FUNDAMENTALS OF PHYTOREMEDIATION

Phytoremediation is an emerging technology that uses various plants to degrade, extract,

contain, or immobilize contaminants from soil and water. This technology has been receiving

attention lately as an innovative, cost-effective alternative to the more established treatment

methods used at hazardous waste sites.

Phytoremediation is a general term used to clean up contaminants using plants, or

remediate sites by removing pollutants from soil and water (Chandra Shekar et al 2003). Plants

can break down or degrade organic pollutants or contain and stabilize metal contaminants by acting

as filters or traps. Phytoremediation involves growing plants in a contaminated matrix, for a

required growth period to remove contaminants from the matrix or facilitate immobilization or

degradation (detoxification) of the pollutants. The plants can be subsequently harvested, processed

and disposed in an environmentally sound manner.

A phytoremediation system capitalizes on the synergistic relationships among plants,

microorganisms, water and soil that have evolved naturally in wetlands and upland sites over

millions of years. In the biological sequences that transform contaminants to neutral compounds,

plants contribute inherent enzymatic and uptake processes that can recycle or sequester the organic

molecules they encounter.

Plants act as hosts to aerobic and anaerobic microorganisms, supplying them with both

physical habitat and chemical building blocks. Plant roots and shoots increase microbial activity

in their direct environment by providing additional colonisable surface area, increasing readily-

degradable carbon substrates by organic exudates and leachates and by decomposition of part of

their mass and creating temporally and spatially varying oxygen regimes. Physically, plants slow

the movement of contaminants in soil (reduced run-off), increase evapotranspiration and adsorbs

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compounds through their roots. Once a wetland or upland phytoremediation system is in place, its

biological components would be naturally self-sustaining (due to photosynthesis).

Phytoremediation can be applied in terrestrial and aquatic environments. It can be used as

a preparative or finishing step for other clean-up technologies. Plants are aesthetically pleasing

and these systems are relatively self-sustaining and cost-effective. Plants have evolved a great

diversity of genetic adaptations to handle the accumulated pollutants that occur in the environment.

Growing, and in some cases harvesting the plants on a contaminated site as a remediation method

is a passive technique that can be used to clean up sites with shallow, low to moderate levels of

contamination. Phytoremediation can be used to clean up metals, pesticides, solvents, explosives,

crude oil, polyaromatic hydrocarbons and landfill leachates. It can also be used in river basin

management through the hydraulic control of contaminants.

APPLICATIONS OF PHYTOREMEDIATION

Phytoremediation applications can be classified based on the contaminant fate:

degradation, extraction, containment, or a combination of these. Phytoremediation applications

can also be classified based on the mechanisms involved. Such mechanisms include extraction of

contaminants from soil or groundwater; concentration of contaminants in plant tissue; degradation

of contaminants by various biotic or abiotic processes; volatilization or transpiration of volatile

contaminants from plants to the air; immobilization of contaminants in the root zone; hydraulic

control of contaminated groundwater (plume control); and control of runoff, erosion, and

infiltration by vegetative covers. A brief explanation of these application categories follows, with

more detailed explanations.

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 1. Degradation

Plants may enhance degradation in the rhizosphere (root zone of influence). Microbial

counts in rhizosphere soils can be 1 or 2 orders of magnitude greater than in non-rhizosphere soils.

It is not known whether this is due to microbial or fungal symbiosis with the plant, plant exudates

including enzymes, or other physical/chemical effects in the root zone. There are, however,

measurable effects on certain contaminants in the root zone of planted areas. Several projects

examine the interaction between plants and such contaminants as trinitrotoluene (TNT), total

petroleum hydrocarbons (TPH), pentachlorophenol (PCP), and polynuclear aromatic

hydrocarbons (PAH).

Another possible mechanism for contaminant degradation is metabolism within the plant.

Some plants may be able to take in toxic compounds and in the process of metabolizing the

available nutrients, detoxify them. Trichloroethylene (TCE) is possibly degraded in poplar trees

and the carbon used for tissue growth while the chloride is expelled through the roots.

