Phytoextraction, also called phytoaccumulation, phytoabsorption, or phytosequestration, refers to the use of plants to absorb, translocate, and store toxic contaminants from soil, sediments, and/or sludge in the root and shoot tissues . Lead is an extremely difficult soil contaminant to remediate because it is a “soft” Lewis acid that forms strong bonds to both organic and inorganic ligands in soil. For the most part, Pb-contaminated soils are remediated through civil engineering techniques that require the excavation and landfilling of the contaminated soil. Soils that present a leaching hazard in the landfill are either placed in a specially constructed hazardous waste landfill, or treated with stabilizing agents, such as cement, prior to disposal in an industrial landfill.

1. Phytoextraction of Lead
By Haider Ali Malik
For more info:
haideralipine@gmail.com

2. Introduction to
Phytoextraction
• Phytoextraction, also called phytoaccumulation, phytoabsorption, or
phytosequestration, refers to the use of plants to absorb, translocate, and
store toxic contaminants from soil, sediments, and/or sludge in the root and
shoot tissues (Salt et al. 1998, Garbisu and Alkorta 2001).
• The idea of using plants to extract metals from contaminated soil was
introduced and developed by Utsunamyia (1980) and Chaney (1983).
• In contrast to organic pollutants, heavy metals do not undergo biodegradation and
therefore persist in soils for thousands of years.
• For example, Pb, one of the most toxic metals, has soil retention time of 150-5000
years and its high concentration may be maintained in the soil for 150 years (Singh
et al., 2016).

3. • Phytoextraction is the most
recognized and applied
phytoremediation technique for the
removal of toxic metals from
contaminated environments (Figure
1.2).
• Phytoextraction requires long-term
maintenance and routine harvesting
of the plants, as well as safe disposal
of polluted plant materials.
• The cost involved in phytoextraction,
when compared with the those of
conventional soil remediation
techniques, is 10-fold lower. This
means that phytoextraction is a cost-
effective technique (Salt et al. 1995).

4. • The only effective way of removal of metallic elements is extraction thereof using
conventional physical or chemical methods (e.g., in situ vitrification, soil
incineration, washing or flushing, but usually soil excavation and replacement - the
so-called ‘dig and dump’), which are usually very expensive and destructive to the
soil ecosystem, or using phytoextraction (Ali et al., 2013; Vangronsveld et al., 2009)
• Phytoextraction represents a cost-effective, efficient, environment-friendly and
ecofriendly alternative to the conventional methods but is also the most
challenging task among all phytoremediation approaches/technologies.

5. • Plants able to accumulate metals are grown on contaminated sites, and the
metal-rich aboveground biomass is harvested, resulting in the removal of a
fraction of the contaminant.
• Phytoextraction is the main and most useful phytoremediation technique for
removal of heavy metals and metalloids from polluted soils.
• It is also the most widespread and promising alternative of soil reclamation
for commercial applications. Metal translocation to shoots is a crucial
biochemical process desirable for an effective phytoextraction because the
harvest of root biomass is generally not feasible.
• Metal translocation from the roots to the shoots for the purpose of harvesting is one
of the key goals of phytoextraction research (Jarvis, Leung, 2001).

6. Ideal Plants for Phytoextraction
• Numerous factors, including pH of wastewater and sediment, mobilization and uptake from soil, compartmentalization and
sequestration within the root, efficiency of xylem loading and transport (transfer factors), distribution between metal sinks in
the aerial parts, sequestration and storage in leaf cells, and plant growth and transpiration rates can also affect the
remediation process of a contaminated site.
• Ideal plants to be used in phytoextraction should have the following characteristics
1. Tolerates high levels of metal concentration
2. Fast growth rate and high biomass production
3. Accumulates high level of metals in harvestable parts
4. Widely distributed and with a deep root system
5. Resistance to disease and pests and is unattractive to animals
6. Easy cultivation, harvesting, and processing
7. Low cultivation requirements
8. Repulsive to herbivores, to avoid food chain contamination

