.023 2.83 0.085
#
N2O fluxes after N fertilizer application, see methods
* post hoc Tukey test: BC 100 and BC 200 tended to be lower than BC 0 (P=0.077 and 0.081, respectively)
X
Repeated measurement ANOVA with all five measurements made during the experiment, Y RM ANOVA with the last two
measurements (day 28 and 51); flux rates were log transformed before testing; means: flux rates averaged over the respective 5 or 2
dates; RM ANOVA (last 2 days), Tukey test results: BC 0 vs. BC 100, and BC 0 vs. BC 200: P=0.042 and P=0.041, respectively.

under good and low water supply, respectively. Increased potassium concentrations in leaves were
Hence, hypothesis #1 was not fulfilled in terms of either due to an improved K+ nutrition via the BC as
absolute biomass increases, but in terms of improved a nutrient carrier, to an increase of the osmolarity in the
relative water use efficiency. soil solution due to BC application or to better binding
Several mechanisms might contribute to the larger and access of the K+ that was repeatedly supplied in
amount of quinoa biomass per water consumption with equal amounts to all treatments during the study.
BC amendments. The water balance of the plants was However, most of the K+ initially introduced with the
improved, likely due to several mechanisms: Biochar BC will likely have been lost when the soil mixture
significantly increased the water holding capacity of the pots were flooded and drained (washed) during initial
sandy soil due to its porous nature (Fig. 1a; Cheng et al. the WHC determination.
2006, Downie et al. 2009, Glaser et al. 2002). A second reason for BC-mediated higher plant
However, in this study control-WHCs (60% and growth might be a reduced transpiration (Table 2).
20%) were used as target values. As a consequence, Together with increased osmolarity this might induce
the absolute water amounts in the BC-amended soils an improved drought tolerance. Biochar-plants used
were not larger than in the control soils. In line with the slightly less water despite larger leaf areas. The higher
findings of Gaskin et al. (2010), BC-addition also total-biomass-WUE was in line with a higher WUE of
increased the overall accumulation of osmotic active photosynthesis (WUEP).
substrances such as K+ in the plant tissues, likely due Significant larger leaf areas of BC-grown plants
to its large cation content, leading to an improved plant allowed higher C gain. They were based on non-
water uptake. Moreover, BC stimulated tap-root significantly larger mean leaf areas and leaf numbers,
growth (i.e. likely also fine root mass) and thus water respectively.
uptake from fine BC pores. Improved plant water The tap root biomass increased strongly with BC,
status with BC was also reflected by lower proline with a shift towards a more pronounced belowground
concentrations and higher osmotic values of the leaves, stimulation: the tap-root-to-shoot ratio showed a highly
indicating a higher tolerance to potential water stress significant effect of BC application (P<0.001), but no
conditions (Barker et al. 1993; Gonzalez et al. 2009). effect of the different water addition levels (P=0.287;
Plant Soil (2011) 345:195–210 207

without interactions). Larger root systems or root of the phytohormone ethylene from either the added
biomass of BC-grown plants have also been reported BC itself, or microbially mediated from the BC-
by e.g. Major et al. (2010). amended soils, a finding recently reported by Spokas
A further contribution could be a more efficient et al. (2010). This would be in line with the linear
leaf-nitrogen use. The nitrogen concentration of 3.3% stimulation response of the tap root biomass with
and 4% in the control leaves indicate that the applied increasing char application rates where the biomass in
N-fertilization was sufficient. However, the BC plants the 100 t ha−1 BC application was double, and that of
seemingly diminished the N-pool by stronger growth, the 200 t ha−1 BC application was three times that of
which leads to lower leaf-N concentrations. The latter the control biomass, respectively.
