Académique Documents
Professionnel Documents
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in the
Rhizosphere
1535/frame/FM Page 2 Tuesday, August 22, 2000 9:08 AM
Trace Elements
in the
Rhizosphere
Edited by
George R. Gobran
Walter W. Wenzel
Enzo Lombi
CRC Press
Boca Raton London New York Washington, D.C.
Library of Congress Cataloging-in-Publication Data
Trace elements in the rhizosphere / George R. Gobran, Enzo Lombi & Walter W.
Wenzel, editors.
p. cm.
Papers from a special symposium held during the Fifth International Conference on
Biogeochemistry of Trace Elements, July 1115, 1999 in Vienna, Austria.
Includes bibliographical references and index.
ISBN 0-8493-1535-2
1. SoilsTrace element contentCongresses. 2. Trace element in plant
nutritionCongresses. 3. Plant-soil relationshipsCongresses. 4.
RhizosphereCongresses. I. Gobran, George R. II. Lombi, Enzo, 1968 III. Wenzel,
Walter W. IV. International Conference on the Biogeochemistry of Trace Elements (5th :
1999 : Vienna, Austria)
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Enzo Lombi was born in Piacenza, Italy, 1968. After graduation in 1993, he earned
a Ph.D. in agricultural chemistry and conducted his research in Italy, Austria, and
the United States. During 19971998 he was Scientific Researcher at the Institute
of Soil Science, University of Agricultural Sciences, Vienna, and at the Austrian
Research Centre, Seibersdorf. His activities focused on the development and appli-
cation of low-input technologies to remediate polluted soils. At present, Dr. Lombi
is Research Scientist at the Soil Science Department of IACR-Rothamsted, UK. His
research includes various aspects of phytoremediation using hyperaccumulator
plants and chemical-assisted phytoextraction. He is interested in physiological
aspects of tolerance, metal transport, and accumulation and compartmentation in
hyperaccumulator plants. Moreover, Dr. Lombis work includes investigations con-
cerning rhizosphere processes involved in metal mobilization and uptake by plants.
A second area of investigation is the in situ fixation of heavy metals and metalloids
using industrial byproducts.
Contributors
Domy C. Adriano Salvatore Deiana
Savannah River Ecology Lab DI.S.A.A.B.A.
University of Georgia Universita degli Studi di Sassari
Drawer E Viale Italia 39, 07100 Sassari
Aiken, South Carolina 29802 Italy
U.S.A.
Bruno Delvaux
Naomi Assadian Unit Sciences du Sol
Texas Agricultural Experiment Universit Catholique de Louvain
Station Place Croix du Sud, 2/10
Texas A & M University 1348 Louvain-la-Neuve
1380 A & M Circle Belgique
El Paso, Texas 79927
U.S.A. Joseph E. Dufey
Unit Sciences du Sol
Universit Catholique de Louvain
Bjrn O. Berthelsen
Place Croix du Sud, 2/10
Norwegian University of Science and
1348 Louvain-la-Neuve
Technology
Belgique
Faculty of Chemistry and Biology
Department of Chemistry
Alain Dufresne
N-7491 Trondheim
Les Laboratoires Savoie-Dufresne Inc.
Norway
C.P. 48808, C.S.P. Outremont
Montral (Qubec), H2V 4V1
Henri Calba Canada
AMIS Programme Agronomie
Centre de Coopration Internationale Lloyd B. Fenn
en Recherche Agronomique Texas Agricultural Experiment Station
pour le Dveloppement Texas A & M University
2477 Avenue du Val de Montferrand 1380 A & M Circle
F-34032 Montpellier Cedex 1 El Paso, Texas 79927
France U.S.A.
Amedeo Palma
DI.S.A.A.B.A.
Universita degli Studi di Sassari
Viale Italia 39, 07100 Sassari
Italy
Contents
SECTION I PHYTOAVAILABILITY OF TRACE ELEMENTS AND
RADIONUCLIDES IN THE RHIZOSPHERE AND
APPLICATION TO PHYTOREMEDIATION
Chapter 1
Dependency of Phytoavailability of Metals on Indigenous and Induced
Rhizosphere Processes: A Review ............................................................................3
Enzo Lombi, Walter W. Wenzel, George R. Gobran, and Domy C. Adriano
Chapter 2
Bioavailability of Trace Elements as Related to Root-Induced Chemical
Changes in the Rhizosphere ....................................................................................25
Philippe Hinsinger
Chapter 3
Rhizosphere Chemical Changes Enhance Heavy Metal Absorption by Plants
Growing in Calcareous Soils...................................................................................43
Naomi Assadian and Lloyd B. Fenn
Chapter 4
Fate of Radiocesium in Soil and Rhizosphere........................................................61
Bruno Delvaux, Nathalie Kruyts, Emmanuel Maes, and Erik Smolders
Chapter 5
Bioavailability of Uranium and Plutonium to Plants in SoilWater Systems
and the Potential of Phytoremediation....................................................................93
Hamid Shahandeh, Jin-Ho Lee, Lloyd R. Hossner, and Richard H. Loeppert
Chapter 6
Interactions and Mobilization of Metal Ions at the SoilRoot Interface .............127
Salvatore Deiana, Bruno Manunza, Amedeo Palma, Alessandra Premoli,
and Carlo Gessa
Chapter 7
Analysis of Metal Speciation and Distribution in Symbiotic Fungi (Ectomycorrhiza)
Studied by Micro X-ray Absorption Spectroscopy and X-ray Fluorescence
Microscopy ............................................................................................................149
Bjrn O. Berthelsen, Geraldine M. Lamble, Alastair A. MacDowell,
and David G. Nicholson
Chapter 8
Bioavailability of Heavy Metals in the Mycorrhizosphere...................................165
Corinne Leyval and Erik J. Joner
Chapter 9
Solid Phase Fractionation of Metals in the Rhizosphere of Forest Soils ............189
Franois Courchesne, Vronique Sguin, and Alain Dufresne
Chapter 10
A Technique for Quantitative Trace Element and Micronutrient Studies
in Plants .................................................................................................................207
Anders Gransson
Chapter 11
Cation Exchange on Plant Roots Involving Aluminium: Experimental
Data and Modeling ................................................................................................227
Joseph E. Dufey, Jos G. Genon, Benot Jaillard, Henri Calba, Gervais Rufyikiri,
and Bruno Delvaux
Chapter 12
Modeling the Dynamics of the Rhizosphere Aluminum Chemistry in Acid
Forest Soils ............................................................................................................253
Heino W. F. Nietfeld
Index......................................................................................................................309
Section I
Phytoavailability of Trace
Elements and Radionuclides in
the Rhizosphere and Application
to Phytoremediation
1 Dependency of
Phytoavailability of
Metals on Indigenous
and Induced
Rhizosphere Processes:
A Review
Enzo Lombi, Walter W. Wenzel,
George R. Gobran, and Domy C. Adriano
CONTENTS
I. Introduction.......................................................................................................4
II. PlantSoilMicrobial Interactions: The Rhizosphere ......................................5
III. Phytoremediation ..............................................................................................5
IV. Biogeochemistry of the Metals in the Rhizosphere.........................................7
A. Rhizosphere Effect on pH ....................................................................7
B. Mineralogy and Soil Composition in the Rhizosphere .......................8
C. Root Growth and Effects on Soil Physical Parameters .......................8
D. Root Exudation and Metal Mobilization in the Rhizosphere..............9
E. Precipitation Phenomena in the Rhizosphere ....................................12
F. RootMicroorganisms Interactions in the Rhizosphere.....................13
G. Volatilization in the Rhizosphere .......................................................13
V. Microtools and Special Setup to Study RhizosphereContaminant
Interactions......................................................................................................14
VI. Future Research ..............................................................................................15
VII. Summary and Conclusions.............................................................................16
Acknowledgments....................................................................................................17
References................................................................................................................17
0-8493-1535-2/01/$0.00+$.50
2001 by CRC Press LLC 3
4 Trace Elements in the Rhizosphere
I. INTRODUCTION
Several metals and metalloids such as Mn, Cu, Zn, Ni, Mo, and B play a dualistic
role in the soil-plant system. They are essential for plant growth but when present
at high concentrations, can result phytotoxic. Their bioavailability is therefore of
fundamental importance. In case of deficiency, an increase in metal bioavailability
can be beneficial for plant growth. On the other hand, in case of pollution, a large
available amount of these elements and other similar nonessential elements (such
as Pb, Cr, As, etc.) can be detrimental to plant growth. Cleanup technologies are
often limited in their efficiency by the mobility of the pollutants. This chapter
will review the rhizosphere aspects related to availability of metals and their role
in plant nutrition and emerging remediation technologies (in particular phytore-
mediation).
At the beginning of this century, several works by Arnon and Stout1,2 and other
authors demonstrated that plants need a number of micronutrients at very low
concentration. One of the first examples of increased plant production as a conse-
quence of micronutrient application is reported by Anderson3 who observed a tenfold
increase in clover production after addition of Mo to an ironstone soil in Australia.
Since then the discovery of the essentiality of other micronutrients has given an
explanation to low plant productions in many areas all around the world. The
bioavailability of micronutrients in the rhizosphere is a key factor controlling plant
nutritional status and therefore plant production.
Soil contamination by heavy metals and metalloids is a phenomenon that has
accompanied human kind throughout its history.4 Examples of contaminated sites
due to past mining activities are common throughout the world. The number of
contaminated sites already identified represents a serious problem and the list of
sites to be treated is most likely to increase along with their remediation costs.5
Soil pollution by trace elements represents a large proportion of the problem.
For instance, trace elements account for 65% of the contaminated Superfund sites
for which the U.S. EPA has signed Records of Decisions.6 The most common
inorganic contaminant is Pb (47% of sites) followed by As (41%), Cr (37%), Cd
(32%), Ni (29%), and Zn (29%).6
The need to treat contaminated soils has created a large market for remediation.
Many technologies have been developed to remediate contaminated soils.4,7-9 The
EPAs Vendor Information System for Innovative Treatment Technologies (VISITT)
lists 66 technologies to treat metal-contaminated soils. Only 9 of the 66 technologies
listed are in situ soil or groundwater treatments.6
Recently, phytoremediation has been thought to provide an alternative, environ-
mentally friendly remediation technology for the treatment of polluted soils. Phy-
toremediation refers to the use of higher plants to remove pollutants from the soil
or to reduce the hazard associated with their presence in the environment.10-12 The
market for phytoremediation in the U.S. is between $1 and 2 million. Glass13
estimated that this market might grow to $4080 million by 2005.
The interactions between soil, metals, and the root system represent key factors
controlling the success of plant production (in case of micronutrient deficiency) and
phytoremediation. Therefore, focal importance can be attributed to the part of the
Dependency of Phytoavailability of Metals 5
soil where these interactions take place: the rhizosphere. Due to the presence of the
root system, the characteristics of the rhizospheric soil differ largely from the bulk
soil.14,15 As a consequence, the biogeochemical processes in the rhizosphere will
play an important role in regulating the mobility and bioavailability of the metals
and, in turn, on its uptake by plants, volatilization, or plant-induced precipitation.
This chapter reviews the role of the rhizosphere in metal uptake, tolerance, and
volatilization. Emphasis is given to the biogeochemical processes in the rhizosphere
and their role in phytoremediation technology and plant nutrition.
III. PHYTOREMEDIATION
In recent years environmental legislation has dealt with the problem of contaminated
or polluted soil and its remediation. The need for soil remediation depends primarily
6 Trace Elements in the Rhizosphere
on the nature of the contaminant (toxicology and ecotoxicology) and the land use.
Moreover, public perception, social costs, and political considerations play decisive
roles in the decision-making process of soil remediation.4 Traditional solutions such
as landfill of contaminated soils, that at the moment cover a large part of the
remediation market, will probably loose their economical and public acceptance.
New technologies based on environmentally friendly and low cost processes are
urgently required.
Phytoremediation refers to the use of higher plants to remove soil pollutants or
to diminish the risk associated with their presence in the environment.11,12,25-27 This
technology meets several of the desirable characteristics of an ideal remediation
process: phytoremediation reduces the hazard associated with the presence of pol-
lutants in the environment, improves soil quality and restores its functionality, and
is a low-input technology. Moreover, its public acceptance is expected to be great
and the cost should be highly competitive ($25100 t1).13 On the other hand, several
years are needed to clean up a contaminated site with the plant material available
at the moment. McGrath et al.28 calculated that it would take 9 years to reduce Zn
concentration in soil from 440 g g 1 to 300 g g 1 using Thlaspi caerulescens.
Phytoremediation can be divided into containment and removal technologies.
Containment processes such as phytoimmobilization and phytostabilization aim to
reduce the mobility, bioavailability, and/or toxicity of the pollutant in the environ-
ment. Removal processes (phytoextraction and phytovolatilization) are technologies
for cleaning up the soil.
Dependency of Phytoavailability of Metals 7
A. RHIZOSPHERE EFFECT ON PH
from the pH of the bulk soil.34 The principal reason for these differences is thought
to be the unequal uptake of cations and anions by plant roots.35,36 A dominant factor
in this process is the form of N that is taken up by plant roots: when N is absorbed
in the form of NH4+ the plants release H+ to maintain electrical neutrality and
therefore the pH in the rhizosphere decreases. On the contrary, when NO3 is taken
up, a net efflux of HCO3 by the roots will cause the pH to increase.34,37,38 Smiley39
reported differences in the rhizosphere pH of wheat up to 2.2 units when different
sources of N where used. The large difference that may exist between rhizosphere
and bulk soil pH can cause processes of adsorption or desorption, precipitation, or
solubilization of trace elements to occur. Heavy metals become more mobile in
acidic conditions whereas the mobility of metalloids follows an opposite trend.
complementary tool able to enhance plant growth and therefore improve phytosta-
bilization (for review see Vangronsveld et al.).53
FIGURE 1.3 Models of Strategy I and II plants: P.M. = plasma membrane; R = inducible
reductase; T1 = transporter for Fe2+; St = stimulated H+ pump for ATPase; X = increased
production/release of chelators/reductants; E = enhanced synthesis and secretion of phytosi-
derophores; PS = phytosiderophores; T2 = translocator for Fe3+ phytosiderophores in the
plasma membrane. (Modified from Marschner, H. et al.140 and Rmheld, V.141 With permis-
sion.)
surface. The chemical nature and the rate of release of phytosiderophores differ
among plant species and even cultivars.46,54-56 Phytosiderophores form chelates not
only with Fe but also with Zn, Cu, and Mn and are therefore able to mobilize other
micronutrients in case of deficiency.57,58 Uptake of ferrated phytosiderophores is
generally considered highly selective.48 More recently, Wiren et al.59 found evidence
that Zn can be taken up in grasses in the form of nondissociated Zn-phytosidero-
phores. The mechanisms of Strategies I and II, effective on Fe oxides or other soil
components may, as a side effect, mobilize metals adsorbed on minerals. Represen-
tations of Strategies I and II are given in Figure 1.3. A third strategy may be identified
in the capacity of microorganisms to release siderophores and the possibility for the
plants to take up these compounds.60 The existence of a microbial siderophore Fe
transport system has been proposed in oat61,62 and maize.63
Exudation of organic acids has been observed in P-deficient plants by Hoffland
et al.64 This mechanism, also reported by other authors,65,66 may have implications
for metal mobility. In fact, mineral weathering and dissolution of P-containing
minerals can increase the mobility of metals.
Plant uptake of trace elements can be enhanced through changes in metal avail-
ability in the rhizosphere. In particular, exudation of phytosiderophores and organic
acids, and changes in pH and redox potential are considered key factors controlling
metal mobility in the rhizosphere.
The ability of root exudates, collected from different plants, to mobilize Cd was
investigated by Mench and Martin.67 They found that root exudates of Nicotiana
tabacum were able to extract more Cd from soil than those of Nicotiana rustica and
Dependency of Phytoavailability of Metals 11
Zea mays. These results are in agreement with the fact that tobacco plants are well
known to accumulate fairly large amounts of Cd in their leaves.
Fenn and Assadian68 reported that Cynodon dactylon was able to mobilize Pb,
Cu, and Mn in the rhizosphere and accumulate them in the leaves. This process was
probably related to changes in rhizosphere pH and the dissolution of carbonates.
Other authors69-71 found that the mobility of Cd, Zn, and Cu increased in the soil
surrounding the roots due to acidification of the rhizosphere. The acidification
process was related to the form of nitrogen taken up by the plants. There is evidence
that the reduction mechanism present in Strategy I plants to mobilize Fe could also
be able to increase Cu and Mn uptake (Welch et al.).72
So far, only a few studies have been conducted to investigate the rhizosphere
of hyperaccumulator species. Redox potential and pH changes in the rhizosphere
of the Ni hyperaccumulator Alyssum murale and the nonhyperaccumulator Rapha-
nus sativus were compared by Bernal and McGrath73 and Bernal et al.74 The results
suggest that the source of N was the main factor responsible for pH changes and
the crop plants were able to reduce the system more effectively than the hyperac-
cumulator. These findings indicate that hyperaccumulation mechanisms may be
related to other rhizosphere processes such as the release of chelating agents and/or
to differences in the number and affinity of metal root transporters. McGrath et
al.75 and Knight et al.76 studied the chemical and physical characteristics of the
rhizosphere of Thlaspi caerulescens. They concluded that the decrease in the
mobile Zn fraction could explain only less than 10% of the total Zn uptake by the
plants. T. caerulescens was able to deplete the mobile fraction of Zn more than
the closely related, but nonaccumulator T. ochroleucum. No significant differences
in pH were observed in the rhizosphere of T. caerulescens. Knight et al.76 proposed
that two possible mechanisms could explain these results: either T. caerulescens
is able to mobilize Zn from the soil or the soil studied had a large capacity to
buffer the concentration of Zn in soil solution. Recently, Hamon and McLaughlin77
used an isotopic dilution method to determine pools of Cd and Zn in contaminated
soils that can be accessed and removed from soil by T. caerulescens and by wheat.
Their results show no difference in specific activity of Cd or Zn taken up by T.
caerulescens or wheat. This indicates that the hyperaccumulator plant was able to
access the same pools of metals that were available to the wheat plants. However,
it has to be pointed out that the Zn added in biosolids was highly labile, and the
population of T. caerulescens used in this experiment acts more as a Cd-tolerant
species than as a hyperaccumulator for Cd. These partly contradictory results
suggest that hyperaccumulator plants seem to take up from the same phytoavailable
metal pools that are accessible to other plants where this pool is large enough.
However, there may exist mechanisms in the rhizosphere of hyperaccumulators,
such as root exudation, that can support metal uptake from less accessible pools.
In field conditions, where metals are returned to the soil via senescence, the easily
available metal pools may typically be large enough to explain the amounts accu-
mulated in the plants.78
Currently, there is not enough information to understand whether mobilization
of metals in the rhizosphere constitutes one of the mechanisms responsible for
12 Trace Elements in the Rhizosphere
can increase soil organic matter, which is very effective in immobilizing toxic
elements such as Cu. Fairly insoluble compounds of Zn and Pb can form in the
rhizosphere for reaction with P. Pyromorphyte [Pb5(PO4)3Cl] has been reported to
form in the rhizosphere of Agrostis capillaris.86 This mineral has a very low solubility
(Ksp = 1084.4) and is thought to maintain Pb at very low concentrations in the surface
environment.87 Cotter-Howells et al. (1999) have localized pyromorphyte granules
in the outer cell walls of the epidermis of Agrostis capillaris.88 Associations between
Cd and polyphosphate granules were found in the mycorrhized roots of Pteridium
aquilinum.89 These precipitation processes may represent active tolerance mecha-
nisms that plants use to reduce heavy metal toxicity.88
experiment, the container was removed and the soil sliced with a piston microtome
to obtain soil samples at defined distances from the rhizoplane.
In laboratory research more complicated techniques, other than the ones
described above, have been used. Different experimental setups (i.e., rhizobox, soil-
packets system, rhizosphere study container) have employed porous membranes to
separate soil at different distances from the roots.69,70,75,119-122 Other experimental
designs have been developed to control plant nutritional conditions and rhizosphere
pH,66,123 or soil redox potential.124 Root-induced changes of pH have been studied
directly in soil using glass microelectrodes,125-127 or by employing agar with a pH
indicator as substrate.128,129 In this latter case, qualitative changes in pH can be
visualized by color changes in the indicator added, and quantitative measurements
are obtained using microelectrodes,33,130,131 spectrodensiometry,132 or a method based
on the buffer curve of the gel used.133
Root growth dynamics and morphology have been studied using rhizotrons or
minirhizotrons.134-136
larger root biomass. An increased soil-binding capacity of the root system can also
be achieved through enhancement of mycorrhizal associations.
Phytoimmobilization processes in the rhizosphere are caused by changes in pH
and redox potential, and by the exudation of inorganic or organic compounds such
as phosphate or phytate. Before we proceed to modify plants to improve phytoim-
mobilization processes, the mechanisms of metal precipitation should be better
understood at the rhizosphere level.
Phytoextraction represents the most studied phytoremediation technology and
many efforts have been directed at the understanding of metal accumulation in plants.
On the other hand, information on rhizosphere processes involved in metal uptake
by hyperaccumulator species is still very poor. It has not been proven that hyperac-
cumulator plants have a need to,78 and would be able to mobilize metals in the
rhizosphere; recent studies rather seem to indicate the opposite.77 However, these
plants have a very high requirement for necessary metals such as Zn and it cannot
be excluded that they have evolved mechanisms to mobilize metals in the rhizo-
sphere. In any case, an improved understanding of these mechanisms will be useful
in both phytoremediation technologies and plant nutrition research. Still the complex
interaction between plant and microorganisms and their potential role in metal
mobilization and plant uptake is largely unknown. The understanding of plantbac-
teria interactions to enhance metal uptake, such as the one recently reported for
Se,106 can represent a major step toward enhancing phytoextraction technologies.
Phytovolatilization of Se has been studied extensively and the increased under-
standing of basic mechanisms in the rhizosphere will, most likely, largely improve
this technology. Further information is required to enhance phytovolatilization of
other elements such as Hg and As.
ACKNOWLEDGMENTS
This study was supported by the University of Agricultural Sciences Vienna, Priority
Research Area Project No. 16 (Rhizosphere processes: Modeling and experimental
assessment of metal interactions with organic ligands exuded by plant roots); the
Fonds zur Frderung der Wissenschaftlichen Forschung (FWF, Project No. P13454-
BIO); and the Austrian Federal Ministry of Science and Transport. It also is part of
an ongoing project that is financed by the Swedish Council for Forestry and Agri-
cultural Research (SFJR No. 23 .0173/98).
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2 Bioavailability of Trace
Elements as Related to
Root-Induced Chemical
Changes in the
Rhizosphere
Philippe Hinsinger
CONTENTS
I. Introduction.....................................................................................................25
II. Bioavailability of Trace Elements as Related to Root-Induced
Accumulation/Depletion of Ionic Species in the Rhizosphere......................26
III. Bioavailability of Trace Elements as Related to Root-Induced
Acidification/Alkalinization of the Rhizosphere............................................30
IV. Bioavailability of Trace Elements as Related to Root-Induced
Oxidation/Reduction in the Rhizosphere .......................................................32
V. Bioavailability of Trace Elements as Related to Root-Induced
Complexation/Chelation in the Rhizosphere .................................................33
VI. Conclusions.....................................................................................................36
References................................................................................................................37
I. INTRODUCTION
Predicting the bioavailability of trace elements to plants is a major agricultural and
environmental issue. Indeed, plants are the prime entry point for trace elements in
the food chain. On the one hand, for trace elements that are essential micronutrients
for plants, animals, and human beings, the need arises for ensuring adequate levels
in agricultural products. Indeed, deficiencies of micronutrients are fairly widespread
in various types of soils worldwide. On the other hand, as a consequence of human
activities, concentrations of potentially toxic trace elements tend to build up in some
environments and ultimately cause contamination of the food chain. Similarly as
described for major nutrients by researchers such as Clarkson1 and Hinsinger,2 a
range of factors and mechanisms are implied in the transfer of trace elements from
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2001 by CRC Press LLC 25
26 Trace Elements in the Rhizosphere
the soil into plant roots.3 In addition to the uptake per se, the acquisition of trace
elements encompasses those chemical processes that occur at the soilroot interface
and that can influence the dynamics of trace elements in the so-called rhizosphere.1,2
Understanding all the mechanisms implied in the process of acquisition of trace
elements by plant roots is thus a prerequisite for assessing their bioavailability.
The rhizosphere effect was first defined by Hiltner4 to account for the increased
microbial biomass and activity that occurs in the immediate vicinity of plant roots.
This phenomenon is largely due to the large flux of C originating from root exudation.
Root exudates include a wide spectrum of organic substances. Some are very reactive
compounds that can directly affect the chemical properties of the rhizosphere and,
in particular, those properties that determine the speciation and ultimately the bio-
availability of trace elements. In addition to root exudation, research conducted in
the past decades has provided ample evidence that plant roots can drastically alter
the physical,5 and chemical properties of the rhizosphere through several other
processes.2, 3, 6-8 These processes have been reported as major steps in the process of
acquisition by plants of major nutrients such as N, P, and K in particular, and of
micronutrients such as Fe and Zn, among others. They can also have a major
influence on the dynamics of trace elements in the soil surrounding roots and are
likely to ultimately determine the actual rates of contamination of plant tissues by
trace elements that are considered as micropollutants.
The aim of this chapter is to give an overview of the possible chemical processes
involved in the rhizosphere that can affect the speciation of trace elements in the
soil solid phase and their bioavailability to plants (i.e., phytoavailability). Although
rhizosphere microorganisms are known to potentially play a role in that respect as
reviewed for mycorrhizal fungi, for instance,9,10 this chapter only addresses the
possible direct effects of plant roots.
TABLE 2.1
Ratio (%) of Mass-Flow to Actual Uptake into
Leaves and Tubers of Radish (Raphanus sativa L.)
for Major Cations and Two Trace Metals in a Sandy
Loam (with Total Cd and Zn Concentrations of
2.8 and 152.5 g g1, respectively).a
Ratio (%) Ca Mg K Zn Cd
elements such as Zn and Cd, (Table 2.1). As a result of this, these elements are
expected to be depleted from the rhizosphere, as reviewed for nutrients by Jungk12
and Hinsinger.2 However, little experimental evidence exists for such a depletion
of trace elements in the rhizosphere, possibly because of the technical difficulty of
accessing measurable, reliable concentrations of trace elements in the soil solution
at this scale. Calculations have alternatively provided a means to support this
hypothesis, as shown for instance for Mn.16 This work reported good agreement
between predicted and observed Mn uptake by wheat, maize, and sugar beet, which
suggests that the calculated values of Mn depletion in the rhizosphere were realistic.
According to this, Mn concentration decreased from 0.20 M in the bulk soil to
0.16, 0.13, and 0.11 M in the rhizosphere of wheat, maize, and sugar beet,
respectively.
Whenever a decrease in the concentration of a trace element occurs, the direct
consequence will be a diffusion of the trace element toward the root along the created
gradient. According to Le Chateliers principle (mass action law), a further conse-
quence will be the shift in the various reaction equilibria involving this trace element,
in order to replenish the soil solution in the rhizosphere. For those trace elements
that are present in the soil as exchangeable cations such as Zn and Cd, their depletion
from the rhizosphere (see Table 2.1) is likely to result in a shift in the adsorption/des-
orption (cation exchange) equilibrium toward an enhanced desorption (toward the
right in Equation (1) that stands for the divalent metal M):
For a range of species grown on a soil spiked with 65Zn and 109Cd, similar specific
activities of Zn were found in the shoots by Hamon et al.,17 demonstrating that none
of the studied species had access to nonisotopically exchangeable pools of soil Zn.
28 Trace Elements in the Rhizosphere
FIGURE 2.1 Values of the Cs adsorption distribution coefficient (Kd) of each rhizosphere
soil relative to that of the bulk (uncropped) soil as a function of the final solution Cs
concentration for a sandy (Soil S) and a clayey (Soil C) soil. The ratios of Kd values obtained
for trace concentrations of Cs are plotted against the geometric average of the Cs solution
concentrations in the rhizosphere and bulk soils. (Adapted from Guivarch, A. et al.31 With
permission.)
Considering this simple equation, proton release by plant roots is thus expected
to result in an enhanced dissolution of such minerals, in application of Le Chateliers
principle. It is noteworthy that most plant species (all but grasses) have been
described as Strategy I plants that respond to Fe deficiency by an increased acidifying
and reducing capacity of their roots.46 Bertrand and Hinsinger47 indeed showed that
goethite could be dissolved significantly in the rhizosphere of Strategy I plants such
as pea, white lupin, and oilseed rape. Their results, however, showed that proton
excretion by plant roots could not account on its own for the measured dissolution,
suggesting that it operated in conjunction with other rhizosphere processes such as
reduction and possibly complexation, too (see below). Many metals occur in trace
amounts in Fe oxides and will thus be released similarly as Fe in Equation (2). In
addition, the process described above for Fe will also apply similarly for other metal
oxides, Mn oxides in particular.48 The following example addresses the case of a
sewage sludge for which a substantial proportion of the Zn was likely to occur as
Zn carbonate (smithsonite).49 This sludge was mixed with quartz sand, with or
without addition of calcite, and cropped with two plant species in a cropping device
designed by Guivarch et al.31 This device enabled them to measure the total amounts
of metal taken up by the plants and to easily collect the rhizosphere material for its
chemical analysis.50 Pecqueux et al.50 showed that the uptake of Zn was by far the
biggest for rape when no calcite was added to the substrate, that is to say in the sole
treatment where a significant decrease in rhizosphere pH was recorded (Table 2.2).
TABLE 2.2
Amounts of Zn Taken up by Plants from a Sewage Sludge and
Changes in Rhizosphere pHa
pH g per pot)
Zn uptake (
Treatment Initial Ryegrass Rape Ryegrass Rape
CaCO3 6.28 0.17 6.25 0.18 6.00 0.20* 8.6 04 24.1 2.7
+CaCO3 6.81 0.08 6.81 0.19 6.88 0.14 6.8 0.9 11.5 1.5
a The sludge, which contained 2.28 g Zn kg1, was mixed with pure quartz sand at a rate
of 20 g sludge kg1. In the +CaCO3 treatment, calcite was added at a rate of 50 g kg1.
The * indicates a significant (at p < 0.05) pH change in the rhizosphere relative to the
initial pH of the substrate.
In comparison, the addition of calcite had little effect on the uptake of Zn by ryegrass,
which did not show any pH change in its rhizosphere. The release of protons by
rape roots and subsequent dissolution of Zn carbonate in the sludge might thus
explain the larger uptake of Zn that this species achieved.
In addition to being involved in the dissolution/precipitation equilibria of metal-
bearing minerals, pH is also considered as a key factor in sorption reactions of trace
metals with soil constituents. This is exemplified in the following equations that
describe the adsorption/desorption of a divalent metal M and concomitant deproto-
nation/protonation of goethite:51
Fe-O
M + 2 H+ 2 (-Fe-OH) + M2+ (3)
Fe-O
FIGURE 2.2 Amounts of trace metals extracted by the root exudates of Zea mays, Nicotiana
rustica, and Nicotiana tabacum and synthetic solutions of glucose, citric acid, and glycine
or deioinized water. Except for controls treated with water, all the exudate and synthetic
solutions contained an equivalent amount of total C, yielding a final concentration of 150 g
C g1 soil. The soil used for the experiment was an acid soil that had been earlier contaminated
with Cd(NO3)2 and contained 3.6, 11, and 485 g g1 of total Cd, Cu, and Mn, respectively.
For each metal, treatments with a different letter were significantly different at p < 0.05.
(Adapted from Mench, M. and Martin, E.66 With permission.)
Several authors60,69,70 have shown the exudation of citrate by the proteoid roots
of white lupin and its possible implication in the mobilization of metals such as Fe,
Mn, or Zn. Dinkelaker et al.69 showed that, when grown in a calcareous soil, citrate
was exuded at such high rates around proteoid roots that it precipitated as discrete
crystals of Ca citrate. They also reported a significant increase in DTPA-extractable
Mn and Zn. The bioavailability of these metals in such conditions as DTPA has been
proposed as a method for predicting the availability of metals in calcareous soils by
Lindsay and Norvell.71 The increase in DTPA-extractable Mn and Zn in this exper-
iment was possibly the consequence of an increased amount of total soluble Mn and
Zn in the soil solution as related to their complexation by citrate ions. It could,
however, also be attributed to the acidification of the rhizosphere that concurrently
Bioavailability of Trace Elements 35
TABLE 2.3
Stability Constants of Complexes Formed between
Fe or Divalent Metals and Two Phytosiderophores
Isolated and Purified from Root Washings of Barley
(Hordeum vulgare L.): Mugineic Acid (MA) and
Deoxymugineic Acid (DMA)
Ca2+ Mn2+ Fe2+ Zn2+ Ni2+ Fe3+ Cu2+
TABLE 2.4
Biomass and Micronutrient Concentrations in the Shoots of Barley
(Hordeum vulgare L.) Grown in a Calcareous Luvisol and Release
of Fe Solubilizing Exudates by Fe-Sufficient (+Fe) or Fe-Deficient
(Fe) Seedlings
(mg kg1 shoot dry weight) Exudate release
Shoot dry weight (nmol Fe solubilized g1
(mg plant1) Fe Mn Cu Zn root DW per 4 hours)
metals is restricted in calcareous soils because of the high pH of the bulk soil, the
bioavailability of metals to plants such as grasses might be larger than expected.
This is because of the enhanced production of phytosiderophores that occur in
response to the low levels of available Fe in the soil solution.
When considering the effects of any organic exudate in the soil, the possible
competition between the various metals, including alkaline earth metals, needs to be
accounted for, as suggested by Jones and Darrah37 for alkaline soils containing large
concentrations of Ca and Mg. Hamon et al.63 showed, for instance, that when increas-
ing Ca concentration from 42 to 1042 mg dm3 in the soil solution, the proportion
of Zn and Cd present as free metal ions increased from 8 to 14% to 83 to 100%, as
a consequence of Ca forming complexes or chelates instead of Zn and Cd. The
competition between various trace metals and Fe for chelate formation has also been
reported to occur for phytosiderophores.81 Owing to the respective stability constants
of the complex that they can form with phytosiderophores (Table 2.3), the competition
between divalent, trace metals and Fe will decrease in the following order: Cu > Ni
> Zn > Mn. In addition, the ecological significance of the impact of the various
exudates on complexation or chelation reaction with various trace metals is still a
question for debate in real soil conditions, as they are likely to be rapidly metabolized
by soil microorganisms that are particularly abundant in the rhizosphere.7
VI. CONCLUSIONS
As drawn schematically in Figure 2.3, a range of mechanisms exists by which plant
roots can alter the chemical conditions in the soil solution in their immediate vicinity.
FIGURE 2.3 Schematic representation of the chemical interactions between plant roots, soil
solution and soil solid constituents that take place in the rhizosphere as a direct consequence
of the biological activity of the root. This diagram depicts how processes occurring at the
solution/root interface can affect (i) a range of soil solution parameters and (ii) thereby the
chemical reactions occurring at the soil solution/soil (solid) interface that ultimately determine
the bioavailability of trace elements for the plant.
Bioavailability of Trace Elements 37
Those changes in ionic and ligand concentrations, pH, and pe can have dramatic
effects on the chemical reactions that occur at the soil solution/soil (solid) interface
and that ultimately determine the bioavailability of trace elements to plants. The
mechanisms involved in these root-induced changes and in the acquisition of trace
elements can vary widely among plant species, and in response to environmental
conditions, too. This diversity needs to be further studied and used in the future,
although it definitively complicates the reliable prediction of the bioavailability
through a single, simple, universal, chemical soil test.
A substantial proportion of the ideas reported herein have been derived from
model experiments, numerical models, or the application of general concepts of soil
chemistry applied to the peculiar chemical conditions of the rhizosphere. Obviously
it is necessary to acquire additional, experimental proofs supporting these ideas and
hypotheses. On the one hand, the refinement of numerical codes by geochemists
and the development of adequate tools and analytical methods by chemists will
certainly provide great help to move forward. On the other hand, the use of new
tools such as those provided by molecular biology will help testing mechanisms
implied at the root-soil interface. This is a challenge for the future as it will help us
improve our capacity to predict and possibly to restrict the contamination of plants
by trace elements.
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77. Cakmak, I., Glt, K. Y., Marschner, H., and Graham, R. D., Effect of zinc and iron
deficiency on phytosiderophore release in wheat genotypes differing in zinc efficiency,
J. Plant Nutr. 17, 117, 1994.
78. Rengel, Z., Root exudation and microflora populations in rhizosphere of crop geno-
types differing in tolerance to micronutrient deficiency, Plant Soil 196, 255260,
1997.
79. Gries, D., Klatt, S., and Runge, M., Copper-deficiency-induced phytosiderophore
release in the calcicole grass Hordelymus europaeus, New Phytol. 140, 95101, 1998.
80. Murakami, T., Ise, K., Hayakawa, M., Kamei, S., and Takagi, S., Stabilities of metal
complexes of mugineic acids and their specific affinities for iron(III), Chem. Lett.
21372140, 1989.
81. Treeby, M., Marschner, H., and Rmheld, V., Mobilization of iron and other micro-
nutrient cations from a calcareous soil by plant-borne, microbial, and synthetic metal
chelators, Plant Soil 114, 217-226, 1989.
3 Rhizosphere Chemical
Changes Enhance Heavy
Metal Absorption by
Plants Growing in
Calcareous Soils
Naomi Assadian and Lloyd B. Fenn
CONTENTS
I. Introduction.....................................................................................................43
II. Materials and Methods ...................................................................................45
A. Site Description ..................................................................................45
B. General Survey ...................................................................................45
C. Sequential Extractions ........................................................................47
D. Statistics ..............................................................................................48
III. Results and Discussion...................................................................................48
A. General Survey ...................................................................................48
B. Vegetation ...........................................................................................49
C. The Rhizosphere .................................................................................51
IV. Conclusions.....................................................................................................56
V. Future Research ..............................................................................................58
References................................................................................................................59
I. INTRODUCTION
Lead (Pb), cadmium (Cd), chromium (Cr), nickel (Ni), zinc (Zn), copper (Cu), and
cobalt (Co) naturally occur in many soils at concentrations less than 300 mg kg1
from weathering of parent materials.1 Elevated metal concentrations often result
from industrial activity. Atmospheric deposition of Pb, Zn, Cd, and Cu from a west
Texas smelter2 operating since the late 1800s was substantial prior to mandatory
environmental laws. For example, airborne emissions of these metals between 1969
and 1971 exceeded 1000, 500, 10, and 1 metric tons for Pb, Zn, Cd, and As,
respectively.3 Adjoining soils contained Pb concentrations exceeding 3000 mg kg1
0-8493-1535-2/01/$0.00+$.50
2001 by CRC Press LLC 43
44 Trace Elements in the Rhizosphere
causing children living near the smelter to experience lead poisoning.3 After stack
emission regulations were in effect, metals were still being emitted. Worthington4
found that native lichens and mosses had been eradicated from the hills surrounding
the smelter. The diversity of winter annual plants had also diminished.
Despite the disappearance of many plants, many have flourished along the banks
of the Rio Grande River, particularly introduced species. It is difficult to generalize
plant responses to heavy metal contaminations.1 Plants are often broadly classified
as metal accumulators, indicators, or excluders.5 Metals accumulate either by foliar
or root absorption. Foliar absorption of metals occurs through leaf cuticles or sto-
mata, an important pathway when atmospheric deposition is high and/or calcareous
soils exist. Foliar absorption may be a contributing factor, but not likely the major
pathway of metal accumulation. More likely, the root and its environment (the
rhizosphere) have the major impact on heavy metal absorption even in a calcareous
environment. Metal mobility and eventual bioavailability in an alkaline calcareous
soil may be dependent on rhizosphere processes.