2. Extraction

Phytoextraction or phytomining, is the process of planting a crop of a species that is known

to accumulate contaminants in the shoots and leaves of the plants, and then harvesting the crop

and removing the contaminant from the site. Unlike the destructive degradation mechanisms, this

technique yields a mass of plant and contaminant (typically metals) that must be transported for

disposal or recycling. This is a concentration technology that leaves a much smaller mass to be

disposed of when compared to excavation and landfilling. This technology is being evaluated in a

Superfund Innovative Technology Evaluation (SITE) demonstration, and may also be a technology

amenable to contaminant recovery and recycling.

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 Rhizofiltration is similar to phytoextraction in that it is also a concentration technology. It

differs from phytoextraction in that the mechanism is root accumulation and harvest using

hydroponic (soil-less) growing techniques. This is useful for separating metal contaminants from

water. Rhizofiltration has been demonstrated on U.S. Department of Energy (DOE) sites for

radionuclides. Volatilization or transpiration through plants into the atmosphere is another possible

mechanism for removing a contaminant from the soil or water of a site. It is often raised as a

concern in response to a proposed phytoremediation project, but has not been shown to be an actual

pathway for many contaminants. Mercury (Hg) has been shown to move through a plant and into

the air in a plant that was genetically altered to allow it to do so. The thought behind this media

switching is that elemental Hg in the air poses less risk than other Hg forms in the soil. However,

the technology or the associated risk has not been evaluated.

3. Containment and Immobilization

Containment using plants either binds the contaminants to the soil, renders them nonbioavailable,

or immobilizes them by removing the means of transport. Physical containment of contaminants

by plants can take the form of binding the contaminants within a humic molecule (humification),

physical sequestration of metals as occurs in some wetlands, or by root accumulation in non-

harvestable plants. Certain trees sequester large concentrations of metals in their roots, and

although harvesting and removal is difficult or impractical, the contaminants present a reduced

human or environmental risk while they are bound in the roots.

Risk reduction may also be achieved by transforming the contaminant into a form that is

not hazardous, or by rendering the contaminant non-bioavailable. EPA and the U.S. Department

of Agriculture (USDA) have ongoing research in this area. Hydraulic control is another form of

containment. Groundwater contaminant plume control may be achieved by water consumption,

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using plants to increase the evaporation and transpiration from a site. Some species of plants use

tremendous quantities of water, and can extend roots to draw from the saturated zone. EPA is

pursuing research in this area at a number of sites, including the SITE demonstrations at Ogden,

UT and Ft. Worth, TX, and the Emergency Response Team (ERT) lead projects at Aberdeen

Proving Grounds (Edgewood, MD) and the Edward Sears Properties Site (New Gretna, NJ).

Private companies have installed trees as a hydraulic control at many sites.

Vegetative cover (evapotranspiration or water-balance cover) systems are another

remediation application utilizing the natural mechanisms of plants for minimizing infiltrating

water. Originally proposed in arid and semi-arid regions, vegetative covers are currently being

evaluated for all geographic regions. The effectiveness in all regions and climates needs to be

assessed on a site-specific basis. If there is potential for gas generation a vegetative cover may not

be an option. For example, a municipal solid waste landfill can produce landfill gas that may be of

concern to human health and the environment. Sites with requirements to collect and control

landfill gas may not meet Federal requirements under the Clean Air Act if a vegetative cover is

used.

Hydraulic control for groundwater plumes and water balance covers are two technologies

that are being applied in the field prior to model development predicting their behavior. Under an

EPA initiative called Alternative Cover Assessment Program (ACAP), several of these field

installations will be monitored carefully and consistently to gather data to both evaluate

performance and to build and verify models to predict the performance of other proposed

installations. Data from a national network of sites that have similar measurement regimes will be

a powerful tool for evaluating the appropriateness of a proposed installation, and help develop the

tools for predicting the efficacy of similar cover systems.

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PHYTOREMEDIATION PROCESSES

At metals contaminated sites, plants are used either to stabilise or remove the metals from

the soil and ground water through mechanisms such as phytoextraction, rhizofiltration, and

phytostabilization (UNEP 2002).

1. Phytoextraction

Phytoextraction also called phytoaccumulation, refers to the uptake and translocation of

metal contaminants in the soil by plant roots into the aboveground portions of the plants.