fits with the significantly reduced proline-, We anticipated that soil-derived CO2 effluxes
chlorophyll- and RuBisCo-concentrations and slightly would be initially stimulated (hypothesis #3) due to
reduced Amax rates in the BC plants. Hence, our priming (Wardle et al. 2008) or initial oxidation
results were in contrast to hypothesis #2 that these (Cheng et al. 2006). However, we found evidence
parameters remain unchanged. against such mechanism, since larger BC application
The reduced dark respiration rates per g of dry mass of 200 t BC ha−1 yielded in significantly lower CO2
were also in good accordance to the lower leaf-N effluxes than 100 t ha−1. In a recent 3.2 year lab study,
concentrations (Reich et al. 2006). Therefore, reduced the 14C-labelled BC lost less than <0.5% per year
leaf-tissue N concentrations could have employed indicating that BC was quite stable against degrada-
another growth-stimulating mechanism: reduced respi- tion (Kuzyakov et al. 2009). In our study, priming of
ratory carbon losses per unit of carbon gain, i.e. a pre-existing soil organic carbon (SOC) by BC
greater efficiency in the use of assimilated carbon. The application must have been too small, too short-
soil respiration tended to decrease in presence of BC. lived, or simply non-existent to be detectable in the
This was surprising and in contrast to our expectation respiration rates (compare Lehmann et al. 2009; Liang
of increased soil respiration with larger plants (hypoth- et al. 2010). In agreement with our findings, signif-
esis #3). In summary, BC application improved quinoa icantly reduced soil CO2 respiration were reported by
growth via several interconnected mechanisms such as Kuzyakov et al. (2009), Spokas et al. (2009) or Novak
the plant water status and its water-, nitrogen- and et al. (2010). However, Kolb et al. (2009) observed
respiratory carbon-use efficiency. increasing basal respiration rates, microbial biomass
Many of the measured parameters were surprising- and activity with increasing rates of charcoal applica-
ly similar to the stimulating effects that elevated tion in four different Wisconsin soils. The authors
atmospheric CO2 concentrations can have on plant attribute this to accelerated old SOC mineralization
growth (Owensby et al. 1999; Nösberger et al. 2006; after charcoal application. In a study in acidic tropical
Nowak et al. 2004). Elevated CO2 improves the WUE plantation soils, Steiner et al. (2007) reported
of plants which could even lead to soil moisture increased basal respiration values but rising microbial
increases (Morgan et al. 2004). Analogous to BC, efficiency (less respiratory CO2 loss per unit of
elevated CO2 usually decreases plant tissue N microbial biomass-C). Hence it is not clear yet, if a
concentrations (e.g. Cotrufo et al. 1998), independent general soil-respiration response pattern to BC addi-
of the N supply, and subsequently protein concen- tion exists or not. Our findings suggest that the plant–
trations such as RuBisCO (Stitt and Krapp 1999). soil system loses less C via respiration per g C of
However, the analogy does not include CO2 gas produced plant biomass, when it is BC-amended.
exchange. While photosynthesis, plant dark respira- Although, one of the most crucial, but to date
tion and soil respiration usually increase under unanswered, questions is, whether reduced soil
elevated CO2 (Long et al. 2006; Ainsworth and respiration after BC application indicates a decrease
Rogers 2007), the opposite seems to occur with BC, in soil fertility, i.e. if it is beneficial or not. Hence,
contradicting hypothesis #4. Hence elevated CO2 and long-term field studies are required.
BC effects have many symptomatic similarities, but As assumed (hypotheses #3 and #4), N2O emis-
likely different mechanisms, which warrant further sions were not immediately reduced by the presence
study. A mechanism involved in the stimulatory effect of BC, although they decreased when plants became
of BC on plant growth may have been the production larger (day 28 and day 51 of the study). A reduction
208 Plant Soil (2011) 345:195–210

in N2O emissions is in agreement with recent reports References

(e.g. Yanai et al. 2007; Lehmann 2007a; Spokas et al.
2009; van Zwieten et al. 2009; Taghizadeh-Toosi et Ainsworth EA, Rogers A (2007) The response of photosynthe-
al. 2011). Reduced water supply strongly reduced sis and stomatal conductance to rising [CO2]: mechanisms
N2O emissions, indicating that denitrification was the and environmental interactions. Plant Cell Environ
dominant N 2 O-generating process (Granli and 30:258–270. doi:10.1111/j.1365-3040.2007.01641.x
Amonette JE, Joseph S (2009) Characteristics of biochar:
Bøckmann 1994). The observed reduction in N2O
microchemical properties. In: Lehmann J, Joseph S (eds)
emissions in the presence of BC could have had Biochar for environmental management—science and
several reasons: (a) a better soil aeration by the porous technology. Earthscan, London, pp 33–52
BC which reduces denitrification (Groffman and Barker DJ, Sullivan CY, Moser LE (1993) Water deficit effects
on osmotic potential, cell wall elasticity, and proline in
Tiedje, 1991), (b) adsorption of ammonium nitrogen
five forage grasses. Agron J 85:270–275
(NH4+) to the charged BC surface in a way that Bates LS, Waldren RP, Teare ID (1973) Rapid determination of
probably reduced nitrification (and hence subsequent free proline for water-stress studies. Plant Soil 39:205–207
denitrification of NO3−), and (c) the increasing plant Blackwell P, Riethmuller G, Collins M (2009) Biochar
application to soil. In: Lehmann J, Joseph S (eds) Biochar
(root) biomass that likely outcompeted microbes for for environmental management: science and technology.