Nutrient (NO3, K+, and Ca2+) mobility into the rhizosphere is partially by mass
flow and a diffusion gradient established by plant water consumption.6 The presence
of excess nutrients brought into the rhizosphere by mass flow can be demonstrated
by surplus calcium (Ca) surrounding plant roots.6 Soil solution supersaturated with
Ca eventually precipitates forming CaCO3.7,8 Increased CO2 (H2CO3) in organic rich
zones, areas close to the roots for example, leads to the supersaturation of Ca in the
soil solution.9 This increased soil solution Ca concentration is governed by the
solubility product of CaCO3. The mass flow of the Ca containing water to the plant
roots and the water use by transpiration continually concentrate the Ca, leading to
continued CaCO3 precipitation. In a desert environment, considerable amounts of
CaCO3 are deposited around active roots and hyphae forming partially cemented
root channels.10 Heavy metals in a calcareous environment coprecipitate with
CaCO3,11 especially those that have ionic sizes similar to Ca, such as Zn, Mg, and
Mn.12 Metals may also adsorb on clay particles or react with hydroxide compounds
at a high pH.13,14 Precipitated and insoluble metals would control phytotoxicity.14,15
Within the rhizosphere, Gobran and Clegg16 recently published the protocols of
examining plant-available nutrients and trace elements. This conceptual model
assumes that plant roots exist in an equilibrium between the bulk soil (no roots),
rhizosphere (roots), and the soilroot interface (soil in contact with roots). The bulk
soil is the principal reservoir of nutrients. Biological activity in the rhizosphere and
soilroot interface contribute to the concentration of organic acids and other organic
compounds. Greater organic matter, weathering, changes in pH, microorganism-
mediated transformations, and chelation may contribute to a larger fraction of plant-
available metals in these soilroot fractions. Those plant species that have the
potential for significant acid production such as grasses and legumes could further
increase the availability of metals in the soilroot interface (the region nearest the
plant root) resulting in greater plant absorption even in calcareous soils.13 Root
exudates increased the solubilities of Mn and Cu in two acid soils, whereas the
solubilities of Ni and Zn were increased only in the least acidic of the two studied
soils.17 It is suggested that the soilroot interface is the most predictive location for
potential ion absorption by the plant.16
Rhizosphere Chemical Changes Enhance Heavy Metal Absorption 45
B. GENERAL SURVEY
River sediments, soils, and plant tissue were collected at the smelter and Canutillo
locations in April and July, 1993. Within 1 km of the smelter, river sediments were
collected from the channel in the lower flood plain of the river to an approximate
depth of 0.3 m. Surface soil (0 to 15 cm) and subsurface soil (15 to 30 cm) samples
were collected from the shoulder terrain along the river with five replications. All
sediment and soil collection sites had no plants growing in the immediate area.
Aboveground plant tissue (leaves and nonbarky stems) from perennial species was
also collected from a 30-m area along the riverbanks, but not necessarily near
sediment or soil sampling areas. Duplicate samples for each plant species were
46 Trace Elements in the Rhizosphere
TABLE 3.1
Identification and Grouping of Riparian Vegetation Collected at Two Sites
along the Rio Grande River, 1993
Riparian Vegetation Collection
Plant Species Common Name Sitea Plant Type Pubescence Microhabitat
collected from ten different perennial plants (Table 3.1). An identical sampling
procedure was followed at the Canutillo site. Plant species were identified using
plant flora publications by Parker20 and Warnock.21
River sediments and surface and subsurface soils were transported and stored
in plastic bags. Sample preservation and preparation included forced air-drying at
105C for 24 hours, sieving with a 2-mm screen, and pulverizing with a ball mill.
An acid-digestion with nitric acid (HNO3) and hydrogen peroxide (H2O2) following
EPA 3050A protocols was used to determine heavy metal concentrations of Cd,
Cu, Pb, Zn, Cr, Ni, Co, V, and Mn, using inductively coupled plasma spectroscopy
(EPA 200.7). Plant tissue samples were transported and stored in plastic bags. Half
of each plant sample was washed twice with deionized water. The other portion
was left unwashed. Washed and unwashed plant tissues were dried in a forced-air
dryer at 65C for at least 72 hours, ground, and passed through a 40-mesh screen.
Samples were acid-digested with nitric and perchloric acids using EPA 200.2
protocols for heavy metal determinations, using inductively coupled plasma spec-
troscopy (EPA 200.7).
Both sites were revisited in late June, 1994. Plant tissue samples of salt cedar
(Tamarix gallica) and Bermuda grass (Cynodon dactylon L.) were recollected to
verify that preparation methods were not affecting analytical outcomes. Plant tissue
samples were either left unwashed, washed with deionized water, or washed with
0.1 N HCL to remove leaf surface contamination prior to drying for heavy metal
analyses (using EPA 200.2 and 200.7 protocols).
Rhizosphere Chemical Changes Enhance Heavy Metal Absorption 47
C. SEQUENTIAL EXTRACTIONS
In 1997, common Bermuda grass and salt cedar from the smelter site were selected
for further evaluation. Both plant species have been introduced to this area and have
aggressively grown on the banks of the Rio Grande River. A soilroot sampling
scheme was selected that follows protocols by Gobran and Clegg.16 Soil samples
were collected from three soil fractions:
1. Bulk soil (BS) soil between plants where few or no roots were expected;
2. Rhizosphere soil (RS) soil that was removed from the roots with gentle
shaking for a minute;
3. Soilroot interface soil (SRI) soil that was strongly adhering to plant
roots.
Soil samples were collected in triplicate for each plant species. Plant tissue was
also collected from each soil-sampling site. All samples were transported in a cold
chest and kept frozen until chemical analysis. Field-moist soil samples were used
for sequential metal extractions using protocols presented by Kersten and Frstner.18
About a 5-g sample was weighed into centrifuge tubes and extracted by the steps
listed in Table 3.2. The samples were done in triplicate at 25C. All samples were
analyzed for Zn, Mn, Pb, Cu, and Ca by inductively coupled plasma spectroscopy.
The exchangeable form (No. 1, Table 3.2) is usually examined by extraction
with NaCl, NH4-acetate, NH4NO3 or KCl. This form is the most available to plants.
The carbonate form of heavy metals (No. 2, Table 3.2) is usually extracted with a
weak acid such as acetic acid and NH4-acetate at a pH value of 5. The carbonate
metals are less available to the plant than the exchangeable metals. The subsequent
extractant category is easily reducible Mn-oxides or metals associated with the
amorphous oxides (No. 3, Table 3.2). These compounds are extracted with a 0.1 M
TABLE 3.2
Sequential Extraction Scheme for Soil from the Root-Zone of Bermuda Grass
and Salt Cedar
Extraction Shaking
Step Source Metals Extraction Chemical Volume Time
NH2OH-HCl in 0.01 M HNO3 extractant and are even less soluble than the carbon-
ates. If soil carbonates exceed 10 to 20%, some CaCO3 residual dissolution occurs
in this step.19 The moderately reducible Fe-oxides (No. 4, Table 3.2) constitute an
amorphous material of some significant extent, but contain metals of usually low
availability to plants. This class is extracted with NH4-oxalate + acetic acid at a pH
value of 3. The metal-associated oxides would become significant if the soil system
is acidic or anaerobic. The organic-sulfidic class (No. 5, Table 3.2) contains the
organic materials consisting largely of humic and fulvic acids that are very insoluble
and only slightly biodegradable. The extractant is H2O2 at acid pH of 2 that destroys
all organic matter and characterizes the strongly held chelated heavy metals. The
last metal fraction in the sequence is the residual fraction, extracted by concentrated
HNO3 heated to 100C (No. 6, Table 3.2).
Samples from all three fractions were also analyzed for total C (carbon), organic
C, soil pH, and electrical conductivity (EC). Representative subsamples from each
soil sample were dried, pulverized, and analyzed for total carbon using a Carlo Erba
NCHS-O analyzer using standard protocols. Subsamples pretreated with a 10% HCl
solution until bubbling (evolution of CO2) ceased were used to determine inorganic
C concentrations. Inorganic C concentration was determined indirectly by subtract-
ing the organic fraction from the total carbon fraction. Saturation extracts were made
from BS and RS samples to determine soil pH and electrical conductivity following
EPA 4500H+B and EPA 2520B protocols, respectively.
D. STATISTICS
Descriptive statistics, including means, correlations, t-tests, and analysis of variance
(ANOVA) for split plot analyses, were used to summarize chemical analyses for the
general survey of riparian vegetation, river sediments, and desert soil samples col-
lected in 1993 and 1994. For soil samples collected in 1997, ANOVA using SAS22
was used to delineate soil differences associated with plant species (Bermuda grass
and salt cedar), and among soil fractions (BS, RS, and SRI). Statistics were based
on a split plot design with plant species as main parameters and soil fraction as sub-
parameters with three replications. Additional ANOVA analyses were used to delin-
eate which heavy metal form dominated the soil rhizospheres between plant species.
Significant differences were delineated at the 0.10, 0.05, and 0.01 level of probability.
TABLE 3.3
Mean Heavy Metal Concentrations in Sediment, Surface Soil, and Subsurface
Soil at Two Sites along the Rio Grande River, 1993
Conventional
Heavy Soil Sediment Surface Soila Subsurface Soilb
Metal Concentrations Smelter Canutillo Smelter Canutillo Smelter Canutillo
of probability.
and Zn were relatively high (34, 27, and 34 mg kg1, respectively) near the smelter
and greater than those at the Canutillo site (Table 3.3). The lack of greater metal
deposition in sediments at the source was most likely due to a combination of factors.
Metal deposition in the river is diluted and transported in river sediment flow. Settling
of particles is dependent on particle size and water velocity according to Stokes
Law. Floating particles may take several days or weeks to settle. There is also greater
sediment disturbance near a riverbank, which would also account for such low metal
concentrations.
Conversely, surface soils near the smelter showed elevated concentrations of
284, 169, and 108 mg kg1 for Cu, Pb, and Zn, respectively (Table 3.3). To a lesser
extent, surface soils also showed Ni contamination. Contamination of these metals
was also found in subsurface soil samples, but at lower concentrations. High metal
levels would be expected in surface soils. Subsurface soil contamination could be
due to vertical movement (e.g., river flooding) over time or possibly to river levee
disturbance in reshaping the river channel and the roads parallel to the river. Soil
metal levels at the contaminated smelter site fell within the high end of conventional
ranges for uncontaminated soils (Table 3.3).
B. VEGETATION
Three plant species, clover, salt cedar, and Bermuda grass, were collected at both
sites (Table 3.4). Metal concentrations based on a water-wash preparation were
greater in leaf tissue from the smelter site than those from the Canutillo site. Cadmium
and Pb levels were at least two times greater in clover and salt cedar leaf tissue
collected from the smelter site than those from the Canutillo site. All other metal
50 Trace Elements in the Rhizosphere
TABLE 3.4
Heavy Metal Concentrations of Water-Washed Plant Tissue Collected at Two
Sites along the Rio Grande River, 1993
Clover Salt Cedar Bermuda Grass
Plant Smelter Canutillo Smelter Canutillo Smelter Canutillo
Metal Concentrations mg kg1
Cu 220 29 aa 14 b 30 NS 34 NS 127 a 16 b
Cd 0.117.6 1a 0b 2a 1b 4a 1b
Pb 0.50 11 a 4b 10 a 51 b 53 a 11 b
Zn 21120 49 a 37 b 68 a 39 b 120 a 31 b
Cr 10% of soil level 2 NS 8 NS 2 NS 1 NS 2 NS 3 NS
Ni 0.15 2 NS 2 NS 2 NS 1 NS 1 NS 1 NS
Co 0.020.5 0 NS 0 NS 0 NS 0 NS 1 NS 1 NS
Mn 1540 26 a 23 b 24 a 16 b 45 a 141 b
concentrations in clover and salt cedar tissue from the smelter site were either similar
to or greater than those metal concentrations from plant tissue collected at Canutillo.
Plant tissue concentrations of Cu, Pb, and Cr exceeded conventional ranges in all
three plant species. In addition to these three metals, Zn and Mn concentrations were
also high in Bermuda grass. Copper, Pb, and Cr concentrations in clover and salt
cedar tissue ranged from 6 to 45% of that found in contaminated surface soil at the
smelter. Metal concentrations in Bermuda grass tissue were 31 to >100% of that of
contaminated soil. Unlike the other two plant species, Bermuda grass appeared to
be an indiscriminate metal accumulator with high tissue Cu, Pb, and Zn concentra-
tions. More importantly, this indicated that riparian vegetation was either preferen-
tially absorbing soil metals or that the soil samples we collected did not reflect the
actual metal concentrations to which the plant roots are exposed.
Plants were also grouped by morphology, location, or tissue preparation cate-
gories for better interpretation of chemical results (Table 3.5). When grouped by
location, plants absorbed the greatest concentrations of metals where the greatest
concentrations were present in the soil in that environment. Plants near the smelter
showed high levels of Cd, Cu, Pb, and Zn and those at Canutillo showed higher
levels of Cr, Mn, Ni, and Co. Neither plant type nor pubescence appeared to affect
metal uptake. However, monocots tended to absorb more Cu, Pb, and Mn relative
to dicots. Plants with hairy leaves tended to accumulate more Pb. Microhabitat
seemed to be the single most critical factor affecting plant uptake of metals. Plant
species living along the river in high, dry areas had higher metal concentrations than
those rooted in low, wet areas. The flow of the river had minimized the effects of
metal deposition at the source and transported contaminated sediments downstream.
Rhizosphere Chemical Changes Enhance Heavy Metal Absorption 51
TABLE 3.5
Average Heavy Metal Concentrations in Water-Washed
Plant Tissue Categorized by Location, Plant Type,
Morphology, Microhabitat, and Sample Preparation at
El Paso, 1993
Cd Cu Pb Zn Mn
Comparisons mg kg1
Location
Smelter 4.0 ab 87 a 26 a 76 a 35 b
Canutillo 0.5 b 20 b 12 b 56 b 142 a
Plant Type
Dicot 2.0 NS 25 NS 16 NS 60 NS 82 b
Monocota 1.7 70 19 68 123 a
Pubescence
Hairy 2.7 NS 26 NS 12 NS 44 NS 26 b
Smooth 1.6 52 19 70 123 a
Habitat
High Dry Bank 2.5 NS 59 NS 22 a 70 NS 122 NS
Low Wet Bank 0.8 24 10 b 54 67
C. THE RHIZOSPHERE
Average calcium carbonate concentrations of 44.1 and 56.1 g kg1 found in the BS
fraction for Bermuda grass and salt cedar, respectively, significantly increased almost
52 Trace Elements in the Rhizosphere
TABLE 3.6
Determined Soil CaCO3 Concentration, Organic
Carbon Percentage, pH, and EC of Soil Fraction
Surrounding Bermuda Grass and Salt Cedar Roots
Organic
CaCO3 Carbon EC pH
Plant/Soil Fraction g kg1 % dS m1 Units
Bermuda Grass
Bulk 44.1 ba 0.56 b 7.35 NS 7.94 NS
Rhizosphere 35.9 b 1.32 b 11.63 7.80
Soilroot interface 80.3 a 3.80 a
Mean 53.4 NS 1.89 Bb 9.49 NS 7.87 NS
Salt Cedar
Bulk 56.1 b 0.41 b 4.86 7.82
Rhizosphere 60.9 b 1.25 b
Soilroot interface 109.6 a 7.40 a
Mean 75.5 NS 3.02 A
twofold to 80 and 110 g kg1, respectively in the SRI fraction (Table 3.6). An
increasing CaCO3 gradient among soil fractions was more evident for salt cedar than
for Bermuda grass. Salt cedars are high water-consuming trees with deep, woody
roots. The fibrous root system of Bermuda grass can be quite extensive, but typically
remains at shallow depths. Calcium carbonate concentrations were much greater
around salt cedar roots. Limited evaluation of electrical conductivity of the saturation
extract showed similar increases to that of CaCO3 for Bermuda grass. Organic C
also increased from BS to RS and from RS to SRI. The increase from BS to SRI
was greater than sixfold for Bermuda grass and about 18-fold for salt cedar. There
were no apparent changes in pH between BS and RS fractions. It was also observed,
but not measured, that the soil texture of the SRI was of a finer composition than
the texture of the other two soil fractions.
Heavy metal concentrations in the BS fraction were similar to metal concen-
trations of the general soil survey of the smelter site (Tables 3.3 and 3.7). However,
metal concentrations were lowest in the BS fraction and increased in the order BS
< RS < SRI (Figure 3.1). Despite increased amounts of CaCO3 in the SRI of salt
cedar, total metal concentrations were greater in the root rhizosphere of Bermuda
grass rather than of salt cedar. ANOVA analyses showed statistical differences in
metal accumulation in the rhizosphere between plant species for the most soluble
metal fractions of exchangeable and carbonate association (data not shown). Low
Rhizosphere Chemical Changes Enhance Heavy Metal Absorption 53
TABLE 3.7
Averaged Bermuda Grass and Salt Cedar Soil Extraction Concentrations of
Heavy Metal Forms According to Soil Fraction
Soil Fraction Cu Pb Zn Mn Ca
Extractant g kg1 mg kg-1
replication likely masked more frequent significant differences between soil fraction
and plant species. It appeared that plant species not only accumulated metals
differently, but also affected the types of metal species within the RS and SRI
(Figure 3.2).
From sequential extraction of root zone soils, we found exchangeable Cu to
reach 8.2 mg kg1 in SRI soil, a value probably in excess of 100 times that needed
for plant nutrition (Table 3.7). Other than Ca and Cu, no other metal was found in
excessive amounts in the exchangeable form. Copper also dominated carbonate-
associated, easily reducible Mn-oxides, moderately reducible Fe-oxides, and the
sulfidic-organic forms. Lead species were mainly associated with carbonate and
residual forms (p > 0.01). Zinc was primarily associated with insoluble forms such
as moderately reducible Fe-oxides, organic, and residual fractions (p > 0.01).
In comparison to bulk soils, metal forms varied in the RS and SRI (Figure 3.2).
With increasing proximity to root, Cu was increasingly associated with Fe-oxides,
organic, and residual forms. Soluble Cu forms increased in the salt cedar rhizosphere,
54 Trace Elements in the Rhizosphere
FIGURE 3.1 Total concentrations of Cu, Pb, Zn, and Mn in the soil from the BS (bulk soil),
RS (rhizosphere soil), and SRI (soilroot interface) of Bermuda grass and salt cedar growing
next to a west Texas smelter site.
but not for Bermuda grass. Similarly, the increased Pb and Zn within the Bermuda
grass rhizosphere were associated with insoluble metal forms (organic and residual
forms), whereas increased soluble metal forms (moderately reducible Fe-oxide and
organic fractions) were found within the salt cedar rhizosphere.
Calcium release at the sequential step for easily reducible Mn-oxides suggested
that not all CaCO3 was dissolved in the carbonate-associated fraction as reported by
Prez-Sirvent et al.19 As a consequence, greater metal quantities may be associated
Rhizosphere Chemical Changes Enhance Heavy Metal Absorption
FIGURE 3.2 Relative change in Cu, Pb, and Zn concentration from bulk soil, rhizosphere soil, and soilroot interface as influenced by Bermuda grass
and salt cedar growth.
55
56 Trace Elements in the Rhizosphere
with carbonates than the data might indicate. In the moderately reducible Fe-oxide
phase, large amounts of Cu and Zn were found, but no Pb or Ca (Table 3.7). Since
the extractant here was NH4-oxalate, the Ca and Pb were evidently precipitated as
oxalates. The organic fraction was high in Zn and Cu concentrations. The highest
concentrations of metals were found in the residual fraction.
IV. CONCLUSIONS
Our results reveal that heavy metals have accumulated in an alkaline desert soil, and
to a lesser degree in river sediment, at the source of smelter contamination. A general
survey of plant vegetation also showed that some plants, in particular Bermuda grass,
might accumulate metals growing on the high dry banks of the Rio Grande River.
From the data, we concluded that metal accumulation in a contaminated desert soil
was not uniform. Both the macro- and microenvironments (those minerals nearest
the root) impacted plant absorption of heavy metals. Heavy metals, Ca content, and
organic C in the soilroot interface were significantly greater than those concentra-
tions in bulk soil. Gobran et al.23 stated that mineral weathering is probably most
severe next to the root, as this is a plants mechanism to obtain required nutrients.
The question is how these metals concentrated in the plant rhizosphere.
It appears that additional abiotic processes such as wind erosion and rain splash
may also contribute to localized metal accumulation in the rhizosphere of desert
plants. Schlesinger and Pilmanis24 found soils N, P, and K were associated with the
presence of shrubs in desert habitats. Intershrub spaces had lower fertility. They
suggested that finer particles were wind-deposited on shrub mounds. Improved soil
stability, soil structure, and water infiltration allowed for greater vertical movement
of nutrients into soil. Banin et al.25 also found that plants act as biofilters of metal-
laden dust. It is possible that metal mobility in this situation was the movement of
precipitated insoluble forms from surface or bulk soils moving into the rhizosphere
as small amorphous particles rather than as ionized forms via mass flow. However,
these high soil concentrations are not excessively absorbed by the biotic species in
the arid-calcareous system (Table 3.4). It is accepted that calcite presence reduced
the potential toxicity of most heavy metals.26
Within the soil profile, the concentration of soil metals increased from bulk soil
to soil in intimate contact with soil roots (the soil rhizosphere and soilroot interface).
The development of variable metal and CaCO3 partitioning among these soil frac-
tions in the desert ecosystem is difficult to answer. Mass flow would seem to be the
more obvious answer.6 Surplus Ca in a CO2-rich environment would cause CaCO3
precipitation as plants reduce the water content of the soil.7 Salt cedar trees have a
greater water consumption potential than Bermuda grass and would account for
greater Ca as CaCO3 deposition within the salt cedar rhizosphere. Suarez et al.9 also
found a two- to threefold increase (a supersaturated Ca solution) in Ca concentration
in solution from an organic-matter-rich zone. The increased CaCO3 in the SRI could
be the result of large increases in unused Ca in the soil solution. In the presence of
high pCO2 we have a reduced precipitation potential, but yet the largest precipitated
fraction in the soilroot system. The observation of CaCO3 shells around old root
channels is frequent in arid regions.10
Rhizosphere Chemical Changes Enhance Heavy Metal Absorption 57
In acid soils, Gobran and Clegg16 found increased nutrients (Ca) and decreased
metals (Al) next to the roots (SRI) suggesting completely different growing condi-
tions than thought from analyses of the bulk soil. In these arid zone soils, proximity
to the roots resulted in up to a threefold increase in all metals (Table 3.7). Increased
organic matter closer to the roots may decrease the tendency for CaCO3 precipita-
tion,9 but yet the largest CaCO3 accumulations were found next to the roots. Bio-
logically active soil organic matter (a heterogeneous mix of living and organic
materials that are readily circulated through biological pools) may also have con-
tributed to metal availability by increasing the acidity of soil microsites. Indirect
evidence of increased exchangeable Mn from our results (Table 3.7) suggested that
Mn released from oxides must have occurred at acidified microsites. Factors affecting
dissolution or preventing precipitation of CaCO3 would affect metal bioavailability.
Boundaries between soil BS, RS, and SRI were diffuse rather than abrupt.
Methodology to partition these soil fractions may have contributed to inaccuracy.
Sand particles were readily separated from the SRI fraction when plant roots were
gently shaken. Only fine, aggregated particles adhered to root hairs (SRI). By
definition of the SRI,16 we likely segregated the most biologically interactive and
root-influenced zone. As a consequence, metal concentrations were elevated. Using
the sequential extraction data to determine bioavailability of metals, we also noted
that much of the Cu, Zn, Pb, and Mn were associated with the CaCO3 phase. In the
easily reducible Mn-oxides fraction, a solubilization of precipitated Ca suggested
that the CaCO3 was solubilized in even greater quantities, a problem in the proper
use of these data (Table 3.7). One cannot be certain of how much of the heavy metals
(Cu, Pb, and Zn) would be coprecipitated with the manganese precipitates. Appar-
ently, a large quantity of the metals must have been surface precipitated on the calcite
crystals as much less was solubilized in the Mn-oxide extraction with even a larger
amount of solubilized Ca (residual CaCO3) than in the carbonate category. Since
this is an equilibrium state, all soil extract values would have some value as potential
reserves. These reserves would be available with localized or general soil pH value
decreases in the SRI.
Concentrations of Cd, Cu, Pb, Zn, and Mn were often many times greater in
Bermuda grass tissue than in salt cedar tissue. Although metal concentrations in
leaf tissue were not proven to be statistically different, we feel that there are real
differences between plant species in metal absorption potential that is possibly
linked to differences in the rhizosphere. The fibrous, shallow root system of Bermuda
grass creates a continuous shallow root mass where metals have accumulated from
atmospheric fallout. Conversely, salt cedar roots are typical of dicotyledonous trees
with deep woody roots with fragmented zones of active root growth. Differences
in root morphology between monocots and dicots, and the shallow soil concentration
of root mass and active absorption sites may affect differences in plant uptake of
heavy metals.
Despite the limitations of soil fractionation and sequential extraction to deter-
mine metal forms in soils, the results demonstrated that not all metals are uniformly
distributed and immobilized in a calcareous desert soil. Calcium carbonate may not
always be a plant barrier to metal uptake in a calcareous soil. Entry of metals into
the food chain is possible from wildlife grazing on grass.
58 Trace Elements in the Rhizosphere
V. FUTURE RESEARCH
This study illuminated a need to examine dominant native flora on the high lands
and plains of the Chihuahua Desert. Dicots such as mesquite (Prosopis glandulosa,
P. julifera, and P. pubescens), acacia (Acacia greggii and A. constricta), four-winged
saltbush (Atriplex canescens), and creosote bush (Larrea tridentata), and monocots
such as yucca (Yucca baccata, Y. elata, Y. torreyi) and Spanish dagger (Agave
lecheguilla) are widespread and commonly grazed by cattle and wildlife. Metal
accumulation patterns in the rhizosphere and in plant tissue needs documentation
to compare with those of introduced species such as Bermuda grass. A broader plant
survey will reveal possible mechanisms of metal mobility into the rhizosphere and
patterns of accumulation in desert environments. It seems likely that our hypothesis
of mass flow of precipitated, clay-sized particles laden with metals could effectively
occur in sandy desert soils, but it is questionable if mass flow would be a primary
metal carrier in soil textures with finer particles commonly found in cultivated lands.
The concept of soil fractionation of the root zone applied to crop fertility needs
reexamination, particularly for irrigated agriculture in arid and semiarid regions.
Pecan, cotton, corn, and wheat might benefit from root-zone nutrient studies. It is
known that Ca in an irrigated arid zone accumulates around the roots. Information
on the movement, deposition, and bioavailability of P and K needs examination.
Our understanding could be expanded by these characterizations. Barber and
coworkers6 began rhizosphere studies many years ago. However, rhizosphere data
were primarily intended for seasonal fertility recommendations. This type of research
might provide information, hopefully, to establish nutrient indices between bulk and
rhizosphere soils or information for improved cultivation techniques for future years.
We also questioned whether our methodology in physical partitioning of root
zone soil samples affected soil particle size fractionation. Did larger particles drop
out of the root zone because of size and weight, leaving small-sized particles
organically cemented to the roots? Or, were fine soil particles interfacing with
roots the result of in situ weathering processes and mass flow. As the roots grow
through the soil, they may proliferate around these smaller-sized particles, but also
be exposed to sand and silt constituents. We hypothesize that a chemical marker
such as CaCO3 in calcareous soils may be used to delineate rhizosphere boundaries
in conjunction with physical methods.
Lastly, the use of sequential extraction protocols in this study suggested that
metal accumulation was greatest in the soilroot interface at root absorption sites.
Yet, metal absorption was low in comparison to soil metal concentrations. The
novelty of insoluble metal compounds being concentrated at the root absorption site,
yet not being absorbed at high rates, leaves many questions unanswered. The phys-
ical, chemical, and biological interactions affecting metal solubilities and precipita-
tion products needs to be more closely examined.
Rhizosphere Chemical Changes Enhance Heavy Metal Absorption 59
REFERENCES
1. Fergusson, J. E., The Heavy Elements, Chemistry, Environmental Impact, and Healthy
Effects, Pergamon Press, Oxford, England, 1970, 614.
2. Woodside, T. J. and Roberts, B., The El Paso smelter, in Nelson, L. A. and Haighs,
B. R., eds, West Texas Geological Society Guide Book, 1958 Field Trip, Franklin and
Hueco Mountain, Texas, The West Texas Geological Society, Midland, TX, 1958,
7577.
3. Landrigan, P. J., Gehlbach, S. H., Rosenblum, B. F., Shoults, J. M., Candelaria, R. M.,
Barthel, W. F., Liddle, J. A., Smrek, N. W., Staehling, N. W., and Sanders, J. F.,
Epidemic lead absorption near an ore smelter. The role of particulate lead, New
England J. Med., 292, 123129, 1975.
4. Worthington, R. D., Effects of El Paso pollutants on the lichen, moss, and winter
annual flora on andesite rock formations, in Ganster, P. and Walter, H., eds., Envi-
ronmental Hazards and Bioresource Management in the United States-Mexico Bor-
derlands, UCLA Latin America Center Publication, University of California, Los
Angeles, CA, 1990, 105115.
5. Whiting, S. N., Leake, J. R., Baker, A. J. M., and McGrath, S. P., Changes in the
phytoavailability of zinc to plants sharing a rhizosphere with the zinc hyperaccumu-
lator Thlaspi caerulescens J. and C. Presi, in Iskandar, I. K., Hardy, S. E., Chang, A.
C., and Pierzynski, G. M., eds., Proceedings of the Fourth International Conference
on the Biogeochemistry of Trace Elements, June 2326, Clark Kerr Campus, Berkeley,
CA, 1997, 469.
6. Barber, S. A. and Ozanne, P. G., Autoradiographic evidence for the differential effect
of four plant species in altering the calcium content of the rhizosphere soil, Soil Sci.
Soc. Am. Proc., 34, 635637, 1970.
7. Miyamoto, S. and Pingitore, N. E., Predicting calcium and magnesium precipitation
in saline solution following evaporation, Soil Sci. Soc. Am. J., 56, 17671775, 1992.
8. Feagley, S. E. and Hossner, L. R., Ammonia volatilization reaction mechanism
between ammonium sulfate and carbonate systems, Soil Sci. Soc. Am. J., 42, 364367,
1977.
9. Suarez, D. L., Wood, J. D., and Ibrahim, I., Re-evaluation of calcite supersaturation
in soils, Soil Sci. Soc. Am. J., 56, 17761784, 1992.
10. Monger, H. C., Daugherty, L .A., and Lindemann, W. C., Microbial precipitation of
pedogenic cacite, Geology, 19, 9971000, 1991.
11. McBride, M. B., Chemisorption of Cd2+ on calcite surfaces, Soil Sci. Soc. Am. J.,
4, 2628, 1980.
12. Xiang, H. F. and Banin, A., Solid-phase manganese fractionation changes in saturated
arid-zone soils: Pathways and kinetics, Soil Sci. Soc. Am. J., 60, 10721080, 1996.
13. Mengel, K. and Kirby, E., Principles of Plant Nutrition, International Potash Institute.
Worblaufen-Bern/Switzerland, 1982, 655.
14. Harter, R. D., Effect of soil pH on adsorption of lead, copper, zinc, and nickel, Soil
Sci. Soc. Am. J., 47, 4751, 1983.
15. Basta, N. T., Pantone, D. J., and Tabatabai, M. A., Path analysis of heavy metal
absorption by soil, Agron J., 85, 10541057, 1993.
16. Gobran, G. R. and Clegg, S., A conceptual model for nutrient availability in the
mineral soil-root system, Can. J. Soil Sci., 76, 125131, 1996.
17. Mench, M. and Martin, E., Mobilization of cadmium and other metals from two soils
by root exudates of Zea mays L., Nicotiana tobacum L. and Nicotiana rustica L.,
Plant Soil., 132, 187196, 1991.
60 Trace Elements in the Rhizosphere
18. Kersten, M. and Frstner, U., Chemical fractionation of heavy metals in anoxic
estuarine and coastal sediments, Wat. Sci. Tech., 15, 121130, 1986.
19. Prez-Sirvent, C., Martinez-Sanchez, J., and Garcia-Rizo, C., Lead mobilization in
calcareous agricultural soils, in Selim, H. M. and Iskander, I. K., eds., Fate and
Transport of Heavy Metals in the Vadose Zone, Ann Arbor Press, Ann Arbor, MI,
1999, 328.
20. Parker, K. F., An Illustrated Guide to Arizona Weeds, The University of Arizona Press,
Tucson, AZ, 1982, 338.
21. Warnock, B. H., Wildflowers of the Guadalupe Mountains and the Sand Dune Country,
Texas, Sul Ross State University, Alpine, TX, 1974, 176.
22. Statistical Analysis System (SAS) Institute, Inc. SAS Users Guide: Statistics, Version
6.03, Cary, NC, 1988.
23. Gobran, G. R., Clegg, S., and Courchesne, F., Rhizospheric processes influencing the
biochemistry of forest ecosystems, Biogeochemistry, 42, 107120, 1998.
24. Schlesinger, W. H., and Pilmanis, S. M., Plant-soil interactions in deserts, Bio-
geochemistry, 42, 169187, 1998.
25. Banin, A., Navrot, J., and Perl, A., Thin horizon sampling reveals highly localised
concentrations of atmophile heavy metals in a forest soil, The Sci. Total Environ., 61,
145152, 1987.
26. Shuman, L. M., Effect of liming on the distribution of manganese, copper, iron and
zinc among soil fractions, Soil Sci. Soc. Am. J., 50, 12361240, 1986.
4 Fate of Radiocesium in
Soil and Rhizosphere
Bruno Delvaux, Nathalie Kruyts, Emmanuel
Maes, and Erik Smolders
CONTENTS
I. Introduction.....................................................................................................61
II. Environmental Fate of Radiocesium ..............................................................63
A. Atmospheric Deposition of Radiocesium ..........................................63
B. Target Ecosystems ..............................................................................63
C. Primary Interception by the Canopy ..................................................64
D. Foliar Uptake, Translocation, and Primary Contamination ...............64
E. Radiocesium Incorporation in the Biogeochemical Cycle ................64
F. Summary .............................................................................................65
III. Retention of Radiocesium in Soil ..................................................................67
A. Introduction.........................................................................................67
B. Mechanisms ........................................................................................67
1. Intrinsic Properties...............................................................69
2. Surrounding Conditions.......................................................73
C. Quantitative Analysis of Radiocesium Retention in Soils.................74
D. Radiocesium Retention and Soil Components...................................76
IV. Root Uptake and Soil-to-Plant Transfer.........................................................78
A. Root Uptake ........................................................................................78
B. Soil-to-Plant Transfer .........................................................................78
V. Mobilization of Radiocesium in the Rhizosphere .........................................79
VI. Conclusion ......................................................................................................82
References................................................................................................................83
I. INTRODUCTION
Since 1945 the radioisotopes 134Cs and 137Cs have been released in the environment
by three main routes: nuclear weapons testing, controlled discharge of waste effluents
from nuclear plants, and accidental release.1 After deposition, much of the radioce-
sium becomes disseminated in aquatic environments, but large quantities remain in
terrestrial ecosystems. From the extensive research on the environmental behavior
of radiocesium in weapons and accidental fallout, a general consensus emerged that
radiocesium exhibits a biogeochemical behavior rather similar to that of potassium,
0-8493-1535-2/01/$0.00+$.50
2001 by CRC Press LLC 61
62 Trace Elements in the Rhizosphere
concentration in plant Bq kg 1 Bq kg 1
TF = 1
or 3
(1)
concentration in soil Bq kg Bq dm
concentration in plant Bq kg 1
Tagg = (2)
concentration in soil Bq m 2
concentration in soil Bq kg 1
Kd = (3)
concentration in solution Bq l 1
The uptake of radiocesium by plant roots and associated microflora is the first
step in plant, and hence food chain contamination. The chemical uptake of trace as
well as major elements is regulated by active and passive mechanisms occurring in
the rhizosphere, at the soil solutionroot interface. These processes and their inter-
action with the soil matrix, were recently reviewed in a state-of-the-art study.14 They
take place in physicochemical conditions that largely depend on ion exchange
reactions between soil components, soil solution, and plant root. The aim of this
chapter is fourfold. First, it will briefly review the environmental fate of radiocesium
and identify the major sinks and pathways for trace Cs. Second, it will focus on the
quantitative analysis and mechanisms of trace Cs retention in soil. Third, it will
briefly review the effect of ionic environment on root uptake of radiocesium as well
as recent approaches modeling the 137Cs soil-to-plant transfer. Fourth, it will consider
the mobilization of radiocesium in the rhizosphere and identify the key factors and
processes in this microenvironment.
B. TARGET ECOSYSTEMS
The contamination of lands with radiocesium rapidly leads to its accumulation in
soil, in both agricultural areas as well as in natural and seminatural environments.
Radionuclide contamination of agricultural lands is obviously highly sensitive,
because of the direct burden on the food chain. Seminatural areas are recognized as
environments showing little interference with man (forests, upland pastures, moor-
land, marshland, peatland, alpine meadows, unimproved meadows, and tundras).
According to Poldini,21 seminatural can be best defined versus natural areas. In the
latter, the flora and fauna are original and spontaneous and the vegetation has not
undergone structural modifications by man. In the seminatural countryside the flora
and the fauna are still mainly native but the vegetation is greatly modified by human
intervention; the relationships between biological forms have been modified. The
dominant biological form has often been replaced and the derived vegetation no
longer belongs to the same formation as the original vegation.21 The seminatural
64 Trace Elements in the Rhizosphere
environments produce human food stuffs as well as various materials. The use of
these ecosystems is important for some population groups.22,23
After the Chernobyl accident, it was observed that the vegetation in natural and
seminatural environments was more highly contaminated with 137Cs than crops from
agricultural lands.10,11,24 Higher 137Cs biological half-lifes were measured in these
environments versus agricultural areas, demonstrating a longer persistence in the
natural and seminatural ecosystems.25 Consequently, one can expect in the forth-
coming years an important increase of radiocesium doses to man from the contri-
bution of these highly complex ecosystems.9,22,26 Among these environments, forest
ecosystems are particularly complex as they involve several vegetative strata as well
as diverse plant communities.
retained in mineral, trace Cs migration rates are quite small and generally less than
2 cm yr1.35,38,39 The major pathway for plant contamination is root uptake. Tradi-
tionally, the assessment of the contamination of above standing vegetation with
radiocesium is commonly based on the determination of Transfer Factor (TF) or
aggregated Transfer Factor (Tagg) coefficients expressing the ratio of radionuclide
concentration in the plant to that in soil. A number of in situ measurements have
been accumulated in several studies, particularly after the Chernobyl accident.
Reported TFs exhibit large seasonal as well as yearly variations in the same eco-
system.40-44 Also, they vary largely between ecosystems, soils, plant, and fungal
species. Figure 4.1 gives a global picture of reported 137Cs Tagg values measured in
agricultural as well as seminatural environments in some case studies.