7. Types of Phytoextraction
• It can be divided into two methods—induced phytoextraction and continuous
phytoextraction (Salt et al., 1998).
• Induced phytoextraction uses plants producing a big amount of biomass in which
metal accumulation is enhanced by addition of a chemical, such as EDTA, NTA,
EDDS, etc.
• Continuous phytoextraction uses plants with natural abilities to accumulate high
levels of metals—hyperaccumulator.
• Hyperaccumulating plants have an extraordinary ability to accumulate heavy
metals, translocate and concentrate them in roots and aboveground shoots or leaves.

8. Continuous Phytoextraction
• Continuous phytoextraction generally depends on the natural ability of
plants to accumulate, translocate, and resist high amounts of metals over
the complete growth cycle.
• Continuous phytoextraction is also environmentally friendly, as it leaves the
site suitable for cultivation of other plants.
• In urbanized areas, continuous phytoextraction may be used in two types of
sites. One type comprises degraded soils in postindustrial areas, while the
other, which is a highly promising future application of phytoextraction, is
connected with soils in the vicinity of transportation routes and in urban
green areas.
• The phytoextraction potentials of ornamental plant species that are most
frequently planted in urban locations are under investigation in many
research centers worldwide. Such species include Tagetes erecta L.

9. Chelate-Assisted Phytoextraction
• Chelate-assisted phytoextraction is also called induced phytoextraction.
• The phytoextraction mechanism has its own limitations, e.g., low mobility and low bioavailability of some heavy
metals (especially Pb) in polluted environments. An increase of heavy metal mobility can be achieved by adding
synthetic chelating agents which are capable of solubilizing and complexing with heavy metals in a soil solution as
well as promoting heavy metal translocation from roots to the harvestable parts of the plant.
• This strategy of phytoextraction is based on the fact that the application of metal chelates to a soil significantly
enhances metal accumulation in plants.
• Several studies have reported that the application of chelating agents, such as ethylene diamine triacetic acid
(EDTA), N-(2-hydroxyethyl)-ethylene diaminetriacetic acid (HEDTA), diethylene–tetramine-pentaacetate acid
(DTPA), ethylenediamine di-ohydroxy phenylacetic acid (EDDS), ethylene glycol-O,O′-bis-(2-amino-ethyl) N,N,N′,N′-
tetraacetic acid (EGTA), and citric acid, can enhance the effectiveness of phytoextraction by mobilizing metals and
increasing metal accumulation in plants.

10. • EDTA for Pb and Cd and citrate for U are normally used for induced phytoextraction. Of all the chelates applied at 5
and 10 mmol kg-1 to soil, EDTA @ 10 mmol kg-1 could induce the phytoextracton of Pb upto 16000 ppm.
Phytoextraction of Pb increases in maize and pea plants by 50% with EDTA. Thousand fold increase in uranium
concentration was observed in citric acid amended soils. Fourteen taxa including Brassica jimea and Zea mays
were reported to be Pb hyperaccumulators with Pb concentration ranging from 1000 to 20000 ug g-1 in the
presence of EDTA (Reeves and Baker 2000). As reported by Ma et al. (2001), Brake fern (Pteris vittata) has removed
22630 ug g-1 Arsenic.
• Huang et al. (1998) estimated that in normal phytoaccumulation method, corn has accumulated uranium (U) to
the extent of 10 ppm and was higher than other test crops, but in case of induced phytoextracton by applying
citric acid to soil, the accumulation could be increased manifold in all crop species and was highest in Brassica
chinensis (1300 ppm) followed by B. juncea (750 ppm) and Amaranthus spp., (600ppm).