mineral N species (Smith and Tiedje 1979). We argue Earthscan, London, pp 207–226
that soil aeration (a) and increasing plant biomass (c) are Chan KY, Van Zwieten L, Meszaros I, Downie A, Joseph S (2007)
the most likely mechanisms. Since WHC was set to 60% Agronomic values of greenwaste biochar as a soil amend-
ment. Aust J Soil Res 45:629–634. doi:10.1071/SR07109
or 20% of the controls, BC-amended soils which could
Chan KY, Van Zwieten L, Meszaros I, Downie A, Joseph S
hold more water than the controls were physiologically (2008) Using poultry litter biochars as soil amendments.
drier i.e. better aerated. More effort is clearly needed to Aust J Soil Res 46:437–444. doi:10.1071/SR08036
understand the mechanisms of N2O-emission reduction Chan KY, Xu Z (2009) Biochar: nutrient properties and their
enhancement. In: Lehmann J, Joseph S (eds) Biochar for
in presence of BC (Clough and Condron 2010).
environmental management: science and technology.
We expected that a very large BC application doses Earthscan, London, pp 67–84
(200 t ha−1) would have negative effects (hypothesis Cheng C-H, Lehmann J, Thies JE, Burton AJ, Engelhard M
#5). We found no evidence for this, only a slight (2006) Oxidation of black carbon by biotic and abiotic
processes. Org Geochem 37:1477–1488
improvement compared to the 100 t ha−1 rate. This is in
Cheng C-H, Lehmann J, Thies JE, Burton SD (2008) Stability
line with findings of Chan et al. (2007, 2008) who also of black carbon in soils across a climatic gradient. J
observed that the positive BC effect exhibited a kind of Geophys Res 113: doi:10.1029/2007JG000642
saturation curve. It may be necessary to investigate Clough TJ, Condron LM (2010) Biochar and the nitrogen
cycle: Introduction. J Environ Qual 39:1218–1223.
dose–response relationships prior to field trials to
Cotrufo MF, Ineson P, Scott A (1998) Elevated CO2 reduces the
roughly assess the most (cost-) effective BC application nitrogen concentration of plant tissues. Global Change
rate which will strongly depend on biochar as well as Biol 4:43–54
soil type and agricultural management practices. Downie A, Crosky A, Munroe P (2009) Physical properties of
biochar. In: Lehmann J, Joseph S (eds) Biochar for
Taken together, BC application reduced the efflux of
environmental management—science and technology.
two potent greenhouse gases from the plant–soil system, Earthscan, London, pp 13–32
while more biomass was produced and carbon seques- Elad Y, David DR, Harel YM, Borenshtein M, Kalifa HB,
tered. We conclude that for quinoa grown in a sandy soil Silber A, Graber ER (2010) Induction of systemic
resistance in plants by biochar, a soil-applied carbon
application of BC can be beneficial and may thus
sequestering agent. Phytopathology 100:913–921
warrant subsequent field trials. Galwey NW (1989) Quinoa. Biologist 36:5
Gaskin JW, Speir RA, Harris K, Das KC, Lee RD, Morris LA,
Fisher DS (2010) Effect of peanut hull and pine chip
Acknowledgements The authors want to thank Christoph biochar on soil nutrients, corn nutrient status, and yield.