The range of the orders of magnitude of the 137Cs Tagg mean values is 102 to
10 m2 kg1 in seminatural environments and 104 to 101 m2 kg1 in agricultural
1
areas. In the latter areas, some Tagg values can be as low as 5 106 m2 kg1. Noteworthy
is the fact that the 137Cs soil-to-plant transfer factor is higher in seminatural envi-
ronments than in agricultural lands. Such high transfer contributes to a much longer
persistence of radiocesium in the seminatural ecosystems, as illustrated in a number
of papers.10,12,13,47,48 In contaminated seminatural environments, mushrooms are well-
known accumulators of radiocesium (Figure 4.1).24,49,50 On a dry weight basis, fungi
accumulate between 10 and 150 times more radiocesium from soil than vascular
plants, as recently reported in the extensive reviews by Avery1 and Nimis.3 Among
the higher plants, heather, Calluna vulgaris, accumulates more radiocesium than
other species.42 The uptake of radiocesium by the understory vegetation largely
contributes to the contamination of the forest phytocenosis.3 Though not illustrated
in Figure 4.1, variations in TF or Tagg values can be related to soil properties in
seminatural environments, as reviewed by Nimis.3 In this respect, vegetation growing
in organic soils appear to be more susceptible to radiocesium contamination than
that growing in clay soils.51 Such a relationship can also appear in agricultural areas,
as a trend in decreasing Tagg values can be observed from peaty to sandy and clayey
soils (Figure 4.1). However, so far, trace Cs TF or Tagg values measured in situ have
never been significantly related to any commonly measurable soil property such as
pH, CEC, organic matter or clay content, or K level and saturation, etc. in a
mechanistic way. Even in controlled pot experiments involving agricultural soils of
widely varying textural classes, 137Cs+ TF is unrelated to clay content.50 Relatively
high radiocesium soil-to-plant transfer has been generally associated with peaty,
organic-rich soils as well as podzols and some sandy soils.51,53-58 In contrast, clay-
rich soils have been generally associated with relatively low 137Cs TF.3
F. SUMMARY
A very high variability is thus generally observed in 137Cs soil-to-plant transfer
patterns, particularly in seminatural environments. That variability requires a com-
prehensive understanding of the microbiogeochemical processes involved in the fate
of radiocesium in soils. Indeed, three main processes will govern the fate of radio-
cesium once accumulated in soil: uptake by living organisms, migration with the
soil solution, and retention on soil particles. A dynamic integration exists between
66
Trace Elements in the Rhizosphere
FIGURE 4.1 Aggregated transfer coefficients for 137Cs+ to food products from seminatural ecosystems and agricultural areas. The solid points represent
the expected values and the vertical lines represent the range of the values. (References: no asterisk:I.A.E.A.45; *Horrill et al.46; **Salt and Mayes41;
***Salt and Mayes42.)
Fate of Radiocesium in Soil and Rhizosphere 67
these processes in soil. More particularly, these processes and their interactions take
place with the highest intensity in the rhizosphere, i.e., the soil volume influenced
by root activity.14 The uptake of major and trace elements takes place in a physico-
chemical environment that depends to a large extent on ion exchange reactions
between the soil matrix, the liquid phase, and the plant root with associated micro-
flora.14,59 In this respect, soil particles and hence soil minerals are key components
of the rhizosphere as they closely interact with biota.14 The next two sections will
be devoted to i) the behavior of radiocesium in soil at the interface solid-solution
and its relationship with soil components, and ii) the uptake of radiocesium by plant
roots at the interface solution-root. The fifth section will involve the mobilization
of radiocesium in the rhizosphere, in which plant roots strongly interact with soil
minerals. Rhizospheric processes are likely to be of major importance in seminatural
ecosystems, because the soil is not fertilized.
B. MECHANISMS
The sorption of trace cesium by soils, sediments, and clay minerals has been widely
studied, as recently reviewed by Cornell.62 It is now generally recognized that
specific adsorption of radiocesium is associated with the presence, even in very
small quantity, of micaceous clay minerals. In particular, the interlayer space of
these phyllosilicates exhibits areas formed by the juxtaposition of nonexpandible
(1.0 nm) and hydrated (1.4 nm) zones. Such areas are called wedge zones and their
associated sites are the frayed edge sites (FES).63-65 The FESs contribute to less than
2% of the overall CEC.65,66 They selectively sorb poorly hydrated alkaline cations
K+, Rb+, Cs+ as well as NH4+ ion. Because they easily loose their hydration water,
these cations are specifically sorbed in the wedge zones. If the concentration of
such cations is sufficient, their specific sorption in the wedge zones induces the
collapse of the interlayer space, and hence their fixation by the layer silicate. This
process is illustrated in Figure 4.2 in which three steps of the collapse are schemat-
ically represented.
68 Trace Elements in the Rhizosphere
FIGURE 4.2 Schematic illustration of the collapsing process of the interlayer space of 2:1
micaceous phyllosilicates with increasing Cs+ saturation of interlayer sites (respective layer
and interlayer thickness are not at scale).
The collapse forms new wedge zones, which are closer to the border of the
particle. The hydrated cations such as Na+, Ca2+, and Mg2+ maintain expansion of
the interlayer space, but are not sorbed in the wedge zones. Thus, the low hydration
energy is the major factor explaining the specific sorption of trace Cs+ by micaceous
minerals.64 For that reason, the decrease in cation selective adsorption by the FESs
follows the increase of the hydration energy (Cs+ > Rb+ > NH4+ > K+). Similarly, the
relatively low hydration energy of Cs+ explains why hydrated cations can readily
induce the expansion of the interlayer space of a micaceous clay mineral exchanged
with K+, but not with Cs+.64
It is generally accepted that the adsorption of radiocesium by micaceous phyl-
losilicates can be described in terms of at least three kinds of sites exhibiting a
decreasing selectivity for poorly hydrated cations.64-68 The location of the three kinds
of sites in a micaceous clay mineral is schematically illustrated in Figure 4.3.
Fate of Radiocesium in Soil and Rhizosphere 69
FIGURE 4.3 Schematic view of the various kinds of sites in an idealized structure of 2:1
micaceous phyllosilicates (interlayer distances are purposely overmagnified for clarity).
First, the frayed edge sites (FES) are located in the wedge zones of the mineral,
as described above, and specifically sorb the poorly hydrated alkaline cations and
NH4+ ion. Second, the hydrated interlayer sites are located in the expanded interlayer
zones, in which the hydrated cations can be exchanged with poorly hydrated cations.
This ion exchange process can, however, induce the collapse of the interlayer space.
Third, the regular exchange sites (RES) are planar sites located on the external
surfaces of the phyllosilicate and regular ion exchange processes occur on such sites.
These processes involve exchange surfaces that are freely accessible to all ions,
hence ion exchange will be governed by valence and hydration effects. Valcke69 as
well as Hird et al.70 have measured a decreasing selectivity for trace Cs+ ions in the
FESs with increasing distance from the collapsed interlayer sites (Figure 4.3), i.e.,
the mica core (1.0 nm), toward the expanded hydrated zone (1.4 nm). Such decreas-
ing selectivity for Cs+ sorption is due to increasing accessibility of competing cations,
which are more hydrated than Cs+. In fact, defining various kinds of sites, each
exhibiting its own and precise selectivity for trace Cs+, is likely a theoretical view
that gives some useful description, but oversimplifies the reality.
The selective adsorption of poorly hydrated cations and their retention by 2:1
phyllosilicates both depend on a number of factors that can be classified in terms
of intrinsic properties of the mineral itself and external conditions related to the
environment of the mineral particles.
1. Intrinsic Properties
The intrinsic properties of the 2:1 phyllosilicates that play a key role in the Cs+
selective sorption are: the layer charge, the octahedral occupancy, the extension
of the wedge zones, the interlayer occupancy, the particle size, and the layer
stacking order.
70 Trace Elements in the Rhizosphere
The selective sorption of Cs+ ions is obviously promoted by a high layer charge
of the 2:1 phyllosilicate. In this respect, the selectivity for Cs+ would decrease in the
order mica (illite) vermiculite smectite.62,63,71 The location of the negative
charge, i.e., its distribution in the octahedral and/or tetrahedral sheet, has often been
evoked as a factor influencing the selective sorption of poorly hydrated cations. The
tetrahedral charge would exert a more direct influence on layer collapse than the
octahedral charge would, because of the proximity of the tetrahedral sheet relative
to the interlayer space.72 In this respect, a larger Cs concentration is required to induce
the collapse of smectites with dominant octahedral charges versus smectites with
dominant tetrahedral charges.73 However, Kodama et al.74 have shown in phlogopite
that the location of the charge was less a determinant than the octahedral occupancy
for the release or extractability of interlayer potassium. These authors suggested that
the location of the charge would play more of a key role on the swelling behavior of
the mineral after K extraction than on the K extractability sensu stricto.
The di- or trioctahedral character of the phyllosilicate measures the octahedral
occupancy of the mineral. It is well known that K+ ions are retained much more in
dioctahedral micaceous phyllosilicates than in the trioctahedral ones.75 The orienta-
tion of the OH dipole has long been considered as the major factor in this differ-
ence.75-77 The orientation of the OH group is quasi-perpendicular in trioctahedral
minerals, whereas it shifts toward the octahedral vacancy in the dioctahedral min-
erals, making an angle of 16 with the (a,b) plane of the layer silicate. In the latter
case, a larger distance between interlayer K+ and the proton of the OH group involves
a lower cation-to-cation repulsion and hence a larger stability in the dioctahedral
minerals versus the trioctahedral ones.75 In fluoride-rich micas, however, the K+H+
repulsion does not occur as the F ion replaces the OH dipole.78 The value of the
rotation angle of the tetrahedrons is another factor influencing the difference in
stability between the di- and trioctahedral minerals because the tetrahedron rotation
is larger in the former minerals.79
The extension of the wedge zones, i.e., the number of specific Cs+ sorbing
sites,71 increases in the order mica (illite) vermiculite.62,71 Using Rb+Ca2+ ion
exchange on a progressive K-depleted biotite (1.0 nm), Le Roux and Rich80 dem-
onstrated that, with increasing K depletion, the selectivity for Cs sorption first
increased, reached a maximum, and further decreased at the full expanded stage of
vermiculite (1.4 nm). Such variation is caused by the extension of the wedge zones
during the process of vermiculitization. The wedge zones are initiated at the border
of the mineral particles and multiplied during the early stage of the transformation
of mica. Their number can be regarded as being steady in the intergrade structure
mica-vermiculite. A further expansion leads to the coalescence of the hydrated
interlayer sites (Figure 4.3), the progressive disappearance of the wedge zones, and
hence the decrease in selectivity for Rb+ ions. Similarly, larger Cs sorption yields
have been measured after K removal from a biotite.81,82
The extension of both the wedge zones and their associated hydrated interlayer
surfaces obviously depends on the interlayer occupancy (Figures 4.2 and 4.3). In
this respect, a full occupancy of the interlayer space by K in mica reduces the
sorption yield of Cs+ ions in a large range of Cs concentration in the equilibrium
solution.81,83 Similarly, aluminium interlayering of vermiculites reduces the CEC of
Fate of Radiocesium in Soil and Rhizosphere 71
FIGURE 4.4 Schematic illustration of the variation in 137Cs+ sorption and retention (after
sequential desorption using first a K+-Ca++-saturated resin and then an H+-resin) of a laboratory
weathering sequence mica vermiculite hydroxy interlayered vermiculite (HIV)
vermiculite. (Adapted from Maes, E. et al.82)
Fate of Radiocesium in Soil and Rhizosphere 73
Similarly, Maes et al.90 demonstrated that increasing the stacking order could form
highly selective Cs-exchange sites in montmorillonites and increase Cs retention by
the clay minerals. Prolonged wetting-drying cycles were shown to largely increase
the retention of radiocesium by soils of widely varying properties, despite their
initially low retention.91
2. Surrounding Conditions
These conditions involve the composition of the solution surrounding the sorbing
sites (concentration of Cs+ and other ions, ionic strength), the solid to solution ratio
as well as the time of contact between the Cs+ sorbing particles and the solution.
Traditionally, the behavior of radiocesium at the solidsolution interface has
long been described in terms of Cs+ solid-liquid distribution coefficient (Kd). The
effect of Cs concentration on Kd has been well studied.62 At Cs concentration less
than 108 M, it seems that Kd is independent of Cs concentration in the equilibrium
solution. At larger concentration, Kd decreases with increasing Cs concentration and
consequently, the Cs sorption is nonlinear.66,92,93 Such nonlinearity is generally inter-
preted as being the expression of the heterogeneity of the Cs sorbing sites.64,90,91
With increasing Cs concentration, the FESs are progressively more and more satu-
rated with Cs+ ions. Consequently, further sorption of Cs+ ions involves exchange
sites with decreasing selectivity (hydrated interlayer sites, RES); as a result, the
mean affinity of the exchanger for Cs decreases, hence Kd diminishes. The Cs
concentration also influences the reversibility of the Cs sorption process. A very
elegant study by Hird et al.70 shows that the KCl-extractable Cs increases with
increasing Cs saturation of the FESs, but abruptly drops at a sorbed Cs level of 10
mmol kg1 in podzolic soils. That drop is attributed to the layer collapse induced by
the increasing Cs loading on the FESs (Figure 4.2).
The electrolyte background of the equilibrium solution is of major importance
in Cs sorption, because of the presence of both competing ions (K+, NH4+) and other
ions (Ca2+). Poorly hydrated cations such as K+ and NH4+ influence the Cs sorption
in two respects: i) they compete with Cs+ ions for sorption on the same exchange
sites66,67,94, and ii) they can induce layer collapse, and hence Cs trapping (Figure
4.2).88,95 Potassium and ammonium salts have long been used to extract exchangeable
Cs+ ions in soils and clays.93,96,97 Increasing concentrations of K+ and NH4+ ions in
the equilibrium solution decreases the Cs sorption yield in micaceous phyllosilicates
and soils.83,92,98-100 As K+ and NH4+ are collapse inducing cations, they can provoke
Cs trapping in micaceous phyllosilicates. The entrapment of Cs is caused by simul-
taneous introduction of Cs+, K+, and NH4+ ions in the surrounding solution,88 whereas
the edge interlayer trapping of radiocesium occurs when K+ and NH4+ ions follow
the Cs+ contamination.95 The K+ and NH4+ ions can therefore promote either the
release of trace Cs or its fixation through the trapping of the trace element. The
predominance of the ion competition effect or the Cs trapping effect over each other
will chiefly depend on the concentration of competing ions.88,95,101,102 Such predom-
inance depends also on the location and extension of the wedge zones95 as well as
on the particle size.88 With respect to the concentration, Absalom et al.102 demon-
strated that trace Cs is reversibly sorbed at PAR < 101.5 mM0.5, whereas it is
74 Trace Elements in the Rhizosphere
entrapped at PAR > 101.5 mM0.5. In other words, with increasing K concentration,
K+ ions first exchange with Cs+ ions to displace them from exchange sites and further
cause Cs trapping through layer collapse. Regarding the extension of the wedge
zones, it can be expected that K+ and NH4+ ions will act as Cs+-competitive ions if
that extension is limited (mica, weakly weathered or not) and as trapping ions if
that extension is large (strongly weathered mica, mica-vermiculite).
Cs sorption in micaceous phyllosilicates pre-equilibrated with Ca2+ ion is larger
and more rapid than that measured in the corresponding K+-saturated clay miner-
als.98,103 In addition, the Ca2+ ion seems to promote the slow migration of Cs in
deeper wedge zones of the mineral.103
Increasing the ionic strength of the equilibrium solution generally decreases
KdCs, as recently reviewed by Cornell,62 likely because of the contraction of the
diffuse double layer. The increase in the ionic strength leads to increasing values of
the Gaines-Thomas selectivity coefficient Kc(Cs+/Mn+) on illite (Mn+ = K+, Na+:
Brouwer et al. 66) and montmorillonite (Mn+ = K+, Na+, Ca2+: Staunton and Roubaud
104); such increase has been attributed to, respectively, the hydration properties of
exchanged cations and, partly, the increase of the size of the clay tactoids.
As KdCs depends on Cs+ concentration below a certain level (108 M), KdCs
decreases with decreasing solid to solution ratio in a nonlinear adsorption sce-
nario.62,105 The contact time between solid and solution seems to play a major role
on trace Cs sorption yield as well as on the reversibility of the sorption process.
Comans et al.103 observed a rapid Cs sorption on illite followed by a slow sorption
phase, the extent of which can last several weeks. Increasing sorption time leads to
an apparent decrease in the reversibility of the Cs sorption process, attributed by
these authors to slow migration of Cs into deeper wedge zones and likely to sorption
onto FESs with very high Cs specificity. Using a multidesorption technique shows
that the effect of sorption time is particularly important in mica-derived smectite,
whereas it is very small on illite and biotite (unpublished data). Wauters et al.106 and
Absalom et al.102 have also observed the time-dependent decrease of release of
radiocesium in the solution. As suggested by Comans et al.,103 strong Cs retention
is thought to occur as radiocesium slowly migrates in the collapsed interlayers of
micaceous clay minerals.95,102 This aging effect seems to reduce the phytoavailability
of radiocesium in agricultural soils.25, 107, 108
quantified the number of FESs in soils by using the silver thiourea (AgTU) complex
to mask the RESs. They measured numbers of FESs contributing 0.01 to 2% of the
overall CEC and quantified the trace Cs+ to K+ selectivity of the FESs (mean
KcFES(137Cs+/K+) 103).65 The same research group57,94,111,112 further developed this
quantitative analysis and demonstrated the following equation:
where RIP, KcFES(137Cs/K), [FES], mK, and KDCs refer, respectively, to the radioce-
sium interception potential, the trace 137Cs to K selectivity coefficient, the number
of FESs, the fixed potassium molar concentration in the equilibrium solution,112 and
the 137Cs solid-liquid distribution coefficient measured in an electrolyte background
confining both Cs+ and K+ ions onto the FESs using AgTU94 or a fixed K+-Ca2+
scenario.112 In a very large majority of soils, the mean value of KcFES(137Cs/K)
amounts to 103.57,61,65,69,94,112 However, some of the FESs can exhibit exceedingly
high Cs to K selectivity (KcFES(137Cs/K) ~ 2 103).65 They are likely the vermiculitic
sites associated with micaceous wedge zones, as suggested by Maes et al.82,113 In
comparison with such high Kc values, the trace Cs to K selectivity in the RES pool
is quite small. Indeed, KcRES(137Cs/K) ranges between 1.0 for humic compounds114
to 5.0 in clay minerals.115 Noteworthy is the fact that lnKcRES(137Cs/K) is about 6.92
0.56 in a wide variety of soils and sediments.112 Such a trend shows that the same
type of Cs specific site is operating in various soils and sediments. Consequently,
the RIP, which combines Cs specific site selectivity and capacity, is an extremely
important property of the system.57,94 That property integrates the quantity-intensity
(Q-I) relationship.116 As such, RIP could be a useful tool to predict the potential
phytoavailability of radiocesium in soils.57
RIP values are presented in Tables 4.1 and 4.2 to illustrate their range in soils
and sediments, respectively.
TABLE 4.1
Ranges in Organic Matter, Clay Contents, and RIP
Values Measured in Various Soil Collections
Number of O.M.a Clay RIPKb RIPNc
Source Samples % % mol g1 mol g1
TABLE 4.2
Ranges of RIP Values Measured in Various Types of
Sediments and Montana Illite; Mean Values of the NH4+
to K+ Selectivity Coefficient of the Frayed Edge Sites
Sediment Type Number of RIPKa RIPNb KCFES (NH4+-K+)
or Mineral Samples mol g1 mol g1 (Mean Value)
The reported values of RIPK [RIPK = KDCs mK: see Equation (1)] cover four
orders of magnitude (10 104 mol g1). Such a range illustrates fairly well the huge
variability of Cs-binding properties in soils and sediments. The range in RIPK values
are quite the same in soils and sediments, providing strong indication that basic
mechanisms in trace Cs retention are similar in both soil and sediment environments.
Table 4.1 also shows that soils rich in organic matter can exhibit substantial RIP, and
hence genuine FESs,69,70 borne by a very small amount of clay particles present in
these soils, as Cs retention by organic matter involves only electrostatic binding.114
As inferred from both Tables 4.1 and 4.2, the values of RIPN (KDCs mNH4+) are
invariably lower than those of RIPK. Such a difference confirms the preference of NH4+
over K+ ions for the sorption on the FESs. The mean NH4+ to K+ selectivity coefficient
KcFES(NH4+/K+), deduced from the ratio [RIPN:RIPK], is about 5 in sediments (Table
4.2). In fact, it generally ranges between 4 and 6 in various soils and sediments.57,94,112
Such a narrow range is an additional fingerprint of the specific action of the FESs and
the common mechanisms of trace Cs retention in soils and sediments.94,112
FIGURE 4.5 Log-log plot of the estimated FES capacity against the number of vermiculitic
sites. Adapted from Maes et al.113 and Delvaux et al.121 (see Table 4.1). Open symbols refer
to the weathering sequence acid brown soil podzol.
retained on vermiculitic sites neighboring micaceous wedge zones. In acid soils, Al-
interlayering of vermiculite leads to the formation of HIV minerals (hydroxy inter-
layered vermiculite), which weakly retain radiocesium. Organic matter can therefore
influence the retention of trace Cs through Al-complexation. Indeed, this process
induces Al-depletion in the soil solution and hence impedes Al-interlayering and
maintains the interlayer sites accessible for Cs retention. In this respect, the role of
soil organic matter on trace Cs+ retention is obviously indirect. In a weathering
sequence acid brown earth podzol (see Figure 4.5), Maes et al.113 further showed
that Cs retention was directly related to the soil vermiculite content, which reflects
to some extent the weathering of mica in soils because vermiculite content was
strongly horizon and depth dependent. These findings largely contribute to under-
standing the marked differences of 137Cs retention properties between the various
horizons in forest soils. Such differences were long not understood in a mechanistic
way.3 The retention of radiocesium can indeed greatly vary within very short vertical
distances in forest acid soils, particularly in those exhibiting thick humus layers.48
The RIP as measured according to Wauters et al.112 in the soil weathering sequence
78 Trace Elements in the Rhizosphere
studied by Maes et al.113 as well as in other soils121 (Table 4.1) leads to computed
[FES] capacity using KcFES(137Cs/K) 103 in equation (1), which correlates fairly well
with the number of vermiculitic sites (r = 0.75, n = 43) (Figure 4.5).
Figure 4.5 shows that the samples representing the weathering sequence brown
soil podzol113 are fairly well distributed over a large range of both estimated FES
capacity and vermiculite content, as do the samples representing a larger collection
of soils with widely varying properties121 (Table 4.1). This observation is consistent
with the conclusion that vermiculitic sites associated with micaceous wedge zones
control, to a very large extent, the retention of radiocesium in soils. In particular,
soil horizons in which the weathering of mica is active would strongly retain
radiocesium in soils.
The available data from clay, soil, and sediment studies indeed converge to show
that weathered mica and/or mica-vermiculite intergrade appear to be the mineralog-
ical structures that best fix radiocesium. The immediate structural environment of
fixed Cs in the frayed edge site exhibiting exceedingly high trace Cs to K selectivity
is, however, not yet fully characterized.
B. SOIL-TO-PLANT TRANSFER
Many reports demonstrate that the 137Cs soil-to-plant transfer largely varies between
soils (see above). Though extensive knowledge has been gathered on the separate
soil and plant factors likely to control 137Cs availability in soil, few studies have
analyzed the soil-to-plant transfer of radiocesium in a mechanistic
way.57,117,124,125,128,129 In meadow upland areas, the RES pool [KcRES(137Cs+/K+)1] con-
trols the concentration of radiocesium in the soil solution in organic-rich soils,
whereas the FES [mean KcFES(137Cs+/K+)103] pool does it in mineral soils.57 Conse-
quently, the 137Cs TF values are much lower in the latter soils than in the former ones.57
In a pot experiment study involving 30 agricultural soils with a wide range of
properties, Smolders et al.117 have shown that the 137Cs soil-to-plant transfer was very
Fate of Radiocesium in Soil and Rhizosphere 79
well predicted by the 137Cs and K concentrations in the soil solution. In this respect,
the bioavailability of radiocesium varies extensively between soils because of differ-
ences i) in 137Cs retention, affecting 137Cs concentration in solution, and hence its
supply to plant roots, and ii) in K availability, affecting the 137Cs root uptake process.
Based on these concepts, a soilplant 137Cs transfer model was developed by Absalom
et al.119 The soil solution concentrations of 137Cs and K were estimated from two soil
characteristics, the percent clay and the exchangeable K content, and from the total
137Cs content in soil and the time after the contamination. The model successfully
TABLE 4.3
Predicted and Observed 137Cs Activity in Wheat Shoots Grown in a
Contaminated Soil at Different K Supply
Bulk Soil Solution Predicted Soil-Root Interface Predicted Observed
K Application K 137
Cs Shoot 137Cs K 137
Cs Shoot 137Cs Shoot 137Cs
mmol pot1 M cpm mL1 cpm g1 (M) cpm mL1 cpm g1 cpm g1
Note: The predictions are based on soil solution composition of the bulk soil or rhizosphere soil and on
137Cs uptake characteristics measured in solution culture. The soil solution composition in the rhizosphere
pH and exudate concentrations are unlikely to affect 137Cs mobility in the rhizosphere
as both factors are known not to alter the solid to liquid distribution in a significant
way. Gradients in the concentrations of cations, mainly K, or 137Cs itself, are likely
to play a significant role on plant availability of 137Cs. This is illustrated in Table
4.3. The 137Cs uptake by young wheat plants was predicted from soil solution
concentrations of 137Cs and K, either those in bulk soil solution data or those in the
rhizosphere. The predictions based on rhizosphere composition were significantly
better than those based on the bulk soil solution composition. The rhizosphere
concentrations were calculated by a solute transfer model and the uptake was pre-
dicted from soil solution composition (137Cs and K concentrations) and data on 137Cs
uptake in wheat in solution culture.124,125
The data in Table 4.3 show that K is more depleted away from the bulk soil than
137Cs. This is remarkable since K is more mobile than 137Cs in soil. This can be
explained by the fact that 137Cs is absorbed less efficiently than K by plant roots.
Indeed, these two cations have been demonstrated to be 4 to 25 times more efficient
in uptake in solution culture studies125 (see also Section IV, A). This uptake efficiency
is expressed relative to solution concentrations and is therefore related to the rate
by which these cations are removed from the soil solution. The root uptake efficiency
becomes high at low K supply, resulting in the well-known K depletion in the
rhizosphere at low K supply of the soil. In contrast, reducing 137Cs concentrations
in the rhizosphere of actively absorbing roots does not induce a stronger sink for
137Cs itself (Cs is not a plant nutrient and its uptake rate is not controlled by nutritional
sandy soils. Small 137Cs RM is associated with high RIP soils, i.e., brown soils,
whatever their clay and organic matter content. Noteworthy are the facts that i) the
RIP values cover four orders of magnitude (see also Table 4.1: unpublished data)
FIGURE 4.6 Plot of the 137Cs+ rhizospheric mobilization against RIP (a) and NaTPB-extract-
able 137Cs+ (b) in a collection of soils of widely varying properties. (Adapted from Delvaux,
B. et al.121; see Table 4.1.)
82 Trace Elements in the Rhizosphere
and ii) the 137Cs RM exhibits a very large range as it varies between 1 and 69% of
the initial 137Cs+ soil contamination. The relationship illustrated in Figure 4.6 suggests
that RIP could serve as a useful tool to predict potential 137Cs+ soil-to-plant transfer
patterns, likely because the RIP concept integrates the quantityintensity relationship
[Equation (1)]. These results further support that the FES pool controls the concen-
tration of radiocesium in the soil solution, and hence its mobility in the vicinity of
active plant roots. However, the absorption of radiocesium by the roots is also likely
driven by K depletion in this experimental device because of K stress (Ca2+ exchanged
soils, K-deficient plant). The selective extraction of 137Cs by NaTPB simulates the
Cs root-induced sink effect due to K depletion135 enhanced by rapid precipitation of
KTPB and CsTPB.136 Indeed, the correlation between 137Cs RM and NaTPB-extract-
able 137Cs (Figure 4.6b: r = 0.94, n = 47) strongly supports that K depletion in the
rhizosphere is a major driving force in the 137Cs uptake by plant roots.
Many soils in seminatural environments are expected to exhibit low RIP because
of their peaty, acidic, sandy, and/or podzolized character. The rhizospheric mobili-
zation of radiocesium can therefore be large in such environments, particularly if K
supply is low. In this respect, intense mobilization of radiocesium in the rhizosphere
can be a key process in the relatively large biorecycling of radiocesium in such areas
(Figure 4.1). This statement is supported by the results reported by Sanchez et al.130
on organic-rich soils from such areas (see Section IV, B). Small RIP, low K avail-
ability, and high NH4+ concentration in the soil solution all contribute to increased
Cs soil-to-plant transfer in these soils.
The relationships described here between soil matrix, soil solution, and plant
roots consider only the chemical interactions involving radiocesium in the rhizo-
sphere. Such chemical interactions should not occult the biological interactions
occurring as well, particularly in the uppermost soil horizons.137 A large part of
radiocesium can be immobilized by the microflora, particularly in fungal mycelia.138
Some major groups of microorganisms living in the rhizosphere are root symbionts,
such as N-fixing bacteria and mycorrhizal fungi. These fungi have a considerable
impact on the accumulation of radiocesium in seminatural environments; they
occupy a central position in the biogeochemical cycle of radiocesium in such envi-
ronments.139,140 However, very few studies have analyzed the role of fungi in the
transfer of radiocesium from soil to plant in a mechanistic way. Some studies have
emerged in the last 5 years, but they exhibit somewhat contrasting conclusions,
partly due to the fact that published results refer to mycorrhizal symbioses involving
distinct structures and species. Entry et al.141,142 observed higher contamination with
radiocesium in plants previously infected with arbuscular mycorrhizal fungi (Glomus
spp.). They suggested that relatively high 137Cs transfer to mycorrhized plants could
be very attractive in phytoremediation. Other studies reported that the infection of
roots of Picea abies by ectomycorrhizal fungi (Hebeloma crustuliniforme) decreases
the plant contamination with radiocesium.143,144
VI. CONCLUSION
The data presented in this chapter show that both clay minerals and plant roots act
as mutual competitive sinks for radiocesium in the rhizosphere. The micaceous 2:1
Fate of Radiocesium in Soil and Rhizosphere 83
clay minerals, which bear selective Cs+ sorbing frayed edge sites, control the con-
centration of radiocesium in the soil solution, and hence its uptake by plant roots.
The strongest trace Cs retention occurs on vermiculitic sites associated with mica-
ceous wedge zones; these sites can be readily quantified. As such, weathered mica
and/or mica-vermiculite intergrade appear to be the mineralogical structures that
best fix radiocesium. The intimate structural environment of the frayed edge site
exhibiting exceedingly high trace Cs to K selectivity is, however, not yet fully
characterized and merits further investigation.
The concentration of K+ ions in the soil solution plays a key role in the root
uptake of radiocesium as K depletion (K concentration < 1mM) increases the trace
Cs uptake rate, and thereby is a principal driving force in the uptake of radiocesium
by plant roots. Because K depletion promotes the weathering of micaceous phyllo-
silicates, one can expect that K depletion may promote, to some extent, trace Cs
retention on newly accessible frayed edge sites with a very high Cs+ specificity.
Such possible interaction needs further investigation.
The soilroot interaction has been presented here in terms of chemical processes
occurring in the rhizosphere. The role of fungi, particularly the mycorrhizal fungal
species in this interaction is largely unknown, from a mechanistic point of view.
Basic research in this topic is required to achieve a fuller understanding of the
bioavailability of radiocesium in soils from natural and seminatural environments.
It is also a prerequisite to lay out the scientific foundations about the use of mycor-
rhizal fungi in the phytoremediation or phytostabilization of contaminated areas.
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90 Trace Elements in the Rhizosphere
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Fate of Radiocesium in Soil and Rhizosphere 91
CONTENTS
I. Introduction.....................................................................................................94
II. Plants Growing at Heavy-Metal-Rich Sites ...................................................95
III. Strategies of Metal Tolerance.........................................................................96
IV. Phytoremediation ............................................................................................97
V. Uranium in Soils and Plants...........................................................................98
A. Uranium and the Environment ...........................................................98
B. Uranium Behavior in Soils.................................................................99
C. Uranium Behavior in Plants .............................................................101
D. Influence of Rhizosphere Processes on Uranium Absorption
by Plants ...........................................................................................102
VI. Plutonium in Plants and Soils ......................................................................103
A. Plutonium and the Environment.......................................................103
B. Plutonium Behavior in Soils ............................................................104
C. Plutonium Behavior in Plants...........................................................106
VII. U Phytoextraction .........................................................................................108
VIII. U Rhizofiltration ...........................................................................................111
IX. Enhancement of U and Pu Uptake by Plants ..............................................112
A. Effect of Synthetic Chelates and Organic Acids on U Uptake .......112
B. Effects of Synthetic Chelates and Organic Acids on Pu
Uptake ...............................................................................................115
X. Conclusions...................................................................................................116
References..............................................................................................................116
0-8493-1535-2/01/$0.00+$.50
2001 by CRC Press LLC 93
94 Trace Elements in the Rhizosphere
I. INTRODUCTION
Large areas of land have been contaminated by radionuclides and fission by-products
from nuclear weapons facilities, aboveground nuclear testing, nuclear reactor oper-
ations, and nuclear accidents.1 The principal contaminants of concern are the radi-
onuclides U, Th, Pu, 60Co, 90Sr, 135Cs, and 99Tc, and toxic metals Ba, Cd, Cr, Ni,
Pb, and Zn. Radionuclides, unlike organic pollutants, cannot be degraded. Radionu-
clides can move to the food chain, and exposure through inhalation and ingestion
of food can result in detrimental health effects, including cancer and genetic muta-
tions. The radionuclides U and Pu and their decay products result in two problems,
specifically radiation dose hazards and chemical toxicity. The radiation hazard intro-
duces the need for special precautions in reclamation beyond that associated with
nonradioactive metals.
Radioactive contamination of the environment that surrounds facilities where U
and Pu have been mined and processed for nuclear weapons and nuclear power has
occurred in many countries. Uranium is reported to be the most frequent radionuclide
contaminant in groundwater and surface soils of United States Department of Energy
(USDOE) facilities.2 One of these facilities, which is now a superfund site, is the
former Feed Materials Production Center at Fernland, OH. The soils around the
Fernland processing plant have become contaminated with U after decades of U
extraction and purification. Efforts are now underway to remediate the site. Various
technologies for removal of U have been evaluated at this site.3-6 Another well-
researched site is Rocky Flats, located north of Golden, CO, where Pu contamination
originated from leakage of drums used to store Pu.7 Plutonium particles entrapped
in the fine fraction of the surface soils have been subsequently transported by winds
and water and deposited across the soils of the site.
The remediation of these and similar areas contaminated with radionuclides
represents a worldwide problem that will require considerable economic resources
of industry and government. For the most part, contaminated soils are remediated
using engineering approaches including thermal, chemical, and physical techniques
(e.g., in situ mobilization, immobilization, degradation, and burial; or removal and
reburial, vitrification, vacuum extraction, steam flooding, pumping and leaching,
electroosmosis, and electroacoustic extraction). Various soil-washing techniques
have been developed for decontamination of radionuclides using organic and inor-
ganic acids, bases, salts, and chelating agents. The merits of these techniques have
been discussed by Francis and Dodge.5 Many of the inorganic chemicals used are
corrosive and alter or damage the soil. Synthetic organic chelating agents used in
decontamination sometimes persist in the environment, causing the radionuclides to
migrate from the disposal site.
In addition to these physical and chemical treatment methods, microbial reme-
diation (bioremediation) has become more common in the last two decades. However,
the success of microbial bioremediation has been largely limited to the degradation
of organic compounds, with less emphasis on metal and radionuclide contaminations.
Bioavailability of Uranium and Plutonium to Plants in SoilWater Systems 95
indicator plants include species of the genera Astragalus. Astragalus species cover
large areas of the western United States and were used by Cannon15 to detect U
mineralization because of the geochemical association of Se and U in the carnotite
ores of the region. Cannon mapped the distribution of Astragalus plants and was
thereby able to pinpoint several economic-grade U deposits. This discovery of U in
the western United States is an example of the successful application of the geobo-
tanical method for metal prospecting.13 Similarly, Dunn16 reported that the twigs of
black spruce (Picea mariana) contained U concentrations >100 mg kg1 ash weight,
in comparison with background levels of <1 to 60 mg U kg1. This relationship was
used to locate a massive U biogeochemical anomaly near Wollaston Lake in Canada.
Plant species that are naturally high in heavy metals have developed a defense
mechanism to tolerate absorption of the metal and, as a result, accumulate a high
concentration of metal in the plant tissue. Since heavy metals are damaging to most
plants at relatively low concentrations, the accumulation strategy requires mecha-
nisms to detoxify the metal.
such as Cu, Zn, and Cd. Phytochelatins are a family of peptides with the general
structure [-GluCys]n-Gly, where n > 1.21 Metallothioneins are similar to PCs in
being Cys-rich, metal-complexing ligands; however, metallothioneins are proteins
synthesized by mRNA translation.
IV. PHYTOREMEDIATION
The term hyperaccumulator was introduced by Brooks et al.18 for plants growing
on serpentine sites that are capable of concentrating nickel (Ni) to more than 1000
g g1 in their leaves on a dry matter basis. A concentration of 1000 g g1 has also
been used to delineate the exceptional uptake of copper (Cu), cobalt (Co), and lead
(Pb). The delineation level is raised to 10,000 g g1 for zinc (Zn) and manganese
(Mn), because of the greater background concentrations of these metals in soil.
The concept of using hyperaccumulator plants to extract and remediate metal-
contaminated sites started in the 1970s,22 and the term phytoremediation emerged.
This term consists of the Greek prefix phyto (plant) and Latin root remedium
(to correct or remove an evil) and is defined as the use of green plants and associated
microbiota, soil amendments, and agronomic techniques to remove or render envi-
ronmental contaminants harmless.23 Chaney et al.10 defined phytoremediation as
using plants to make soil contaminants nontoxic. Green remediation, bioremediation,
and botanical-bioremediation are other names used for the process of using plants
to remediate the environment.24
Phytoremediation requires that the target metal must be sorbed by the roots and
either (1) translocated from the root to the shoot for subsequent removal of the
biomass, or (2) rendered harmless by biochemical or chemical processes associated
with the plant. In the case of U and Pu, the metals cannot be rendered harmless,
hence phytoremediation must involve absorption, translocation, and subsequent
removal. The metal is removed from the site by harvesting the plant material. After
harvesting, the biomass is processed to either recover the metal or further concentrate
the metal to facilitate disposal.
Phytoremediation has been categorized into five techniques: (1) phytoextraction
(use of plants to remove contaminants, e.g., Zn and Cd from soils);25,26 (2) phyto-
volatilization (use of plants to volatilize soil elements like Hg and Se);27-29 (3)
rhizofiltration (use of plant roots to remove contaminants like U and Pb from flowing
water);30-32 (4) phytodegradation of organic compounds by rhizosphere biodegrada-
tion;33,34 and (5) phytostabilization (use of plants to transform soil metals to less
toxic forms, e.g., to reduce Cr(VI) to Cr(III) but not to remove the metal from the
soil).10,35
Some of the successful pilot projects in phytoremediation have been: (1) Se and
boron (B) removal from saline soils in California using Brassica juncea and Festuca
arundinacea,36 (2) Zn removal from metal-contaminated soils using hyperaccumu-
lator plants,37,10 (3) lead (Pb) cleanup from a contaminated industrial site in New
Jersey using Brassica juncea,38 and (4) removal of 137Cs and 90Sr from contaminated
soils at Chernobyl.39,40
Fundamental to the environmental and economic success of phytoremediation
of heavy metals and radionuclides is the existence of high biomass plant genotypes
98 Trace Elements in the Rhizosphere
FIGURE 5.1 Species distribution for U(VI) hydrolysis and carbonate species at 2.38 mg
U(VI) L1 and 31.1 total inorganic C L1. (From Duff, M. C. and Amrhein, C.57 With
permission.)
Uranium (VI) is the most mobile form of U.55 It exists in solution predominantly
as the stable linear ion UO22+ and as soluble carbonate complexes, (UO2)2CO3(OH)3,
UO2CO3, UO2(CO3)22, UO2(CO3)34, and possibly (UO2)3(CO3)66.56-59 Within the
pH range of 4.0 to 7.5 and in the absence of dissolved inorganic ligands (carbonate,
fluoride, sulfate, and phosphate), the hydrolysis species UO2OH+, UO2(OH)2o, and
(UO2)2(OH)22+ dominate U(VI) speciation.60 The equilibrium distribution of U(VI)
hydrolysis and carbonate species is shown in Figure 5.1. Uranium in soil and water
forms complexes with sulfate and phosphate as well as with carbonate and hydroxide,
and these complexes increase the total solubility of U.
Bunzl et al.61 found that a significant portion of U contaminant in soils exposed
to vented air from U ore mine shafts was associated with soil clay minerals. The
least amount of U was found in the exchangeable and soluble forms; therefore, the
amount of U available for plant uptake and subsequent phytoremediation was limited.
