Review Paper Nisha

EnvironmentalScienceProcesses & Impacts

CRITICAL REVIEW

Publ

ishe

d on

12

Nov

embe

r 20

13. D

ownl

oade

d by

RSC

Int

erna

l on

23/1

2/20

13 1

0:52

:49.

View Article OnlineView Journal

A review with rec

School of Biotechnology, Rajiv Gandhi Pro

India. E-mail: [email protected]; T

Cite this: DOI: 10.1039/c3em00491k

Received 26th September 2013Accepted 12th November 2013

DOI: 10.1039/c3em00491k

rsc.li/process-impacts

This journal is © The Royal Society of

ent advancements onbioremediation-based abolition of heavy metals

Nisha Gaur,* Gagan Flora, Mahavir Yadav and Archana Tiwari

There has been a significant rise in the levels of heavy metals (Pb, As, Hg and Cd) due to their increased

industrial usage causing a severe concern to public health. The accumulation of heavy metals generates

oxidative stress in the body causing fatal effects to important biological processes leading to cell death.

Therefore, there is an imperative need to explore efficient and effective methods for the eradication of

these heavy metals as against the conventionally used uneconomical and time consuming strategies that

have numerous environmental hazards. One such eco-friendly, low cost and efficient alternative to

target heavy metals is bioremediation technology that utilizes various microorganisms, green plants or

enzymes for the abolition of heavy metals from polluted sites. This review comprehensively discusses

toxicological manifestations of heavy metals along with the detailed description of bioremediation

technologies employed such as phytoremediation and biosorption for the potential removal of these

metals. It also updates readers about recent advances in bioremediation technologies like the use of

nanoparticles, non-living biomass and transgenic crops.

Environmental impact

Levels of heavy metals are rising in the environment due to increased industrial usage causing severe damage to all spheres of life. Commonly followed methodslike ion exchange, chemical precipitation, reverse osmosis, bio-piles, bio-slurries and land-lling are not only expensive but their byproducts are hazardous tothe environment. Bioremediation is an upcoming technique which utilizes eco-friendly agents like enzymes, microorganisms and plants and can prove to be asuitable alternative for the elimination of these heavy metals. It is imperative to carry out conclusive research which can rene and improve this process to a levelwhere it can be accepted universally. On this note this review throws light on the technology of bioremediation and discusses recent additions to this area.

Introduction

Pollution refers to the state of existence of undesirablesubstances (pollutants) in the environment beyond a permis-sible limit which can harmfully affect every sphere of life.Sources of pollution can be both natural and anthropogenic.Natural sources include geothermal activities, comets, spacedust and volcanic activities. Whereas, anthropogenic sourceshave arisen mainly on account of rapid industrialization andextensive use of chemical substances such as hydrocarbons,pesticides, chlorinated hydrocarbons and heavy metals.1 Thelatter mentioned source is the major contributor to pollution incontrast to the former.2 Out of a large number of aforemen-tioned anthropogenic sources, toxicological manifestationscaused by heavy metals are well known and are considered ashighly detrimental.

Lead (Pb), cadmium (Cd), arsenic (As) and mercury (Hg) arethe major pollutants that bring about heavy metal toxicity. Thenon-biodegradable nature of these metals is the principle

udyogiki Vishwavidyalaya, Bhopal, M.P.,

el: +91 8234884887

Chemistry 2014

reason that leads to their prolonged presence in the environ-ment. Moreover, these metals can enter into the food chain andover a period of time become accumulated in the human body.This accumulation can cause many health effects which mightbe irreversible in nature.3

Chelation therapy is the mainstay of the treatment regimefollowed so far for curing heavy metal poisoning. However, thistherapy is coupled with severe side effects as apart from theremoval of toxic metals it also eliminates important mineralsand metals from the body like iron (Fe), calcium (Ca), zinc (Zn)etc. which directly affects normal biological processes of thebody.4 Thus, rather than a curative approach using chelation totreat heavy metal poisoning, a preventive approach can be aneffective alternative focusing on the eradication of these heavymetals from the environment itself.

Conventional methods like ion exchange, chemical precipi-tation, reverse osmosis, bio-piles, bio-slurries, and land-llingare used conventionally for the remediation of heavy metalspresent in water and soil.5 However, they suffer from a majordrawback of being expensive owing to the requirement ofsophisticated infrastructure. Moreover, they also generate toxic

Environ. Sci.: Processes Impacts

http://dx.doi.org/10.1039/c3em00491k

http://pubs.rsc.org/en/journals/journal/EM

Environmental Science: Processes & Impacts Critical Review

Publ

ishe

d on

12

Nov

embe

r 20

13. D

ownl

oade

d by

RSC

Int

erna

l on

23/1

2/20

13 1

0:52

:49.