Forreiter for critical reading of the manuscript and Judy Libra Agron J 102:623–633
for proof reading. The authors acknowledge the technical Gaunt JL, Lehmann J (2008) Energy balance and emissions
assistance of Nicol Strasilla and Gerlinde Lehr with proline associated with biochar sequestration and pyrolysis bio-
and RuBisCO extractions and greenhouse gas analyses and energy production. Environ Sci Technol 42:4152–4158
Gerhard Mayer for his assistance at the ion-chromatograph. Glaser B (2007) Prehistorically modified soils of central
Thanks to Johanna Kreiling for technical assistance, and to the Amazonia: a model for sustainable agriculture in the
Department of Applied Microbiology, in particular to Stefan twenty-first century. Phil Trans R Soc London B 362:187–
Ratering, for help with the GC analyses. 196
Plant Soil (2011) 345:195–210 209

Glaser B, Haumaier L, Guggenberger G, Zech W (2001) The Kim J-S, Sparovek G, Longo RM, De Melo WJ, Crowley D
‘Terra Preta’ phenomenon: a model for sustainable (2007) Bacterial diversity of terra preta and pristine forest
agriculture in the humid tropics. Naturwissenschaften soil from the Western Amazon. Soil Biol Biochem
88:37–41 39:684–690
Glaser B, Lehmann J, Zech W (2002) Ameliorating physical Kimetu JM, Lehmann J, Kinyangi JM, Cheng CH, Thies J,
and chemical properties of highly weathered soils in the Mugendi DN, Pell A (2009) Soil organic C stabilization
tropics with charcoal—a review. Biol Fertil Soils 35:219– and thresholds in C saturation. Soil Biol Biochem
230 41:2100–2104
Golluscio RA (2007) On the link between nitrogen productivity Kolb SE, Fermanich KJ, Dornbush ME (2009) Effect of
and residence time: two opposite nitrogen use strategies? J charcoal quantity on microbial biomass and activity in
Arid Environ 68:165–169 temperate soils. Soil Sci Soc Am J 73:1173–1181.
Gonzalez JA, Gallardo M, Hila LM, Rosa M, Prado FE (2009) doi:10.2136/sssaj2008.0232
Physiological responses of quinoa (Chenopodium quinoa Kuzyakov Y, Subbotina I, Chen H, Bogomolova I, Xu X (2009)
Willd.) to drought and waterlogging stresses: dry matter Black carbon decomposition and incorporation into soil
partitioning. Bot Stud 50:35–42 microbial biomass estimated by 14C labeling. Soil Biol
Graber ER, Harel YM, Kolton M, Cytryn E, Silber A, David Biochem 41:210–219. doi:10.1016/j.soilbio.2008.10.016
DR, Tsechansky L, Borenshtein M, Elad Y (2010) Biochar Laemmli UK (1970) Cleavage of structural proteins during the
impact on development and productivity of pepper and assembly of the head of bacteriophage T4. Nature 27:680–
tomato grown in fertigated soilless media. Plant and Soil 685
337:481–496 Laird DA, Brown RC, Amonette JE, Lehmann J (2009) Review
Granier F (1988) Extraction of plant proteins for two- of the pyrolysis platform for coproducing bio-oil and
dimensional electrophoresis. Electrophoresis 9:112–718 biochar. Biofuels Bioprod Biorefin 3:547–562
Granli T, Bøckmann OC (1994) Nitrous oxide from agriculture. Lal R (2004) Soil carbon sequestration impacts on global
Norweg J Agr Sci Supp 12:1–128 climate change and food security. Science 304:1623–1627.
Groffman PM, Tiedje JM (1991) Relationships between doi:10.1126/science.1097396
denitrification, CO2 production and air-filled porosity in Lal R (2009) Challenges and opportunities in soil organic
soils of different texture and drainage. Soil Biol Biochem matter research. Eur J Soil Sci 60:158–169
23(3):299–302 Lehmann J (2006) Bio-char sequestration in terrestrial ecosys-
Hansen J, Sato M, Kharecha P, Beerling D, Berner R, Masson- tems—a review. Mitigat Adaptat Strat Glob Chan 11:403–
Delmotte V, Pagani M, Raymo M, Royer DL, Zachos JC 427. doi:10.1007/s11027-005-9006-5
(2008) Target atmospheric CO2: where should humanity Lehmann J (2007a) Bio-energy in the black. Front Ecol
aim? Open Atm Sci J 2:217–231 Environ 5:381–387
Hutchinson GL, Mosier AR (1981) Improved soil cover Lehmann J (2007b) A handful of carbon. Nature 447:143–
method for field measurement of nitrous oxide fluxes. 144
Soil Sci Soc Am J 45:311–316 Lehmann J, Czimczik C, Laird D, Sohi S (2009) Stability of
Ibarra-Caballero J, Villanueva-Verduz C, Molina-Galan J, biochar in soil. In: Lehmann J, Joseph S (eds) Biochar for
Sanchez-de-Jimenez E (1988) Proline accumulation as a environmental management—science and technology.