In another study, it was observed that in soils spiked with U(VI) as UO2(NO3)2
6H2O and analyzed shortly afterwards, the dominant proportion of U(VI) was found
in association with the Fe-Mn oxide portion of the soil.62,63 Sorption of U(VI) onto
soil surfaces tends to increase with increasing pH (up to pH 7) and is readily
reversible by decreasing the pH.64
Bioavailability of Uranium and Plutonium to Plants in SoilWater Systems 101
239 and -238 (239Np and 238Np) are produced by reaction on 238U and decay by
emission to plutonium. The neutron arises from fission of 235U:79,80
( n, ) 239 239
238
U 239 U Np Pu (4)
on the basis of equal weight or equal CEC. Plutonium extractability from montmo-
rillonite was markedly higher than that from kaolinite and illite between pH 4.0 and
10.5 on both an equal weight and CEC basis. However, on an equal weight basis,
the Pu extractability from illite was slightly greater than that from kaolinite below
pH 2.8 and between pH 7.3 and 11.6, but it was lower than that of kaolinite between
pH 2.8 and 7.3. On an equal CEC basis, the Pu extractability curves for kaolinite
and illite were reversed. Rhodes100,101 investigated Pu adsorption on a calcareous soil
containing mainly montmorillonite and kaolinite and little or no organic matter.
Strong adsorption of Pu (>98%) began at pH 2 and extended to pH 8.5. In the region
between pH 8.5 and 12.5, adsorption was lowered to a maximum of about 80%.
The ability of humic compounds (humic and fulvic acids), present in soils and
natural waters, to enhance the solubilities of metal ions is well known.102 Cleveland
and Rees103 investigated the solubilization of Pu and Am by humic compounds in
contaminated soil. They reported that the solubilization of Pu and Am by fulvic acid
was slight, though some of the radionuclide was associated with the aqueous phase
as a result of surface complexation with suspended colloids. They also found that
the resulting suspension was unstable, and over a period of a few days most of the
radionuclide was flocculated either as a result of colloid coagulation or hydrolysis.
Chelation by naturally occurring soil organic constituents was proposed as the
most likely mechanism of mobility of Pu and subsequent uptake by plants.104,105
Assumptions were made that Pu associated with low molecular-weight molecules
was more mobile and thus readily available for plant uptake.106 Plutonium associated
with higher molecular-weight molecules was assumed to be nonmobile and not
plant available.
Nishita and Haug107 studied the effects of organic (humic and fulvic acids) and
inorganic (free iron oxides, free silica, and free alumina) fractions of soil on the
sorption and extractability of 239Pu(IV) as a function of pH. They found that in
mixtures of pure humic and fulvic acids with soil inorganic fractions, the extract-
ability of Pu was substantially influenced by the organic-inorganic interaction. The
Pu(IV) was strongly sorbed on the inorganic fraction of the soil under soil pH
conditions normally encountered in natural environments. Also, they reported that
the influence of the inorganic fractions was most evident in the lower pH range,
whereas that of the organic materials was most evident in the higher pH range, i.e.,
above pH 6. The mobility of Pu in soil under the leaching action of water depended
on the type of organic matter present. If both soluble and insoluble forms of organic
matter are present in the soil under either alkaline or acidic conditions, Pu is likely
to show relatively high mobility, due to soluble complexes or chelated forms of Pu,
respectively. Nishita and Hamilton108 investigated the influence of CaCO3, organic
matter, and the inorganic fraction of a calcareous soil on the chemical extractability
of 239Pu(IV). In the system containing humic acid with CaCO3, the extractability of
Pu ranged between 0.1 and 1.3%. Free CaCO3 in all treatments drastically reduced
the Pu extractability.
Nishita et al.109 investigated the effect of 5 organic (Na-formate, -acetate, -tar-
trate, -citrate, and -oxalate) and 21 inorganic (Na2SO4, NaNO3, Na2Cr2O7, NaI,
Na2HPO4, Na4P2O7, NaOH, NH4OH, etc.) compounds on the extractability of 239Pu
from a contaminated soil. They reported that among the organic compounds, the
106 Trace Elements in the Rhizosphere
order of 239Pu extractability was Na-acetate < Na-formate < Na-oxalate < Na-tartrate
< Na-citrate. Also, among the 21 inorganic compounds, soil treated with Na2HPO4
had the least 239Pu extractability (0.07% of the dose), and soil treated with Na4P2O7
had the greatest extractability (28.6% of the dose). Other compounds that had
relatively high 239Pu extractability included NaF, Na2B4O7, NaCO3, Na2O2, NaOH,
and NH4OH.
Jacobson and Overstreet,110 who worked with an aqueous suspension of elec-
trodialized Wyoming bentonite adjusted to pH 5.5 with Ca(OH)2, reported that the
order of adsorption was Pu(III) > Pu(IV) > Pu(VI). Murray and Fukai111 also showed
that the adsorption of Pu was Pu(III) > Pu(IV) > Pu(VI) at pH 3 in sediments of
the Var River, but at pH 8, the adsorption was Pu(VI) > Pu(III) > Pu(IV). Prout112
found that Pu adsorption by a Savannah River facility soil containing mainly kaolin-
ite was >90% from solutions of Pu(III) and Pu(IV) between pH 1.2 and 2.5.
However, from solutions of Pu(VI), more than 90% adsorption was not attained
unless the pH values were >pH 6. Bondietti et al.104 found that the adsorption of
239Pu by the clay fraction of a Miami silt loam was >99.4% regardless of initial
Vyas and Mistry134 studied plant uptake of 239Pu and Americium-241 (241Am)
from different clay mineral (montmorillonite and kaolinite)-organic matter-sand
mixtures simulating contrasting soil types. They reported a marked reduction in
uptake of both 239Pu and 241Am with an increase in organic matter as well as clay
content of the mixtures. Reduction in the uptake of 241Am with increasing clay
content was more pronounced in the montmorillonite-organic matter mixtures com-
pared to the kaolinite-organic matter mixtures. They also observed that increasing
organic-matter content and the interaction of organic matter with montmorillonite
and kaolinite reduced the uptake of 239Pu and 241Am by plants.
In general, the information available on this topic has been from a dosimetry
standpoint, with little emphasis on developing an understanding of the mechanisms
involved in the uptake of Pu by plants. Agricultural plants were investigated for their
ability to accumulate Pu and the potential migration of Pu into humans due to
ingestion of agricultural products. More Pu was found in the seeds of soybean
(Glycine max)135 than would normally be expected. This was likely due to harvesting
practices in which the suspension of airborne particles and subsequent deposition
onto the seed contributed to the quantity of Pu present.
As with U, it is likely that rhizosphere processes could strongly influence
solubility of Pu. The predominant factors that might be expected to influence Pu
absorption are plant and microbially induced pH changes that occur in the rhizo-
sphere and the release of organic metal-complexing agents by plant roots and
microbes. It is very unlikely that specific channels or transporters exist in the root
plasma membrane for the absorption of U or Pu. Absorption likely occurs by mass
flow through nonspecific channels or plasma membrane defects. Therefore, rhizo-
sphere processes that influence root plasma membrane structure and selectivity (e.g.,
chemical or enzymatic damage to the membrane) might also contribute to the
absorption of U or Pu.
VII. U PHYTOEXTRACTION
Recently, a few U phytoextraction investigations have been reported.4,75,76,31 Some
reports on the high bioavailability of U to agriculture plants are also available.75
Results of U accumulation by some of the plant species used in the experiment of
Hossner et al.77 are shown in Figure 5.3. Uranium uptake and accumulation by plant
species grown in a soil contaminated with 100 mg U(VI) kg1 differed significantly
(Figure 5.2). Uranium concentrations in plant shoots ranged from 3.5 to 23 mg U
kg1. Indian mustard and sunflower had the highest U concentrations in the shoot,
while wheat had the lowest U concentration (3.5 mg U kg1). Sheppard et al.70 found
that U concentration of alfalfa and chard ranged from 0.0 to 27.6 mg kg1 when 50
and 100 mg U kg1 were applied to the soil as uranyl nitrate. Saric et al.67 also found
a large variation in U uptake among plants and concluded that sunflower and leafy
vegetables accumulated more U than other plants. Shoot concentrations of U in plant
species from the Brassica family were higher than from grasses77 (Figure 5.2).
Mustard green, turnip, and Chinese cabbage from the Brassica family and Swiss
chard (Beta vulgaris), a vegetable root crop, were better U accumulators than grain
crops. Sheppard et al.70 reported that Swiss chard U concentration ranged from 0.0
Bioavailability of Uranium and Plutonium to Plants in SoilWater Systems
FIGURE 5.2 Uranium concentration in shoots and roots in plant species grown in a Weswood soil (pH 7.55, CaCO3 equivalent 2.45%; fine-silty, mixed,
thermic Fluventic Ustochrept) contaminated with 100 mg U kg1 as uranyl nitrate.
109
110 Trace Elements in the Rhizosphere
FIGURE 5.3 Uranium concentration in shoots and roots of sunflower (H. annuus) grown on
soils contaminated with 300 mg U(VI) kg. Chelates were applied as solutions to soils after
plant emergence. Control denotes U concentration plants grown in soils in the absence of
chelates.
to 27.6 mg kg1 with soil contamination with uranyl nitrate at levels of 50 to 100
mg U kg1.
Uranium accumulates mainly in the roots. Uranium concentrations in roots of
different plant species were markedly different and varied from 69 to 789 mg U kg1
of root dry weight (Figure 5.2). Indian mustard and sunflower had the highest root
U concentration, and wheat and rye the lowest concentration. Sunflower and Indian
mustard are reported accumulators of U.30,38
Bioavailability of Uranium and Plutonium to Plants in SoilWater Systems 111
VIII. U RHIZOFILTRATION
Radionuclide contamination in water is usually removed by ion exchange, reverse
osmosis, microfiltration, precipitation, or flocculation. These methods are of limited
use for large water volumes, low metal concentration, and saline water, and are
expensive. Recently, Gu et al.138 used Fe fillings as reductants in removing U from
contaminated water. Results indicated that Fe fillings were much more effective
than adsorbents such as peat, iron oxide, or organic cation exchangers in removing
uranyl (UO22+) from aqueous solution. Nearly 100% of U was removed by reaction
with Fe. This is an inexpensive and effective method, though reduced U(IV) pre-
cipitated on the Fe surface could be reoxidized to U(VI) and remobilized.
Rhizofiltration is an alternative technology of remediation in aqueous systems
that might provide a cost-effective method to treat contaminants at very low con-
centrations.31,32,139 Rhizofiltration is a technique in which plants are grown in polluted
water for subsequent sorption of the target element(s) by the live roots. Sunflower
has been shown to have some potential for rhizofiltration.30 The high root biomass
has been effective in removing heavy metals from polluted water. Sunflower grown
hydroponically was able to drastically reduce the concentration of Cr and U(VI) to
low levels within 24 hours.32 In tests carried out in Ashtabula, OH, sunflower roots
were submerged in water contaminated with 100 to 400 g U L1.140 The U content
was decreased to below the EPA standard of 20 g U L1. Sunflower was shown to
be very effective in recovering U from contaminated water.1 Uranium was accumu-
lated mainly in the roots with a final concentration 5,000 to 10,000 times greater
than the concentration in water. Both sunflower and Indian mustard tend to concen-
trate metals in the roots and translocate only a small portion of the metal to the shoots.
Scientists used sunflower plants to clean a pond at Chernobyl contaminated with
cesium and strontium, the most common radionuclides left from the fallout of the
1986 nuclear reactor disaster, which polluted more than 4.6 million hectares.141,142
Dushenkov et al.31 tested the ability of two sunflower cultivars, bean and Indian
mustard to remove U from an Ashtabula, OH, site with a U concentration of 56 g
L1. Sunflower cultivars removed more than 95% of the U from solution in 24 hours.
Almost all of the U removed from water in a laboratory experiment was concentrated
in the roots. The bioaccumulation coefficients based on the ratio of U concentration
in the roots to U concentration in the aqueous phase reached 30,000. Shoot U
concentration was <5 g g1 and root concentration was >15,000 g g1. The best
environment for U removal using rhizofiltration was a moderately acidic pH (approx-
imately 5.5). Dushenkov et al.31 concluded that root U absorption is a combination
112 Trace Elements in the Rhizosphere
best chelator for the leaching of Pb from the soil and enhancing Pb uptake by plants.
In the soil, the applied chelate acts first to complex the soluble metal in the soil
solution. As the free-metal activity decreases, dissolution of bound or precipitated
metal ions begins to compensate for the shift in equilibrium. This process continues
until the equilibrium saturation of the chelator ligand with the metal is achieved, the
supply of metal from solid phases is exhausted, or the equilibrium solubility of the
metal ion with respect to the solid phase is achieved. The uptake of metal by the
plant must then involve either: (1) the release of the metal from the chelate and
absorption of free metal by the plant, (2) the absorption of intact chelate-metal
complex, or (3) the exchange of metal between the chelate ligand and a plant
metabolic ligand. In the case of U and Pu, these processes are not well understood.
The use of chelates in soils to increase metal uptake by plants has a substantial
impact on the phytoextraction of heavy metals. Chelates have been used extensively
to enhance plant nutrient uptake and the uptake of insoluble elements by plants.147-
149 Recently Huang and Cunningham144 and Huang et al.150 have shown that the
helps to reduce the possible movement of U-citric acid complexes within the soil
profile. Citric-acid triggered U accumulation in plants is very rapid, which helps
reduce the risk of having plants with high U levels in the field for a long period of
time. Another advantage of using citric acid is its relatively low cost.
Hossner et al.77 studied the effect of different chelators and rates of application
on U accumulation by two potential U accumulators grown in four different soils.
They found that the degree of U uptake and accumulation among soils was different
even before chelate addition (Figure 5.3). Uranium concentrations in sunflower
shoots of the control treatment of soils contaminated with 300 mg U(VI) kg1 and
without chelate addition varied between 21 and 165 mg U kg1. Sunflower grown
in calcareous Weswood soil (pH = 7.6) had the highest U uptake, and that grown in
very acid Crowley soil (pH = 4.85) had the lowest U uptake. Chelate addition
enhanced U accumulation in both shoots and roots of sunflower in each of the four
soils (Figure 5.3). However, the increase was not as dramatic as reported by Huang
et al.,76 except in sunflower plant roots grown in Crockett and Weswood soils.
Addition of EDTA, citric acid, or oxalic acid to the acid soils, Labelle and Crowley,
did not influence the U concentration in sunflower roots (Figure 5.3). Chelator
addition increased shoot U concentrations two- to fivefold to values no greater than
500 g g1. Citric acid was the most effective of the chelates applied to Crockett
and Weswood soils. In Crowley and Labelle soils, EDTA was the most effective
chelate. The highest U accumulation was in plants growing in soils with CaCO3 or
high pH. Uranium concentration in the roots of sunflowers grown in Crockett soil
(pH = 7.1) in the presence of oxalate was more than 5500 mg kg1. The sunflower-
root U concentration of the control treatment of Crockett soil [(300 mg U(VI) kg1
of soil with no chelate addition)] was also very high (about 2700 mg U kg1). The
source of U contamination in soils in this study was uranyl nitrate which is soluble
in water at the concentrations used. Uranium in its oxidized form, U(VI), is more
soluble than the reduced U form; however, oxidized U(VI) can vary in solubility
depending on the physicochemical properties of the soil.59 In acid soils and in the
absence of complexing ligands, the uranyl cation (UO2)2+ would be readily adsorbed
onto exchange sites of clay minerals. However, in alkaline soils containing carbonate
minerals, the uranyl ion is complexed with the carbonate ion, forming highly mobile,
anionic complexes [i.e., (UO2)(CO3)22, (UO2)(CO3)34]. The processes of uranium
adsorption and complexation might have influenced the uptake and accumulation of
U in acid Labelle and Crowley soils and the calcareous Weswood and limed Crockett
soils. Uranium (VI) carbonate complexes would dominate the U(VI) speciation in
Crockett and Weswood soils. These complexes apparently did not translocate to the
shoots but seemingly precipitated either inside or outside of the roots. Uranium
solubility and mobility were limited in Labelle and Crowley soils due to the presence
of highly adsorptive Fe and Mn oxides.
In growth chamber experiments conducted by Dushenkov et al.31 more than 99%
of the U was found in the roots. The concentration of U in sunflower roots in the
rhizofiltration system was more than 10,000 g g1. Dushenkov et al.31 concluded
that the ability of roots to remove U from solution was plant species dependent.
Results of the experiments of Hossner et al.77 show that U transport from roots to
the shoots of sunflower is limited (Figures 5.2 and 5.3). Addition of synthetic chelate
Bioavailability of Uranium and Plutonium to Plants in SoilWater Systems 115
or organic complexing agents increased U concentration in the plant root but did
not contribute significantly to U accumulation in the plant tops.
The concentration ratio for tissues increased from 6 103 after 1 hour of uptake
to 0.3 after 24 hours. The concentration of Pu in xylem exudates increased from
about 0.03 ng Pu mL1 of exudate at 1 hour to 1.6 ng Pu mL1 at 24 hours. When
Pu was supplied as Pu(NO3)4 under these experimental conditions, less than 1 pg
Pu mL1 was found in exudates at 24 hours.
X. CONCLUSIONS
Most experimental results have shown that U and Pu accumulate mainly in the roots
of plant species. No plant species have been identified as hyperaccumulators of U
or Pu, however, there are differences in U and Pu accumulation in plants. U(VI)
uptake and translocation, and shoot and root concentrations among plant species,
vary by more than 100-fold.
It is unlikely that a phytoremediation strategy using hyperaccumulating plants
will be feasible for U and Pu remediation. However, there are plant species that can
accumulate substantial amounts of metals by changing their root environment and
altering metal availability. It has been suggested that hyperaccumulation may not be
necessary for remediation of U contaminated soils in the U.S.157
The effectiveness of remediation of soil is strongly influenced by soil type. The
use of plants to extract U and Pu from contaminated soils will require the identifi-
cation of high biomass U and Pu accumulating plant species. Two plant species,
sunflower and Indian mustard, have been identified as potential accumulators of Pu
and U. They may be especially effective in the decontamination of radionuclides in
aqueous systems, where contaminants can be accumulated in the roots using hydro-
ponic techniques. For the successful phytoremediation of soil, U and Pu must be
available to plant roots, absorbed by plant roots, and translocated from roots to
shoots. Further research to maximize both the absorption and translocation of radi-
onuclides in plants is needed.
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Health Physics, 22, 559, 1972.
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icium-241 in three major Indian soils, J. Nuclear Agric. Biol., 9, 85, 1980.
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Section II
Interactions of Trace Elements
with Root Microbial Associations
6 Interactions and
Mobilization of Metal
Ions at the SoilRoot
Interface
Salvatore Deiana, Bruno Manunza, Amedeo
Palma, Alessandra Premoli, and Carlo Gessa
CONTENTS
I. Introduction...................................................................................................127
II. Role of the SoilRoot Interface in Uptake Processes .................................129
A. Interaction Reactions ........................................................................129
B. Mobilization......................................................................................135
C. Degradation Products and Their Activity in the Rhizosphere.........143
III. Conclusions...................................................................................................144
Acknowledgment ...................................................................................................145
References..............................................................................................................145
I. INTRODUCTION
The rhizosphere can be considered as an accumulation phase of protons, reductants,
and organic ligands produced by biological activity.1-5 Redox and complexation
reactions that take place in the rhizosphere can appreciably increase the solubility
of the soil mineral components, which leads to an increase of the macro- and
micronutrient concentration in the soil solution.6-8 During their life cycles, plants
release root exudates, which affect the mobility and uptake of nutrients as well as
the physical, chemical, and biological properties of the rhizosphere. The release of
the root exudates takes place mainly at the proximal region of the root hairs, which
are also responsible for the uptake of nutrients.
The continuous exchange of material between the soil and the root is essential
for plant nutrition, soil fertility, and environmental protection.9 The soil particles
and the root surface are joined by a mucilaginous interface, mostly made up of acid
polysaccharides.10-13 Figure 6.1 shows a transmission electron micrograph of an ultra
thin section of garlic root. The cell wall consists of two layers: the outer layer (the
soilroot interface), which is tenuous and largely mucilaginous and is characterized
0-8493-1535-2/01/$0.00+$.50
2001 by CRC Press LLC 127
128 Trace Elements in the Rhizosphere
FIGURE 6.1 Transmission electron micrograph of an ultra thin section of a garlic root. C =
cuticle. Magnification 20,000. (From Gessa, C. and Deiana, S.14 With permission.)
by a network of electrodense zones; and the inner layer, which is compact and mainly
made up of cellulose.14 The mucilaginous layer is discharged into the rhizosphere,
but is continuously regenerated by the Golgi apparatus.
The soilroot interface, which is generally 1 to 10 m thick,15 is characterized
by interlaced fibrils that define a porous system, known as free space volume. The
interface is highly hydrated and, acting as a selective accumulator for nutrients and
as a filter for chemicals, regulates their movement through and out of the cell. It
may support nitrogen-fixing bacteria as well as microorganisms, which produce
substances that antagonize the root pathogens.16
The absorption of nutrients by plants involves their transfer from the root surfaces
to the cell wall, hence to the plasmalemma, and finally to the cytoplasm. The soilroot
interface is involved in cation exchange reactions and often contains precipitates of
iron, manganese, and other elements. In fact, ion adsorption, precipitation, chelation,
reduction, and other reactions occur within the free space.17-21
Plant species differ widely in their ability to mobilize the sparingly soluble soil
compounds. As an example, iron efficient plants have adopted both morphological
modifications and chemical mechanisms for the mobilization and uptake of iron
from the rhizosphere. These include acidification of the rhizosphere, formation of
Interactions and Mobilization of Metal Ions at the SoilRoot Interface 129
A. INTERACTION REACTIONS
A Calcium-polygalacturonate (Ca-PGA) network, which resembles the natural root
mucilages and thus constitutes a good model of natural systems, was used to study
the interaction with metals.14,31 The micrograph (Figure 6.3) shows the fibrillar
structure of the Ca-PGA network. From a molecular point of view the fibrillar
arrangement of the PGA chains seems to result from the dissociation of the COOH
groups and the formation of ionic bridges. The carboxylic groups are arranged as
far apart as possible so that monomers are positioned at an angle of 120 degrees
(Figure 6.4). Interchain interaction is assured by Ca(II) bridges, each formed of two
COO groups. This structure is supposed to be stabilized by H-bonds formed
between the -OH groups of the polymers and the first layer of the water molecules.
Molecular Dynamics (MD) experiments confirm that the aggregation of the PGA
polymers is mainly due to interchain ionic bridges32 as evidenced in Figure 6.5 where
four MD cells of the Ca-PGA system, each with only two chains, are represented.
On the other hand, experiments carried out on a system formed by three PGA
chains in equilibrium with 16 Ca(II) ions, 20 Na+ ions, 12 Cl, and 1024 H2O
molecules demonstrated that calcium has a stronger affinity toward the polymer
130 Trace Elements in the Rhizosphere
FIGURE 6.2 Schematic representation of the root mucilage. SP: free space, S: soil particle,
L: ligand, ML: complex, PS: phytosiderophores, NFA: fulvic acids, NHA: humic acids, TR:
transport of Fe(II), TL: translocator for metal complexes, F: fibrils of Ca-polygalacturonate,
PM: plasma membrane.
matrix compared to Na+ ions. Calcium ions are complexed in the reticulating sites
forming interchain bridges, while Na+ ions diffuse in the free space (Figure 6.6).33
The porous system, which results from the fibrillar arrangement of the polymers,
defines the free space volume that mainly depends on the type of reticulating ion.
Calcium is an excellent reticulating agent as it induces high hydration of the network
and stabilizes the fibrillar arrangement of the system over a wide pH range.34 One
gram of Ca-PGA retains about 250 g of water; this amount increases as the Ca(II)
concentration in solution increases. On the contrary, Cu(II) destabilizes the porous
Interactions and Mobilization of Metal Ions at the SoilRoot Interface 131
FIGURE 6.3 Scanning electron micrograph showing the fibrils and the free-space volume of
the Ca-PGA network. Magnification 100,000. (From Gessa, C. and Deiana, S.14 With
permission.)
system, as will be shown later, notably reducing the degree of hydration and then
the free-space volume. One gram of Cu-PGA retains 100 g of water and decreases
as the Cu(II) concentration increases.34
Ionic competition experiments35 between Ca(II) and Cu(II) ions indicate that:
1. The affinity of Cu(II) for the organic matrix is so high that at very low
concentrations of copper, the Cu(II) ions are nearly all complexed by the
polymer. This condition characterizes the soil liquid phase where the
calcium concentration is very high with respect to that of Cu(II).
2. The free-space volume of the Cu-Ca network decreases slowly as the
amount of Cu(II) increases up to 40%. Then a dramatic reduction occurs,
suggesting that the fibrillar system could be deeply modified if reticulated
Ca is lower than about 60%.
FIGURE 6.4 Schematic representation of the fibril structure: conformation of two polyga-
lacturonate chains projected on the xy plane (A); the three different positions of the carboxylic
groups () refer to the molecular structure created by three rotations of 120 degrees around
the molecular axis; a, a, b, b, and c, c represent the carboxylate groups inside, above, and
below the plane, respectively. Structure of the polygalacturonic acid (B).
Interactions and Mobilization of Metal Ions at the SoilRoot Interface 133
FIGURE 6.5 Four MD cells of the Ca-PGA system, with only two chains represented. (From
Manunza, B. et al.32 With permission.)
(6.0 mM). They indicate that Pb(II) has a slightly higher affinity toward the functional
groups of the network fibrils than Cu(II). The absorption of Pb(II) and Cu(II) at pH
3.5 were equal to 52 and 43 moles /180 moles of COOH groups, respectively,
whereas at pH 5.5 they were equal to 49 and 41 moles, respectively. The absorption
kinetics performed at the same pH values in the presence of both ions indicate that
the amount of Pb(II) absorbed does not vary considerably, whereas that of Cu(II)
decreases to 25 and 15 moles at pH 3.5 and pH 5.5, respectively (Figure 6.7). This
behavior can be attributed to both hydrolysis reactions of the metal ions and the
structural modifications of the network.
The same systems studied in the presence of Cd(II) and Zn(II) show that these
ions compete with Pb(II) and Cu(II) for the same sites.
The collapse of the fibrillar structure of the network induced by the Cu(II) ions
is characterized by the formation of microspheres, which are easily distinguishable
in the SEM microphotographs (Figures 6.8a and 6.8b), with a high Cu(II)
concentration as shown by X-ray mapping (Figures 6.9a and 6.9b). The formation
of microspheres does not occur when the collapse of the fibrillar structure is induced
by Pb(II) ions (Figures 6.10a and 6.10b). The IR spectra of the Pb(II)-PGA and
Cu(II)-PGA complexes suggest that the carboxylate groups coordinate the ion
through a sole oxygen atom. In contrast, the spectra of the Cd(II)-PGA and Zn(II)-
PGA complexes, similar to those of the Ca-polygalacturonates, indicate a bidentate
structure of the carboxylate groups in these complexes. The electron spin resonance
measurements at room temperature of the Cu(II)-PGA network show that the Cu(II)
absorption is accompanied by the formation of an inner-sphere complex. These
findings suggest that a purely electrostatic interaction between the Ca(II), Mg(II),
Mn(II), Zn(II), and Cd(II) ions and the carboxylate groups occurs (i.e., outer-sphere
complexes). On the contrary, the Cu(II), Fe(III), Cr(III), Al(III), VO(IV), and rea-
sonably Pb(II) ions form inner-sphere complexes with the carboxylate groups (Figure
134 Trace Elements in the Rhizosphere
FIGURE 6.6 Snapshots of the polygalacturonate chains from the end of the equilibrated MD
trajectory. The Ca(II) (large) and Na(I) (small) ions are represented as hard spheres. Water
molecules are omitted to make vision easier. (From Manunza, B. et al.32 With permission.)
Interactions and Mobilization of Metal Ions at the SoilRoot Interface 135
FIGURE 6.7 Absorption kinetics at pH 3.5 and 5.5 of Pb(II) and Cu(II) by the Ca-network
in the presence of calcium acetate 6.0 mM. The initial metal concentrations were both 0.6 mM.
6.11). This justifies the markedly higher affinity of these latter ions for the polysac-
charidic matrix and the important role of the mucilages as an accumulation phase
of these metals.36,37
The results of this study have a great biochemical significance in the transfer of
heavy metals through the soilroot interface. Covalently bound ions are very dan-
gerous as they tend to induce the collapse of the fibrils of the root mucilage thus
destroying its porous structure.
The functionality of the PGA matrix depends on the metal concentration. In
fact, it acts as an accumulator of metals until their concentration is under a threshold
that is a property of the metal considered. Above this threshold the porous structure
can be seriously damaged so that it gives up acting as a selective filter, and toxic
ions can easily reach and injure the cellular biological structures. Probably, the ability
of several plants to tolerate the presence of toxic metals can be related to their
capacity to regenerate the mucigel at a high rate, allowing it to maintain its ability
to act as a filter.
The interaction studies of PGA with Fe(III), V(V), and Cr(VI) have shown that
redox reactions also occur in these systems.38 Mssbauer and Elecronic Spin Res-
onance (ESR) measurements evidenced the presence of Fe(II), VO(IV), Cr(V), and
Cr(III) ions. The redox reaction for Fe(III) and VO(V) leads to the formation of
formic acid as a decomposition product of the polymer (Figure 6.12).
B. MOBILIZATION
The metal complexed on the reticulating sites of the mucilage can be mobilized
by the root exudates. An important example of this process is represented by the
136 Trace Elements in the Rhizosphere
FIGURE 6.8 Scanning electron micrograph of the Ca-PGA network containing 10% (a) and
50% (b) Cu(II).
Interactions and Mobilization of Metal Ions at the SoilRoot Interface 137
FIGURE 6.9 X-ray mapping of the Ca-PGA network containing 10% (a) and 50% (b) Cu(II).
Magnification 6,000.
138 Trace Elements in the Rhizosphere
FIGURE 6.10 Scanning electron micrograph of the Ca-Pb-PGA network (a) and its X-ray
mapping (b). Magnification 6,000.
Interactions and Mobilization of Metal Ions at the SoilRoot Interface 139
FIGURE 6.11 Schematic representation of outer-sphere (a) and inner-sphere complexes (b).
mobilization of iron, which can occur through both ligand exchange or redox
reactions.
In the first case, complexing agents such as mugineic acid or compounds
synthesized by microorganisms such as DFOB (desferrioxamine-B) can free Fe(III)
reticulated on the polymer.39 Desorption kinetics carried out with CaCl2 solutions
show that this treatment does not remove the metal from the reticulating sites,
whereas DFOB, which had little effect on the amorphous iron hydroxide, forms
140 Trace Elements in the Rhizosphere
very stable chelates with Fe(III).40 Caffeic acid (CAF) is particularly active in the
redox processes.41-44
What happens when a reductant such as caffeic acid exuded by roots runs through
the interface? In aqueous phase, the redox reaction is pH-dependent and, at pH <
3.5, it proceeds through two steps: the first is faster and involves five electrons, while
the second involves four electrons.41 The redox reaction is characterized by the
formation of compounds able to reduce Fe(III). At pH > 3.5, the redox activity
dramatically decreases. Hydrolyzed Fe(III) species interact with CAF giving rise to
the formation of soluble complexes that successively transform into insoluble iron-
CAF polymers (Figure 6.13). We wish to point out the great chemical flexibility of
CAF, which can activate redox, complexation, polymeriation, and adsorption reac-
tions depending on the environmental conditions.41
Caffeic acid also shows a strong redox activity toward the Fe(III)-PGA com-
plexes, but the mobilization of the metal undergoes a different mechanism.27 In fact,
Interactions and Mobilization of Metal Ions at the SoilRoot Interface 141
the kind of iron coordination in the complex varies with pH so that a different redox
activity of CAF is observed. At acid pH values from 2.4 to 4.0, the metal coordinates
five or four water molecules and one or two oxygen atoms of the COOH group,
which prevents iron from undergoing hydrolysis and precipitation processes. At pH
values from 4.7 to 7.0, where the coordinated water molecules start to deprotonate
and form nonreducible hydroxylated species, the redox reaction is less effective so
that the Fe(II) yield decreases as pH increases. As the hydrolysis of chelated iron
starts at pH values higher than that of Fe(H2O)63+, the amount of reducible iron
bound to the mucilage is much higher in the pH range of 4 to 7.
The Fe(II) produced can be separated into two fractions: diffusible and exchange-
able with CaCl2 6.0 mM. The Fe(II) mobility, in addition to electrostatic effects that
are established between Fe(II) and the PGA carboxylic groups, can be notably
conditioned by the interaction of the CAF oxidation products with the metallic center
in the reticulation sites. The fibrillary structure that characterizes the calcium network
is altered following the substitution of Ca(II) with Fe(III) ions and is restored by
the reduction of Fe(III) by CAF at pH 4.0. This arises from two distinct interaction
mechanisms, which are established between Fe(III), or Fe(II), and the carboxylic
groups of the fibrils. Iron(II), like Ca(II), forms outer sphere complexes, whereas
Fe(III) forms more stable complexes. This has important implications in plant nutri-
tion because the porous structure of the interface favors the transfer of nutrients to
the plasmalemma.
The CAF oxidation products with high molecular weight are hypothesized to
be trapped in the fibrils, whereas those with low molecular weight, such as esculetin
(6,7-dihydroxycoumarin, ESC), can diffuse in the free space (Figure 6.14).44-46 These
products were found to possess reducing and complexing activity and, therefore,
can be considered active in the mobilization of nutrients in the rhizosphere. In
CH CH COOH
OH
Free space OH
Fibril
Free space
C
C C
O O
O O O O
2+ +
Fe(II)-,Fe(III)-CAF Polymers Fe(III) FeOH Fe(OH)2
O O O O
O O
C C C
Free Space
FIGURE 6.14 Schematic representation of Fe(III) reduction by CAF inside the fibrils.
142 Trace Elements in the Rhizosphere
FIGURE 6.15 Structure and probable formation mechanism of products following the reduc-
tion of Fe(III) by esculetin. Fe(III)/ESC molar ratios < 3 (A) and > 3 (B).
particular, esculetin is able to reduce Fe(III) to Fe(II) in the pH range 3.0 to 4.0
through the formation of two main products whose structure and probable formation
mechanisms are reported in Figure 6.15. Their formation can be explained by
considering that the phenoxylic radicals (ArO.), well represented in the systems with
an Fe(III)/ESC molar ratio < 3, can couple or react with other radicals to form
esculetin oligomers, whereas when the Fe(III)/ESC molar ratio is > 3, esculetin is
oxidized to organic acids. At pH > 4 the reducing activity decreases and the formation
of Fe(III)-ESC complexes, which tend to precipitate, is observed. Organic acids
present in the root exudates, such as malic, pyruvic, citric, and oxalacetic acid did
not affect the redox reaction but kept these complexes in solution. This is important
as Fe(III)-ESC complexes can thus reach the plasmalemma where they can be
absorbed or subjected to Fe(III)-chelate reduction. The same molecule is able to
buffer the toxic effects of Cu(II) by reducing this ion to Cu(I), which, in the presence
of chloride ions, tends to precipitate as sparingly soluble CuCl. Other CAF oxidation
products obtained by electrochemical methods or by reactions in an alkaline medium
Interactions and Mobilization of Metal Ions at the SoilRoot Interface 143
with molecular oxygen were found to possess both reducing and complexing activity
toward Fe(III) and a marked complexing activity toward Fe(II).43
O O O O O O
C M C M C
M
C O C OH C O
OH O -H+ O
M + GAL C C C C C C
H H
C C C C C C
OX
FIGURE 6.16 Mechanism of the D-galacturonic acid oxidation by Fe(III), Cr(VI), and V(V).
144 Trace Elements in the Rhizosphere
turonate complexes do not induce a significant change in the Fe(II) yield with respect
to the simple Fe(III)-D-galacturonate system.
At the soilroot interface, the D-galacturonic acid can also reduce toxic-heavy
metals such as Cr(VI).52 This redox reaction can be considered a Cr(VI) detoxifica-
tion mechanism because CrO42-, a very dangerous and mobile anion in the soil-plant
system, transforms into the Cr(III) cation, which tends to precipitate as Cr(OH)3, to
form organic complexes or to be immobilized on particle surfaces. The reduction
of Cr(VI) is slow even at low pH values, but proceeds much faster in the presence
of Cr(III) or other metals which, through their reaction with D-galacturonic acid,
form reducing metal complexes. It is important to note again the particular behavior
of the redox system, which can be activated by the same reduction product.
III. CONCLUSIONS
The soilroot interface, as well as the apoplast, can be represented as an interlacing
of channels that controls the flow of water, ions, organic molecules, and complexes
with different charges through and out of the cells. This system of channels is
stabilized by ionic bridges, mainly composed of Ca-complexes. A substantial sub-
stitution of Ca(II) with ions such as Al(III), Fe(III), Cr(III), Cu(II), VO(II), and Pb(II)
in the reticulation sites can damage the interface, thus exposing the absorbing cells
to the action of dangerous environmental agents. Nutrient mobility inside the inter-
face can be notably different depending on the affinity for biopolymers. In this sense,
it is possible to distinguish:
1. Chemical species with low affinity like potassium, sodium, and several
anionic and molecular species that flow rapidly through the interface;
2. Chemical species such as Ca(II), Mg(II), Fe(II), Mn(II), Zn(II), and Cd(II)
that interact electrostatically in the reticulation sites forming outer-sphere
complexes and can be mobilized by ionic exchange;
3. Chemical species with a high affinity that interact in the reticulating sites
as inner-sphere complexes such as Al(III), Fe(III), Cr(III), Cu(II), and
Pb(II). They show low solubility and are accumulated at the interface and
apoplast from which they can be released following redox and/or com-
plexation reactions. These ions can be very dangerous for plants because
they can damage the root surface. For example, in acid soils, aluminum
can present serious problems for several crops;
4. Chemical species unable to cross the free-space system due to their high
molecular size.
Therefore, our future work will focus on improving the model of the soilroot
interface by using other components present in the mucigel and extending the study
in vivo of this matrix in the plant mineral nutrition.
ACKNOWLEDGMENT
The financial support of 40% from Ministero dellUniversit e della Ricerca Scien-
tifica e Tecnologica (MURST) is gratefully acknowledged.
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7 Analysis of Metal
Speciation and
Distribution in Symbiotic
Fungi (Ectomycorrhiza)
Studied by Micro X-ray
Absorption Spectroscopy
and X-ray Fluorescence
Microscopy
Bjrn O. Berthelsen, Geraldine M. Lamble,
Alastair A. MacDowell, and David G. Nicholson
CONTENTS
I. Introduction...................................................................................................150
II. Ectomycorrhiza Structure and Function .................................................150
III. Heavy Metal Accumulation and Binding in Mycorrhizal Biomass ............152
A. Uptake and Accumulation of Heavy Metals ....................................152
B. Metal Localization ............................................................................152
C. Binding Mechanisms Speciation of Metals ................................153
IV. Techniques and Approaches for Studying Metal Speciation in Fungal
Biomass.........................................................................................................154
A. Available Approaches and Techniques.............................................154
B. X-ray Absorption Spectroscopy (XAS) ...........................................155
V. Zn Speciation in Ectomycorrhiza from Contaminated Forest Soils
Studied by Micro-XAFS ..............................................................................156
A. Materials and Methods .....................................................................156
1. Sampling and Sample Pretreatment ..................................156
2. Instrumentation and Analyses ...........................................157
B. Results and Discussion.....................................................................158
VI. Conclusions...................................................................................................161
References..............................................................................................................162
0-8493-1535-2/01/$0.00+$.50
2001 by CRC Press LLC 149
150 Trace Elements in the Rhizosphere
I. INTRODUCTION
This chapter mainly focuses on a new technique for examining metal binding and
speciation in microscopic structures, micro X-ray Absorption Fine-Structure Spec-
troscopy (mXFAS), a technique based on conventional X-ray Spectroscopy (XAS).
mXFAS may prove to be useful for analysis with respect to trace element binding
and speciation in several types of microorganisms or specific parts of biological
structures other than ectomycorrhiza, which is addressed in this chapter.