View Article Online

sludge which affects environment and might not completelyremove the metals.6

Bioremediation is one such eco-friendly and sustainableprocess that can prove to be much more effective and efficientfor eliminating heavy metals present in different spheres of theenvironment. This strategy aims to clean up the environmentwhile maintaining the normal biological processes associatedwith it.7 According to Glazer and Nikaido, bioremediation isdened as a process that uses microorganisms, green plants orenzymes to treat the polluted sites for regaining their healthycondition.8 This technique is highly favored as it provides muchbetter results through the application of low cost and economicinputs in comparison to conventional means. Bioremediationcan thus be regarded as a highly cost-effective, eco-friendly andmore pronounced solution to the problems arising due to theuse of transition metals.

Heavy metal pollution in soil and water

Heavy metals such as Pb, As, Cd and Hg are ubiquitous innature and cause an unfavorable result on the surroundingsparticularly at high concentrations. Even though the heavymetals biochemical equivalence and geochemical cycles arenormal components of the earth's crust, their concentration hasbecome remarkably exacerbated following the advent of theindustrial revolution which resulted in a manifold rise in thelevel of usage of these metals.9

Nisha Gaur received her M.Techin Biotechnology from RajivGandhi Technological Univer-sity, Bhopal, India. She iscurrently a research trainee atR&D department of Kilpest IndiaPrivate Limited. Her researchinterests include bioremediationof organic and inorganic wastes.

Gagan Flora received his M.Techin Biotechnology from RajivGandhi Technological Univer-sity, Bhopal, India. He will verysoon be starting his doctoratestudies. His research interestsinclude toxicity of heavy metals.


These heavy metals are known to facilitate phytotoxicitythrough contamination of soil, a problem that has called forconsiderable attention over the past few decades. The pres-ence of heavy metals in soil can considerably decrease the sizeof the microbial community along with reduction of environ-mental and biological activities such as organic mattermineralization and leaf litter decomposition.10 The level ofcontamination however depends on factors such as chemicalcomposition, toxicity, mobility and varying bioavailability ofthe metal.11 As soon as these heavy metals come in contactwith the soil surface they initially become readily adsorbed,which is followed by slow adsorption and distribution inthe soil.12,13 When the plants grow on metal-polluted soil theytend to accumulate these heavy metals, which greatly affecttheir growth and development. This not only threatenstheir survival but also affects the life which consumes them.Owing to this contamination, a large amount of terrain hasturned out to be dangerous and non-arable for humans andanimals.

In a manner similar to soil, both surface water and groundwater can easily become contaminated by heavy metalsthrough natural sources (leaching of ore, erosion of mineralswith sediments and volcanic extruded products) or humanactivities (chemical fertilizers, pesticides, solid waste disposal,industrial and domestic wastes). Due to its polarity andhydrogen bonds, it may adsorb, dissolve and absorb manydifferent compounds.

Mahavir Yadav received hisPh.D in Molecular Biology at theInstitute of Microbial Tech-nology, Chandigarh, India. He iscurrently working as an Assis-tant Professor in the School ofBiotechnology at Rajiv GandhiTechnological University, Bho-pal, India. His research interestsinclude biodiesel productionand bioremediation of heavymetals.

Archana Tiwari received herPh.D in Environmental Sciencesfrom Barkatullah University,Bhopal, India. She is currentlyworking as an AssociateProfessor and Head of theSchool of Biotechnology, RajivGandhi Technological Univer-sity, Bhopal, India. Her researchinterests include production ofbioplastics from biologicalsources.

This journal is © The Royal Society of Chemistry 2014


Critical Review Environmental Science: Processes & Impacts

Publ

ishe

d on

12

Nov

embe

r 20

13. D

ownl

oade

d by

RSC

Int

erna

l on

23/1

2/20

13 1

0:52

:49.