11. Mechanism of phytoextraction
• Five things need to happen for a plant to extract a heavy metal from water or soil
1) The metal needs to be dissolved in something the plant roots can absorb
2) After dissolution, metal ions are chelated with a specific metal transporter or specific agents
secreted by plant roots and then transport the metal over the cell wall. Some examples of
chelators are: phytosiderophores, organic acids or carboxylates.
3) The plant needs to chelate the metal in order to both protect itself and make the metal more
mobile.
4) The root-to-shoot transport of heavy metals is strongly regulated by gene expression. These
genes code for the transport systems of heavy metals and are constantly over-expressed in
hyper-accumulating plants when they are exposed to heavy metals. These transporters are
known as heavy metal transporting ATPases (HMAs). One of it is HMA4, which belongs to the
Zn or Co or Cd or Pb HMA subclass and is localized at xylem parenchyma plasma membranes.
5) Finally, the plant adapts to any damages the metals cause during transportation and storage.

12. Lead Phytoextraction
• Lead (Pb) is a naturally occurring element and, as a result of
anthropogenic activities, a ubiquitous environmental contaminant.
• Lead is an extremely difficult soil contaminant to remediate because it is a
“soft” Lewis acid that forms strong bonds to both organic and inorganic
ligands in soil.
• For the most part, Pb-contaminated soils are remediated through civil
engineering techniques that require the excavation and landfilling of the
contaminated soil. Soils that present a leaching hazard in the landfill are
either placed in a specially constructed hazardous waste landfill, or treated
with stabilizing agents, such as cement, prior to disposal in an industrial
landfill.

13. • Many innovative site decontamination techniques have been tried, including
electroreclamation and several variations of soil washing, but, with the exception
of soil washing, most of these techniques appear ineffective for the remediation of
Pb-contaminated soils in a cost-effective manner.
• All remediation techniques currently available for Pb-contaminated soil are
expensive and disruptive to the site. Additionally, they require further steps to
return a healthy ecosystem to the remediated area.

14. Sources of Lead
• Frequent use in many industrial processes is the main
reason for lead contamination of the environment.
• There are a variety of industrial processes that involve
the use of lead such as mining, smelting, manufacture
of pesticides and fertilizers, dumping of municipal
sewage and the burning of fossil fuels that contain a
lead additive.
• Many commercial products and materials also contain
lead including paints, ceramic glazes, television glass,
ammunition, batteries, medical equipment (i.e., x-ray
shields, fetal monitors), and electrical equipment. The
uses of lead for roofing and the production of
ammunition has increased from previous years.
• Lead battery recycling sites, of which 29 have been
labelled Superfund sites, and manufacturers use more
than 80% of the lead produced in the United States.
On average, recycled lead products only satisfy half of
the nation’s lead requirements.

15. Forms of Lead
• Ionic lead (Pb2+), lead oxides and hydroxides and lead-metal oxyanion complexes are the
general forms of lead that are released into the soil, groundwater and surface waters.
• The most stable forms of lead are Pb2+ and lead-hydroxy complexes. Pb2+ is the most
common and reactive form of lead, forming mononuclear and polynuclear oxides and
hydroxides.
• The predominant insoluble lead compounds are lead phosphates, lead carbonates (form
when the pH is above 6) and lead (hydr)oxides.
• Lead sulfide (PbS) is the most stable solid form within the soil matrix and forms under
reducing conditions when increased concentrations of sulfide are present. Under
anaerobic conditions a volatile organolead (tetramethyl lead) can be formed due to
microbial alkylation

16. Lead and the Soil Matrix
• Once introduced into the soil matrix, lead is very difficult to remove.
• The transition metal resides within the upper 6-8 inches of soil where it is strongly
bound through the processes of adsorption, ion exchange, precipitation, and
complexation with sorbed organic matter
• Lead found within the soil can be classified into six general categories: ionic lead
dissolved in soil water, exchangeable, carbonate, oxyhydroxide, organic or the
precipitated fraction. All of these categories combined make up the total soil lead
content .
• Water soluble and exchangeable lead are the only fractions readily available for
uptake by plants. Oxyhydroxides, organic, carbonate, and precipitated forms of
lead are the most strongly bound to the soil .