symptom of drought stress in maize: a tissue differentia- Earthscan, London, pp 183–205
tion requirement. J Exp Bot 39:889–897 Liang B, Lehmann J, Sohi SP, Thies JE, O"Neill B, Trujillo L,
IPCC (2007a) Climate change 2007: Climate change impacts, Gaunt J, Solomon D, Grossman J, Neves EG, Luizão FJ
adaptation and vulnerability. Working Group II Contribu- (2010) Black carbon affects the cycling of non-black
tion to the Intergovernmental Panel on Climate Change carbon in soil. Org Geochem. doi:10.1016/j.orggeo
Fourth Assessment Report—Summary for Policymakers. chem.2009.09.007
IPCC, Bern Liang B, Lehmann J, Solomon D, Kinyangi J, Grossman J,
IPCC (2007b) Climate change 2007: the physical science O"Neill B, Skjemstad JO, Thies J, Luizao FJ, Petersen J,
basis. Contribution of Working Group I to the Fourth Neves EG (2006) Black carbon increases cation exchange
Assessment Report of the Intergovernmental Panel on capacity in soils. Soil Sci Soc Am J 70:1719–1730.
Climate Change. Cambridge University Press, Cam- doi:10.2136/sssaj2005.0383
bridge Loftfield N, Flessa H, Augustin J, Beese F (1997) Automated
Ishida H, Nishimori Y, Sugisawa M, Makino A, Mae T (1997) gas chromatographic system for rapid analysis of the
The large subunit of ribulose-1,5-bisphosphate carboxyl- atmospheric trace gases methane, carbon dioxide, and
ase/oxygenase is fragmented. Plant Cell Physiol 38:471– nitrous oxide. J Environ Qual 26:560–564
479 Long SP, Zhu X-G, Naidu SL, Ort DR (2006) Can improve-
Jacobsen S-E, Stølen O (1993) Quinoa—morphology and ment in photosynthesis increase crop yields? Plant Cell
phenology and prospects for its production as a new crop Environ 29:315–330
in Europe. Eur J Agron 2:19–29 Major J, Lehmann J, Rondon M, Goodale C (2010) Fate of
Jensen CR, Jacobsen S-E, Andersen MN, Núñez N, Andersen soil-applied black carbon: downward migration, leaching
SD, Rasmussen L, Mogensen VO (2000) Leaf gas and soil respiration. Global Change Biol 16:1366–1379.
exchange and water relation characteristics of field quinoa doi:10.1111/j.1365-2486.2009.02044.x
(Chenopodium quinoa Willd.) during soil drying. Eur J Marris E (2006) Putting the carbon back: black is the new
Agron 13:11–25 green. Nature 442:624–626
210 Plant Soil (2011) 345:195–210

Mathews JA (2008) How carbon credits could drive the Sperry JS, Hacke UG (2002) Desert shrub water relations with
emergence of renewable energies. Energ Pol 36:3633–3639 respect to soil characteristics and plant functional type.