View Article Online

Toxicity of heavy metals

Metals having high atomic weight and a density of more than5 g cm�3 are regarded as heavy metals or transition metals.More than 20 different kinds of heavy metals are found innature but only a few of them are of concern to human health.According to the Agency for Toxic Substances and DiseaseRegistry (ATSDR), lead, cadmium, arsenic and mercury are wellknown for showing a toxicity prole upon exposure. Althoughnumerous cellular, intracellular and molecular mechanismshave been reported to underpin heavy metal toxicity, generationof oxidative stress is the well accepted mechanism whichexplains most of the arising symptoms14 (Fig. 1). Chiey, humanexposure to these metals occurs from industries and toxic wastesites. The non-biodegradable nature of these heavy metalsfurther results in their prolonged persistence in the environ-ment. It is now known that existence of transitionmetals even ata very low concentration (picomolar) in humans can result infatal health effects.15

1. Lead. Lead is a widely known ubiquitously presentxenobiotic heavy metal. Its unique properties like high ductility,highly malleability, low melting point and soness makes it animportant metal in industries such as automobiles, paint,ceramics, plastics etc.16 Due to this widespread usage, humanshave become vulnerable targets for its exposure. No level of leadhas been considered to be safe or benecial to living beings.Upon exposure it affects many organs like the nervous system,renal system, hematopoietic system, reproductive system andcardiovascular system, and shows some effects on bone. Thenervous system is the most sensitive target compared to theothers for lead-induced toxicity.17,18

High exposure of lead may cause fatal consequences likeconvulsions, lack of coordination, delirium and paralysis. Italso affects the hematopoietic system which inhibits the

Fig. 1 Generation of oxidative stress in cell owing to exposure ofheavy metals such as Pb+2, As+2, Cd+2, and Hg+2, leads to formation ofreactive oxygen species and impairs anti-oxidant defense causing celldeath.


synthesis of hemoglobin and thus causes anemia. Renaldysfunction has also been reported on account of lead-inducedtoxicity.19,20

The major mechanism of lead-induced toxicity is inductionof oxidative stress which occurs as a result of imbalancebetween pro-oxidant and anti-oxidant ratio. This imbalancebrings about protein oxidation, lipid peroxidation and nucleicacid peroxidation making a cell prone to cell death.21 The ionicmechanism is the other mode of action of lead toxicity. In thisprocess, lead mimics and substitutes other monovalent andbivalent ions like Na+, Ca2+, and Mg2+, and hinders many bio-logical process like intracellular signaling, cell adhesion,protein folding, ionic transportation etc.17

Chelation therapy has been regarded as the mainstay treat-ment which involves introduction of chelating agents likecalcium disodium ethylenediaminetetraacetic acid (CaN-a2EDTA), D-penicillamine (DPA), Dimercaprol (BAL) and Suc-cimer into the organism. These chelating agents then bind tothe lead ions forming a complex known as a chelate which isexcreted out of the body mainly through urine. Many naturalanti-oxidants like vitamins (B, C and E), avonoids, and herbalavonoids have also been used for curing lead-induced toxicity.4

2. Cadmium. Cadmium is an extremely toxic metal havingdistinctive properties such as good lustre, high ductility,malleability and soness that have led to its extensive usage indiverse industries like Ni–Cd batteries, coatings and plating,and as stabilizers for plastics.22

It causes many adverse health effects by damaging kidney,liver, bone and cardiac tissues. The kidneys and liver are thechief targets for cadmium-induced toxicity. Nephropathy is themost common renal abnormality that occurs owing to cadmiumexposure. Renal vitamin D metabolism is also affected whencadmium accumulates in the kidney.23 This signicantly bringsabout a calcium imbalance which leads to osteoporosis andosteomalacia as well as increased excretion of calcium (medicalcondition referred to as Itai-Itai).24 Cadmium is also regarded asa potent human carcinogen that is associated with high risk ofrenal and prostate cancer. It is also known to act as a robustmutagen and can cause multi-locus deletions.

Cadmium weakens the antioxidant defense by severelyreducing the intracellular glutathione levels. It also inhibits theactivity of various antioxidant enzymes like superoxide dis-mutase and catalase along with generation of ROS.25 Thecombinatorial effect of these processes renders cells into a stateof oxidative stress. The increased level of ROS causes damage toDNA and inhibits DNA repair resulting in mutation.23

3. Arsenic. According to the ATSDR, arsenic is regarded asthe most common cause of acute heavy metal poisoning inadults and children.26 Arsenic is a ductile metalloid which existsin three allotropic forms: metallic grey, yellow and blackarsenic. It is broadly used to make insecticides, fungicides,weed killer, antifouling agents and in preserving woods.

Clinical manifestation of arsenic is referred to as Arsenicosisand is caused by the prolonged exposure of arsenic in humans.Pigmentation (development of spotty rain drop patches over thefront of chest) and keratosis are the toxic aermaths on theskin.27 Arsenic toxicity leads to many respiratory diseases like




Publ

ishe

d on

12

Nov

embe

r 20

13. D

ownl

oade

d by

RSC

Int

erna

l on

23/1

2/20

13 1

0:52

:49.