Through the years international literature has provided overwhelming documen-
tation on the ability of a great many types of microorganisms, including fungal
species with a wide variety of growth strategies, to accumulate large amounts of
trace metals. Soil fungal biomass, in addition to its numerous and important roles
in most ecosystems, makes up a significant part of the organic matter in surface
soils, with biomass values that may exceed 50 g kg1 as measured by the ergosterol
method.1 According to Bth et al.,2 FDA-active fungal length in forest humus may
exceed 400 m g1 dry soil. Thus soil fungal biomass may constitute an important
metal reservoir in surface soils. In addition to measurements of total metal content
in soil fungal biomass, knowledge on mechanisms of metal binding and speciation
is a key factor in fully assessing the role of fungal biomass in the biogeochemical
cycling of trace metals.
This chapter reviews ectomycorrhizal structures and functions with emphasis on
metal accumulation, localization, and binding mechanisms, followed by a detailed
description of the background and approaches provided by the mXFAS technique.
Other techniques for studying metal localization, distribution, and binding at a small
spatial scale are also summarized. Finally, the application of mXFAS to metal
localization and speciation in biological tissues is demonstrated in a case study on
Zn binding in ectomycorrhizal structures.
B. METAL LOCALIZATION
Metal-binding sites and distribution in mycorrhizal biomass have been extensively
studied for several years. However, caution is needed in such studies since microbial
structures are fragile and may be subject to structural damages upon sample prep-
aration and analysis. This may cause both a redistribution of metals as well as
changes in the metal speciation within the fungal structures. For instance, observa-
tions on metal association with polyphosphate granules as observed by Turnau et
Analysis of Metal Speciation and Distribution in Symbiotic Fungi 153
and Winge.27 who stated with respect to fungi: Glutathione appears to provide the
first line of defense and may be the only detoxifying molecule under certain condi-
tions. Esser and Brunnert28 studied Cd speciation in fruiting bodies of Agaricus
bisporus. Even though they were studying individuals exhibiting high Cd concen-
trations (>70 ppm dry weight), they detected no presence of metallothioneins.
Through advanced Sephadex and Sephacel chromatography they did, however, find
three Cd-binding complexes, but these were not subject to further characterization.
By a combination of EDAX analyses and capillary electrophoresis, Turnau et
al.29 indicated that heavy metal binding to inorganic constituents may take place in
ectomycorrhiza. They concluded that oxalate and/or carbonate may be involved in
complexation of Cd and Pb in mycorrhizal structures.
FIGURE 7.2 Simplified EXAFS spectrum, displaying the four distinct regions pre-edge,
edge, XANES region, and EXAFS region.
tips were stored between humidified tree leaves surrounded by soil material in closed
plastic cups, and then cleaned once more prior to analysis. The intact and still living
mycorrhizal tips were then encapsulated in Kapton tape (used for mounting samples
because of its low absorptivity for high-energy radiation). Cross-sections of ecto-
mycorrhiza, approximately 1 mm thick, were prepared with a scalpel and air-dried
on Kapton tape before being covered with tape.
Data were collected using the X-ray microprobe on the bending magnet beamline
10.3.2 at the Advanced Light Source, Lawrence Berkeley National Laboratory,
U.S.A. A Si (III) four-bounce monochromator was used to monochromatize and
scan the synchrotron X-ray energy for the XANES spectra. Following the mono-
chromator are two grazing incidence mirrors in the Kirk-Patrick Baez arrangement
that image the synchrotron source with high demagnification to an X-ray spot
typically of size 1.5 2 micrometers. High-order harmonics are not present due to
the low energy of the ring (1.9 GeV) and the grazing incidence cutoff energy set by
the grazing incidence mirrors. XANES data were collected at the zinc K-edge
(lambda = 1.28330, energy = 9 661 eV). The beam currents ranged from 300 to
400 mA at 1.9 GeV. The energy resolution of the Si (III) monochromator was
approximately 1.4 10-4 (E/E).
The spectral energy calibration was checked by measuring the transmission
spectrum of a Zn-foil (thickness 5 micrometers; the energy of the first inflection
point being defined as the edge 9661 eV).
The focused dimensions of the beam were checked by carrying out a knife edge
test.
The intensity of the incident X-rays was measured by an ionization chamber.
Excited fluorescence X-rays from the sample were recorded using a solid state SiLi
detector (effective area 55 mm2) placed in the electron orbit plane of the storage
ring and mounted 90 degrees to the incident X-rays to minimize background caused
by scattering.
A two-dimensional elemental map for Zn and Cu was produced by scanning the
sample through the microbeam and recording the emission of K-alpha fluorescence
X-rays. From this elemental map, points of interest were chosen and the microbeam
positioned at these points. XANES spectra were collected at the chosen points by
scanning the incident X-ray energy from ca. 50 eV below the edge to ca. 300 eV
above the edge (i.e., into the EXAFS region) and measuring the K-alpha fluorescence
X-rays from the sample at each energy step. The step size of the data points was
0.5 eV/step. The XANES spectra were calculated by normalizing the intensities of
the fluorescent X-rays to that of the incident X-rays, and plotting against the X-ray
energy. Several spectra were collected and summed.
X-ray fluorescence mapping with respect to Zn and Cu was carried out on fresh,
intact mycorrhiza to identify possible metal hot spots or anomalies for speciation
studies, and on dried cross-sections to study the metal distribution within the
mycorrhiza.
158 Trace Elements in the Rhizosphere
Several XANES spectra extending into the EXAFS region with respect to Zn
were collected from a 5 m zinc particle in the mantle of the Cortinarius mycorrhiza.
Similar spectra were collected from a Zn-oxalate standard.
FIGURE 7.4 Optical micrograph of intact Cortinarius sp. (above) and XRFM of a 400
400 micrometer part of the mycorrhiza (below). Values are shown on the legend bar as counts
per 0.5 seconds.
eV
FIGURE 7.5 Micro-XAFS spectra extending into the EXAFS region of a 5 m Zn particle
in Cortinarius sp. and a Zn-oxalate standard.
our conclusion supports that of Sarret et al.42 who showed, using conventional XAFS,
that for Zn sequestered by lichens under contaminant exposure, the dominant product
is zinc-oxalate. Some uncertainty may be raised concerning such an interpretation,
because Zn binding to acetate produces a similar spectrum.43 Zn-acetate is, however,
highly soluble while Zn-oxalate has a very low solubility. This fact, in addition to
the consistency of the obtained XAFS spectra, thus points to binding of Zn as oxalate
as an important means of Zn immobilization in Cortinarius mycorrhiza.
Several types of fungi produce oxalate, including Basidiomycetes,44,45 mycor-
rhiza of Monotropa uniflora,46 and mycelium of Mucorales.7 Oxalate, among several
other important features,41 may precipitate calcium and thus may reduce calcium
toxicity in calcium-rich soils. The fact that Cortinarius mycorrhiza binds Zn as
oxalate may imply that oxalate precipitation is an important means for the fungus
to buffer and reduce excessive Zn uptake into the host. Based on EELS analyses
Turnau et al.16 observed a gradual decrease of Cd and Al in Rhizopogon roseolus
mycorrhiza from the mantle, along the Hartig net toward the inner region of the
hosts roots, suggesting that the fungal symbiont may act as a filtering system for
toxic elements. Cd and Al, as well as other toxic metals such as Ni and Hg (in the
+I state), form sparingly soluble oxalates. Our findings, together with the findings
of Turnau et al.16 with respect to Zn, suggest that precipitation of oxalate salts may
be involved in metal detoxification in mycorrhizal systems, and perhaps in other
organisms that produce oxalate.
Analysis of Metal Speciation and Distribution in Symbiotic Fungi 161
VI. CONCLUSIONS
The case study presented here clearly demonstrates the capability of direct chemical
speciation at microscales. Micro X-ray spectroscopy has proven to be a feasible
method for studying Zn binding in ectomycorrhiza. There are, however, some factors
restricting the feasibility of this method:
1. The sample needs to contain high levels of the metal studied, or anomalies
within the structure that are sufficiently rich in the metals of interest.
Small metal concentration can cause a decrease in the signal-to-noise
ratio, which means that structural information that otherwise may be
retrieved from the EXAFS region cannot be obtained.
2. If a metal is bound by several mechanisms and no specific binding form
is dominant, there may be difficulties in interpreting the resulting data.
3. An appropriate model compound is needed, especially at low concentra-
tions of the investigated metal, from which extraction of useful structural
data from the EXAFS region is limited.
4. At present only a few synchrotrons fitted with micro X-ray facilities are
available. Accordingly, a shortage of beam-time, which affects planning
and implementation of research projects, is prevalent. In addition, there
is the problem of conservation and storage of sample material.
5. Some metals, e.g., Cd, which are important in terms of environmental
concern, are difficult to study using XAS.
162 Trace Elements in the Rhizosphere
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14. Colpaert, J. V. and van Assche, J. A., Zinc toxicity in ectomycorrhizal Pinus sylvestris,
Plant Soil, 143, 201211, 1992.
Analysis of Metal Speciation and Distribution in Symbiotic Fungi 163
15. Colpaert, J. V. and van Assche, J. A., The effects of cadmium and the cadmium-zinc
interaction on the axenic growth of ectomycorrhizal fungi, Plant Soil, 145, 237243,
1992.
16. Turnau, K., Kottke, I. and Dexheimer, J., Toxic element filtering in Rhizopogon
roseolus/Pinus sylvestris mycorrhizas collected from calamine dumps, Mycol. Res,
100, 1622, 1996.
17. Berthelsen, B. O., Olsen, R. A. and Steinnes, E., Metal concentrations in ectomyc-
orrhiza as affected by sample pre-treatment and storage conditions, Soil Biol. Bio-
chem., in press.
18. Wilkins, D. A., The influence of sheathing (ecto-) mycorrhizas of trees on the uptake
and toxicity of metals, Agric. Ecosyst., Environ., 35, 245260, 1991.
19. Heyser, W. and Donner, B., X-ray microanalytical studies on element uptake and
deposition indifferent tissues of beech mycorrhizae, Agric. Ecosyst. Environ., 28,
175179, 1989.
20. Kumpfer, W. and Heyser, W., Zinc accumulation in beech mycorrhizae a mecha-
nism of zinc tolerance, Agric., Ecosyst., Environ., 28, 279283, 1989.
21. Wasserman, J. L., Mineo, L. and Majumdar, S. K., Detection of heavy metals in oak
mycorrhizae of northeastern Pennsylvania forests, using X-ray microanalysis, Can.
J. Bot., 65, 26222627, 1987.
22. Leyval, C., Turnau, K. and Haselwandter, K., Effect of heavy metal pollution on
mycorrhizal colonization and function: physiological, ecological and applied aspects,
Mycorrhiza, 7, 139153, 1997.
23. Dehn, B. and Schepp, H., Influence of VA mycorrhizae on the uptake and redistri-
bution of heavy metals in plants, Agric. Ecosyst. Environ., 29, 7983, 1989.
24. Howe, R., Evans, R. L. and Ketteridge, S. W., Copper-binding proteins in ectomy-
corrhizal fungi, New Phytol., 135, 123131, 1997.
25. Morselt, A. F. W., Smits, W. T. M. and Limonard, T., Histochemical demonstration
of heavy metal tolerance in ectomycorrhizal fungi, Plant Soil, 96, 417420, 1986.
26. Galli, U., Meier, M. and Brunold, C., Effects of cadmium on non-mycorrhizal and
mycorrhizal Norway spruce seedlings [Picea abies (L.) Karst.] and its ectomycor-
rhizal fungus Laccaria laccata (Scop. Ex. Fr.) Bk. & Br.: Sulphate reduction, thiols
and distribution of the heavy metal, New Phytol., 125, 837843, 1993.
27. Mehra, R. K. and Winge, D. R., Metal ion resistance in fungi: Molecular mechanisms
and their regulated expression, J. Cell. Biochem., 45, 3040, 1991.
28. Esser, J. and Brunnert, H., Isolation and partial purification of cadmium-binding
components from fruiting bodies of Agaricus bisporus, Environ. Pollut., Series A,
41, 263275, 1986.
29. Turnau, K., Dexheimer, J. and Botton, B., Heavy metal sequestration and filtering
effect in selected mycorrhizas from calamine dumps EDAX microanalysis, in
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Frstner, U. And Knchel, A., Eds., CEP Consultants, Edinburgh, 1995, 317320.
30. Johansson, T. B., Akselsson, R. and Johansson, S. A. E., X-ray analysis: Elemental
trace analysis at the 1012 g level, Nuclear Instr. Meth., 84, 1970, 141143.
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Analysis, Wiley, Chichester UK, 1988.
32. Kottke, I. and Martin, F., Demonstration of aluminium in polyphosphate of Laccaria
amethystea (Bolt. Ex Hooker) Murr. by means of electron energy-loss spectroscopy,
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1986, 349 pp.
164 Trace Elements in the Rhizosphere
34. Koningsberger, D. C. and Prins, R., Eds., X-ray absorption: Principles, Applications,
Techniques of EXAFS, SEXAFS and XANES, John Wiley & Sons, New York, 1988,
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imaging using elliptically bent mirrors, Proc. SPIE, 3152, 126135, 1997.
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natural soil. Identification of contributions from different sources, Analyst, 120,
14791483, 1995.
37. Steinnes, E., Allen, R. O., Petersen, H. M., Rambaek, J. P. and Varskog, P., Evidence
of large scale heavy metal contamination of natural soils in Norway from long-range
atmospheric transport, Sci. Tot. Environ., 205, 255266, 1997.
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atmospheric deposition using naturally growing moss as biomonitor, Chemosphere,
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46. Whitney, K. D. and Arnott, H. J., The effect of calcium on mycelial growth and
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80, 707715, 1988.
8 Bioavailability of
Heavy Metals in the
Mycorrhizosphere
Corinne Leyval and Erik J. Joner
CONTENTS
I. Introduction...................................................................................................165
II. Effects of Heavy Metal Pollution on Mycorrhizae......................................167
A. Field Observations ............................................................................167
B. Use of Mycorrhizal Fungi for the Diagnosis of Metal
Toxicity in Soil .................................................................................169
C. Effect of Heavy Metals on the Diversity of AM Fungi ..................169
D. Metal Tolerance of Mycorrhizal Fungi and Mycorrhizal
Plants.................................................................................................170
III. Effects of Mycorrhizal Fungi on the Bioavailability of Heavy
Metals and Their Transfer to Plants.............................................................171
A. Sorption of Metals by Mycorrhizal Fungi .......................................172
B. Accumulation of Metals in Fungal Structures .................................173
C. Solubilization of Minerals Containing Metals.................................174
D. Transport of Metals to Plants by Mycorrhizal Fungi ......................175
E. Application for Land Remediation...................................................179
References..............................................................................................................179
I. INTRODUCTION
Mycorrhizae are symbiotic interfaces between soil fungi and plant roots that permit
plants to acquire nutrients from soil with a far lower carbon expenditure than would
be necessary if all nutrients were to be taken up by the roots alone. Mycorrhizae
may also represent other advantages to plants, such as improved water relations,
enhanced resistance against certain pathogens, and enhanced tolerance to salt and
metal stress.1,2 The majority of both wild and cultured plants form mycorrhizae of
one type or another. In most cases mycorrhizae can determine plant successions and
botanical compositions of terrestrial ecosystems and are crucial for their normal
functioning.2 Evidence of their importance is also provided by their possible role
0-8493-1535-2/01/$0.00+$.50
2001 by CRC Press LLC 165
166 Trace Elements in the Rhizosphere
during plant colonization of land 420 million years ago and their persistence during
plant evolution through the present time.3
The different types of mycorrhizae have quite different host plant ranges and
are thus largely confined to different terrestrial ecosystems. A crude and general
classification4 distinguishes between ericoid mycorrhizae found in heathland and
alpine vegetation (often in high latitudes on highly organic and acid soils), ectomy-
corrhizae (ECM) found in most boreal to subarctic forest trees (intermediate acidity
and organic matter content), and arbuscular mycorrhizae (AM) that are formed by
most herbaceous plants (including agricultural plants) and with the majority of trees
in mediterranean and tropical ecosystems (near neutral soil pH and low organic
matter content).
A common feature to all types of mycorrhizae is the production of extraradical
hyphae, which explore the soil outside the root depletion zone, up to 10 cm from
the root in AM fungi.5 The biomass of mycorrhizal fungi in soil varies greatly with
mycorrhizal type, and may be in the range of 12 to 138 mg g1 soil (organic layer
of forest soil) for ECM6 and 0.1 to 1 mg g1 for AM (mineral soil).7,8 ECM often
form aggregated fungal structures (rhizomorphs, mantles, etc.), whereas AM and
ericoid mycorrhizal fungi form simple, branched hyphae in soil. Extraradical hyphal
biomass or lengths of ericoid mycorrhizae have, to our knowledge, not been reported,
but they are likely to be in the same range as AM when calculated on a soil volume
basis, i.e., around 20 m cm3. The external hyphae of mycorrhizal fungi are respon-
sible for the improved nutrient uptake in mycorrhizal plants, which is greater for
elements with narrow diffusion zones around plant roots such as P, NH4, and most
heavy metals.9
The mycorrhizosphere may be defined as the rhizosphere of mycorrhizal plants.
As most plants are mycorrhizal (unless one deals with plants from the families
Chenopodiaceae, Carophyllaceae, Brassicaceae, and Urticaceae, or plants in ster-
ilized or heavily P-fertilized soil), rhizosphere and mycorrhizosphere are thus syn-
onymous, but the latter emphasizes that the presence of mycorrhizae has been
verified. The rhizosphere is modified in several respects depending on the pres-
ence/absence of mycorrhizae. Thus, microbial communities10 and activity11 are
altered; soil pH, exchangeable acidity, and quantity of exchangeable cations may be
influenced;12 root exudation is commonly reduced;13 and root architecture, morphol-
ogy, and longevity are modified.14 Such observations, along with the often dramatic
effects of mycorrhizae on growth and nutrient uptake of plants and their ubiquitous
distribution, has led mycorrhizal researchers to state that the study of plants without
mycorrhizae is a study of artifacts.15
Heavy metal concentrations in soil can reach toxic concentrations and pose a
major environmental and health problem, mainly due to man-made pollution through
the burning of fossil fuels, mining and smelting of metalliferous ores, municipal
wastes, sewage sludge, and the use of pesticides and fertilizer amendments.
The term heavy metal strictly refers to metallic elements with a specific mass
>5 g cm3 able to form sulfides,16 but is commonly used for some metals that do
not fit this definition. Trace metals would be a more correct term to use since the
concentrations are <0.1% in soil or 100 mg kg1 in dry matter of biological samples.
Bioavailability of Heavy Metals in the Mycorrhizosphere 167
However, the term heavy metal is generally used and accepted in environmental
studies, and it will be used here.
Metals in soil are present as free metal ions, soluble metal complexes (seques-
tered to ligands), exchangeable metal ions, organically bound metals, and precipi-
tated or insoluble compounds such as oxides, carbonates, and hydroxides, or they
may be part of the structure of silicate minerals (indigenous soil content). Some of
the metals are essential for plant growth, such as Zn, Cu, Mn, Ni, and Co,17 while
others have no apparent function such as Cd, Pb, and Hg. However, all exhibit
toxicity above a certain concentration, which depends on the metal, the organism,
and the physicochemical soil properties.
Toxicity of metals in soil depends on their bioavailability, defined as their ability
to be transferred from a soil compartment to a living organism.18 This is a function
not only of their total concentration, but also of physicochemical (pH, Eh, organic
matter, clay content, etc.) and biological factors (e.g., biosorption, bioaccumulation,
and solubilization).19 As pointed out by Giller et al.,20 bioavailability cannot be
measured because it can only be assessed by the growth of an organism of interest
and an evaluation of uptake and toxicity of a metal after the fact.
Soil microorganisms including mycorrhizal fungi are affected by the presence
of high metal concentrations in soil (see reviews by Gadd21 and Giller et al.20). But
the organisms in turn influence the availability of metals in soil either directly,
through alterations of pH, Eh, biosorption, or uptake, or indirectly in the rhizosphere
through their effect on plant growth, root exudation, and resulting rhizosphere
chemistry. A unique trait of mycorrhizal fungi is that they form a direct link between
the soil and the root, providing a route for movement of elements in the rhizosphere
(Figure 8.1). These aspects of the interactions between mycorrhizae and heavy metals
in soil are the topic of the present nonexhaustive review. For full coverage, the reader
should also consult other recent reviews.22-26
A. FIELD OBSERVATIONS
Field observations have shown that AM as well as ECM fungi and ericoid fungi
occur on heavy-metal-polluted sites. For ectomycorrhizal fungi, such studies have
focused mainly on fruiting bodies and shown a strong decrease in fruit produced
and of fruiting species with increasing pollution along a complex heavy metal
pollution gradient.27 Such studies have also shown a decrease in the abundance of
fruit for some species and an increase for others, which sometimes is rare in
unpolluted ecosystems.27
The presence of AM fungi in metal-contaminated soils has been observed by
counting spore density, estimating root colonization, or determining mycorrhizal
infectivity potential (MPN counts). Results are not always consistent, probably due
to different metal concentrations and availability in the soils. In agricultural soils
contaminated with metals through atmospheric deposition from a smelter28,29 and
sludge amendments,28 low to high levels of mycorrhizal colonization and spore
168 Trace Elements in the Rhizosphere
numbers were observed. These effects were attributed not only to metal concentra-
tions, but also to soil pH, P content, and organic matter content, factors that influence
metal availability and mycorrhizal colonization. The effect of heavy metals on
mycorrhizal occurrence and infectivity seemed to be more a function of availability
rather than total soil concentrations.29
Mycorrhizal fungi have also been found in many mine spoils heavily polluted
with metals, where plant establishment was greatly improved by the presence of
AM and ECM.26,30 Indeed, the absence of AM colonization in some mine spoils has
been attributed to poor or absent mycorrhizal inoculum.31
AM spore numbers, root colonization (estimated by nonvital staining), and the
abundance of ECM fruiting bodies may not necessarily reflect the actual activity of
these organisms in the soil system, or even their functional structure. Mycorrhizal
infectivity, i.e., the number of infective propagules (including viable spores and other
infective units) has also been estimated in polluted soils. For example, along a
gradient of heavy metals on a metallicole site in northern France, where only
metallophytes and pseudometallophytes were able to grow, mycorrhizal infectivity
was reduced with increasing metal concentrations compared to control uncontami-
nated soil, but it was never completely suppressed. The vitality of hyphae in soil,
as evaluated by enzyme staining techniques (vital staining using alkaline phosphatase
or succinate dehydrogenase), is another way to estimate toxicity of metals to myc-
orrhizal fungi. For AM fungi, this parameter is reported to be negatively affected
by metals.32 However, the growth of such hyphae seemed to be less sensitive to
metals than plant roots, entering soil volumes amended with up to 100 mg Cd kg1.33
Most of the reports cited above do not contain estimates of metal availability,
or differences in extraction methodology have made it difficult to compare the results
Bioavailability of Heavy Metals in the Mycorrhizosphere 169
from different studies. Still, the presence of mycorrhizae in many soils polluted with
metals suggests metal tolerance of mycorrhizal fungi (see below).
The ability of microorganisms to survive metal toxicity has been defined as resis-
tance or tolerance21 depending on the mechanism involved. Metal resistance
corresponds to an induced mechanism in response to heavy metals, such as the
production of metallothioneins. Metal tolerance is due to intrinsic properties and/or
modification of the environment to reduce toxicity, including binding or precipitation
via, for example, extracellular polysaccharide secretion. Other authors distinguish
between heavy metal avoidance, where the organism is able to restrict metal uptake
(reduced uptake or increased efflux, formation of complexes outside cells, organic
acid release, etc.), and tolerance, where the organism survives in the presence of
high internal metal concentrations (intracellular chelation by synthesis of ligands
such as metallothioneins, polyphosphates, and/or compartmentation within vacu-
oles).40,41 The ability of an organism to survive heavy metal toxicity may involve
more than one mechanism.
Ericoid mycorrhizal fungi grow without any sign of toxicity at 50 g Cu ml1
or 100 g Zn ml1 in liquid medium.42 Higher concentrations lead to no growth in
the case of Cu (100 g ml1) and a 60 to 80% reduction in growth with Zn (at 500
g Zn ml1). For ECM fungi, metal tolerance has been tested by their growth response
in axenic cultures exposed to increasing metal concentrations, and showed large
differences between species. Fifty percent inhibition of growth occurred at Cu and
Zn concentrations ranging from 10 to 200 g ml1, and Ni and Cd concentrations
from 1 to 10 and 0.1 to 10 g ml1, respectively.43 Among the five fungi tested, there
were no interfungal differences regarding tolerance to Pb, Hg, and Cr. For the other
metals, the most tolerant fungi were Pisolithus tinctorius and Hymenogaster sp. In
another study, isolates from metal-contaminated sites were more tolerant than cor-
responding fungi from uncontaminated sites.44 However, metal tolerant fungi have
also been isolated from nonmetal polluted soils, and a wide genetic variation exists
for metal tolerance in ECM fungi.45
Different mechanisms have been suggested for metal tolerance in ECM fungi
involving adsorption or accumulation of metals due to ion exchange, formation of
complexes, precipitation, or crystallization.46 Specific metal binding proteins were
involved in the accumulation of Cd, Cs, and Th in different ECM fungi.47,48 Copper-
binding proteins were detected in the culture media of tolerant isolates of Paxillus
involutus, but not in other fungi also regarded as tolerant, nor in any of the less
tolerant P. involutus isolates.49 Galli et al.50 did not detect metallothioneins in the
mycelium of Laccaria laccata, although Cd induced an increase in sulfate assimi-
lation and cysteine formation.
Components of fungal cell walls, such as chitin and melanins, and bind metals,
have been suggested as the main barrier against uptake of toxic metals.21,23 Denny
and Wilkins51 suggested that Zn was adsorbed to electronegative sites in the hyphal
cell wall and extrahyphal polysaccharide slime of P. involutus. Using electronic
microscopy and associated methods such as energy loss spectroscopy (EELS) and
electron spectroscopy imaging (ESI) microscopy, heavy metals were found to be
Bioavailability of Heavy Metals in the Mycorrhizosphere 171
localized in crystals within ECM mycelium,52 and in P-rich material in the vacuoles,26
suggesting immobilization mechanisms involving sequestration/compartmentation
and precipitation/complexation.
AM fungi cannot be grown without a host plant. Their tolerance to metals has
thus been tested regarding spore germination53,54 and root colonization. One isolate
of a very common AM fungal species, G. mosseae P2 (BEG 69), isolated from a
metal-contaminated soil was more tolerant to heavy metals than another isolate of
the same species (G. mosseae Gm, BEG 12) with no history of exposure to metals.53
AM isolates are often regarded as tolerant, because they are collected from metal-
contaminated soils without checking their effective tolerance. Furthermore, very few
isolates from uncontaminated soils have been checked for metal tolerance.
The mechanisms of heavy metal tolerance in AM fungi have not been studied
yet, nor has the stability of the tolerance of either AM or ECM fungi. Metal tolerance
of mycorrhizal fungi in axenic culture may be different from the tolerance of the
fungi in symbiotic conditions.55 The mechanisms of plant tolerance17 are comparable
to microbial mechanisms and differ between plants as well. Metal tolerance of
mycorrhizal plants may depend on the host plant, on fungal tolerance, and on the
compatibility between them.51 In most studies concerning metal tolerance of mycor-
rhizal plants, only the effect of the fungi has been considered, but possible plant
protective effects against heavy metal toxicity to the fungus have not been studied.
The protective effect of ectomycorrhizal fungi against heavy metal toxicity to
plants has mainly been attributed to the extramatrical mycelia binding metals and
excluding metals from the host,23,56,57 to a filtering effect of the fungal mantle,58,59
and to the whole fungal biomass reducing heavy metal availability to the host plant.56
Binding of the heavy metals by the ericoid mycorrhizal fungi60 and production
of extrahyphal slime61 explained the improved tolerance of ericoid mycorrhizae to
heavy metal toxicity.
In soils heavily contaminated with heavy metals and nutrient deficient, where
nonmycorrhizal plants died, a Glomus isolated from a metal tolerant plant, Viola
calaminaria, allowed plants like maize, barley, and alfalfa to grow.62 Using micro-
beam analysis, it was shown that heavy metals were immobilized in root tissues
containing fungal structures.63 Using compartment devices and a metal tolerant G.
mosseae, fungal immobilization of 109Cd within AM roots and reduced translocation
to the shoots were observed.33
A possible indirect effect of mycorrhizal fungi on plant metal tolerance via the
modification of the rhizosphere, suggested by Dixon and Buschena,58 was never
investigated.
The fungi forming the three types of mycorrhizae have a capacity to protect
their host plants against excessive metal uptake that generally decreases from ericoid,
to ECM and AM fungi. This seems to be the result of adaptation to soil pH and thus
metal availability in their native environments, but it is also related to the morphology
and biomass of the fungal structures characteristic of each group of mycorrhizae.
The ectomycorrhizae are exceptional in this respect as they form an often dense and
thick sheet of fungal tissue (the mantle) that covers the root surface completely so
that any compound entering the root has to pass through the fungal mantle. This
gives ectomycorrhizae the unique ability to filter ions that enter the plant, which
the other types of mycorrhizae do not have.
FIGURE 8.2 Sorption of Cd and Zn on arbuscular mycorrhizal hyphae of four fungi from
solutions containing 0.089 mM Cd(NO3)2 or 0.15 mM Zn(NO3)2. Bars represent S.E.M. (n = 3).
of a fungus may differ between fungi grown in vitro and in symbiosis with a plant,
with higher sorption having been found in the former.68 Though the metal sorption
capacity of AM fungi is unusually high7 and may exceed that of ECM fungi, the
total immobilization capacity of ECM would still be 1 to 2 orders of magnitude
higher than that of AM due to differences in biomass. Lower soil pH and thus higher
metal solubility does, however, potentially impose higher quantities of metals on
the former (see above).
The outcome of sorption studies like those above depends greatly on ion con-
centrations in the incubation solution and the presence of competing cations. In an
experiment on Cd sorption of AM hyphae the presence of 50 g Ca ml1 or 100 g
Zn ml1 in a solution containing 1 g Cd ml1 reduced fungal Cd sorption by 87 and
92%, respectively.7 Such considerations show that the impressive values cited above
are disproportionately high considering the low ion concentrations commonly found
in the soil solution. However, these investigations do serve to illustrate the important
capacity of fungal biosorption, and further show that at ion concentrations < 1M
more than 93% of Cd2+ may be removed from solution by these fungi.7
Cd (g/flask) P (mg/flask)
5 0,5
P
4 Cd 0,4
3 0,3
2 0,2
1 0,1
0 0
0 5 10 20 30 days
FIGURE 8.3 Growth, acidification of the medium, and mobilization of cadmium and phos-
phorus from North Carolina rock phosphate by Suillus granulatus in liquid culture.
sion of Cd was demonstrated with five varieties of lettuce and four of maize, whereas
the same plants showed variable effects of AM on uptake of Zn.96 The consistent
exclusion of Cd and Zn at high soil Zn levels were later verified for lettuce in
symbiosis with three different AM fungi in two soils with contrasting metal con-
tents.85 Contrary to these reports, Killham and Firestone97 found mainly enhanced
uptake of Cu, Ni, and in some cases Pb and Zn in mycorrhizal bunchgrass at three
metal loads and three pH levels. Cd uptake was either not affected or enhanced in
mycorrhizal plants in an experiment with maize and two AM fungi in sand culture.98
The differences in metal uptake were, however, correlated to differences in plant
size in the two latter reports, which complicates the interpretation.
Mycorrhizal transport of some heavy metals has been demonstrated for AM
using compartmented pots. In these pots, roots are restricted to one part of the pot
without metals, whereas external hyphae cross a fine mesh and grow into a root-
free soil volume where metals (sometimes as radioisotopes) have been placed. Thus,
the transport of Zn,99,100 Cu,100,101 and Cd33,100 has been demonstrated, and it has been
established that Ni is not transported by AM hyphae.100 These experiments have not
used more than one fungus each. Thus interspecific variation, which may be con-
siderable with respect to P transport,102,103 remains unresolved regarding AM trans-
port of metals.
AM effects on nutrient uptake in plants depend on root length densities in soil.104
This is because mycorrhizal functioning is based on fungal uptake of nutrients from
outside the rhizosphere and their transport back to the plant, rather than an enhanced
affinity for nutrients in soil already exploited by roots. Elements found in the
rhizosphere are taken up by the root itself and root densities high enough to make
rhizospheres overlap preclude any additional uptake by the fungi. Pot experiments
with AM and heavy metals have differed largely in pot volumes and plant growth
periods, resulting in different root length densities (data for the latter are rarely
reported, but they may be deduced from root weight data). In our laboratory, we
found different effects of the same mycorrhizal fungus on metal uptake by maize
from the same soil at contrasting root length densities. In two experiments with
similar pot size, but with plant size ten times larger in one than in the other, no
effect of AM was found in the experiment with the larger plants. The smaller plants,
however, showed reduced accumulation of Cd.92 In a recent experiment, we com-
pared metal uptake in plants as affected by pot size and AM, using an industrially
polluted soil. The results show that at high root length densities (>25 cm cm3) the
effect of mycorrhizae in reducing plant metal uptake is lower than in plants that had
three times larger soil volume available (Figure 8.4). With these results in mind we
reexamined some of the reports on AM effects on metal uptake, and found that those
reporting no difference or enhanced metal uptake by AM plants seem to have used
higher plant densities, longer growth periods and/or smaller pot volume than those
where the presence of mycorrhizae resulted in decreased metal uptake in plants. For
example, Dueck et al.105 reporting reduced uptake of Zn, used one plant per pot (1 L)
grown for 7 weeks; Heggo et al.,94 who reported reduced Zn, Cd, and Mn in AM
plants, used one plant per pot (2 L) grown for 6 weeks, and Killham and Firestone,97
who reported enhanced uptake of Cu, Ni, Pb, and Zn in AM plants, used 10 plants
per pot (1 L) and a 14-week growth period.
178 Trace Elements in the Rhizosphere
FIGURE 8.4 Zn concentration in shoots of maize grown in an industrially polluted soil (see
Weissenhorn92 for details on soil) as affected by pot size and presence (Myc) or absence (NM)
of an arbuscular mycorrhizal fungus (Glomus mosseae P2). Bars represent S.E.M. (n = 3).
may be valid only under certain conditions. For example, Killham and Firestone97
demonstrated that reduced pH enhanced metal uptake in AM plants, with no effect
of AM on uptake of Cu, Ni, Pb, and Zn by Erharta calycina at pH 5.6, but with a
considerably increased mycorrhizal uptake at pH 3.0.
REFERENCES
1. Gianinazzi, S. and Schepp, H., Impact of Arbuscular Mycorrhizas on Sustainable
Agriculture and Natural Ecosystems, Birkhuser Verlag, Basel, 1994, 226.
2. Smith, S. E. and Read, D. J., Mycorrhizal Symbiosis, 2nd edition, Academic Press,
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Section III
Experimental Techniques and
Modeling Approaches to Study
the Fate of Trace Elements in
the Rhizosphere
9 Solid Phase Fractionation
of Metals in the
Rhizosphere of Forest
Soils
Franois Courchesne, Vronique Sguin,
and Alain Dufresne
CONTENTS
I. Introduction...................................................................................................189
II. Materials and Methods .................................................................................191
A. Study Sites ........................................................................................191
B. Separation of Soil Components........................................................192
C. Soil Chemistry and Metal Fractionation ..........................................192
III. Results and Discussion.................................................................................193
A. Soil pH and Organic Carbon............................................................194
1. pH in H2O ..........................................................................194
2. Organic Carbon..................................................................194
B. Metal Fractionation...........................................................................196
1. BaCl2 Extraction ................................................................196
2. Pyrophosphate and Oxalate Extractions............................201
3. Acid Extraction..................................................................202
IV. Conclusions...................................................................................................203
References..............................................................................................................204
I. INTRODUCTION
The fractionation of metals in the solid phase is controlled by a series of environ-
mental factors such as the pH, redox potential, mineralogy, organic matter, clay, and
oxide content of a soil. Sequential extraction procedures have been used extensively
to estimate the distribution of metals among different chemical forms (e.g.,
exchangeable, organically complexed, oxide bound) in soils.1 One of the overriding
aims of these fractionation studies is to assess the bioavailability of metals in order
to evaluate their potential limiting effect on crops or their harmful impacts on biota
and humans.2 In this context, empirical relationships between metal forms in soils
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2001 by CRC Press LLC 189
190 Trace Elements in the Rhizosphere
and uptake by plants have been developed in agriculture and forestry as a tool to
predict the response of terrestrial ecosystems to environmental changes (e.g., acid-
ification, metal contamination) or management practices (e.g., fertilization, flood-
ing). These relationships are based on the premises that the fractionation of metals
in the bulk soil provides a realistic indication of the chemical conditions faced by
plant roots and of the pool of bioavailable metals. This seems to be the case for a
range of environments where strong relationships between metal fractions in the
bulk soil and metal content in plant parts have been established.3,4 However, this
type of link is either weak or nonexistent in many other soilplant systems.5-8
Although numerous reasons could be invoked to explain these discrepancies, we
believe that the microscale heterogeneity of soil materials needs to be considered to
better assess the relationships between the components of the soilplant system.
The soil is an extremely heterogeneous system and the chemistry of metals has
been shown to vary considerably between horizons and from one microsite to another
within a given horizon. For example, Wilcke and Kaupenjohann9 observed that
greater proportions of total metal (Cd, Ni, Zn, Cu) were found in the bioavailable
fractions at the surface than in the core of soil aggregates. Recent work on the
rhizosphere, the soil microvolume surrounding living plant roots, further documented
the microscale heterogeneity of dissolved metals in soils. Soil solution displacement
studies showed that Zn and Cd were depleted from the rhizosphere of radish (Rapha-
nus sativus cv. Crystal Ball) compared to nonrhizosphere soil.10 The results sug-
gested that Zn and Cd were taken up by plants at a higher rate than they were
supplied by mass flow. A chemical speciation analysis indicated that dissolved Zn
and Cd were mostly found in the uncomplexed ionic form (Zn2+, Cd2+) in nonrhizo-
sphere soil.11 The proportion of free Zn and Cd ions decreased in the rhizosphere
solution in response to an increase in the concentration of dissolved organic sub-
stances produced by the growth of radish. Working with heavy-metal-contaminated
soils, Lorenz et al.12 further demonstrated that the Cd concentrations in plant parts
were more closely correlated to the total or free ion concentrations in the rhizosphere
solutions than with the dissolved Cd concentrations in the nonrhizosphere soil.
However, they found no such relationship for Zn.
Microscale analyses of the solid phase of soils also emphasized the heteroge-
neous spatial distribution of metals. In a red spruce (Picea rubens Sarg.) forest, total
Al concentrations exhibited a strongly descending gradient from the bulk soil to the
root surface, while total Zn followed the inverse trend.13 Total Fe was invariant across
the rhizosphere. Energy dispersive X-ray microanalyses indicated that Al accumu-
lates in the soil material of the root surface zone of oilseed rape (Brassica napus
cv. Rafal) relative to the whole soil.14 Much of the Al taken up by plants was further
retained in the root cell walls. Using acid ammonium oxalate, Chung and Zasoski15
and Courchesne and Gobran16 observed that extractable Al and Fe were systemati-
cally higher in the rhizosphere than in the bulk fraction of agricultural and forest
soils. Gobran and Clegg17 demonstrated that the amount of NH4Cl-exchangeable Al
was higher in the rhizosphere than in the bulk fraction of a Spodosol developed
under Norway spruce (Picea abies (L.) Karst). Yet, Ruark et al.18 found no difference
Solid Phase Fractionation of Metals in the Rhizosphere of Forest Soils 191
TABLE 9.1
Selected Characteristics of the Study Sites
Parent
Site Horizona Soil Vegetation Material Mineralogyb
Coniferous (CON) BC Podzol Abies balsamea Glacial till Qz, Pla, K-f, Amp
Lac Croche (CRO) Bf, BC Podzol Abies balsamea Glacial till Qz, Pla, K-f, Amp
LaTuque (LAT) Bf, BC Podzol Abies balsamea Glaciofluvial Qz, Chl, K-f, Amp
Parc des Grands-Jardins Bf, C Brunisol Picea mariana Glaciofluvial Qz, K-f, Pla, Int
(PGJ)
a Horizon designations and soil orders according to Soil Classification Working Group.25
b Minerals in the clay fraction (< 2 m) are in order of relative abundance where: Qz = quartz, K-f = K-
feldspar, Pla = plagioclase, Int = expandable interstratified minerals, Chl = chlorite, Amp = amphibole.