McHenry MP (2009) Agricultural bio-char production, renew- Funct Ecol 16:367–378
able energy generation and farm carbon sequestration in Spokas KA, Koskinen WC, Baker JM, Reicosky DC (2009)
Western Australia: certainty, uncertainty and risk. Agric Impacts of woodchip biochar additions on greenhouse gas
Ecosys Environ 129:1–7 production and sorption/degradation of two herbicides in a
Morgan JA, Pataki DE, Körner C, Clark H, Del Grosso SJ, Minnesota soil. Chemosphere 77:574–581
Grünzweig JM, Knapp AK, Mosier AR, Newton PCD, Spokas KA, Baker JM, Reicosky DC (2010) Ethylene: potential
Niklaus PA, Nippert JB, Nowak RS, Parton WJ, Polley key for biochar amendment impacts. Plant Soil 333:443–
HW, Shaw MR (2004) Water relations in grassland and 452
desert ecosystems exposed to elevated atmospheric CO2. Steiner C, Glaser B, Teixeira WG, Lehmann J, Blum WEH,
Oecologia 140:11–25 Zech W (2008) Nitrogen retention and plant uptake on a
Mosier AR, Mack L (1980) Gas chromatographic system for highly weathered central Amazonian Ferralsol amended
precise, rapid analysis of nitrous oxide. Soil Sci Soc Am J with compost and charcoal. J Plant Nutr Soil Sci 171:893–
44:1121–1123 899. doi:10.1002/jpln.200625199
Nösberger J, Long SP, Norby RJ, Stitt M, Hendrey G, Blum H Steiner C, Teixeira M, Zech W (2007) Soil respiration curves as
(2006) Managed ecosystems and CO2: case studies, soil fertility indicators in perennial central Amazonian
processes, and perspectives, vol 187. Ecological studies. plantations treated with charcoal, and mineral or organic
Springer, Berlin, pp 457 fertilisers. Trop Sci 47:218–230. doi:10.1002/ts.216
Novak JM, Busscher WJ, Watts DW, Laird DA, Ahmedna MA, Stitt M, Krapp A (1999) The interaction between elevated
Niandou MAS (2010) Short-term CO2 mineralization after carbon dioxide and nitrogen nutrition: the physiological
additions of biochar and switchgrass to a Typic Kandiu- and molecular background. Plant Cell Environ 22:583–
dult. Geoderma 154:281–288 621
Nowak RS, Ellsworth DS, Smith SD (2004) Functional Taghizadeh-Toosi A, Clough TJ, Condron LM, Sherlock RR,
responses of plants to elevated atmospheric CO2—do Anderson CR, Craigie RA (2011) Biochar incorporation
photosynthetic and productivity data from FACE experi- into pasture soil suppresses in situ nitrous oxide emissions
ments support early predictions? New Phytol 162:253–280 from ruminant urine patches. J Environ Qual, in press
Owensby CE, Ham JM, Knapp AK, Auen LM (1999) Biomass (open access) https://www.soils.org/publications/jeq/view/
production and species composition change in a tallgrass 40-42/q10-0419.pdf
prairie ecosystem after long-term exposure to elevated van Zwieten L, Singh B, Joseph S, Kimber S, Cowie A, Chan
atmospheric CO2. Global Change Biol 5:497–506 KY (2009) Biochar and emissions of non-CO2 greenhouse
Reich PB, Tjoelker MG, Machado J-L, Oleksyn J (2006) gases from soil. In: Lehmann J, Joseph S (eds) Biochar for
Universal scaling of respiratory metabolism, size and environmental management—science and technology.
nitrogen in plants. Nature 439:457–461 Earthscan, London, pp 227–249
Rondon M, Lehmann J, Ramírez J, Hurtado M (2007) van Zwieten L, Kimber S, Morris S, Downie A, Berger E, Rust
Biological nitrogen fixation by common beans (Phaseolus J, Scheer C (2010) Influence of biochars on flux of N2O
vulgaris L.) increases with bio-char additions. Biol Fertil and CO2 from Ferrosol. Aust J Soil Res 48:555–568
Soils 43:699–708 Wardle DA, Nilsson M-C, Zackrisson O (2008) Fire-derived
Schulte PJ, Brooks JR (2003) Branch junctions and the flow of charcoal causes loss of forest humus. Science 320:629.
water through xylem in Douglas-fir and ponderosa pine doi:10.1126/science.1154960
stems. J Exp Bot 54:1597–1605. doi:10.1093/jxb/erg169 Yanai Y, Toyota K, Okazaki M (2007) Effects of charcoal
Smith MS, Tiedje JM (1979) The effect of roots on soil addition on N2O emissions from soil resulting from
denitrification. Soil Sci Soc Am J 43:951–955. rewetting air-dried soil in short-term laboratory experi-
doi:10.2136/sssaj1979.03615995004300050027x ments. Soil Sci Plant Nutr 53:181–188

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