17. Uptake of lead in
roots and shoots
• Lead is only sparingly soluble in solution, and even at highly contaminated
sites, Pb in the soil solution is often less than 4 mg/L.
• At such low concentrations, many plants can effectively assimilate soil
solution Pb into root tissues. However, very little of the Pb is translocated to
aerial plant tissues which can be harvested.
• When Pb enters the plant root, it immediately comes in contact with high
phosphate concentrations, relatively high pH, and high carbonate-
bicarbonate concentrations in the intercellular spaces.
• Under these conditions, Pb precipitates out of solution in the form of
phosphates or carbonates. Plant roots contain inclusion bodies of these
forms of Pb in the tissue, reduce Pb translocation in plants.
• Soluble Pb present in the growing media may not be a key factor
governing Pb uptake. Other factors such as presence or absence of
complexors or chelators are important in Pb uptake. Chelators not only
provide more absorption by the root but facilitate translocation to the
upper plant parts.
• Several chelating agents such as EDTA, CDTA, DTPA, and citric acid have
been used for fortification of Pb-contaminated soils in order to increase
Pb bioavailability and uptake.
• Among these chelating agents, EDTA was shown to be the most
efficient enhancing shoot Pb concentrations by 120-fold in corn growing
in soils containing 2500 mg/kg Pb.[EDTA addition resulted in a 150 fold
increase in Pb concentration in Indian mustard grown in soils

18. Lead uptake in plants mechanism
• The upper layers of the radicular cortex (rhizoderm and collenchyma/parenchyma) constitute a physical barrier against Pb
penetration into the root. Initially, Pb present in the soil solution is adsorbed on root surface. It may bind to the carboxylic
groups of uronic acids which composed the root mucilage or directly to the polysaccharides of rhizodermic cell surface.
• Once it has fixed to the rhizoderm, Pb could penetrate the root system passively and follows the water conduction system. Such
absorption is not uniform throughout the root, since a Pb concentration gradient is observed in the tissues, starting from the
apex, which is the area of highest concentration. The young tissues, and the apical area in particular (excluding the root cap) in
which the cell walls are still thin, are the parts of the plant that absorb the most Pb. This apical region also corresponds to the
area in which the rhizospheric pH is more acidic. This low pH favours metal solubility and a locally high Pb concentration in
the soil solution.
• At the molecular level, the mechanisms, through which the metal manages to penetrate the roots, have not yet been explained.
Pb may benefit from the nonselectivity of some channels/transporters and the very high potential in membrane that can exceed -
200 mV in the rhizoderm cells. Pb absorption is therefore a passive absorption, but requires the cell to expend energy in order to
maintain this very negative potential through the excretion of protons into the external environment via H+/ATPase pumps.

19. • Among non-selective cation channels, depolarization-activated calcium channels
(DACC), hyperpolarization activated calcium channels (HACC) and voltage
insensitive cation channels (VICC) are thought to be one of the principal routes of Pb
entry into root cells (White 2012).
• Other transporters, such as the families of Cation Diffusion Facilitator (CDF),
ZRT/IRT-like Protein (ZIP) or the Natural resistance-associated macrophage proteins
(Nramps), associated with the transport of copper, zinc, cadmium and manganese
(see, for review, Hall and Williams 2003), could also play a role in Pb transport.
• Finally, Krzesłowska et al. (2010) hypothesized that Pb could enter the cell protoplast
endomembrane system as a pectin—Pb complex during internalization of low-
methyle sterified pectins from the cell wall

20. Radial Diffusion in the Root
Apoplastic Pathway
• Many histological studies have shown that Pb is essentially transported in the apoplast and that it follows
water movements within the plant.
• Once inside the apoplast, Pb can migrate relatively quickly. Although Pb can diffuse within the root, only a
small fraction present in the root is mobile. More than 90 % of Pb is found in insoluble forms and is strongly
linked to external components of the cells. Pb is mainly linked to the cell walls but can be found associated
with the middle lamella or the plasma membrane. It may also be precipitated in the intercellular space.
• This distribution that is very specific to Pb, can be explained by its particular affinity for the carboxyl groups
and pectins, and, to a lesser extent, to hemicellulose and cellulose molecules and lignin of the cell walls.