TABLE 9.2
Total Metal Content of the Bulk
Component of the Seven Soil Horizons
Site Horizon Al Fe Mn Cu Zn
g kg1 mg kg1
(24 to 32% weight), Al, and Fe (Table 9.2). For a given profile, the observed increase
in total Al and Fe from the BC or C to the Bf horizon reflects the accumulation of
sesquioxides in the upper portion of these podzolic soils. The total Mn, Cu, and Zn
contents are much lower than those of Al and Fe and show no systematic pattern,
although Mn tends to increase with depth in the profile.
The pH in H2O of soil materials is significantly ( < 0.05) lower in the rhizosphere
(mean pH = 4.84) than in the bulk (mean pH = 5.01) component of six of the seven
horizons (missing data for the LAT BC). The difference for individual horizons ranges
from 0.05 to 0.50 pH units and is somewhat more pronounced under black spruce
(PGJ) than balsam fir (Figure 9.1). Changes in pH close to roots are documented for
a variety of agricultural and forest soils and include both pH increases and decreases
with respect to the bulk soil.34,35 The main factors responsible for the change in
rhizosphere pH are the exudation and secretion of organic acids by roots and by
rhizosphere microorganisms and the relative contribution of H+ and HCO3 efflux to
the soil following the uptake of nutrients by plants.8 Hedley et al.36 demonstrated
that a greater uptake of cations than anions by plants, with the associated excess H+
release, was closely related to the decrease in rhizosphere pH under rape (Brassica
napus var. Emerald). Petersen and Bttger 37 subsequently showed that the acidi-
fication of the rhizosphere of maize (Zea mays L. cv. Goldprinz) was predominantly
controlled by H+ efflux from roots, not by the secretion of organic acids.
2. Organic Carbon
The amount of organic C is systematically higher ( < 0.05, missing data for the
LAT BC horizon) close to the root surface (mean = 20.8 g kg1) than in the bulk
soil (mean = 5.0 g kg1). The organic C content of the soils covers a wide spectrum
of values and so does the difference between rhizosphere and bulk materials (0.5 to
Solid Phase Fractionation of Metals in the Rhizosphere of Forest Soils 195
FIGURE 9.1 Soil pH in H2O for the rhizosphere and bulk components of seven forest soil
horizons. Error bars are standard deviations of duplicates. The legend for soil names is: PGJ
= Parc des Grands-Jardins, LAT = LaTuque, CRO = Lac Croche, and CON = Coniferous.
The horizons are designated according to Soil Classification Working Group.25
70 g kg1) for a given horizon (Figure 9.2). The average concentration of dissolved
organic C (DOC), as estimated by water extracts, is also significantly higher ( <
0.05) in the rhizospheric environment (28.6 mg L1) than in soil materials further
from the rootsoil interface (7.2 mg L1). This accumulation of organic matter in
the rhizosphere may result from root exudation and decay, microbial activity, and
preferential water flow along root channels and has been observed in a number of
studies.17,38 On a horizon basis, the extent of organic C enrichment in the rhizosphere
is not significantly correlated to the decrease in soil pH, suggesting that the accu-
mulation of organic substances is probably not the main acidifying mechanism in
the rhizosphere of these forest soils.
The growth and activity of roots create a microenvironment that is both acidified
and enriched in organic substances relative to the bulk soil. Moreover, the abundant
C supply of the rhizosphere increases the establishment of microbial colonies (fungi,
bacteria), which in turn, act as a source of organic matter to the rhizosphere.19,39
These specific properties of the rhizosphere are known to accelerate the weathering
of minerals at the rootsoil interface.40,41 Courchesne and Gobran16 showed that the
abundance of weatherable minerals was consistently lower in the rhizosphere than
in the bulk soil and that the magnitude of the mineralogical differences was related
to the resistance to weathering of soil minerals. For example, the amount of amphib-
oles was found to be significantly lower ( < 0.10) near the roots of Norway spruce
[Picea abies (L.) Karst] than in the bulk materials of six horizons collected from
196 Trace Elements in the Rhizosphere
FIGURE 9.2 Organic C content (log scale) in the rhizosphere and bulk components of seven
forest soil horizons. Error bars are standard deviations of duplicates. The legend for soil names
and horizon designations is the same as in Figure 9.1.
B. METAL FRACTIONATION
1. BaCl2 Extraction
Table 9.3 presents both the average amounts of metals dissolved by the various
extractants and the results of the Wilcoxon signed rank test. For all metals (Al, Fe,
Mn, Cu, and Zn) the amounts extracted by the BaCl2 solution are significantly higher
( < 0.05) in the rhizosphere than in the bulk soil as shown in Figure 9.4 for Zn.
The Wilcoxon test uses the size of rank order differences within pairs of data
(rhizosphere versus bulk) and does not rely on the average values. Therefore, the
averages are presented in Table 9.3 to illustrate the extent of differences between
rhizosphere and bulk materials, although they are not the information used to estab-
lish whether the properties of the two components are statistically different. The
mean metal concentrations in the BaCl2 extracts lie within the range for salt extrac-
tions of uncontaminated forest soils.2,42 It should be noted that the standard deviation
values in Table 9.3 reflect the high heterogeneity between soil samples rather than
analytical variation. On a profile basis, the amounts of Al and Fe in the rhizosphere
Solid Phase Fractionation of Metals in the Rhizosphere of Forest Soils 197
FIGURE 9.3 Abundance of amphiboles [expressed as a ratio between the intensity of the
amphibole peak (I) and the intensity of the 100 peak for quartz (IQZ)] in the rhizosphere and
bulk components of six horizons collected from two Podzols (P1 and P2) formed under
Norway spruce [Picea abies (L.) Karst.]. The legend for horizon names is: A = 010 cm, B1
= 1025 cm, and B2 = 2550 cm.
TABLE 9.3
Average Metal Concentrations in the Rhizosphere (R) and Bulk (B)
Components of Seven Forest Soil Horizons as Measured by Four Chemical
Extractions
Extraction
Metal Component BaCl2 Pyrophosphate Oxalate Acid
FIGURE 9.4 BaCl2-extractable Zn in the rhizosphere and bulk components of seven forest
soil horizons. Error bars are standard deviations of duplicates. The legend for soil names and
horizon designations is the same as in Figure 9.1.
and bulk materials decrease with depth and follow the trend observed for organic C
(Figure 9.2) while the other three metals, Mn, Cu, and Zn (Figure 9.3) do not.
The BaCl2 salt is thought to displace the cations that are adsorbed, in an
exchangeable fashion, on the negative charges of the soil.26 In acidic forest soils,
the cation exchange capacity (CEC) increases with organic matter content and is
pH-dependent, decreasing with soil pH.43 The exchangeable metal cations are loosely
held at the soilsolution interface and can easily be replaced by other cations with
their subsequent displacement toward the soil solution. In this context, the exchange-
able form represents one of the fractions, among all the metal fractions on the soils
solid phase, that is the most readily available for uptake by the plants and biota.3
The analysis of the rhizosphere thus shows that the roots are exposed to a larger
pool (up to fivefold) of available, potentially toxic metals than what would have
been estimated from bulk soil analysis. For the seven horizons, the significant
accumulation of BaCl2 exchangeable metals in the rhizosphere parallels the observed
enrichment in organic C (Figure 9.2). Also, because the mineral horizons studied
contain very little phyllosilicate minerals, their CEC is mostly controlled by organic
matter content.44 This suggests that the larger pool of organic matter found in the
rhizosphere, through its known positive effect on the CEC of rhizospheric materi-
als,17,24 favors the retention of exchangeable metals at the rootsoil interface. The
reaction occurs even though the pH of the rhizosphere is lowered by root activity
Solid Phase Fractionation of Metals in the Rhizosphere of Forest Soils 199
FIGURE 9.5 Cumulative amounts of Cu, Zn, Fe, Mn, and Al dissolved (log scale) by the
four chemical extractions for the rhizosphere (R) and bulk (B) components of the Parc des
Grands-Jardins C horizon. Metal contents for each extractant are expressed as percentages of
the total metal content measured by X-ray fluorescence (XRF) in the bulk soil. Error bars are
standard deviations of duplicates for the acid extraction.
(Figure 9.1), a condition that should diminish the CEC of the rhizosphere and the
amount of exchangeable metals.
The exchangeable metal pool in the rhizosphere and bulk components generally
represent less than one percent of the total metal content in the soil as illustrated in
Figure 9.5 for the PGJ C horizon. The exchangeable fraction is known to constitute
a small proportion of total metals in soils.4 However, the amount of BaCl2-exchange-
able Cu reaches 13% of total Cu in the PGJ C horizon and varies from 10 to 24%
when all horizons and components are considered. Exchangeable Zn also accounts
for 2 to 8% of total Zn in six of the seven rhizosphere materials, although its
contribution remains below 2% in the bulk component. These exchangeable Zn
percentages are similar to those reported by Johnson and Petras42 for a Spodosol
and by Ma and Rao45 for contaminated soils. Up to 40% of total Zn was associated
with the exchangeable fraction of Spodosols under citrus production.46 As for Cu,
the high proportion measured in the exchangeable form is in contrast with data for
coarse-textured soils where the bulk of soil Cu is in unavailable forms, either in the
residual fraction or bound to organic matter.47 We suggest that the BaCl2 salt was
effective in displacing some of the Cu present in the weaker Cu-organic matter
complexes found at these low soil pH values.48 Under such conditions, a fraction of
the Cu that would otherwise be strongly bound to organic compounds in a less acidic
environment behaves more like an exchangeable metal.
200 Trace Elements in the Rhizosphere
TABLE 9.4
Average Metal/Organic C Molar Ratio
in the Rhizosphere (R) and Bulk (B)
Components of Seven Forest Soil
Horizons
Metal Rhizosphere Bulk
Molar Ratioa
Al 130.0 (103) 192.0 (321)
Fe 8.2 (9.0) 10.9 (21.2)
Mn 0.9 (0.9) 0.8 ( 1.1)
Cu 1.8 (2.4) 2.3 ( 3.1)
Zn 0.6 (0.6) 0.5 ( 0.6)
a [Amount of metal in the BaCl2 extraction
(M)/amount of organic C in the horizon (M)] 104.
Note: Standard deviation values are in parentheses.
The ratio of the amount of metal displaced by the BaCl2 extraction to the amount
of organic C is presented in Table 9.4. This molar ratio was calculated (conversion
of concentration units from g kg1 to M kg1) for each horizon and metal and averaged
through horizons for a given metal. The computation of this simple ratio is based
on the assumptions that organic matter constitutes the dominant source of cation
exchange sites in these soils and that organic C content represents a good analogue
of the relative abundance of such sites.17,49 The results indicate that the amounts of
exchangeable metal do not differ between the rhizosphere and bulk soil when the
metal concentration is normalized with respect to the amount of organic C (Table
9.4), even though the absolute exchangeable metal concentrations are much higher
in the rhizosphere (Table 9.3). If we accept the assumptions of the molar ratio, then
the data suggest that the proportion of exchange sites occupied by metals (per unit
mass of soil) at the surface of organic substances is virtually equal in both compo-
nents. Only the total number of organic C containing exchange sites differs, being
higher in the rhizosphere.
When interpreting the molar ratio values, consideration should be given to the
fact that the concentration of negative charges per unit mole of organic C probably
differs between the rhizosphere and the bulk soil. Because the charges carried by
the organic matter are pH-dependent, the charge density of organic substances should
be lower in the rhizosphere where the pH is lower. Also, differences in the compo-
sition of organic matter between the two soil components19,21,38 will reflect on the
nature and abundance of the functional groups capable of attracting cations. None-
theless, the observations provided by the molar ratio draw attention to the dynamic
role of organic matter enrichment in the rhizosphere on the accumulation of
exchangeable metals at the rootsoil interface. However, further work is needed on
the composition, properties, and reactivity of the organic substances present in the
rhizosphere of forest soils before their reaction with metals is fully understood.
Solid Phase Fractionation of Metals in the Rhizosphere of Forest Soils 201
FIGURE 9.7 Acid ammonium oxalate-extractable Cu in the rhizosphere and bulk compo-
nents of seven forest soil horizons. Error bars are standard deviations of duplicates. The legend
for soil names and horizon designations is the same as in Figure 9.1.
3. Acid Extraction
Only Cu concentrations were found to differ significantly between the bulk and
rhizosphere components following the HNO3HCl extraction (Table 9.3). The results
of the acid extraction emphasize the fact that, in these forest soils, the rhizosphere
Solid Phase Fractionation of Metals in the Rhizosphere of Forest Soils 203
effect is probably confined to the smallest, most available metal pools, like the BaCl2-
exchangeable and pyrophosphate-extractable fractions (Table 9.3). It seems that the
metal contents estimated for the larger acid-soluble (Figure 9.5) and residual (average
of 63, 58, 87, 48, and 59% of the total metal content for Al, Fe, Mn, Cu, and Zn,
respectively) fractions are too extensive to be significantly affected by the develop-
ment of the root network in soils. This conclusion somewhat parallels observations
concerning the mineralogical composition of rhizospheric and bulk materials where
only the most weatherable minerals of the assemblage (e.g., amphiboles, phlogopite)
were measurably affected by the increased weathering activity generated by root
growth.53,54 Also, the component separation procedure used in this study is macro-
scopic in nature and does not assure the collection of pure rhizosphere, because
the contamination of the rhizosphere by materials from the bulk soil can occur during
manipulations. This fact may limit the detection of changes or gradients in an
environment as small as the rhizosphere. Techniques allowing the in-place charac-
terization of the chemical composition of the rootsoil interface need to be developed
to circumvent this problem.
IV. CONCLUSIONS
The major findings provided by the study of the rhizosphere and bulk components
of uncontaminated forest soils are as follows:
Evidence presented here from forest soils shows that the analysis of the rhizo-
spheric component of soils yields a more complete picture of the microenvironment
created and faced by the roots of coniferous trees. Indeed, the impact of the rhizo-
204 Trace Elements in the Rhizosphere
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10 A Technique for
Quantitative Trace
Element and
Micronutrient Studies
in Plants
Anders Gransson
CONTENTS
I. Introduction...................................................................................................207
A. Historical Rsum.............................................................................207
B. Nutrient Supply in Plant Nutrition Research ...................................211
II. Microelement Nutrition ................................................................................214
III. Toxicity of Elements.....................................................................................218
IV. The Root SubstrateRoot Interface..............................................................220
V. Concluding Remarks ....................................................................................222
References..............................................................................................................222
I. INTRODUCTION
A. HISTORICAL RSUM
Different ways of increasing harvest yield have been discussed for as long as crops
have been cultivated. For example, the peoples of ancient Egypt and China were
aware of the beneficial aspects of flooding on crop yield and used the approach
extensively. Among the first documented philosophers of plant nutrition, Aristotle
(384322 BC) and Theophrastos (371286 BC) recognized the importance of soil
as a source of materials from which plants are made.1 The influence of Aristotle on
the science of plant nutrition persisted, as evidenced by Linn2 (1751) who, in his
Philosophia Botanica, quoted Aristotles philosophy that the soil is the stomach
of plants: plantarum ventriculus est terra .
Two centuries ago, the possibility that plant growth was in any way dependent
on mineral nutrients was still being debated.3 One of the first mineral elements to
be recognized as a plant nutrient was nitrogen. This was demonstrated in 1804 by
0-8493-1535-2/01/$0.00+$.50
2001 by CRC Press LLC 207
208 Trace Elements in the Rhizosphere
TABLE 10.1
Knops culture solution, developed in 1868.4
Note the inaccurate specification, especially of the
Fe-salt, of the amounts to be added per liter of
distilled water for healthy-looking plants.
Chemical Salt Amount (g)
the Swiss scientist T. de Saussure.4 The science of mineral nutrition took advantage
of developments in chemistry, and as purer chemicals were developed, the essential
nature of other mineral elements was discovered. For example, the Frenchman E.
Gris5,6 documented the effects of iron deficiency on plant appearance, although it is
J. Sachs who is normally credited with having, in 1860, conclusively proven the
essential nature of iron.3 By the end of the 19th century, ten mineral elements were
known to be essential to plant growth and culture solutions were being used for
experimental purposes. An example of one of these early solutions is shown in Table
10.1. In 1915, Maz was one of the first to seriously challenge the belief that only
those elements listed in Table 10.1 were required by plants.7 It had become clear
that trace amounts of different elements were being added as impurities in experi-
ments with solution cultures and that some of these might well be important to plant
growth. In 1905, the French scientist G. Bertrand, recognized manganese as being
fundamental to plant growth4 and, in 1922, J.S. McHague showed that manganese
was essential to the metabolism of higher plants.3 Manganese was found in much
smaller quantities than the other elements, and was the first element to be defined
as a micronutrient, in contrast to the other macronutrients.8 By the mid-19th
century, it was clear that plants required several different elements in balanced
proportions to maintain healthy growth, and today it is general practice to list six
mineral nutrient elements that are required for plant growth as macronutrients, and
six trace elements as micronutrients (Table 10.2). Other elements are often consid-
ered as beneficial to plant growth (Table 10.3), but their true importance is debated.
At the same time other elements were recognized as being potentially growth-
retarding or toxic for several plant species when present in higher concentrations in
the rooting substrate (Tables 10.4 and 10.5). During evolution, some plants species
have developed the ability to accumulate large amounts of metals without showing
toxic responses (Tables 10.4 and 10.5). These species are often referred to as
chemoecotypes, and have developed a genetically based heavy metal resistance,
which may be modified by adaptation. Despite the current level of knowledge
regarding which elements are essential to plant growth, the quantitative requirements
and proportions of these different nutrients are much less well appreciated. For
Quantitative Trace Element and Micronutrient Studies in Plants 209
TABLE 10.2
Essential plant macro- and micronutrient
mineral concentrations considered to be
adequate for most higher plants81 and for
Betula,36 Pinus, and Picea species34 at free
access to all nutrients.
Concentration in Dry Matter
Element Higher Plants Betula Pinus Picea
g g1 dm)
Macronutrients (
N 15 40 26 22
K 10 26 12 12
P 2 5 5 4
Ca 5 2.8 3 3
Mg 2 3.5 2 1
S 1 3.6
g g1 dm)
Micronutrients (
Fe 100 280 180 160
Mn 50 160 100 90
B 20 80 50 45
Zn 20 25 15 15
Cu 6 10 10 10
Mo 0.1 3 2 2
Cl 10 10 10
example, it may be noted that, in recent times, iron has still been listed as a
macronutrient,4,9 although the proportion of iron required for healthy growth may
not be much greater than that required for many micronutrients. The distinction
between macro- and micronutrients on the basis of amounts found in plant tissues
can be misleading in a functional context. From a physiological point of view, other
definitions may be proposed, according to the function of the element in the plant
(Tables 10.6 and 10.7).
TABLE 10.3
Some elements regarded as beneficial for
higher plants.3
Element Example of Function in Higher Plants
TABLE 10.4
Examples of elements and concentrations regarded as toxic
for higher plants. Note that some of the micronutrient
elements are very toxic in slightly raised concentrations.82, 83
Concentration
Element Typical Response or Effect g g1)
(
TABLE 10.5
Examples of concentrations in some chemoecotypes.
Typical Concentrations
Species Accumulated Element g g1)
(
Astragalus preussi U 70
Astragalus racemosus Se 15.000
Jasione montana As 31.000
Minuartia verna Cu 1.0501.850
Minuartia verna Pb 11.40026.300
Minuartia verna Cd 350400
Thlaspi caerulescens Zn 11.30025.000
TABLE 10.6
Classification of essential mineral nutrients according to photosynthetic
function.3 Parentheses denote that function is unclear or mainly based on
indirect evidence.
Constituents of Activators of Enzymes,
Elements Process Organic Structure Osmoregulation
TABLE 10.7
Classification of essential mineral nutrients according to the general
function.84 Parentheses denote that function is unclear or mainly based
on indirect evidence.
Group Function Elements
FIGURE 10.1 The dependence of relative addition rate of nutrients, RA (day1) and photo-
synthetic active radiation (PAR) on the relative growth rate, RM (day1), of plants. The equation
of the fitted line is y = 0.003 + 0.997 with r2 = 0.945. The symbols represent; = Betula
pendula, PAR 450 M s1 m2; = B. pendula, PAR 250 M s1 m2; = B. pendula,
PAR 50 M s1 m2; = Pinus sylvestris, PAR 250 M s1 m2; = Pinus contorta, PAR
250 M s1 m2; = Picea abies, PAR 250 M s1 m2.
1 dn
RA = (1)
n dt
Quantitative Trace Element and Micronutrient Studies in Plants 213
When using RA to control the relative uptake rate of nutrients, stable relative
growth rates, RM, may be attained. Thus, the relative growth rate equals the relative
addition rate of nutrients, which may be defined as:
1 dM 1 dn
RM = = = RA (2)
M dt n dt
where M (g) is the plant mass. This is the steady-state approach and has been
used extensively in studies of plant nutrition during recent years.26 At steady-state
growth, the plant nutrient concentration, n/M, remains constant for a given genotype,
growth environment, and the chosen value of RA.27
n
d
M
=0 (3)
dt
When nutrients are added at different RA, any submaximal or maximal RM will
be in a one-to-one relationship with the corresponding RA regardless of plant species
or external environmental conditions.21,28-37 The conductivity of the culture solution
is usually very low, less than 50 S cm1 at submaximum supply rate as RA = RM.
Solution conductivity at maximum RM may be kept at about 50 S cm1, but is
normally about 100 to 200 S cm1, to ensure free access to all nutrients. The RA
of nutrient elements controls plant characters other than growth, e.g., biomass par-
titioning to above- and belowground plant parts (Figure 10.2), leaf or needle area
development, and the uptake rate of nutrient elements. Uptake rates of different
elements at the growing root tip may thus be calculated as nutrient uptake rate per
unit root growth rate, defined as dn/dMr (mol g1 root DM):
dn n M
= (4)
dMr M Mr
where n is the nutrient amount in the plant, and M and Mr are plant and root dry
matter, respectively. (Related theoretical considerations concerning the technique
have been discussed elsewhere).10,27,37,38 The use of nutrient addition rate as a treat-
ment variable has proved to be a useful tool for a number of research areas related
to plant growth and soil acidification and pollution.13,39-46
The fertility of a soil depends highly on weathering and mineralization of
inorganic nutrient elements. It is possible, by using the steady-state technique, to
mimic low concentrated fluxes of nutrients, which may occur in the rhizosphere.
Consequently, by using the technique as a tool to mimic nutrient fluxes in the
rhizosphere in field trials, soil fertility and the growth of Pinus sylvestris and Picea
abies increased with only small nutrient leakage to the soil solution.47-48
214 Trace Elements in the Rhizosphere
FIGURE 10.2 Root mass ratio (percent root DM of plant DM) at different relative addition
rates of Cu, Fe, Mn, and Zn (day1). FA denotes free access to all nutrient elements. The root
mass ratio of N at different RA (day1) is included as reference.
FIGURE 10.3 Calculated increase of internal zinc amount in Betula pendula plants growing
at a maximum RM of 0.25 day1 () and at a submaximum RM of 0.10 day1 (). The addition
of a constant external concentration of 5 M day1 is included (dotted line). The latter results
in growth decline and decrease in internal concentrations of nutrients within a few days. The
calculations are based on a seedling dry mass of 1 g at time = 0 and an internal plant [Zn]
of 25 g g1 DM.
FIGURE 10.4 Traditional view of the relationship between growth of plants and nutrient
status. Region I denotes severe limitation and Region II mild limitation. It is in these two
regions that the Piper-Steenbjerg effect occurs. Region III consists of more or less healthy-
looking plants. In Region IV, luxury consumption of nutrients occurs, and in Region V, toxicity
is encountered. Redrawn from Bouma.49
FIGURE 10.5 The relationship between the relative growth rate, RM day1, and plant con-
centrations of iron, manganese, copper, and zinc in small Betula pendula seedlings. The
concentrations are given as g g1 DM. Open symbols are for controlled exponential supply
rates; closed symbols are for free access to all nutrients.
TABLE 10.8
Growth response of Picea abies (L.) Karst. to
different aluminum concentrations in culture solution
experiments.
[Al], (mM) Plant Growth Reaction Reference
mM in all treatments. The RM of the plants was close to RMg or RCa and was not
affected by the concentration of Mg2+, Ca2+, or Al3+ in the solution. Uptake of Ca,
at Ca-limitation, increased significantly in the roots after addition of Al, and
decreased in all plant parts when the Mg concentration of the culture solution was
raised. Fewer clear effects were found on Mg uptake at Mg-limitation. Thus it seems
as if Al interferes with the uptake of Ca and Mg in excess of growth requirements
and to a much lesser extent with the uptake of these elements when they are in low
supply. Van Oene46 developed a mechanistic model for the study of interactions
between Ca, Mg, and Al. The model is based on surface complexation of the
membrane with both specific adsorption reactions and exchange reactions included.
The behavior of the model agreed with reported inhibiting effects of Al, and showed
that effects of Al might be reversed by increased nutrient solution concentrations of
cations. However, antagonism between Ca and Mg is a well-known phenomenon in
plants3 and was clearly seen in this investigation.
The balance between Ca and Al in the root substrate is regarded as playing an
important part in the growth of trees.60,63,68 A similar approach has been advocated
by Sverdrup et al.69,70 who used molar ratios of (Ca + Mg + K/Al) as a diagnostic
tool when examining literature data about growth responses to Al. Sverdrup et al.
proposed that a molar ratio above 1 is required for normal growth of several tree
species. Such ratios do not offer a deeper physiological understanding of the
observed nutrient imbalances in forest trees. The molar ratios in the experiments
were all far below the critical value of 1 (often 0.1) in all investigations performed
with the steady-state approach, except in the highest Ca treatment in the Mg-limited
experiment.13,40,43-44 Consequently, Al-induced inhibition of cation uptake cannot
simply be explained from the balance between these elements in the soil solution.
According to Van Oene,46 the ratio is rather an expression for a mixture of pH and
Ca effects.
elongation) increase for N, P, S, and Fe but decrease for K, Mn, and Cu.10,24,28,29,31,32,73
Similar results have been found with respect to Cu-limitation in Hordelymus,20 but
with no control of the stability of biomass partitioning.
The nature of a hydro- or aeroponic culture differs widely from a soil culture.
Nevertheless, the regulation of nutrient intake in the roots is the same regardless of
rooting substrate. The use of hydro- or aeroponic systems may thus, because of their
relative simplicity, be advantageous for the study of root morphology or nutrient
uptake. As the nutrients are delivered to the roots, the plant response may be
interpreted as an inherent plant property. By controlling the RA of nutrient elements,
plant characters such as biomass partitioning to the roots may be controlled and kept
stable during an experiment. Plant responses, which are treatment specific and thus
not time dependent, may then be studied more thoroughly. In experiments with the
micronutrient elements Cu, Fe, Mn, and Zn, biomass partitioning to the roots was
found to be highly dependent on the chosen value of RA and the micronutrient
element in question.24,25,31,32 The relative proportion of root biomass was lower at
severe Cu-, Mn- and Zn-limitation than at moderate or no limitation, and the ability
of plant roots to penetrate a new soil volume becomes less evident at severe Cu-,
Mn-, and Zn-limitation. By cultivating the birch plants at steady-state nutrition,
it was possible to calculate uptake rates of the micronutrient elements with respect
to the growth rates of the roots.24,25,31,32 The range in dn/dMr [Equation (4)] (excluding
FA) was much less than the range of plant RM, which implies that partitioning to
roots, or changes in root RM, and not necessarily changes in specific uptake rate, are
important components for the further acquisition of these nutrient elements. Under
field conditions, root growth can be one of the most important factors in acquisition
of nutrients,72 and apparently there is no evidence that growth response at Cu-, Mn-,
or Zn-limitation will result in a relatively improved acquisition of these elements,
because the partitioning pattern does not favor root growth. It is evident, from the
examples given above, that the steady-state technique may be a useful tool to model
and predict processes in the rhizosphere under a variety of different environmental
conditions. Studies of the rhizosphere are important for the understanding and
modeling of nutrient and trace element acquisition in forest ecosystems. Because of
the use of bulk soil chemistry and culture solution input data in computer models,
it has been questioned whether they predict the nutritional conditions of the soil
surrounding the root.47,74,75 Mass flow and diffusion are often assumed to be the most
important ion transport processes in supplying roots with nutrients,76,77 although the
magnitude of these are difficult to quantify and vary in different ecosystems.78 Rapid
plant growth can deplete nutrients from the rhizosphere, particularly if additional
nutrient supply is mediated by weathering.79 It may therefore be argued that these
variables in controlling P and K availability have been overemphasized and that
other processes, such as nutrient flux density, control nutrient availability in the
rhizosphere.47,80 Ingestad37 and coworkers12,22,24,25,28-30,31,32 showed that it was possible
to grow plants at stable and high growth rates, and at extremely low external nutrient
concentrations, by using the steady-state technique as a tool to mimic rhizospheric
inorganic nutrient fluxes. Thus, the nutrient acquisition of the plants is not depending
on diffusion and mass flow at the root surface, but on nutrient fluxes and root growth
rate. The fertility of a soil depends highly on weathering and mineralization of
222 Trace Elements in the Rhizosphere
V. CONCLUDING REMARKS
It has been demonstrated that steady-state nutrition can be achieved where the supply
of a micronutrient is regulated in an appropriate manner. In such studies, it is essential
that contamination by micronutrient impurities is avoided, and appropriate measures
for achieving this have been outlined. Over the range of supply for which either Cu,
Fe, Mn, or Zn was limiting, plant growth rate was positively correlated with the
amount of limiting nutrient in the plant. Because the plants were grown at steady-
state nutrition, it was possible to discuss growth response in terms of structural and
functional aspects of root and shoot growth, which may be considered to characterize
the response to micronutrient limitation.
It may be concluded from the small, direct effects of aluminum concentration
on tree seedling growth rate, that this variable (at the concentrations normally
encountered in the fine-root zone) is unlikely to account for the extent of forest
dieback that has been observed on many sites. The measured effects of aluminum
concentration on the uptake of calcium and magnesium did not result in growth-
limiting proportions of these nutrients in plant tissues. However, the proportion of
magnesium was approaching a critical level in spruce.
The findings have illustrated the feasibility and usefulness of the steady-state
approach in plant nutrition studies of potentially toxic substances and growth
response to micronutrients. As the steady-state technique has been used as a tool to
mimic rhizospheric inorganic nutrient fluxes, it has been found that nutrient acqui-
sition by plants is not dependent on diffusion and mass flow at the root surface, but
on nutrient fluxes and root growth rate.
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69. Sverdrup, H., Warfvinge, P., Frogner, T., Hya, A. O., Johansson, M., and Andersen,
B., Critical loads for forest soils in the nordic countries, Ambio 21, 348, 1992a.
70. Sverdrup, H., Warfvinge, P., and Rosn, K., A model for the impact of soil solution
Ca:Al ratio, soil moisture and temperature on tree base cation uptake, Water, Air, and
Soil Pollut. 61, 365, 1992b.
226 Trace Elements in the Rhizosphere
71. Barber, S. A., A diffusion and massflow concept of soil nutrient availability, Soil Sci.
93, 39, 1962.
72. Clarkson, D. T., Factors affecting mineral nutrient acquisition by plants, Ann. Rev.
Plant Physiol. 36, 87, 1985
73. Ericsson, T., Rytter, L., and Linder, S., Nutritional dynamics and requirements in
short rotation forests, in Ecophysiology of Short Rotation Forest Crops, Mitchell, P.,
Ford-Robertson, J. B., Hinkley, T., and Sennerby-Forsse, L., eds., Elsevier Applied
Science, London, 1992, 3565.
74. Hgberg, P. and Jensn, P., Aluminium and uptake of base cations by tree roots: A
critique of the model proposed by Sverdrup et al., Water, Air, and Soil Pollut. 75,
121125, 1994.
75. Gobran, G. R. and Clegg, S., A conceptual model for nutrient availability in the
mineral soil-root system, Can. J. Soil Sci. 76, 125, 1996.
76. Nye, P. H. and Tinker, P. B., Solute Movement in the Soil Root System, Blackwell
Scientific Publications, Oxford, 1977.
77. Barber, S. A., Soil Nutrient Bioavailability. A Mechanistic Approach, John Wiley &
Sons, New York, 1984.
78. Binkley, D., Forest Nutrition Management, John Wiley & Sons, New York, 1986, 25.
79. Nilsson, L. O. and Wiklund, K., Influence on nutrient and water stress on Norway
spruce production in south Sweden The role of air pollutants, Plant Soil 147,
251265, 1992.
80. Clegg, S. and Gobran, G. R., Rhizospheric P and K in forest soil manipulated with
ammonium sulfate and water, Can. J. Soil Sci. 77, 525533, 1997.
81. Pettersson, S., Mikronringsmnena ur vxtfysiologisk synpunkt upptagning,
funktion och samspel, in Micronutrients Uptake, Function and Interactions, Skogs,
K., ed., o. Lantbr. akad. tidskr. Suppl. 16, 7, (English summary). 1984.
82. Balsberg Phlsson, A.-M., Toxicity of heavy metals (Zn, Cu, Cd, Pb) to vascular
plants, Water, Air, and Soil Pollut. 47, 287, 1989.
83. Larcher, W., Physiological Plant Ecology, Springer, Berlin, 1995.
84. Mengel, K. and Kirkby, E. A., Principles of Plant Nutrition, 3rd Ed., Int. Potash
Institute, Bern, 1984, 13.
85. Rost-Siebert, K., Untersuchungen zur H- und Al-ionen-toxizitt an keimpflanzen von
Fichte (Picea abies Karst.) und Buche (Fagus sylvatica L.) in Lsungskultur, Berichte
des Forschungszentrums Waldkosysteme/Waldsterben, Bd. 12, 1985.
86. Abrahamsen, G., Effects of acidic deposition on forest soil and vegetation, Phil.
Trans. R. Soc. Lond. B 305, 369, 1984.
87. Evers, F. H., Ein Versuch zur Aluminium-Toxizitt bei Fichte, Forst- und Holzwirt
12, 305, 1983.
88. Wilkins, D. A. and Hodson, M. J., The effects of aluminium and Paxillus involutus
Fr. on the growth of Norway spruce [Picea abies (L.) Karst.], New Phytol. 113, 225,
1989.
11 Cation Exchange on
Plant Roots Involving
Aluminium:
Experimental Data
and Modeling
Joseph E. Dufey, Jos G. Genon,
Benot Jaillard, Henri Calba, Gervais Rufyikiri,
and Bruno Delvaux
CONTENTS
I. Introduction...................................................................................................228
II. Origin, Measurement, Variability, and Implications of the
Cation Exchange Capacity of Roots (CECR)..............................................228
A. Origin ................................................................................................228
B. Measurement.....................................................................................229
C. Variability..........................................................................................229
D. Implications ......................................................................................230
III. Aluminium Exchange on Roots ...................................................................231
IV. Modeling Cation Exchange on Roots ..........................................................235
A. Review of Existing Models ..............................................................235
B. Description of the Model .................................................................237
1. Chemical Speciation ..........................................................237
2. Ion Exchange .....................................................................238
a. General Equation for Ion Exchange................239
b. The Donnan Volume ........................................241
C. Estimation of the Model Parameters and Simulations ....................242
1. Calcium-Aluminium Exchange for Clover and
Ryegrass Roots ..................................................................242
2. Cation Exchange Data for Banana and Maize
Roots ..................................................................................246
V. Conclusion ....................................................................................................247
Acknowledgments..................................................................................................247
References..............................................................................................................248
0-8493-1535-2/01/$0.00+$.50
2001 by CRC Press LLC 227
228 Trace Elements in the Rhizosphere
I. INTRODUCTION
The uptake of major and trace elements by plants is regulated both by active and
passive mechanisms at the soilroot interface. Among the active mechanisms, the
release of protons and organic substances in the rhizosphere is extensively studied
at present as they play key roles in the mobilization of nutrients and toxic elements.
A state-of-the-art paper on these processes was recently published by Hinsinger.1
These active mechanisms take place in a physicochemical environment, which
depends to a large extent on ion exchange reactions between soil matrix, solution
phase, and root. The interface between the living part of root cells (the cytosol
delimited by the plasmalemma) and the soil solution is made up of a macroreticulated
network of cell walls, which reduces solute mobility by both mechanical friction
and electrostatic interactions. The composition of the solution close to the plasma-
lemma and the functioning of proteins and enzymes included in the cell walls are
deeply influenced by ion exchange processes.
The aim of this chapter is to briefly recall the origin, sources of variability, and
practical implications of root exchange capacity, to present results of exchange
experiments involving aluminium as a model trace element, and to show how a
mathematical model can be used to simulate ion fixation on root exchange sites.
A. ORIGIN
Although exchange equilibria evidently take place inside any plant cell, the
exchange sites that can react directly with the soil solution are essentially located
in the apoplast. Carboxylic groups of uronic acids from pectin and hemicellulose
in cell walls, and proteins to a lower extent, are mostly responsible for the exchange
properties of roots.2,3 The accessibility of the exchange sites for cations from the
soil solution can be limited to the cortex when the endodermis is suberized;
exchangeable trace cations are sometimes found only in peripherical cell layers,4,5
likely because thermodynamic equilibrium throughout the apoplast is slow to
become established when the concentrations in solution of high affinity cations
are low. Also direct observation of element localization in roots (e.g., using
microprobes) does not distinguish between exchangeable cations and other forms
of accumulation.
Phospholipid head groups from plasma membranes, which are in close contact
with cell walls, and membrane proteins also exhibit cation exchange properties that
influence transport processes from apoplast to symplast,6,7 but their contribution to
total root exchange capacity is expected to be low when considering the relative
weights of plasma membranes and cell walls. Ishikawa and Wagatsuma8 reported a
strong positive correlation between CECR (i.e., including membranes) and the cation
exchange capacity of cell walls among four plant species.
Cation Exchange on Plant Roots Involving Aluminium 229
TABLE 11.1
Comparison of CECR Values (cmolc kg1) Measured by
Different Methods on Clover and Ryegrass Roots
Titration in the Presence of
Cu Fixation Ca + Mg + Na Na Mg Ca
B. MEASUREMENT
The cation exchange capacity of roots can be measured in different ways, which
can influence the results and consequently the interpretation of data. Basically, as
CECR mainly originates from carboxylic groups, parameters such as pH, reference
cation, and ionic strength, used in the measurement procedures should be controlled
and mentioned. A method based on Cu adsorption followed by acid extraction was
proposed as a reference procedure by Dufey and Braun.9 Because copper is a trace
element in roots, Cu content after extensive leaching with CuSO4 can be attributed
to Cu binding with exchange sites. In addition, carboxylic groups have a very high
affinity for Cu ions, so that most native exchangeable cations are easily displaced
by CuSO4 leaching. Table 11.1 compares CECR values estimated by Cu fixation to
the sum of base cations displaced by Cu (clover and ryegrass roots were first
equilibrated with mixed CaCl2-MgCl2-NaCl solutions). Titration of acidified roots
can also provide CECR values; the data reported in Table 11.1 were obtained by
titrating protonated roots with NaOH in the presence of 0.1M CaCl2, MgCl2, or NaCl
up to pH 6, i.e., a pH value at which all carboxylic groups should be deprotonated.
These data support the validity of the Cu method for CECR measurements. This
method was recently used by Pintro et al.10 and Calba et al.11 to speciate Al forms
in maize roots.
Strontium or Ba were used as extracting cations by other authors;12-14 like Cu,
they have high affinity for root exchange sites, so they can leach the native exchange-
able cations.
C. VARIABILITY
As the uronic acid content in roots is first related to the plant species, CECR also
depends first on plant species and even on varieties or cultivars of the same species.