Contaminants are generally removed by harvesting the plants. This concentration technology

leaves a much smaller mass to be disposed than excavation of the soil or other media. Certain

plants, called hyperaccumulators, adsorb unusually large amounts of metals in comparison to other

plants. One or a combination of these plants is selected and planted at a particular site based on

the type of metals present and other site conditions. After the growth period of the plants, they are

harvested and either incinerated or composted to recycle the metal!). This procedure may be

repeated if necessary, to bring soil contaminant levels down to permissible limits. If plants are

incinerated, ash is disposed in a hazardous waste landfill. The volume of ash will be less than

10%of the volume that is created, if the contaminated soil itself were dug up for treatment.

Metals such as nickel, zinc and copper are the best candidates for removal by

phytoextraction because it has been shown that they are preferred by a majority of the

(approximately400) plants that uptake and absorb unusually large amounts of metals. This

technology is most often applied to metal-contaminated soil. Phytoextraction is primarily used in

the treatment of soil, sediments, and sludges. It can be used to a lesser extent for treatment of

contaminated water.

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Advantage

The plant biomass containing the extracted contaminant can be a resource. For example,

biomass that contains selenium (Se), an essential nutrient, has been transported to areas that are

deficient in Se and used for animal feed (Bañuelos 1997a).

Disadvantages

Phytoextraction has the following disadvantages:

a. Metal hyperaccumulators are generally slow-growing with a small biomass and shallow

root systems.

b. Plant biomass must be harvested and removed, followed by metal reclamation or proper

disposal of the biomass. Hyperaccumulators may accumulate significant metal

concentrations — e.g., Thlaspi rotundifolium grown in a lead-zinc mine area contained

8,200 g/g Pb (0.82%) and 17,300 g/g zinc (Zn) (1.73%), and Armeria maritima var. halleri

contained 1,300 g/g Pb, dry weight basis (Reeves and Brooks 1983).

c. Metals may have a phytotoxic effect (Nanda Kumar et al. 1995).

d. Phytoextraction studies conducted using hydroponically grown plants, with the

contaminant added in solution may not reflect actual conditions and results occurring in

soil. Phytoextraction coefficients measured under field conditions are likely to be less than

those determined in the laboratory (Nanda Kumar et al. 1995).

2. Rhizofiltration

Rhizofiltration is the adsorption or precipitation onto plant roots, or absorption into the

roots of contaminants that are in solution surrounding the root zone, due to biotic or abiotic

processes. Plant uptake, concentration, and translocation might occur, depending on the

contaminant. Exudates from the plant roots might cause precipitation of some metals.

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 Rhizofiltration is similar to phytoextraction, but the plants are used primarily to address

contaminated ground water rather than soil. The plants to be used for cleanup are raised in

greenhouses with their roots in water rather than in soil. To acclimatize the plants, once a large

root system has been developed, contaminated water is collected from a waste site and brought to

the plants where it is substituted for their water source. The plants are then planted in the

contaminated area where the roots take up the water and the contaminants along with it. As the

roots become saturated with contaminants, they are harvested. Rhizofiltration first results in

contaminant containment, in which the contaminants are immobilized or accumulated on or within

the plant. Contaminants are then removed by physically removing the plant.

Extracted groundwater, surface water, and waste water can be treated using this

technology. Rhizofiltration is generally applicable to low-concentration, high-water-content

conditions. This technology does not work well with soil, sediments, or sludges because the

contaminant needs to be in solution in order to be sorbed to the plant system.

Advantages

Rhizofiltration has the following advantages:

a. Either terrestrial or aquatic plants can be used. Although terrestrial plants require support,

such as a floating platform, they generally remove more contaminants than aquatic plants.

b. This system can be either in situ (floating rafts on ponds) or ex situ (an engineered tank

system).

c. An ex situ system can be placed anywhere because the treatment does not have to be at the

original location of contamination.

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Disadvantages

Rhizofiltration has the following disadvantages:

a. The pH of the influent solution may have to be continually adjusted to obtain optimum

metals uptake.

b. The chemical speciation and interaction of all species in the influent have to be understood

and accounted for.

c. A well-engineered system is required to control influent concentration and flow rate.

d. The plants (especially terrestrial plants) may have to be grown in a greenhouse or nursery

and then placed in the rhizofiltration system.

e. Periodic harvesting and plant disposal are required.

f. Metal immobilization and uptake results from laboratory and greenhouse studies might not

be achievable in the field.