21. Symplastic Pathway
• In non-lethal doses, Pb only penetrates the symplast in areas in which cells are dividing actively, such
as in the apical area or in the protoderm. Young cells do not yet possess a secondary wall and their
primary wall is very thin. Access to the cell membrane is thus facilitated in these parts of the root.
• Symplastic Pb may be found confined to certain cell compartments, such as vacuoles, dictyosomal
vesicles, and vesicles of the endoplasmic reticulum.
• In lethal doses, Pb penetrates all the radicular tissues, and the cell membranes no longer appear to play
their role of physical barrier. In such concentrations, Pb causes the disorganisation of the membranes.
It is then able to penetrate massively into the cytoplasm, the nucleus and the various organelles,
including those with double membranes, such as the mitochondria.

22. Translocation to the Aerial Parts of the Plant
• Certain plant species are capable of transferring large amounts of Pb to the aerial parts. However, slight metal
translocation seems to be quite a common phenomenon in other plant species.
• As previously mentioned, this phenomenon is largely due to the very large amount of Pb immobilised in
insoluble forms. However, it appears that the physical barrier constituted by the endodermis also plays an
important role.
• In fact, the Pb that mostly passes along the apoplastic pathway is blocked by the Casparian strips in the
endodermis.
• In order to go with the water flow, it must travel along the symplastic pathway through the filter constituted
both by the permeability of the cell membranes and the cytoplasm sequestration/detoxification systems.

23. • Pb absorbed by the epidermis and root hairs penetrates into cortical tissues, but does not seem to be capable of
passing through the endodermis.
• Pb mostly penetrates into the central cylinder via the apex, a region in which the endodermis has not yet formed.
• Consequently, it would appear that in non-lethal doses, Pb translocation to the aerial parts originates solely from the
radicular apex. Once Pb has penetrated into the central cylinder, it can once again resume its route through the
apoplastic pathway.
• Pb then uses the vascular system by following the water flow to the leaves, in which water evaporates and where it
accumulates.
• When it passes into the xylemic sap, Pb may be compounded with amino acids such as histidine and with organic
acids such as citric, fumaric and malic acids or with the PCs, as discussed previously. It may also be transferred,
mostly in an inorganic form, as is the case for cadmium.

24. Phytoextraction of Lead
• Pb is mostly confined to the roots, with minimal transport to the green
parts of plants.
• Certain plants have been identified which have the potential to uptake
lead. Many of these plants belong to the following families:
Brassicaceae, Euphorbiaceae, Asteraceae, Lamiaceae, and
Scrophulariaceae.
• Brassica juncea, commonly called Indian Mustard, has been found to
have a good ability to transport lead from the roots to the shoots,
which is an important characteristic for the phytoextraction of lead.
• Thalspi rotundifolium ssp. Cepaeifolium, a non-crop Brassica,
commonly known as Pennycress, has been found to grow in soils
contaminated with lead (0.82%) and zinc from a mine. Bench scale
studies have also shown that certain crop plants are capable of
phytoextraction. Corn, alfalfa, and sorghum were found to be effective
due to their fast growth rate and large amount of biomass produced.

25. • In phytoextraction, soil Pb is taken into plant roots,
translocated into the top of the plant, and removed by
plant harvesting (Figure 19.1). Several technical
parameters affect how efficiently this process functions.
• First, soil Pb must be in a form that is available to the
plant root. Often at a site, the total Pb level is quite
high, but the fraction of Pb availability for root uptake is
exceedingly low.
• Second, the plant must be able to transfer Pb in the
roots to the xylem stream to be carried to harvestable
plant tissues. In most plants, this is difficult due to the
chemical and physical environment in their tissues.
• Finally, the Pb-containing harvested plant material must
be processed. Presently, the technical and logistic points
regarding post-harvest treatment are mostly
conceptual. The plant may be viewed as a solar driven
extracting system that moves Pb from a silica-, iron-, or
aluminum-based matrix (the soil), into a primarily
carbon-, hydrogen-, or oxygen-matrix (the plant).