Wacquant15 listed CECR for a few hundred herbaceous species: monocots usually
have low CECR in the range 10 to 20 cmolc kg1 (dry matter basis), whereas most
dicots have CECR in the range 20 to 50 cmolc kg1. Experiments on forest tree
rootlets showed that coniferous trees had higher CECR than deciduous trees;16 CECR
values (titration method, pH 6) of 13.5 and 14.0 cmolc kg1 were found for beech
and oak, respectively, whereas CECR of 18.0, 20.5, 23.0, 23.5, and 26.0 cmolc kg1
were found for larch, nobel fir, pine, spruce, and douglas fir, respectively.
230 Trace Elements in the Rhizosphere
cmolc kg-1
The age of roots or root parts also influences CECR.17-18 Indeed, secondary cell
walls very poor in pectin are formed in older root parts so that the increase of dry
weight is not accompanied by a proportional increase of exchange sites. For pine-
apple roots collected in the field or grown in nutrient solution (Figure 11.1, data
from F. Ikans thesis, UCL Louvain-la-Neuve, not published), we observed CECR
values ranging from 25 cmolc kg1 near the apex down to less than 10 cmolc kg1 at
20 cm from the apex.
We also noticed that CECR changes according to root order. For example, in
banana roots, we measured CECR 23 cmolc kg1 for main roots and CECR 34
cmolc kg1 for lateral roots (mean values of five cultivars, data not published).
The nutritional status of roots or the presence of toxic elements were also
reported to influence CECR. This might result from changing the average age of
the root system. Indeed toxic elements inhibit root growth and, therefore, increase
the average root age, whereas adequate nutrient supply results in healthy root systems
with well-developed young roots.
D. IMPLICATIONS
Many authors attempted to find relationships between CECR and various plant
characteristics such as adaptation to different stresses (salinity, Al toxicity, low or
high soil pH, etc.). These studies are not easy to generalize, because they deal almost
always with very limited series of plant species or plant varieties. For example, it
seems that halophytes have high CECR, which can keep enough Ca on cell walls
in Na rich soils.19 Woodward et al.20 observed that in natural plant communities, the
CECR of monocots was correlated to the cation exchange capacity of soil. According
to the authors, this might result from a competitive adaptation for the uptake of
Cation Exchange on Plant Roots Involving Aluminium 231
cation nutrients. Plants with low CECR should be handicapped for cation uptake in
soils with high CEC. Solubilization of sparingly soluble phosphorus (AlPO4, FePO4)
was hypothesized by Ae et al.21 to be related to the chelating ability of pectic
substances, i.e., to CECR; however, by comparing different plant species, these
authors demonstrated that the relationship between CECR and phosphorus solubi-
lization was not straightforward.
Tolerance to aluminium was tentatively related by many authors to CECR. Plants
with high CECR should be more sensitive to Al, because Al has high affinity for
exchange sites and impairs physical properties of roots.3,12,22 However, other mech-
anisms of Al toxicity, e.g., alteration of plasma membrane properties can also take
place,8,23-25 so that the relationship between CECR and Al tolerance is not obvious.26,27
Nevertheless, in a series of five banana cultivars (recent data, not published), we
observed that the most Al-sensitive one had CECR 30 cmolc kg1 compared to
CECR 19 to 23 cmolc kg1 for the other four cultivars. Recent studies on maize
showed that the CECR values of Al-sensitive and Al-tolerant cultivars were not
significantly different, although the total amounts of Al accumulated in roots were
greater for Al-sensitive than for Al-tolerant cultivars.11,28
and consequently to plant species. Figure 11.2 shows results obtained on roots
equilibrated in an acid soil suspension prepared by repeated washing of a mineral
soil sample with CaCl2-HCl at pH 4. Exchangeable Al expressed in moles of charge
(extracted using CuSO4) is higher than the CECR values at pH 4 (measured by
titration in 0.1 M CaCl2) and close to the CECR values measured at pH 6. At least
three reasons can explain this observation: (1) the affinity of Al for exchange sites
is higher than the affinity of Ca, and consequently the fixation of Al induces more
deprotonation of carboxylic groups than the fixation of Ca; (2) Al might be fixed
on exchange sites not only as monomeric Al3+, but also as less charged ionic forms
such as Al(OH)2+, Al(OH)2+; (3) Al extracted by CuSO4 might come not only from
exchange sites, but also from other forms such as Al-precipitates.11,31
Other experiments were done by equilibrating roots with solutions from differ-
ent soil horizons and different soil types. Furthermore, measurements of root
exchangeable cations were carried out after adding acid or lime to soil samples.
Figure 11.3 shows typical results obtained for three horizons of a forest acid soil
from the Belgian Ardens.
The fraction of sites occupied by Al depends on the activity and on the fraction
of free Al in solution. Roots equilibrated with solutions coming from organic soil
horizons do not fix much Al even at low pH. The reason is that Al activity in solutions
from organic horizons is not controlled by Al-bearing minerals, but by exchange
and chelating processes with organic compounds. In such conditions, the root
exchange sites compete with the exchange sites of organic matter for available
Al, and the solution is not replenished with Al coming from mineral dissolution as
Cation Exchange on Plant Roots Involving Aluminium 233
cmolc kg-1
cmolc kg-1
cmolc kg-1
FIGURE 11.3 Exchangeable cations of roots equilibrated with soil solutions from three
horizons (O, A, B, from top to bottom) of a forest acid soil (Vielsalm, Belgium).
cmolc kg-1
FIGURE 11.4 Exchangeable cations on banana roots in nutrient solution and in two soils
from Cameroon.
FIGURE 11.5 Exchangeable Al on maize roots and isolated cell walls as a function of pH.
(as assessed by CuSO4 extraction) was observed at pH 4.4. Below that value, Al
fixation decreased due to H-Al competition. This behavior is valid only for fixed Al
concentration in solution, keeping in mind that in mineral soils a decrease in pH is
normally accompanied by an increase in Al3+ activity.
Exchange experiments were also conducted in well-controlled synthetic solu-
tions in order to produce reference data that can be used to calibrate models. Roots
of clover and ryegrass were equilibrated with solutions at 2 Ca concentrations (0.5
and 5 mM), 2 pH values (3.5 and 4.0), and 4 Al concentrations (0, 25, 50, 100 M).
The experimental data were reported by Dufey et al.,16 and a comparison with
simulated values is presented in the next section of this chapter. Another series of
data was recently produced with banana roots (not published). Root equilibration
was carried out at 3 Ca concentrations (0.2, 1, and 5 mM), 3 Mg concentrations
(0.02, 0.1, and 0.5 mM), 3 K concentrations (0.2, 1, and 5 mM), 5 Al concentrations
(0, 10, 25, 50, and 100 M), and at 2 pHs (3.75 and 4.25). The model parameters
obtained from this very extended range of solution composition are given below.
because of the variety of exchangers among soil constituents (including roots) and
because of the multiplicity of processes that influence soil solution composition.
Two approaches can be distinguished when dealing with ion exchange models: either
a macroscopic approach formally expressed as mass-action laws, or a molecular
approach expressed by double diffuse layer formulations.
The first mathematical formulation as mass-action law was issued in 1913 and
developed by Kerr in 1928.33,34 Simultaneously, Vanselow35 and Gapon,36 and later
Gaines and Thomas,37 proposed various expressions of the exchange constant for
binary cation exchange. If such thermodynamic treatment of cation exchange has
proven helpful in interpreting the exchange processes,38-40 it has unfortunately no
general predictive power. The cause for this is the lack of a general formulation for
the surface activity coefficients like the Debye-Hckel or Davies formulae for solu-
tion activity coefficients.34,41
Up to the beginning of the 1980s, few applications were made beyond binary
exchange.42-50 In many studies, multication exchange is treated as simultaneous
binary exchange,51-53 and most of the time, the studies were limited to model com-
pounds like smectitic clays or zeolites, but not often extended to soils.
Many authors were in search of constants for characterizing cation
exchange.32,34,54,55 But it was soon recognized that the exchange constants generally
vary with ionic strength and pH, and with the composition of the surface
phase.34,36,37,49,50,56,57 Indeed even a given clay mineral may have distinct exchange
sites and cannot be considered as a homogeneous exchanger.38,58-59 A fortiori a soil
system is a more complex mixture of various exchangers that show a different affinity
for ions.36,60-63 Furthermore, the charge itself of a series of exchangers varies with
pH, edge sites of clay minerals, oxihydroxides of Al, Fe, etc., and organic exchangers,
including root cell walls. The mass-action law alone is not sufficient to describe the
behavior of such variably charged materials.
Cation exchange may also be described in terms of statistical mechanics as
Helmholtz and Gou and Chapman did with the diffuse double layer (DDL) model.
A Stern layer may be added to the DDL model for taking into account the real size
of ions and possible specific interactions with the exchanger surface. The DDL
approach as been widely developed and applied to clay minerals,64-67 and also to
biomembranes.23,68-70 In particular, Kinraide et al. (1992) clearly demonstrated that
phytotoxicity is not related to concentrations, but to ion activity close to the plasma
membranes computed according to the DDL model.23
When dealing with exchangers such as soil organic matter or roots, a major
limitation of this model resides in the fact that DDL equations are derived for well-
defined geometry; planar surfaces received more attention than cylindrical or spher-
ical geometry, because the reference exchangers in soils have been clay minerals in
most studies. Exchange sites in roots are not confined in a simple geometrical frame,
a difficulty that is avoided by Donnan-type models. Indeed, the Donnan theory
considers mean electrostatic effects in a phase or a compartment that includes a
charged substance that cannot diffuse throughout the system due to the existence of
a barrier, be it physical (semipermeable membrane) or energetic (charged surface).
Furthermore Donnan-based models, like DDL models, can include, in addition to
electrostatic effects, specific interactions between the fixed charge and the counte-
Cation Exchange on Plant Roots Involving Aluminium 237
1. Chemical Speciation
system of equations. In this chapter we will not develop the equations used for
solving the chemical speciation, because this part of the model is well covered in
other papers.86
2. Ion Exchange
Thus exchangeable ions, either bound or in the free space, are not expressed on the
basis of exchanger weight as molc kg1, but on the basis of the Donnan phase volume
as molc dm3.
a. General Equation for Ion Exchange
Definitions:
The subscript D refers to the Donnan free space, whereas no subscript refers to
the bulk solution, i.e., the solution that remains unaffected by the electrical potential.
The relationship between the activity of any ion in the Donnan free space, (XDi),
and its activity in the bulk solution, (Xi), is given by the Maxwell-Boltzmann
equation:
z F
( X Di ) = ( X i ) exp i D (1)
RT
or, for the sake of simplification, if we define a new variable, r, function of the
Donnan potential D as follows
F D
r = exp (2)
RT
( X Di ) = ( X i )r Z i (3)
The relationship between surface-bound ions and the Donnan free space is
expressed by dissociation reactions between the cations and multivalent exchange
sites, R(zi), of the same valence as the cations considered:
R zi X i R (zzi i ) + X D
240 Trace Elements in the Rhizosphere
All the exchange sites are supposed to be able to cluster, with the result that the
concentration in such multivalent sites is equal to the CECR divided by cation
valence. Therefore, the concentration in dissociated multivalent groups is calculated
as follows:
[R ]
[R (zzi i ) ] = (4)
zi
[R ]( X Di )
K dint,i = (5)
z i [ R zi X i ]
Equations (3) and (5) are the basic equations of the Sentenac and Grignon73
model for cell walls.
Equation (5) implies that only neutral surface complexes are formed. Here we
introduce an extension to the original model. Charged surface complexes occur when
all the charges of a multivalent ion do not bind to an equivalent number of reactive
groups. This has been reported by Baes and Bloom87 for peat, and by Miyajima et
al.71 in linear and cross-linked polymers. Tipping63 also considers monodentate
complexes with polyvalent cations, and bidentate complexes with trivalent cations.
The formation of charged surface complexes is illustrated by the following reaction
with aluminium, where only two of the three positive charges are bound to negatively
charged surface groups:
R 2 + Al 3+ {R = Al}
[R ]( X Di )
K dint,i = (6)
l i [R li X i ]
Cations
[R]Tot = [R ] + l [R i li Xi ] (7)
Anions
[R ] + | z | [A i Di ]=
(8)
Cations Cations
(z l )[R
zi li
i i li Xi ] + z [C i Di ]
By grouping the surface charges on the left-hand side and the charges in the Donnan
solution on the right-hand side, Equation (8) may be condensed to
Cations Ions
[R ] (z l )[R
zi li
i i li Xi ] = z [X i Di ] (9)
Combining Equations (6) and (9), factoring by [R], and introducing the expression
of [R] in Equation (7) results in
Cations
(C Di )
1 +
Ions K dint,i
[R]Tot =
z i [X Di ] Cations
(10)
1 (C )
(z i l i ) Did
l i K int,i
zi li
where I is the ionic strength and b 0.43 for humic acid. In their study of bacterial
cell walls, Plette et al.76 also used an inverse relationship between the Donnan volume
and ionic strength and found good agreement with other experimental data. The
most elaborate model for the Donnan volume is probably found in the WHAM
computer code for fulvic and humic acid particles.63 Briefly, the Donnan volume,
dependent on ionic strength, is computed from a spherical geometry with Debyes
length as radius. The volume is then restricted by a Michaelis-Menten-type function
of the dissociated charge of the organic particle. Only tentative values of the param-
eters have been obtained up to now.
242 Trace Elements in the Rhizosphere
In the present work, the Donnan volume was estimated for each set of data as
outlined below. Typical values reported for the Donnan volume of ion exchangers
are mentioned in Section C.1 and compared with the values found in our own
experiments.
TABLE 11.2
Analysis of Variance, Plant-Independent
Dissociation Constants
Source DFa Sum of Squares Mean Squares
TABLE 11.3
Analysis of Variance, Plant-Dependent Dissociation Constants
for Clover
Source DFa Sum of Squares Mean Squares
95% C.I.a
Parameter Estimate Standard Error Lower Upper
i.e., that the fit is better with plant-dependent dissociation constants. Figure 11.7
shows the whole set of experimental data and the fitted curves, with plant-dependent
dissociation constants.
The parameter estimates, standard errors, and 95% confidence intervals are given
in Tables 11.3 and 11.4. The estimated intrinsic dissociation constants for H are
close to the 103.36 deprotonation constant of galacturonic acid, a main component
of pectin, used by Sentenac and Grignon,73 and also close to the constants of some
humic acids sites of the WHAM model.63
The intrinsic dissociation constants for Ca are greater than the values estimated
by Sentenac and Grignon73 and used by Amory and Dufey.89 Furthermore, the
constant for ryegrass is greater than for clover. Thus, according to the statistical
estimation, the association between Ca ions and the root exchange sites is looser in
ryegrass than in clover. Indeed Whitehead et al.91 reported a higher proportion of
pectin in clover roots and of hemicellulose in ryegrass. Pectin consists of longer,
more organized chains of uronic acids than hemicellulose, and this may allow easier
and closer association with cations.
For Al, the most coherent results were obtained, for both plant species, with
surface complexes of the form R3-Al0 and no association of AlOH++ with the
exchanger. The intrinsic dissociation constant for Al+++ is 102.41 for clover, with a
244 Trace Elements in the Rhizosphere
TABLE 11.4
Analysis of Variance, Plant-Dependent Dissociation Constants
for Ryegrass
Source DFa Sum of Squares Mean Squares
95% C.I.a
Parameter Estimate Standard Error Lower Upper
95% confidence interval ranging from 103.67 to 101.14, and 101.95 for ryegrass, with
a 95% confidence interval ranging from 103.40 to 100.51.
The estimated CECR is 57 cmol kg1 for clover and 33 cmol kg1 for ryegrass.
These values are 25 to 60% higher than the CECR measured by Dufey and Braun9
from titration with Na, Ca, and Mg, and from copper adsorption, but they are very
close to the 63 and 31 cmol kg1 observed by Amory and Dufey89 from Ca adsorption
at pH 5.2.
The estimated Donnan volumes are listed in Table 11.3. The volumes are 4 to
5 times smaller for ryegrass than for clover. This is probably related to differences
in the cell wall structure of the plant species. For both plants the volume decreases
with increasing ionic strength. The smaller volume at pH 3.5, Ca 0.5 mM, as
compared to pH 4.0, Ca 0.5 mM, may be attributed to contracted Donnan space
resulting from higher protonation, hence lower electrical potential. A slightly smaller
potential (modulus) is indeed observed at pH 3.5 (not shown). As expected the
volumes are the smallest at the highest ionic strength, i.e., in the Ca 5 mM treatment.
The Donnan volumes estimated for the 0.5 mM Ca treatments (lower ionic
strength) range from 0.5 to 2.5 cm3g1. These values are in agreement with the
Donnan volumes reported by Sentenac and Grignon73 for root cell walls, by Benedetti
et al.75 for humic acid gels, or by Plette et al.76 for bacterial cell walls. Values even
Cation Exchange on Plant Roots Involving Aluminium 245
as high as the upper confidence limits for clover (9.1 and 12.4 cm3g1) have been
reported by Ritchie and Larkum92 for algal cell walls. Benedetti et al.75 obtained
even higher values for fulvic acid gels, from 10 to 80 cm3g1 at similar ionic strength
ranging from 0.010 to 0.001, but, according to the authors, the highest values might
be suspect. On the contrary, the volumes estimated at the higher ionic strength (5
mM Ca treatments) are very small, but nonetheless close to the 0.15 cm3g1 reported
by Benedetti et al.75 for humic acids, even if the latter was established at a far higher
ionic strength.
Table 11.5 presents the sensitivity of the model response to a variation of 20%
of each parameter. For clover, the sensitivity is the highest for the dissociation constant
for the proton, then for Al, and finally Ca, 4 times lower than that for the proton. The
sensitivity to Donnan volume is of the same order of magnitude as to the dissociation
246 Trace Elements in the Rhizosphere
TABLE 11.5
Sensitivity of the Model Response to
a 20% Variation of the Parameters
Mean Square Difference
Parameter Clover Ryegrass
constant of calcium. The dissociation constants for ryegrass rank in the same order
as for clover, but their influence is 3 times smaller for H and Al, and about 15 times
smaller for Ca. The model is rather insensitive to the dissociation constant of Ca for
ryegrass. The sensitivity to Donnan volume for ryegrass is smaller than for clover,
and more comparable to the sensitivity to the dissociation constants. The model is
most sensitive to the root cation exchange capacity, especially for clover. The root
cation exchange capacity is the most easily measured parameter of the model.
Estimation of the model parameters was also carried out for banana roots and for
maize root cell walls. The experimental conditions used for the measurements are
described briefly in Section III above. Table 11.6 presents the estimates of the
parameters from our work and from Sentenac and Grignon73 for horse beans and
lupin cell walls. In these studies, good agreement was found with experimental
data using high values of dissociation constants for monovalent ions, which shows
that the complexation of root exchange sites with these ions can be neglected in
modeling. They are confined mostly in the Donnan free phase. As for ryegrass and
clover, the intrinsic dissociation contants of protons found for banana and maize
are close to the value reported for horse beans and lupin, and all these values are
comparable with the deprotonation constant of galacturonic acid. The dissociation
constants estimated for Ca cover 2 orders of magnitude (100 to 102), which can
possibly reflect structural differences in root cell walls of the studied species. As
far as Al is concerned, satisfactory fits were obtained for clover, ryegrass, and
banana assuming only surface complexes with Al3+. However, no definitive conclu-
sion can be drawn concerning the existence of complexes with hydroxy-Al species,
because models are very poor tools for establishing detailed physicochemical mech-
anisms. Indeed, Calba et al.11 found that by considering surface complexes with
Cation Exchange on Plant Roots Involving Aluminium 247
TABLE 11.6
Intrinsic Dissociation Constants Estimated for Roots or Cell
Walls of Different Plant Species (log10 of values)
Horse Bean and Clover Ryegrass Maize Cell Banana
Complexes Lupin Cell Wallsa Rootsb Rootsb Wallsc Rootsb
Al(OH)2+, they were able to improve the agreement of model data with measured
exchangeable Al.
V. CONCLUSION
The data reported in this chapter show that exchange properties of roots must be
considered in studies dealing with cations that have high affinity for root cell walls.
Indeed, drastic changes in the chemical properties of the apoplast can result from
changes in the composition of root exchange sites. It is also shown that a model based
on intrinsic characteristics of rootion interactions can account for this composition
in a wide range of solution compositions. Further data are needed to improve the
reliability of the model parameters, although those available already allow one to
make reasonable simulations. The theoretical approach developed in this model is
immediately applicable to other trace elements without any supplementary algorithmic
work, because the number of ionic species can take any value in the computer code.
Understanding the functioning of the rhizosphere is not an easy task, mainly
because of considerable technical difficulties. However, in acid conditions, the
increasing solubility of Al and of most trace elements strengthens the need for
mechanistic approaches at the rhizosphere scale. Only limited studies have been
reported so far,93,94 and the development of numerical tools including kinetic aspects
is a promising research field.11,95 We might expect that such studies will lead to better
understanding of plant adaptation in problem soils and will suggest appropriate
cultural practices. Implementation of adequate countermeasures against toxic trace
elements and phytoremediation should also derive benefit from numerical tools
describing the soilroot interface.
ACKNOWLEDGMENTS
We thank the European Commission for supporting part of this work through the
project REPITISC contract EV5V-CT93, and the project INCO-DC contract CEE
CT 970208.
248 Trace Elements in the Rhizosphere
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12 Modeling the Dynamics
of the Rhizosphere
Aluminum Chemistry in
Acid Forest Soils
Heino W. F. Nietfeld
CONTENTS
I. Introduction...................................................................................................254
II. Development of the Model Approach..........................................................256
A. Model Assumptions and Hypotheses of Rhizosphere Processes.....257
B. Mathematical Model.........................................................................258
1. Ionic Flux Equations .........................................................259
2. Cation Exchange................................................................262
3. Transpiration-Induced Water Flux.....................................263
4. System of Mass Balance Equations ..................................263
5. Root Ion Uptake and H/OH Excretion..............................265
6. Boundary and Initial Conditions .......................................265
III. Materials and Methods .................................................................................266
A. Spectrum of Parameter Values Used in Model Calculations...........266
B. Numerical Solution Procedure .........................................................269
IV. Results...........................................................................................................270
A. Influence of Soil Conditions: Low NO3 Concentration...................270
1. Influence of Daily-Patterned Water Flux ..........................270
2. pH Changes at Various Bulk pH and Mb Cation
Concentrations ...................................................................273
3. Al3+ Concentration Changes at Low Bulk pH ..................277
4. Al3+ and A1-Hydroxo Complexes Concentration
Changes at High Bulk pH and Various Mb Cation
Concentrations ...................................................................278
B. Influence of Soil Conditions: Increasing NO3 Concentrations .......281
1. H+ and Aluminum Concentration at Fluctuating
Daily-Patterned Water Fluxes............................................281
2. pH Changes at Various Mb and Ma Cation
Concentrations and Bulk pH Values .................................286
3. Aluminum Concentration at Various Mb and Ma Cation
Concentrations and Different Bulk pH Values..................287
0-8493-1535-2/01/$0.00+$.50
2001 by CRC Press LLC 253
254 Trace Elements in the Rhizosphere
I. INTRODUCTION
Solution chemistry of acid soils of forest ecosystems with high concentrations of
acid salts (Ma-cations: Al3+, Mn2+, Fe3+) and protons, and low concentrations of
neutral salts (Mb-cations: Na+, K+, Ca2+, Mg2+)119 has been regarded as a stress
potential for forest tree development.82,120 A wide number of recent experimental
studies have demonstrated that aluminum (Al) has adverse effects on the growth,
root structure,20,115 and nutrient uptake31,61 of forest trees. But transferring the results
of experiments with Al-enriched solution cultures to field observations is problem-
atic, although root toxicity symptoms discovered in field observations are quite
similar to those examined in solution cultures.
First of all, solution culture experiments have shown that Al generates root
damage and plant growth reduction only at high Al concentrations. In many inves-
tigations, a maximum Al concentration threshold of about 500.0 mol L1 has been
found to be tolerable for growth of European beech and spruce.22,33,116 However, this
threshold is not actually reached in soil solutions or if so, only temporarily. These
results contrast with the results of field observations and growth chamber experiments
in which much lower Al solution concentrations caused root damages.70,120 Generally,
it can be established, that the distribution patterns of the degree of forest tree vitality
within forest stands deduced from crown damages are very heterogeneous and dam-
aged trees occur in the direct neighborhood to symptom-free trees.93 These distur-
bance patterns may be considered as evidence that stress-related and stress-free
chemical conditions can occur in close proximity to one another and may be
explained, among other things, by the spatial and temporal heterogeneity of the soil
chemistry98 and physiological and morphological differences of the fine root system.
Indeed, with about the same ionic soil solution composition, Al rhizotoxicity
must be assessed differently for mycorrhizal and nonmycorrhizal plants. Nonmyc-
orrhizal trees seem to be more susceptible to unfavorable acid soil conditions.
Cumming and Weinstein17 demonstrated clearly that nonmycorrhizal pitch pine
seedlings react more sensitively to Al (50.0 to 200.0 mol L1) than mycorrhizal
ones. But the risk of Al toxicity to mycorrhizal plants cannot be clearly determined,
because reports on the role of ectomycorrhizaes in the response of Al treatment have
often been contradictory. On the one hand, depression of the mycorrhizal develop-
ment was limited to declining trees.78 These observations are supported by results
of solution culture experiments showing a potential risk of toxicity to mycorrhizae,
e.g., high Al concentrations also caused root damage and reduced root growth of
mycorrhizal spruce seedlings.55 On the other hand, the higher tolerance of
Modeling the Dynamics of the Rhizosphere Aluminum Chemistry 255
Accumulations of Al3+ and SO42 around roots have been detected;24 consequently
the concentration of Al-sulfate complexes in the rhizosphere may be higher as in the
bulk soil. It is well known that due to the H+/OH root efflux, which is mainly
determined by imbalance of cation/anion uptake of all major nutrients,71 the rhizo-
sphere pH may change considerably.74 Root-induced pH increases in acid soils caused
by anion uptake excess, which is usually due to high NO3 uptake, may increase the
fraction of Al hydroxocomplexes59 in the total Al concentration. In the opposite case
at a higher bulk pH, a pH decrease may cause a transformation of Al hydroxocom-
plexes in the bulk soil solution to the Al3+ ions. Furthermore, the H+/OH root efflux
rate may change with time because the root surface concentration of cations and
anions involve changes19 and therefore, the equilibrium between hydroxo-Al species
may shift. These chemical transformations are also affected by the transport charac-
teristics of the ions involved. Al complexes have their own self-diffusion velocities,
which are different from those of the underlying dissociated ions.2 Moreover, the
cation exchange between soil solution and soil exchanger may considerably modify
the chemical reactions in soil solution. Root-induced soil solution depletions can
cause a release of nutrients from the soil exchanger58 or, more generally, root-induced
decreases of cations and shifts of proton concentrations may lead to a new equilibrium
between soil solution and exchanger composition. This may lead to an additional
buffering or a release of Al3+ ions from soil exchanger sites.97
The few direct measurements of Al in the rhizosphere also show depletions and
accumulations of Al,24 which may reflect the briefly outlined possible dynamics of
the Al chemistry in the rhizosphere. It may be concluded that in the rhizosphere
there may exist a dynamic equilibrium between the various Al complexes of different
risks of toxicity. In order to evaluate risks of Al rhizotoxicity originating from
complex, interrelated mechanisms, the coupled processes of the Al chemistry in the
rhizosphere must be described by appropriate modeling approaches.
It is obvious that this can only be achieved by an extension of the known one-
component rhizosphere models18,21 to a multicomponent model approach linking
transport mechanisms and simultaneous chemical reactions.12
In this theoretical study a multicomponent model is presented that includes non-
Fickian diffusion transport and cation adsorption as the competition process for the
exchanger sites. It is the purpose of this study to use model calculations to demon-
strate various scenarios of the dynamics of the Al species in the rhizosphere as
affected by root uptake of water and nutrients and the physicochemical conditions
of the bulk soil. In detail, model calculations are introduced based on various initial
concentrations of Mb cations, NO3, and Al at various bulk pH values, soil water
conditions, and different water and nutrient uptake rates. Taking into account the
heterogeneity of the absorbing root system,113 the calculations distinguish between
different uptake patterns and root diameters of the apical and basal zones of usually
nonmycorrhizal long roots26 and mycorrhizal short roots.63
B. MATHEMATICAL MODEL
It is assumed that each root, of radius r0, exerts its influence on a cylindrical volume
of soil (rhizosphere) starting from root surface with a radius of r1, i.e., the rhizosphere
is a hollow cylinder with a radius of r1 r0. The following equations are formulated
for a general set of species, = {M1, , Mn}, which includes m + 1 dissociated
(basic) cations and anions (species subsets cation and anion) that belong to 1, and
n-(m + 1) ion complexes that belong to the subset complex. Table 12.8 (Appendix)
shows a summary of the model parameters and variables.
Modeling the Dynamics of the Rhizosphere Aluminum Chemistry 259
The flux of each ion Mi is described by the Nernst Planck equation38 that is
extended by a water flux term and is defined by
c F
J i = Di i + zc q r ci [1]
r RT i i r
where Ji is the flux of Mi, Di = f1 Di0, its soil diffusion coefficient, Di0 its self
diffusion coefficient, ci its solution concentration, zi its charge, is the volumetric
water content, and f1 is the soil-impedance factor, which accounts for the tortuosity
of the diffusion path. is the diffusion potential, T is the absolute temperature, R
is the ideal gases constant, F is Faradays constant, qr is the transpiration-induced
water velocity toward the root, and r is the radial position in the rhizosphere.
Starting out from the assumption that no net flux of electrical charge (condition
of current density) occurs in the system, that is
z J
i =1
i i =0 [2]
n n
F c i
z c
ziDi qr
= [3a]
RT r n
r n i i
i =1
j=1
z j D jc j
2
z
j=1
j
2
D jc j i =1
n
c c k
z D
Dizici
J i = Di i + q r ci ; 1 i n [3b]
r n k k
r
j=1
z j2 c jD j k =1
Note that the second term in Equation [3b] represents a correction term due to the
existing Columb forces between the ions. Furthermore, the electroneutrality condition
allows one to express the concentration of, e.g., Mn as cn = n1
i=1 (z i /z n)c i ; extracting
cn/r from Equation [3b] and replacing it by the derivative of the expression above
yields, for each Mi \{Mn} in the modified flux expression
260 Trace Elements in the Rhizosphere
n 1
c j
Ji = D*
j=1
ij
r
q r ci ; 1 i n 1 [4a]
where
and
zici Di
fij = n (D jz j D n z j ) [4c]
z
k =1
k
2
Dkck
and ij is the Kronecker symbol. Equations [4a], [4b], and [4c] show that the flux
of each ion is coupled with the concentration gradients of all ions.
For each ion complex Mi complex there exist ions M1, Mk 1 with
a il M l zl + a ik M k z k M i zi . The reaction is characterized by an equilibrium constant
Ki, which may be written according to the law of mass action
c Mi
Ki = [5a]
c Ml a il
c M k a ik
In this equation c Ml , c M k are the concentrations of the basic ions, c Mi the concen-
tration of the ion complex, and ail, aik are the stoechiometric coefficients.
A possibility for treating chemical ion complex reactions is by differentiating
[5a] with respect to spatial coordinates, by solving the equations thus obtained, and
by substituting the expression for these derivatives into the flux equations. The result
is what Rubin103 called the the partial differential equations option; e.g., using
Equation [5a] the differentiation of an ion complex Mi complex yields
c Mi c M k c Ml
= K i a ik c Ml a il c M k a ik 1 + a il c M k a ik c Ml a il 1 [5b]
r r r
Thus, numbering the n 1 ions from 1 to m for the independent basic ions and from
m + 1 to n 1 for the dependent ion complexes the flux Equation [4a] may be
expressed by
n 1
c c j c j
m
J i = Dc i i +
r j=1
fij
r
+
j= m +1
fij
r
q r ci
[5c]
m
c j
= D* r q c ;
j=1
ij r i 1 i n 1
Modeling the Dynamics of the Rhizosphere Aluminum Chemistry 261
which is forced by the weighted gradients of the independent basic ions as well as
by the ion pairs, where
n 1
c
D ij *: = ij D i fij +
k = m +1
fik k
c j
[5d]
for i = 1, , m and
n 1
c i c
D ij *: = D i fij +
c j
k = m +1
fik k
c j
[5e]
K W = c H c OH [6a]
where pKw = 14.069 and which may be termed here as conditional ion product because
its definition is based on concentrations instead of activities. The protolysis of H2CO3,
expressed by the equilibrium reaction H+ + HCO3 H2CO3, is according to
Equation [5a]
c H2CO3
KS = [6b]
c H c HCO3
where pKS = 6.46.112 The equilibrium of vapor CO2 and H2CO3* = CO2(aq) + H2CO3
in the soil solution, CO2(g) + H2O H2CO3*, is described by
where KH = 3.47 107 MPa1 is the Henry constant and pCO2 is the CO2 partial
pressure.112
By setting pCO2 equal to a constant, the relation between H+ and HCO3 is
given by
1
c H c HCO3 = K = K H pCO 2 [6d]
KS
262 Trace Elements in the Rhizosphere
Hence c HCO3 and cOH are considered as a function of cH. The relations [6a] and [6d]
are used to eliminate all concentration gradients of HCO3 and OH ions in the flux
Equations [5c]. This is realized by differentiation [6a] and [6d] with respect to spatial
coordinates and by substituting the expression for these derivatives into the flux
equations, e.g., cOH/r = (cOH/cH) cH/r.
Therefore, the elimination of the concentration gradients of the hydroxyl ion
and bicarbonate ion in flux Equation [5c] (assume M1 = H+, Mm1 = HCO3 and Mm
= OH) yields
c m 1 c c c c
D i*,m 1 + D i*,m m = D i*,m 1 m 1 + D i*,m m 1 [7a]
r r c1 c1 r
m 2
c j
Ji = D
i =1
ij
r
q r ci [7b]
where i = 1, , n 1 and
c m 1 c
D *ij D i*,m 1 + D i*,m m for j = 1
D ij = c1 c1 [7c]
D ij* elsewhere
2. CATION EXCHANGE
In the model the stoichiometric cation exchange reactions between the soil solution
and the soil exchange phase may be presented in accordance with thermodynamic
convention. If there are two cations Mi and Mj with charges zi and zj, the exchange
reactions can be described by
1 1 1 1
M + zi + M j X z j M j + z j + M i X zi [8a]
zi i zj zj zi
where X denotes one unit of charge of the cation exchanger with a fixed exchange
capacity CT.92 In analogy to the law of mass action one defines the selectivity
coefficient for the equation above11 as
c i1/ zi m j1/ z j
K Mseli / M j = [8b]
c j1/ z j m i1/ zi
Modeling the Dynamics of the Rhizosphere Aluminum Chemistry 263
n1
m
i =1
i =1 [8c]
r0
qr = q [9a]
r 0
where q0 is the constant water uptake rate of the root. Alternatively, according to
the diurnal cycle of transpiration, a cyclic water uptake rate may be defined,
where a = 0, 24, 48, and q0max is the maximum water uptake rate and t is the time
in hours. The time-dependent mass flow term defines a water flow period of 16 hours
and a subsequent 8-hour diffusion period.
m 2
c i 1 c j c
t
=
r r j=1
r D ij
r
+ q 0 r0 i ;
r
1 i n 1 [10a]
where ci is the concentration of the ion Mi in a unit of soil volume, i.e., for basic
cations ci = ci + ci and for both basic anions and ion complexes ci = ci, where
ci is the soil exchanger concentration, is the soil bulk density and is the volumetric
water content, and the other parameters are the same as in the equations above.
The mass balance of an ion complex, Mi, is expressed in gradients of the basic
ions (see Equation [5b]).
264 Trace Elements in the Rhizosphere
c i c c c c k
= i l + i =
t
tc t c k t
[10b]
m 2
1 c j c c c c k
r r j=1
r D ij
r
+ q 0 r0 i l + i ;
c l r c k r
m +1 i n 1
ci = Fi (c1 , c 2 , , c n1 ); i = 1, 2, , n1 [10c]
Applications of [10a] under consideration of Equation [10c] gives the mass balance
equation for basic cations
n1
c i c c j
t
= i +
t g j=1
ij
t
=
[10d]
m 2
1 c j
r r j=1
r D ij
r
+ q 0 r0 c i ;
1 i n1
where
ci Fi
g ij = = [10e]
c j c j
c( t, r0 ) c min
J = J max [11a]
K + (c( t, r0 ) c min )
where Jimax is the maximum influx rate, ci(r0,t) the solution concentration at the root
surface, cmin the minimum concentration below which no influx occurs, and K the
MichaelisMenten affinity constant. For Al ions, it is JAl = 0.
Based on these definitions, a cation-anion balance of the uptake rates of various
ions can be calculated. Let C = and A =
M i cation \{H}z i J i M i anion \{OH , HCO3} | z i | J i
be the sum of cation and anion fluxes across the root surface. As a result, the fluxes
of the H+ or OH-ions at the root surface are defined as
(C A), if C A > 0
J H =
0 elsewhere
[11b]
( A C), if A C > 0
J OH =
0 elsewhere
c Mi ( r, t ) = c i ; 1 i n; t = 0
[12a]
cMi ( r, t ) = c i ; 1 i n1 ; t = 0
where c i and c i are the initial concentrations in soil solution and soil exchanger,
which are constant for the entire rhizosphere soil. These initial concentrations are
consistent with the equilibrium relations [5a], [6a], [6d], and [8b].
If no competition between neighboring roots is assumed the concentration at the
outer boundary remains on initial concentration, i.e.,
c M i ( r, t ) = c i ; i = 1, , n [12b]
r = r1
Ji r = r0
= J i ; i = 1, , n [12c]
266 Trace Elements in the Rhizosphere
where Ji is the parameter expression defined according to [11a] and [11b]. The
defined boundary conditions guarantee the electroneutrality.
TABLE 12.1
Self Diffusion Coefficients
D0 D0
Ion [10 m2s1]
9 Ion [10 m2s1]
9
Cl 2.03 b H + 9.34b
NO3 1.92b Na+ 1.33b
SO42 1.07b K+ 1.98b
OH 5.5a Mg2+ 0.705b
H2PO4 0.846a Ca2+ 0.793b
Al3+ 0.559a NH4+ 1.98b
Fe3+ 0.846a Mn2+ 0.688a
AlSO4+ 1.5c Al(OH)2+ 1.0c
Al(OH)2+ 1.5c HCO3 1.33a
a Li and Gregory.68
b Nye.86
c According to charge (cf Lasaga66).
Modeling the Dynamics of the Rhizosphere Aluminum Chemistry 267
TABLE 12.2
Equilibrium Constants of Ion Complexes
Ion Complex pK
H2CO3 6.45
AlSO4+ 3.2
AlOH2+ 5.02
Al(OH)2+ 9.3
pKsel values used in the calculations correspond to the range of pK values determined
by Prenzel and Schulte-Bisping.92
The assumed range of volumetric soil water content values () of 0.225 to 0.4
corresponds to a range of water suction values of 100 kPa to 10 kPa (Beese, p.
13 ff).6 The calculations use values of 0.225, 0.3, and 0.4 that imply soil impedance
factors (f1) of 0.0325, 0.08, and 0.22, respectively, (for calculations see Beese, p.
36 ff).6 The soil density () is assumed to be 1.19 Mg m3.
Table 12.6 shows assumed diameter values of the various root zones.
TABLE 12.3
Initial Concentrations in Soil Solution
Concentration in Soil Solution
[mmol L1]
Case I Case II Case III Case IV
Ion A B A B A B
TABLE 12.4
Initial Concentrations on Soil Exchanger
Concentrations on Soil Exchanger
[mmolc kg1]
Case I Case II Case III Case IV
Ion A B A B A B
TABLE 12.5
Selectivity Coefficients
Soil Chemical Cases
sel
Case I Case II Case III Case IV
pK M / M
i j A B A B A B
TABLE 12.6
Diameter Values of Root Absorption Zones
Root Diameter Values [mm]
Range of Values Used
Root Absorption Zone Measurements in Simulations
TABLE 12.7
Potential Uptake Rates (J max-Values)
Jmax-Values
[105 mmol m2s1]
Ion Apical Zone Basal Zone Mycorrhizae
Potential uptake rates ( Jmax-values) of the three different root absorption zones
have to be established (Table 12.7). The MichaelisMenten constant is 103 mmol
L1 and the cmin value is set to 2.0 103 mmol L1; both values are assumed to be
the same for all ions.