3. Phytostabilization

Phytostabilization is defined as (a) immobilization of a contaminant in soil through

absorption and accumulation by roots, adsorption onto roots, or precipitation within the root zone

of plants, and (b) the use of plants and plant roots to prevent contaminant migration via wind and

water erosion, leaching, and soil dispersion.

Phytostabilization occurs through root-zone microbiology and chemistry, and/or alteration

of the soil environment or contaminant chemistry. Soil pH may be changed by plant root exudates

or through the production of CO2. Phytostabilization can change metal solubility and mobility or

impact the dissociation of organic compounds. The plant affected soil environment can convert

metals from a soluble to an insoluble oxidation state (Salt et al. 1995). Phytostabilization can also

occur through sorption, precipitation, complexation, or metal valence reduction (EPA, 1987).

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 Plants can also be used to reduce the erosion of metal contaminated soil. This process

reduces the mobility of the contaminant and prevents: migration to the ground water, and it reduces

bio-availability of metal into the food chain. This technique can also be used to re-establish

vegetation cover at sites where natural vegetation fails to survive due to high metals concentrations

in surface soils or physical disturbances to surface materials. Metal-tolerant species is used to

restore vegetation at contaminated sites, thereby decreasing the potential migration of pollutants

through wind erosion and transport of exposed surface soils and leaching of soil contamination to

groundwater.

The term phytolignification has been used to refer to a form of phytostabilization in which

organic compounds are incorporated into plant lignin (Cunningham et al. 1995b). Compounds can

also be incorporated into humic material in soils in a process likely related to phytostabilization in

its use of plant material. Phytostabilization is used in the treatment of soil, sediments, and sludge.

Advantages

Phytostabilization has the following advantages:

a. Soil removal is unnecessary.

b. It has a lower cost and is less disruptive than other more-vigorous soil remedial

technologies.

c. Re-vegetation enhances ecosystem restoration.

d. Disposal of hazardous materials or biomass is not required.

Disadvantages

Phytostabilization has the following disadvantages:

a. The contaminants remain in place. The vegetation and soil may require long-term

maintenance to prevent rerelease of the contaminants and future leaching.

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b. Vegetation may require extensive fertilization or soil modification using amendments.

c. Plant uptake of metals and translocation to the aboveground portion must be avoided.

d. The root zone, root exudates, contaminants, and soil amendments must be monitored to

prevent an increase in metal solubility and leaching.

e. Phytostabilization might be considered to only be an interim measure.

f. Contaminant stabilization might be due primarily to the effects of soil amendments, with

plants only contributing to stabilization by decreasing the amount of water moving through

the soil and by physically stabilizing the soil against erosion.

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MICROPHYTES
Microphytes or microalgae are microscopic algae, typically found in freshwater and marine

systems, living in both the water column and sediment (Thurman, 1997). They are unicellular

species which exist individually, or in chains or groups. Depending on the species, their sizes can

range from a few micrometers (µm) to a few hundred micrometers. Unlike higher plants,

microalgae do not have roots, stems, or leaves. They are specially adapted to an environment

dominated by viscous forces. Microalgae, capable of performing photosynthesis, are important for

life on earth; they produce approximately half of the atmospheric oxygen and use simultaneously

the greenhouse gas carbon dioxide to grow photoautotrophically. Microalgae, together with

bacteria, form the base of the food web and provide energy for all the trophic levels above them.

Microalgae biomass is often measured with chlorophyll a concentrations and can provide a useful

index of potential production. The standing stock of microphytes is closely related to that of its

predators. Without grazing pressures the standing stock of microphytes dramatically decreases

(Thrush et al, 2006)

The biodiversity of microalgae is enormous and they represent an almost untapped

resource. It has been estimated that about 200,000-800,000 species in many different genera exist

of which about 50,000 species are described (Starck, 2012). Over 15,000 novel compounds

originating from algal biomass have been chemically determined (Cardozo, 2006). Most of these

microalgae species produce unique products like carotenoids, antioxidants, fatty acids, enzymes,

polymers, peptides, toxins and sterols.

Microalgae is an important source of nutrition and is used widely in the aquaculture of

other organisms, either directly or as an added source of basic nutrients. Aquaculture farms rearing

larvae of molluscs, echinoderms, crustaceans and fish use microalgae as a source of nutrition. Low

12
bacteria and high microalgal biomass is a crucial food source for shellfish aquaculture. Microalgae

can form the start of a chain of further aquaculture processes. For example, microalgae is an

important food source in the aquaculture of brine shrimp. Brine shrimp produce dormant eggs,

called cysts, which can be stored for long periods and then hatched on demand to provide a

convenient form of live feed for the aquaculture of larval fish and crustaceans.