26. Hyperaccumulators for lead
• The term hyperaccumulator was coined by Brooks et al. (1977) to describe plants that contain greater than 0.1% nickel (Ni) in their dried leaves. Since
then, threshold values have also been established for other metals, such as zinc (Zn), lead (Pb), cadmium (Cd), copper (Cu), chromium (Cr), iron (Fe),
manganese (Mn), etc. (Brooks 1998).
• Over 450 plant species, including trees, vegetable crops, grasses, and weeds, comprising 101 families (including Asteraceae, Brassiciaceae,
Caryophyyllaceae, Cyperaceae, Cunnoniaceae, Fabiaceae, Flacourtiaceae, Lamiaceae, Poaceae, Violaceae, and Euphobiacea), have been identified as
hyperaccumulators
• Hyperaccumulators of Pb are defined as naturally accumulating 0.1% or more of their dry biomass as Pb (1000 mg kg-1) (Raskin et al. 1997; Pilon-Smits
2005; Saifullah et al. 2009).
• Chemical analysis on foliage of plants naturally growing in contaminated areas have shown a few possible Pb hyperaccumulators such as Stellaria vestita,
Sonchus asper, Festuca ovina, Arenaria rotumdifolia, Arabis alpinal Var parviflora, Oxalis corymbosa, Eupatorium adenophorum, Crisium chlorolepis,
Taraxacum mongolicum, and Elsholtzia polisa (Yanqun et al. 2005).
• Different genera of Brassicaceae (mustard family), Asteraceae (sunflower family), Celosia cristata pyramidalis (an ornamental plant), and Pelargonium
(scented geraniums) have also been identified as possible hyperaccumulators of Pb
• Reeves and Baker (2000) reported some examples of plants which can accumulate Pb
– Minuartia verna and Agrostis tenuis (Pb hyperaccumulators)

27. • Some plants, such as corn, may accumulate at most a few hundred mg Pb/kg in the roots. Other plants grown
under identical controlled conditions, such as Thlaspi rotundafolium L., may accumulate over 3% in their roots.
Shoot concentrations for most plants are disappointingly low. Relatively little Pb is transferred from the root to
shoot..
• Nouri et al. (2011) collected 12 native plant species grown on a Pb- and Zn-contaminated mine area and
analyzed the plants for accumulation of Pb, Zn, Mn, and Fe. They found that among the analyzed plants,
none of them was suitable for phytoextraction of Pb, Zn, Mn, or Fe or for phytostabilization of Fe, but
Scrophularia scoparia was the most suitable plant for phytostabilization of Pb (BCF, 1.43; TF, 0.09);
– Thlaspi rotondifolium(Pennygrass) from Brassicaceae - Pb (Reeves and Brooks1983)
– Drosera rotundifolia (Common sundew) from Droseraceae - Pb (Fontem Lum et al. 2015)
– Eleusine indica (Goose grass) from Poaceae - Pb (Fontem Lum et al. 2015)
– Euphorbia cheiradenia from Euphorbiaceae - Pb (Chehregani and Malayeri 2007)