Calculations with diurnal-patterned water flux (Equation [9b]) use values for the
maximum water uptake rate, q0max, of up to 7.5 105 L m2 s1 (apical zone of long
roots). For the subsequent basal zone a lower value of 2.5 105 L m2 s1 is assumed.73
For mycorrhizal root zones the model calculations assume much lower values of 1.0
105 L m2 s1. For calculations with constant water flux, unchanged during the
whole simulation period (Equation [9a]), q0 is defined as 2.5 105 L m2 s1.
alternative approach, split operator technique,79 is used here, which keeps the alge-
braic equations that define the chemistry separate from the differential equations.
Therefore, a solution is obtained by iterating between the two sets of equations:
solving in turn the physical transport and chemical reactions until the two results
are consistent. The Galerkin method has been applied for solving the transport
equations using the piece-wise linear hat-functions. The resulted set of ordinary
differential equations is converted into a set of nonlinear algebraic equations with
the aid of the Crank-Nicolson approximations for the unknown variables and their
derivatives.1 A linearization of the algebraic equation systems has been achieved by
means of the predictor-corrector technique. Based on the total concentrations, the
concentrations of the individual species are calculated separately by a Newton
iteration algorithm.111 A final solution at each time step is obtained by iterations
between the two sets of equations until the results are consistent.
IV. RESULTS
The following scenarios have been introduced based on various soil physical (vari-
ation of soil water content) and various soil chemical conditions (various bulk
concentrations of Mb and Ma cations and NO3 and different pHb values) for different
transport conditions (e.g., for diffusion transport only, sinusoidal water fluxes (Equa-
tion [9b]) or constant water flux unchanged during the whole simulation period
(Equation [9a])) and for various root absorption rates of nutrients. The distribution
dynamics of H+ and Al ions concentrations are presented as radial concentration
patterns in the rhizosphere soil and, alternatively, temporarily as concentration devel-
opment on the root surface. If not explicitly stated, the calculation examples are
based on the potential ion uptake rates of the apical root zone, its diameter value r0
= 0.5 min, and a soil water content of = 0.3.
This calculation example shows the concentration changes of Al3+ and H+ ions based
on the soil chemical case I-A as influenced by daily-patterned water fluxes defined
according to Equation [9b]. The intensity of the maximum water flux rate, q0max,
determines the accumulation of Al3+ at the root surface. Figure 12.1A shows that,
depending on the particular determined maximum water flux, the Al3+ concentrations
reach an accumulation level between 0.4 mmol L1 (q0max = 2.5 105 Lm2s1) and
1.4 mmol L1 (q0max = 7.5 105 Lm2s1); also a slight increase to 0.22 mmol L1 is
calculated for diffusion transport only. Increases and decreases of Al3+ soil solution
concentration on the root surface are calculated according to the fluctuating water
fluxes (Figure 12.1B). Parallel to an increase in the soil solution, an increase in the
Al3+ equivalent fraction on the soil exchanger is observed (Figure 12.1C) with
Modeling the Dynamics of the Rhizosphere Aluminum Chemistry 271
FIGURE 12.1A Distribution of Al3+ ions depending on intensity of daily-patterned water flux
rates toward the root (soil chemical case I-A, t = 4.66d).
FIGURE 12.1B Temporal dynamics of Al3+ root surface concentration depending on intensity
of water flux rates (soil chemical case I-A).
272 Trace Elements in the Rhizosphere
FIGURE 12.1C Distribution of equivalent fraction of Al3+ ions on the soil exchanger depend-
ing on intensity of daily-patterned water flux rates (soil chemical case I-A, t = 4.66d).
increasing water flux rates. Cations that are taken up in high quantities by the root
(Ca2+, Mg2+, NH4+) leave the soil exchanger sites. A new equilibrium between soil
solution and soil exchanger is established, so that vacant soil exchanger sites are
occupied by cations that have an accumulation or low depletion in soil solution
(Al3+, Mn2+, Fe3+, K+, Na+) caused by little or no root uptake, or in the case of H+,
by root efflux (Figure 12.2B). The soil exchanger sites previously covered by nutrient
cations are occupied mainly by Al3+ ions, for the most part near the root surface
because of the high affinity of Al3+ ions for the soil exchanger sites.
For diffusion transport alone there is already a steep pH decrease; calculations
with cyclic water flux show that H+ ions additionally accumulate on the root surface
during the water flux periods, and particularly encompass a larger spatial area around
the root (Figure 12.2A). This is caused, among other things, by the root H+ excretion
rates. Figure 12.2B shows the temporal development of the root H+ excretion rates,
depending on the water fluxes. The course of the curves reflects the actual transport-
influenced ion uptake rates. First, in all cases the H+ excretion rates decrease with
increasing depletion of highly absorbed cations. Second, rising water fluxes cause,
in this case, rising cation uptake rates and consequently an excess of H+ ions.
Together with the convective H+ ion input, a cyclic pH pattern is created (not
demonstrated; compare Figure 12.1B), which is characterized by additional pH
decreases during water flux periods and increases (with respect to the previous pH
level) during the subsequent diffusion periods. Increases in soil solution correspond
to decreases in the H+ exchanger concentration and vice versa. The occupation of
soil exchanger sites by H+ ions is highest with diffusion transport only (Figure
12.2C). With water transport competing cations, particularly Al3+ ions (Figure
12.1C), acting in the simulation course previously adsorbed protons to be desorbed
Modeling the Dynamics of the Rhizosphere Aluminum Chemistry 273
again during the water flux periods. Hence, for water flux periods an input of protons
into the rhizosphere soil solution consists of the influxes caused by root excretion
and convective flux and of H+ desorption from the soil exchanger.
The following calculations have been introduced with a constant water flux rate
unchanged during the whole simulation period (q0 = 2.5 105 L m2 s1). The
simulation results represent the distribution of H+ ions and Al components at various
soil chemical bulk conditions (soil chemical cases I, III, and IV) at low and high
pHb (subclasses A and B).
FIGURE 12.2B Temporal development of the H+ root excretion rates depending on intensity
of water flux rates (soil chemical case I-A).
FIGURE 12.2C Distribution of the equivalent fraction of H+ ions on the soil exchanger
depending on intensity of water flux rates (soil chemical case I-A, t = 4.66d).
Modeling the Dynamics of the Rhizosphere Aluminum Chemistry 275
High pHb
Low pHb
on the soil exchanger (Figures 12.3B and 12.3C). At pHb 4.0 the accumulated H+
ions on the soil exchanger distribute over a large space (Figure 12.3B) while at pHb
4.9 the soil exchanger just around the root, to a maximum of 2.0 mm distance from
the root surface, is covered though with higher amounts of H+ ions (Figure 12.3C).
FIGURE 12.3D Temporal development of the H+ root excretion rates depending on Mb cation
concentrations and initial pH values (soil chemical cases I-A (low Mb cation concentration,
pHb = 4.01); III-A (medium Mb cation concentration, pHb = 4.0); IV-A (high Mb cation
concentration, pHb = 4.0); III-B (high Mb cation concentration, pHb = 4.91); q0 = 2.5 105
Lm2s1).
In the soil cases III-A and IV-A the Al3+ soil exchanger accumulation is higher just
around the root compared with the cases III-B and IV-B but distribute over a smaller
space; in cases III-B and IV-B the Al3+ soil exchanger increases cover a larger space
(Figure 12.4C and Figure 12.5C).
Figure 12.4A shows that the water-flux induced Al3+ accumulation is modified
by the availability of Mb cations. High uptake rates of Mb cations produce depletions
in soil solutions as well as on the soil exchanger. Consequently, high bulk concen-
trations of Mb cations on the soil exchanger cause high amounts of free exchanger
sites. These exchanger sites are occupied, among other cations, by Al3+ ions producing
a lower concentration increase of Al3+ ions in soil solution. In contrast, small amounts
of Mb cations on the soil exchanger produce only small free exchanger sites during
the root uptake process and therefore have a small buffering-effect of Al3+ ions, e.g.,
a higher amount of the accumulated Al3+ ions remain in the soil solution.
But under these soil chemical conditions, in which the amount of cations taken
up by the root equals nearly the amount of excreted H+ ions, the competition between
H+ and Al3+ ions for the free exchanger sites defined by the pKsel values determines
the distribution of Al3+ and H+ ions in soil solution and exchanger. Documented by
the pKsel values in all soil chemical A-cases there is a high affinity of Al3+ ions for
the soil exchanger sites compared with H+ ions. In case IV-A the assumed higher
278 Trace Elements in the Rhizosphere
affinity of Al3+ ions (in relation to H+ and compared with cases I-A and III-A) for
the soil exchanger sites leads to a higher exchanger phase concentration and a low
Al3+ solution accumulation just around the root only. This is also documented at the
clear concentration decrease at diffusion transport only compared with, e.g., case
I-A (Figure 12.4B).
Although the initial Al3+ concentrations are equal in all bulk soil concentrations
of subcases A, the temporal development of the accumulation levels on the root
surface are different (Figure 12.4B).
FIGURE 12.4B Temporal distribution of Al3+ ions at the root surface at various high Mb
cation concentrations and convective and diffusive transport (pHb 4.0; soil chemical cases
I-A, III-A, IV-A; q0 = 2.5 105 Lm2s1).
FIGURE 12.4C Distribution of equivalent fraction of Al3+ ions on soil exchanger at various
Mb cation concentrations (pHb 4.0; soil chemical cases I-A, III-A, IV-A; q0max = 2.5
105 Lm2s1).
280 Trace Elements in the Rhizosphere
The following calculation results are based on soil chemical case I-A but progres-
sively replace Cl concentration by an increasing NO3 concentration; q0max; a daily-
patterned water flux with a q0max = 7.5 105 Lm2s1 is used.
Due to its high root uptake rate, increasing NO3 availability causes a reduced
H or an increased OH root excretion rate (Figure 12.6C), resulting in a smaller
+
FIGURE 12.5C Distribution of the equivalent fraction of Al3+ ions on the soil exchanger at
various Mb cation concentrations and high initial pH (pHb 4.9; soil chemical cases I-B, III-
B, IV-B; q0 = 2.5 105 Lm2s1).
excretion and negative for OH root excretion). Increasing NO3 availability shows
reduced or increased OH root excretion rates. It is seen that during the water flux
periods increased OH excretion and higher H+ excretion rates are calculated.
At increasing NO3 availability, a decreasing Al3+ ion accumulation level at maximum
water fluxes is calculated; Figure 12.7A shows a reduction in the Al3+ ion root surface
concentration from 0.95 mmol L1 (NO3 = 0.25 mmol L1) to 0.55 mmol L1 (NO3
= 0.75 mmol L1) on the root surface. This is due to the increased buffering of Al3+
ions by the soil exchanger; an increased amount of soil solution Al3+ occupies the
soil exchanger sites, which becomes free through the desorption of nutritional cations
and in this situation, additionally due to desorption of H+ ions from the soil
exchanger.
But at the highest NO3 concentration (1.0 mmol L1) the concentration of
previously accumulated Al3+ on the soil exchanger decreases again (Figure 12.7B).
At the given NO3 concentration, the initial high H+ desorption rate from the soil
exchanger at the beginning of the calculation process continuously decreases during
the course of the process. Therefore, the continuous pH increases as a response of
the root-released OH ions causing a desorption of protons from the soil exchanger.
Consequently, at the highest NO3 concentration, the resulting high pH increase
Modeling the Dynamics of the Rhizosphere Aluminum Chemistry 283
(Figure 12.6A) causes a change in the chemical bonding form of the Al3+ ions. An
increasing amount of solution Al3+ ions is transformed into hydroxo-Al complexes
by their convective transport toward the root. Thus the accumulated Al ions at the
root surface are not only present as Al3+ ions, but rather to a larger extent, as Al-
hydroxo complexes.
Furthermore, this continuous reduction of Al3+ ion soil solution concentration
causes a desorption of previously adsorbed Al3+ ions just around the root as a result
of the cation competition for the soil exchanger sites. Also, this input of desorbed
Al3+ ions is transformed into hydroxo-Al complexes and is transported via retrodif-
fusion.
But as the pH in the rhizosphere has a clear spatial gradient, so are the Al
hydroxocomplexes partially transformed back to dissociated Al3+ ions; consequently,
the Al3+ ions are partially adsorbed by the soil exchanger sites. The spatial concen-
tration distribution of Al3+ on the soil exchanger in Figure 12.7B shows the described
Al3+ depletion just around the root as well as a slight increase of Al soil exchanger
fraction behind this small depletion zone.
The following calculations have been introduced with a constant water flux rate
(q0 = 2.5 105 L m2 s1). The simulation results represent the distribution of H+
ion and Al components at various soil chemical bulk conditions at low pHb (soil
chemical cases I-A, II, III-A and IV-A).
284 Trace Elements in the Rhizosphere
FIGURE 12.6B Distribution of equivalent fraction of H+ ions on the soil exchanger depending
on NO3 concentration (soil chemical case I-A, t = 3.54d; q0max = 7.5 105 Lm2s1; compare
Figure 12.6A).
FIGURE 12.6C Temporal development of the H+ (positive values) and OH (negative values)
root excretion rates depending on the NO3 concentration (soil chemical case I-A; q0max = 7.5
105 Lm2s1).
Modeling the Dynamics of the Rhizosphere Aluminum Chemistry 285
FIGURE 12.7B Temporal dynamics of Al3+, hydroxo-Al complexes, and AlTotal on the root
surface at an NO3 concentration of 1.0 mmol L1 (soil chemical case I-A, q0max = 7.5 105L2s1).
286 Trace Elements in the Rhizosphere
Figure 12.8A shows that high nitrate concentrations at various soil chemical bulk
conditions cause various pH increases (Figure 12.8A and 12.8B). First of all the
different NO3 concentrations produce various rates of OH excretions. At the nitrate
concentration values of 1.125 mmol L1 and 1.4 mmol L1 in the soil chemical case
I (low Mb cation concentration) and II (high Ma cation concentration), respectively,
the highest OH rates are calculated (Figure 12.8C). Increased Mb cation availability
reduces the OH excretion rates also at high nitrate initial concentrations (cases III-A
and IV-A). At the beginning of calculation, even an H+ ion excretion at high Mb
cation availability is produced (case IV-A). Hence it can be stated that high NO3
concentrations alone do not necessarily produce an OH excretion (e.g., in case IV-A
an NO3 concentration of 1.0 mmol L1 causes an H+ excretion during the whole
calculation period; not documented) but the changing uptake ratio of anions, in
particular of NO3, and cations determine the amount of excreted H+/OH ions
(Figure 12.8C).
Second, the calculated root surface pH resulting from OH root excretion has a
clear temporal pattern (Figure 12.8B). The first partly drastic pH increase in all soil
Modeling the Dynamics of the Rhizosphere Aluminum Chemistry 287
chemical cases is caused by the initial adsorption on the soil exchanger, e.g., also
at the initial H+ root excretion in case IV-A there is a pH increase. The subsequent
period is characterized by the transport-influenced formation of gradients of all ions
involved. During this period a continuous increase in pH is observed. Depletion of
H+ ions in soil solution due to the neutralization by root excreted OH ions leads to
a desorption of protons from the soil exchanger (Figure 12.8D). In case II the
depletion of H+ ions on the soil exchanger includes a large spatial extent; this H+
ion input into the soil solution prevents a higher pH increase also at the highest OH
root excretion rate. Also in soil chemical case I-A the calculated highest pH increase
corresponds to a clear H depletion on the soil exchanger (Figures 12.8A and 12.8D).
there is nearly no significant accumulation of Al3+ on the root surface even at high
Al3+ bulk concentrations. The amount of Al ions transported via water flux toward
the root are lower as the adsorption rates on the soil exchanger and the transformation
rates of Al3+ into complexed Al bonding forms.
Figure 12.9A shows the gradients of Al3+ ions concentration on the root surface
at various soil chemical conditions (cases I-A, II, III-A, IV-A). As already observed
from the proton concentration distribution, the Al3+ concentration in soil solution in
all soil chemical cases drops immediately at the beginning of the uptake process.
This is due to the buffering effect of the soil exchanger. Also at high Ma cation
concentrations the Al3+ solution concentration decreases continuously within the
whole rhizosphere. These patterns are correlated with the development of the pH
values in the rhizosphere and particularly on the root surface (Figure 12.8A). At a
high H+ concentration on the soil exchanger (and a low occupation with Mb cations,
soil chemical case II) the occupation of exchanger sites, which became free due to
H+ desorption (Figure 12.8D), constitutes the only buffering effect other than the
release of Mb cations from the soil exchanger. In the soil chemical cases III-A
Modeling the Dynamics of the Rhizosphere Aluminum Chemistry 289
FIGURE 12.8D Distribution of equivalent fraction of H+ ions on the soil exchanger at various
concentrations of Mb and Ma cations and NO3 (t = 4.66d; soil chemical cases I-A (NO3 =
1.125 mmol L1), II (NO3 = 1.4 mmol L1), III-A (NO3 = 1.15 mmol L1), and IV-A (NO3
= 1.25 mmol L1); q0 = 2.5 105 Lm2s1).
the pH profile at low Mb cation concentrations). Also the desorbed Al3+ ions are
transformed into complexed Al bonding forms. The back transformation of Al
hydroxocomplexes into Al3+ ions, according to the pH gradient, causes an Al3+ ion
accumulation in the soil and solution exchanger.
The following calculations have been introduced with soil water contents of =
0.225 and = 0.4. From calculation results with one component models it is well
known that at low soil water content the diffusive flux per unit of conducting surface
is smaller. The diffusion velocity is reduced by a smaller soil impedance factor and
the contribution of the water flux to the total flux is higher. With high soil water
content there is a higher contribution of diffusion flux to the total transport rate.
Under dry soil conditions only steep concentration gradients and under wet condi-
tions even small gradients cause a diffusion transport.
Modeling the Dynamics of the Rhizosphere Aluminum Chemistry 291
FIGURE 12.9B Distribution of equivalent fraction of Al3+ ions on the soil exchanger at
various concentrations of Mb and Ma cations and NO3 (soil chemical cases I-A (NO3 = 1.125
mmol L1), II (NO3 = 1.4 mmol L1), III-A (NO3 = 1.15 mmol L1), and IV-A (NO3 = 1.25
mmol L1); t = 4.66d; q0 = 2.5 105 Lm2s1).
Figure 12.10A shows that wet soil conditions produce moderate Al3+ accumu-
lations even at high (constant) water flux rates of q0 = 5.0 105 L m2 s1 (compare
Figures 12.4A and 12.9A). Even at high Ma cation concentrations (soil case II) only
a moderate concentration increase is calculated and at high NO3 concentration a
depletion is calculated. At high Mb cation concentrations (cases IV-A and IV-B) the
concentration increase at low NO3 concentrations are low in both cases (compare
Figures 12.4A and 12.5A).
Figure 12.10B shows that already at moderate and constant water flux rates of
1.0 105 L m2 s1, an Al3+ accumulation level of up to about 1.0 mmol L1 has
been calculated (soil chemical case I-A; low NO3 concentration). At high Mb cation
concentrations the Al3+ accumulation pattern is slightly higher as calculated at =
0.3. But at a high NO3 concentration, even a high Al3+ bulk concentration (soil
chemical case II), decrease at diffusion and a very low water flux rate occur (q0 =
0.5 105 Lm2s1) due to the decreased buffering of the soil exchanger. The release
of H+ ions from the soil exchanger caused by root excreted OH ions is restricted
292 Trace Elements in the Rhizosphere
FIGURE 12.9C Distribution of various Al species in soil chemical case I-A (for details see
text).
to a smaller space just around the root. First, water fluxes of 1.0 105 L m2 s1
(and higher) produce Al increases also at high NO3 concentrations.
FIGURE 12.10A Distribution of Al3+ concentration depending on various soil chemical con-
ditions and high soil water content ( = 0.4) (soil chemical case II (low NO3 = 0.025 mmol
L1, high NO3 = 1.25 mmol L1), IV-A (NO3 = 0.025 mmol L1), and IV-B (NO3 = 0.025
mmol L1), t = 4.66d; q0 = 5.0 105 Lm2s1).
cations, which compete with Al3+ ions for the exchanger sites. Consequently, a higher
affinity of Mb cations for the soil exchanger sites as in Case IV-A (compared with
the other soil chemical cases) results in a considerably reduced fraction of Al3+ ions
on the soil exchanger and a higher concentration in the soil solution. At higher
potential root uptake rates of Mb cations these are exchanged by less competitive H+
ions, which results in a higher Al fraction on the soil exchanger. In the soil chemical
case IV-B protons have a higher affinity for the soil exchanger sites compared with
Mb cations. Consequently, a considerable increase of proton concentration (at low
NO3) caused by high nutrient uptake reduces buffering of Al3+ ions. At smaller
potential root uptake rates the Al3+ compete with the less competitive nutrient cations
for the exchanger sites; consequently, the Al exchanger fraction is higher.
Figure 12.11C shows that the zone-related differences in the potential ion uptake
rates (Table 12.7) of the defined root absorption zones and differences in root
diameters (Table 12.6) produce differences in Al3+ concentrations in the rhizosphere
soil solution. The calculations are based on daily-patterned water fluxes and show
the Al3+ accumulations at maximum water flux rate (for details see the caption for
Figure 12.11C). Generally, it can be established that larger root diameters at a given
water flux rate produce larger Al3+ accumulations in soil solution. Influences of the
root diameter are documented by various Al3+ accumulation levels around the apical
294 Trace Elements in the Rhizosphere
FIGURE 12.10B Distribution of Al3+ concentration depending on various soil chemical con-
ditions and low soil water content ( = 0.225) at soil chemical cases I-A (low Mb cation
concentration, NO3 = 0.025 mmol L1, q0 = 1.0 105L2s1), case II-A (high Ma cation
concentration, NO3 = 1.25 mmol L1), at medium (q0 = 1.0 105L2s1), and low (q0 = 0.5
105L2s1) water fluxes, case III-A (NO3 = 0.025 mmol L1, q0 = 1.0 105L2s1), and case
IV-A (high Ma cation concentration, NO3 = 0.025 mmol L1, q0 = 1.0 105L2s1); t = 4.66d.
root zone calculated at different diameter values. The Al3+ accumulation level around
mycorrhizal roots is lowest due to small root diameter and small water flux rate. In
contrast, the Al3+ accumulation around the apical root zone of small diameter is still
over 0.5 mmol L1. The basal zone of long roots takes a middle position.
demonstrated between Al3+, H+ ions, and nutrient cations (e.g., Figures 12.1C and
12.2C). This is attributed to the fact that the reaction between soil solution and soil
exchanger is treated as competitive cation exchange and to the consideration of the
diffusion potential that cannot be discussed here in detail. The importance of cation
exchange reactions in the rhizosphere is pointed out by Gobran et al.;30 the necessity
to consider the diffusion potential in multi-ion systems is supported by the results
of appropriate diffusion experiments in soil columns.98
The summarizing result of the simulations presented is that the aluminum
dynamics in the rhizosphere created by the root uptake activity as driving force
includes both clear depletions and drastic accumulations in the soil solution. The
establishing Al3+ solution concentration on the root surface is a result of source-sink
relationships, e.g., it depends on the amount of Al3+ ions transported via water flux
toward the root (Figures 12.1A, 12.4A, and 12.7A), the transformation rate of
hydroxo-Al complexes into Al3+ ions and vice versa (Figures 12.5A, 12.5B, 12.7C,
and 12.9A), and the amount of Al3+ ions buffered by the soil exchanger (Figures
12.4C, 12.5C, 12.7B, and 12.9B). It should be noted that high SO42 bulk concen-
trations additionally may lead to a fixation of accumulating Al3+ ions into sulfate-
Al complexes (in preparation). The extent of Al buffering is determined by the
amount of soil exchanger sites becoming free during the root uptake process and
296 Trace Elements in the Rhizosphere
FIGURE 12.11B Distribution of the equivalent fraction of Al3+ ions on the soil exchanger
depending on various potential root uptake rates (soil chemical cases I-A, III-A, and IV-A; t
= 4.66d; q0 = 2.5 105 Lm2s1).
the concentration of competing cations and their affinity for the soil exchanger sites.
Based on the ionic composition of the rhizosphere bulk soil, the intensity of the
water fluxes and the selectivity and heights of the potential uptake rates are the root-
induced driving forces that determine the ionic concentration changes near the root
surface. Simultaneously, they induce soil chemical reactions that determine the
concentration ratios of Al3+, H+, and nutrient cations in soil solution, usually distin-
guishable from the bulk soil. A high or low root-driven exchange rate of nutrient
cations for H+ ions at low NO3 availability may increase or decrease the buffered
amount of Al3+ ions because the change of nutrient cations for H+ ions is connected
to a change of the affinity for the soil exchanger sites. The transformation rate of
Al3+ ions into hydroxo-Al complexes is low at high Al buffering rates which are
caused by high H+ and Mb cation release rates (Figure 12.9A). On the other hand,
it may be predicted that soil chemical bulk conditions, characterized by a small
cation exchange capacity and a low occupation of the soil exchanger with H+ and
Mb cations, lead at low NO3 concentration to highest Al3+ accumulations (Figures
12.1A and 12.4A); the transformation of Al3+ ions into hydroxo-Al complexes at
high NO3 uptake rates is, in this case, the only quantitatively significant Al3+ ion sink.
Rhizosphere pH measurements have been introduced comprehensively and often
have been represented for various root zones. Shifts of cation/anion uptake ratio of
Modeling the Dynamics of the Rhizosphere Aluminum Chemistry 297
FIGURE 12.11C Distribution of Al3+ concentration around various root absorption zones of
various root diameter values at daily-patterned water fluxes (soil chemical case I-A; NO3 =
0.025 mmol L1; t = 4.66d; q0max = 7.5 105 Lm2s1 (apical root zone); q0max = 2.5 105
Lm2s1 (basal root zone); q0max = 1.0 105 Lm2s1 (mycorrhial root)).
different root zones and different rates of depletion and replenishment of various
cations and anions in the various zones, especially of NO3, are assumed to be the
main factors responsible for these pH changes27,28 (Figures 12.3A, 12.4A, 12.8A,
12.10A, and 12.10B). It is obvious that pH changes due to root excretion between
rhizosphere and bulk soil tend to be clearer the lower the H soil initial concentration
(compare 28 and 101). Hence in more acid soils, the pH changes reported are usually
smaller and do not extend over several pH units.102 In acid soils the pH shifts of
0.051.5 pH units have been observed.27,28,44,74 The measurements of Marth74 (bulk
soil pH 5.25; pH decrease at the rhizosphere of about 0.5 pH units) demonstrate the
small spatial extent of pH changes; in a 1.5 mm distance from the root surface the
pH drop is only 0.2 pH units and at 2.5 mm from the root surface no changes have
been observed which agree with the simulations (Figures 12.3A and 12.4A).
The pH increases a maximum of 1.4 pH units, calculated here at 1000.0 mol
L1 NO3 (Figure 12.8A). At low NO3 concentrations a considerable acidification,
also around the apical root zone, of 0.25 pH up to 1.35 pH units is calculated (Figures
12.3A and 12.4A) which, on the one hand, contrasts with the measurement results
of several reports27,71 that always show an alkalization around the root tip, but on
the other hand is supported by measurements of Marth74 showing a considerable
acidification around the root tips. Furthermore, Marths growth chamber experiments
298 Trace Elements in the Rhizosphere
with spruce seedlings reveal only very slight pH increases also at NO3 fertilization,
findings that match the simulation results (Figures 12.10A and 12.10B).
The temporal development of the root H+/OH excretion rate with its deceleration
(Figures 12.2B, 12.3B, 12.4D, 12.8C, and 12.10C) after a few hours reflects the
influence of transport upon root uptake rates. During the development of the deple-
tion zones of some ions their root surface concentration is changed, hence uptake
rate and H/OH excretion rate, thus confirming the predictions by Darrah.19
It is not only the quantity of H/OH root excretions that determines the rhizo-
sphere pH, but also the H initial concentration (Figures 12.8A and 12.8B), the
adsorption and desorption of protons on and from the soil exchanger (Figures 12.3C,
12.6C, and 12.8D), and the input of protons via water flux (Figures 12.10A and
12.10B). Even at high NO3 concentrations, high desorption rates of H+ ions from
the soil exchanger may lead to moderate pH increases only (based on compensation
by the increased OH root excretion). This is significant because in the upper layers
of acid soils with high proton concentrations on the exchanger (e.g., upper soil layer
of Solling plot)7 possible high OH root excretions may lead to only slight pH
increases. On the other hand at a higher initial pH, a high availability of Mb cations
can cause a drastic pH drop (Figure 12.3A), a situation which is comparable with
and has been observed in liming experiments.44
Al accumulations around the root in both soil solution and exchanger have been
measured also by Ohno,89 Hussling,44 Fritz and Knoche24 and Knoche,62 whose
measurements include all relevant ions on various forest sites. The X-ray measure-
ments of rhizosphere chemistry of spruce roots at the Solling site by Fritz and
Knoche,24 and Knoche62 showed accumulations and depletions of Al (not differen-
tiated into apical and basal root zone). Hussling,44 Hussling et al.,45 Fritz and
Knoche,24 and Knoche62 determined only slight concentration shifts around mycor-
rhizal roots for most ions.
The identification of the water flux rate as the driving force for Al3+ accumulation
is supported by measurements of Hamza and Aylmore.39 Their measurement results
identify Na+ accumulation at the root surface as a function of the intensity of
transpiration-induced water fluxes in the rhizosphere. They demonstrate that Na+
concentration increases continuously with time of transpiration as it is calculated
here for Al3+ accumulations. Just as the slight Na+ accumulation, they put down to
the high back diffusion rate, the high Al3+ accumulation can be put down to the
lower back diffusion rate. Furthermore, the water fluxes used in the calculations here
are higher than the ones measured by Hamza and Aylmore.39 Their measurement
results support the hypothetical conclusion that the high Al3+ accumulations are most
probably around long roots with highest values around the apical root zone. Thus
risk of Al3+ toxicity is highest to long roots. This is supported by the investigations
of spruce by Murach.81 Also at much lower initial Al solution concentration in the
rhizosphere toxicologically relevant accumulations are possible at high water flux
rates. Therefore, the inconsistency of the toxic effects of Al3+ between soil grown
and solution grown plants is explainable.116 The simulation results are also consistent
with the reports by Cumming and Weinstein.17 The discussed protection of mycor-
rhizal roots was deduced from the fact that the mycorrhizae act as a physical and
chemical barrier restricting uptake and transport of Al. This result is completed by
Modeling the Dynamics of the Rhizosphere Aluminum Chemistry 299
the model results demonstrating that smaller root diameter and smaller water flux
rates produce smaller Al accumulations at the root surface. It is well known that
mycorrhizal roots and their hyphaes represent an extension of the absorbing root
surface41 causing smaller ion depletions and accumulations around them. Since for
the majority of the investigation dates Al depletions around mycorrhizal roots have
been detected this gives support to the calculation results. Also decreases of water
conductivity of ectomycorrhizal roots67,110 may induce smaller water fluxes in the
surrounding soil hence smaller Al3+ accumulation at the root surface. Although
differences of the Al accumulation levels are confirmed by the previous measurement
results, a correlation between transpiration and root Al enrichment at their surface
and root Al toxicity symptoms have not yet been reported.
But while high water fluxes cause Al3+ accumulations to decrease with increasing
NO3 availability (Figure 12.7A) they may still be above the threshold of tolerance
formulated by Thornton et al.116 The risk of toxicity also at high NO3 availability
is supported by Godbold et al.,32 who determined root damages also at high NO3
concentrations. But very high NO3 concentrations may lead to high pH increases
and consequently to a transformation of Al3+ ions in both soil solution and soil
exchanger into hydroxo-Al complexes (Figures 12.7B, 12.9B, and 12.9C). Kinraide60
concluded the likelihood that the toxicity of Al hydroxocomplexes must be rejected.
In addition to Al accumulations, depletions of Al concentration in the rhizosphere
were also measured.24,108 Also Ohno89 measured a depletion of soil water extracted
Al and an accumulation of KCl-extractable Al around honeylocust roots that,
together with the accumulations around red oak roots, fit into the simulation results.
Changing preference for Al3+ in the rhizosphere soil, compared with the one in the
bulk soil,15 may change the level of Al3+ accumulation. High affinity of Al3+ ions for
soil exchanger sites may additionally reduce Al3+ solution concentration, also at high
initial concentrations. On the other hand, a lower affinity of Al3+ ions for the soil
exchanger sites may reduce Al3+ buffering and hence the level of solution Al3+ is
increased. Small Al3+ bulk concentrations (0.025-0.06 mmol L1) as in the soils used
in the experiments of Ljunstrm and Stjernquist70 could therefore lead to an accu-
mulation level at the root surface higher than the cited tolerance threshold causing
the reported root damage.
Furthermore, one of the most sensitive indicators is the soil water content. At
the given high water fluxes, the considerably increased Al3+ accumulations at low
water content and the considerably decreased Al3+ accumulations at high water
content are in agreement with general field observations. The occurrence of tree
decline symptoms are reduced at humid vegetation periods and, during dry summer
periods, increasing damage is observed in the tree crown as well as in the fine root
system (Figure 12.10B).75 But the results presented here offer a more differentiated
picture. During acidification pulses, due to high nitrification characterized by a large
release of H+, Al3+, and NO3 concentrations, regardless of whether they occupy
larger soil compartments14,76 (pH = 3.6 and c NO3 800.0 mol L1) or whether they
only appear in spots,65 the calculated Al3+ depletions illustrate reduced or no stress
for the tree roots if, in dry soil conditions, transpiration-induced water fluxes are
strongly reduced or zero. This limited condition seems to be valid because during
drought experiments using roof installations it could be determined that trees react
300 Trace Elements in the Rhizosphere
with decreasing to zero water uptake with increasing soil drought124 (Xu, personal
communication). If there is a surplus production of NO3 and consequently of protons
and Al3+ ions during rewetting periods subsequent to drought periods, the risk of
Al3+ toxicity is reduced also at high rhizosphere water fluxes.
But beyond the direct toxic effects of Al3+ to plants and roots, Al3+ concentrations
at nontoxic concentrations also have a negative impact on root uptake of Ca2+ and
Mg2+. In solution culture investigations the application of Al3+ in a nontoxic quantity
may cause reductions of the root uptake rate to about 50%,35 which is particularly
demonstrated by concentration increases of Mb cations in the applied solutions.23
Transferring these results to the chemical situation in the rhizosphere of soil-grown
plants may lead to accumulations of Mb cations on the root surface and the inten-
sification of the competitive situation of Al3+ ions for the soil exchanger sites (Figures
12.11A and 12.11B). Consequently, a firstly moderate Al3+ accumulation may
increase to toxic accumulation levels.
These outlined relationships also demonstrate a deficiency of the model in its
presented version. Modeling of the ion uptake process of the root merely according
the MichaelisMenten kinetics ignores the complex ion dynamics within the root
apoplast (Dufey et al., Chapter 11, this volume), e.g., the antagonism between Ca2+
and Mg2+ and the competitive or noncompetitive effects of Al3+.23,31 An extended
model version should clearly distinguish between physical (passive) and physiolog-
ical (active) uptake mechanisms. One way to achieve this is to extend the reaction
range of the rhizosphere by the root apoplast, that is coupling of two porous media
with different properties. In addition, an improvement of the model should take into
consideration the enrichment of organic matter in the rhizosphere, i.e., complexation
and adsorption of Al3+ on mobile and nonmobile root exudates.56,57
Most important is the development of a measurement method in order to quantify
the Al dynamics in the rhizosphere as calculated here. Although the presented
simulation results agree with the previous observations, the theoretical results con-
nected with the assignment of the underlying processes can only be examined by
direct measurements, e.g., causal connection of pH pattern to rhizosphere transport
processes is missing until now. Some measurement approaches have been imple-
mented.25,37 But, the model results show that a great demand of spatial resolution of
the measurement method is necessary. Ionic selective microelectrodes, which offer
this demand of resolution, have been introduced successfully in other porous media.88
Furthermore, in order to be able to identify harmful soil conditions for tree
development, it is necessary to quantify the ion dynamics of the rooting space,125
having particular regard to smaller-scale differences. But the treatment of the rhizo-
sphere here and in most other investigations is characterized by a static approach.
The ion concentration gradients in the rhizosphere may be superimposed by root
growth,54 the ion dynamics in the whole rooting space; e.g., quickly growing roots
and fast ion transport in the soil column may modify any occurrence of gradients
in the rhizosphere. Therefore, in this regard it is most important for the quantification
of tree responses to soil-induced stress conditions100,123 to characterize the chemical
conditions of the potential rhizosphere pore space, i.e., of the pore space preferen-
tially occupied by roots because root-induced changes are based on these chemical
Modeling the Dynamics of the Rhizosphere Aluminum Chemistry 301
ACKNOWLEDGMENTS
I would like to acknowledge J. Hattenbach (GWDG, Gttingen), and G. Striker
(MPI, Gttingen) for their generous support, and an anonymous reviewer for his
valuable comments.
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Appendix
TABLE 12.8
Variables and Parameters Used in the Model
Symbol Meaning Unit
sel
K M /M Selectivity coefficient
i j
309
310 Trace Elements in the Rhizosphere
W
Y
Wacquant studies, 229
Wagatsuma, Ishikawa and, studies, 228 Youssef and Chino studies, 26
Wallace, Hale and, studies, 115 Yucca baccata (yucca), 58
Wallace studies, 115
Warnock studies, 46
Wassermann studies, 154 Z
Water content models, 290292
Water flux, daily-patterned, 270273, 281286, Zasoski, Chung and, studies, 190
298299 Zea mays L. (maize)
Wauters studies, 74, 77 cadmium uptake, 177
Weathering cation exchange capacity, 229, 246
brown earth model, 7778 complexation/chelation, 3334
laboratory model, 71, 76 crop rotation, 107
metals biogeochemistry, 7 manganese uptake, 27
mica, 71, 7778 metal intolerance, 171
mineral concentration in soils, 43 rhizosphere acidification, 194
mineral transformation, 8 root exudation, 1011, 191
Wedge zones, 6775 uranium accumulation, 112
Weinstein, Cumming and, studies, 254255, 298 zinc uptake, 178
Welch studies, 11 Zinc
Wheat, see Triticum (wheat) acid extractions, 203
White clover, see Clover complexation/chelation, 3336
Whitehead studies, 243 deficiency, 9
White lupin, 31, 35, 246 ectomycorrhiza fungi speciation, 156162
Wikstrm studies, 216 fractionation, 190191, 193, 196199
Wilcke and Kaupenjohann studies, 190 ion accumulation/depletion, 2728, 30
Wildung and Garland studies, 107 metal mobilization, 911
Wilkins, Denny and, studies, 152153, 158, 170 metal tolerance, 170, 172173
Willow, 102 microelement nutrition, 215216, 222
Winge, Mehra and, studies, 153154 mycorrhizal biomass, 152153
Wiren studies, 10 nuclear contaminants, 94
Wollaston Lake, Canada, 94 phytosiderophores, 10
Woodward studies, 230 precipitation, 1213
Woof, Tipping and, studies, 237 pyrophosphate extractions, 201202
reduction, time requirements, 6
rhizosphere acidification/alkalinization,
X 31
root growth, 8
XANES spectra, 155, 157158, 161162 root substrate-root interface, 221
XAS, see X-ray Absorption Spectroscopy (XAS) smelter study, Texas, 4357
X-ray Absorption Spectroscopy (XAS), 150, transport, 175179
155156, 161162 uptake regulation, 174
X-ray Fluorescence Microscopy (XRFM), 155, Zinc carbonate (smithsonite), 31
158 Zygomycotina, 150