Other applications of microalgae within aquaculture include increasing the aesthetic appeal

of finfish bred in captivity. One such example can be noted in the aquaculture of salmon, where

microalgae is used to make salmon flesh pinker. This is achieved by the addition of natural

pigments containing carotenoids such as astaxanthin produced from the microalgae

Haematococcus to the diet of farmed animals.

CHARACTERISTICS AND USES

The chemical composition of microalgae is not an intrinsic constant factor but varies over

a wide range, both depending on species and on cultivation conditions. Some microalgae have the

capacity to acclimate to changes in environmental conditions by altering their chemical

composition in response to environmental variability. A particularly dramatic example is their

ability to replace phospholipids with non-phosphorus membrane lipids in P-depleted

environments. It is possible to accumulate the desired products in microalgae to a large extent by

changing environmental factors, like temperature, illumination, pH, carbondioxide supply, salt and

nutrients. Microphytes also produce chemical signals which contribute to prey selection, defense,

and avoidance.

These chemical signals affect large scale tropic structures such as algal blooms but

propagate by simple diffusion and laminar advective flow. Microalgae such as microphytes

constitute the basic foodstuff for numerous aquaculture species, especially filtering bivalves.

13
Photosynthetic and chemosynthetic microbes can also form symbiotic relationships with host

organisms. They provide them with vitamins and polyunsaturated fatty acids, necessary for the

growth of the bivalves which are unable to synthesize it themselves.

In addition, because the cells grow in aqueous suspension, they have more efficient access

to water, carbondioxide, and other nutrients. Microalgae play a major role in nutrient cycling and

fixing inorganic carbon into organic molecules. While fish oil has become famous for its omega-

3 fatty acid content, fish don't actually produce omega-3s, instead accumulating their omega-3

reserves by consuming microalgae. These omega-3 fatty acids can be obtained in the human diet

directly from the microalgae that produce them.

TECHNIQUES FOR MASS CULTIVATION OF MICROPHYTE

Opens systems: open ponds
Normally, microalgae and cyanobacteria large scale mass cultivation is done in shallow

open ponds tanks, of circular or raceway type, with solar light. One of the major advantages of

using open systems is that they are easy to build, operate, and they have lower costs than closed

systems. Even though it has been demonstrated that open pond culture is economically feasible,

they still have some disadvantages and limitations, they use light in a very inefficient manner, have

evaporation water loses, low carbondioxide mass transfer rate from the atmosphere, due to its

inefficient mixing mechanisms; open ponds also require a large area of land for the culture due to

its shallow depth. Additionally, open systems can be contaminated with predators or fast growing

microorganisms like bacteria that can compete with microalgae for nutrients, this is why open

systems are only used for organisms that can tolerate extreme conditions, like high salinity or pH.

The scaling up of open ponds culture systems can only be performed by increasing the area,

because increasing depth will not increase light penetration leading to lower productivities.

14
Closed systems: photobioreactors
To overcome the problems detected in open systems it has been proposed the use of closed

photobioreactors (PBRs). The former are more appropriate for strains that cannot tolerate extreme

environments or when final product is highly susceptible to degradation or contamination. Closed

systems also allow the prevention of contamination, allowing the operation in culture modes like

photoautotrophic, heterotrophic, or mixotrophic. Also, closed systems can obtain up to three times

more biomass than open systems, thus reducing harvesting costs. Despite the great advances that

have been achieved in the construction and operation of PBRs, its technology is still in

development. Around 90% of current biomass production worldwide is obtained in open systems,

despite the fact that PBR technologies offer greater potential in terms of productivity, control of

culture conditions, and applicability to cultivate various strains.

15
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Symposium, April 28 - May 1, 1997, New Orleans, LA. 3:303.

Bañuelos, G. S., H. A. Ajwa, B. Mackey, L. L. Wu, C. Cook, S. Akohoue, and S. Zambrzuski.

1997b. Evaluation of Different Plant Species Used for Phytoremediation of High Soil
Selenium. J. Environ. Qual. 26:639-646.
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