28. Role of Chelates in enhancin Pb uptake
• Lead is only sparingly soluble in solution, and even at the most contaminated sites, Pb in the soil solution is often less than 4 mg/l. At these
low concentrations, many plants can effectively remove soil solution Pb into root tissues; however, very little Pb is translocated to aerial plant
tissues that can be harvested.
• The chemistry of plants provides a logical reason for poor Pb translocation in most plants. When Pb enters the plant root, it immediately
comes in contact with high phosphate concentrations, relatively high pH, and high carbonate-bicarbonate concentrations in the intracellular
spaces. Under these conditions, Pb precipitates out of solution as phosphates or carbonates.
• Phytoextraction appears to be complicated by three factors:
– (1) the low solubility of Pb in soils causing Pb to be unavailable for plant uptake,
– (2) the poor Pb translocation in plants to harvestable plant portions, and
– (3) the toxicity of Pb to the plant tissue.
• One major factor limiting the potential for lead phytoextraction is low metal bioavailability for plant uptake. To overcome this limitation,
synthetic chemical chelators may need to be added to the contaminated soil to increase the amount of lead that is bioavailable for the plants.
The use of synthetic chelates in the phytoremediation process is not only to increase heavy metal uptake by plants through increasing the
bioavailability of the metal, but also to increase micronutrient availability, which decreases the possibility of plant nutrient deficiencies.
• Based on scientific studies, it has been shown that only 0.1% of the total amount of lead in contaminated soils is in solution and bioavailable to
plants for remediation. With the addition of synthetic chelators, the total amount of lead in solution can be increased up to 100 times

29. • No effective hyperaccumulating plant, with high Pb uptake and high
biomass essential for efficient phytoextraction, has been reported so
far for Pb, one of the most widespread and important metal
contaminants
• In non-hyperaccumulating plants, factors limiting their potential for
phytoextraction include small root uptake and little root-to-shoot
translocation of heavy metals. Chemically enhanced phytoextraction
has been shown to overcome the above problems.
• Common crop plants with high biomass can be triggered to accumulate
high amounts of low bioavailable metals, when their mobility in the soil
and translocation from the roots to the green part of plants was
enhanced by the addition of mobilizing agents when the crop had
reached its maximum biomass. The feasibility of chemically enhanced
phytoextraction has been primarily studied for Pb and chelating agents
as soil additives, less attention has been given to other metals and
radionuclides or their mixtures.

30. • Plants can regulate metal solubility by acidification of the rhizosphere due to the extrusion
of H+ from roots and by exuding their own chelating agents, phytosiderophores and
organic acids; for example, malic and citric acids.
• A chelating agent is a substance whose molecules can form several coordinative bonds to a
single metal ion (Figure 2).
• A chelating agent is therefore a multidentate metal ligand and forms complexes with
metals. The localized excretion of plant chelating agents mobilizes nutrients such as Zn, Fe,
Mn and other metals. Water-soluble chelating agents increase and maintain metal
concentration in the soil aqueous phase.
• To make plants take up Pb and other low bioavailable metals, which are much more
strongly bound to the soil solid phase than Cd, strong synthetic chelating agents have been
used to bring metals into the soil solution.
• For example, the addition of Na salt of ethylenediamine-tetracetic acid (EDTA) to soil
shifted Pb mostly from the carbonate fraction to the fraction soluble in the soil solution, in
which the Pb concentration increased from 3 to 362 mg kg–1

31. • The addition of synthetic chelators has been shown to increase soil Pb
mobility and plant uptake. Huang and Cunningham showed N-(2-
hydroxyethyl)-ethylenediaminetriacetic acid (HEDTA) addition enhanced
Pb accumulation.
• Blaylock showed that chelator supplements increased uptake of Pb, Cd,
Cu, Ni, and Zn.
• Huang et al. also reported that among the five chelators,
Ethylenediaminetriacetic acid (EDTA) was the most efficient in increasing
shoot Pb concentration in both pea and corn, followed by HEDTA.
• They found the order of the effectiveness in increasing Pb accumulation:
EDTA > HEDTA > diethylenetrinitrilopentaacetic acid (DTPA) >
ethylenegluatarotriacetic acid (EGTA) > Etheylenediaminedinitrilodiac acid
(EDDTA).

32. • Increasing the mobility and bioavailability of lead in the soil through
certain chelators, organic acids, or chemical compounds, allows for
the hyperaccumulation of metals in some plants.
• For lead, a number of different chelators have been tested: EDTA
(ethylene-dinitrilotetraacetic acid), CDTA (trans-1,2-cyclohexylene-
dinitrilo-tetraacetic acid), DTPA (diethylenetrinitrilo-pentaacetic
acid), EGTA (ethylebis[oxyethylenetrinitrilo]-tetraacetic acid), HEDTA
(hydroxyethyl-ethylene-dinitrilo-triacetic acid), citric acid, and malic
acid.
• Addition of the chelates resulted in enhanced shoot lead
concentrations. EDTA proved to be the best and least expensive,
costing around $1.95 per pound.
• In soils with a pH of 5 and amended with EDTA, plants accumulated
nearly 2000 mg/kg more lead in their shoots when compared to
other treatments in soil limed to a pH of 7.5. EDTA, DTPA, and CDTA
all achieved shoot lead concentrations of more than 10,000 mg/kg

33. • In order for substantial lead accumulation (> 5,000mg/kg) to
occur in the shoots, the concentration of synthetic chelates
(EDTA, DTPA, CDTA) exceeded 1 mmol/kg.
• It was also noted that plants grown in soils amended with
chelators varied in their lead concentration uptake.
• For example, the lead concentration in peas (Pisum sativum L.
cv Sparkle) was 11,000 mg/kg compared to corn, which
accumulated 3,500 mg/kg in soils receiving equivalent
amounts of EDTA.
• The major concern associated with using chelates to enhance
phytoremediation and increase the bioavailability of the toxic
metals is the fear of lead leaching or running off into the
ground or surface water. By making the metals more soluble in
the soil matrix, leaching is more probable, threatening the
contamination of nearby water sources

34. Efficiency of chelating agent enhanced phytoextraction
• The efficiency of phytoextraction is determined by two key
factors: biomass production and the metal
bioconcentration factor.
• The bioconcentration factor is defined as the ratio of metal
concentration in the plant shoot to metal concentration in
the soil.
• For phytoextraction to be feasible, the bioconcentration
factor of the plant must be greater than 1, regardless of
how large the achievable biomass is.
• Another way to measure the efficiency of phytoextraction is
by using the phytoextraction potential. This can be
calculated from soil and plant metal concentrations and dry
biomass plant yield, as the total amount of metal extracted
per ha of soil, in a single phytoextraction cycle, and
expressed as kg ha –1
• In order to be economically viable, plants for Pb
phytoextraction should be able to accumulate at least
10,000 mg Pb kg–1 in their green parts (harvesting the roots
is not practical) and achieve a dry biomass of 20 t ha–1.
Figure 4: Percentage Pb distribution in root, stem, and leaf of the studied plants
grown in hydroponic system at the concentration of 2.5 mM Pb(NO3)2 in the
presence and absence of EDTA during the fourth week of Pb exposure. (A) H.
annuus. (B) N. tabacum

35. Molecular structures of EDDS binding to a metal ion (M) (Smokefoot
2013).
Molecular structures of EDTA binding to a metal ion (M)
(Yikrazuul 2010

36. Role of Miccorhiza in phytoextraction
• Research to date indicates that the effectiveness of phytoextraction depends on
agronomic practices, plant selection, chelate selection, soil pH, ionic balance of the
soil solution, climatic factors, and the use of fertilizers.
• Mycorrhizal infection of the plant roots may also influence phytoextraction. Under
low Pb levels, mycorrhizae increase Pb uptake into the plant, but they actually
decrease Pb uptake under higher Pb levels.
• Arbuscular mycorrhizal fungi (AMF) are symbiotic, asexual organisms of the
rhizosphere which are dependent on vascular plants as their carbon sources, while
providing the plants with nutrients from the soil (mainly phosphorus), assistance
with water absorption, and protection against root pathogens. Around 80% of all
known vascular plants have symbiotic associations with AMF in the rhizosphere.