View Article Online

reduced pulmonary function, lung cancer, chronic cough orchronic bronchitis. Peripheral neuritis, black foot disease, liverbrosis and gastroenteritis is also caused due to the uptake ofarsenic-contaminated water.28

Unlike lead and cadmium, the molecular mechanismunderlying arsenic toxicity is multi factorial. It involves gener-ation of oxidative stress, suppression of DNA repair, inhibitionof cell cycle check points and induction of apoptosis.29

Chelation therapy is considered as the most preferredapproach to control the toxic effects of arsenic. Numerousavonoids, vitamins and herbal extracts have also been repor-ted for curing and preventing arsenic-mediated cellular andmolecular damage.26,30

4. Mercury. Mercury is a naturally occurring metal that canexist in inorganic, organic (ethyl-, methyl-, alkyl-, or phenyl-mercury) and vapor states, with the organic state consideredmore hazardous in comparison to other forms. It has bothindustrial (batteries, fossil fuel emission, paints, cosmeticproducts etc.) as well as clinical applications (thermometers,sphygmomanometer, barometers etc.).31 In the environment, itsexposure occurs through the erosion of mercury-containingores and in the form of gases dissipating from volcanic erup-tions which are rich in mercury.

The route of absorption of all forms of mercury is different.95–100% of organic mercury (methylmercury) is absorbed in theintestinal tract and almost 100% gets inhaled through vapor.The absorption rate of elemental mercury is less as compared toorganic mercury and is found to be around 75–85% occurringmainly through inhalation. Inorganic mercury is absorbed atmuch lower rates (7–15%) of ingested dose and 2–3% of dermaldose.31,32

Two forms of mercury, i.e. organic and elemental, are lipo-philic in nature and become distributed throughout the body.Both forms can cross the blood brain barrier (BBB) and even theplacental barrier and nally accumulate in the brain andkidneys. Whereas, inorganic mercury is not capable of crossingthe BBB or placental barrier. It is found in the brain neonatesand accumulates in the kidneys.31,33

Mercury is a powerful neurotoxin which primarily affects thecentral nervous system.34 It may lead to lack of coordination ofmovements, impairment of speech and hearing, and muscleweakness. The lungs absorbs metallic mercury through thebreathing process which can lead to respiratory impairment. Ithas diverse mechanisms through which it can cause biochem-ical damage to tissues and genes. Mercury induces toxicity byforming free radicals and generating oxidative stress.35 It alsobind to thiol-containing enzymes and inhibits them.31 Methyl-mercury forms complexes with cysteine, a thiol-containingcompound, which helps in intracellular absorption.36

Fig. 2 Different mechanisms involved in phytoremediation.

Phytoremediation: a robust strategy forthe eradication of toxic heavy metals

Phytoremediation is as an emerging technology that involvesapplication of selected plants to degrade, assimilate, metabolizeor detoxify undesirable substances like heavy metals, pesticides,


hydrocarbons, organic solvents and crude oil from soil andwater to improve its quality.37,38

Although every plant has the capability to remove contami-nants, only a few selected or engineered plants are used exten-sively to remove contaminants efficiently such as Clerodendruminfortunatum, Croton bonplandianus, Pistia stratiotes, Thlaspicaerluescens, Brassica junceae, Alysum lesbiacum, etc.39 Phytor-emedial strategies applied in context to heavy metal eliminationtake into consideration any of the following methods depend-ing upon the nature of the contaminant:

� Complete removal of the accumulated heavy metals.� Degradation or containment of heavy metals.� Combination of these.Compared to conventional strategies being followed (in situ

vitrication, soil incineration, excavation and landll, soilwashing, soil ushing and solidication) phytoremediation isan aesthetically pleasing, efficient and eco-friendly process inremoving contaminants from low to moderate levels.40,41 It isalso an economical method which reduces the cost to less thanthe half the price of the conventional methods. Moreover, itrequires low installation and maintenance costs. It alsoprovides an added advantage by not only cleaning polluted soilbut by also preventing soil erosion and metal leaching.42

Mechanism of phytoremediation

Specically, the process of phytoremediation is broadly dividedinto two phases for the sequestration of heavy metals from soiland water: ex situ and in situ. The ex situ bioremediation processfor soil and water is a two-step method. Firstly, it involves theexcavation of contaminated soil or pumping out the ground-water for treatment. The soil and water is then subjected toseveral chemical and physical methods like chemical reduction/oxidation, dehalogenation, soil washing, uid vapour extrac-tion, stabilization/solidication, and solvent extraction toeradicate the contaminants.43 Thereaer, the treated soil orwater which is free from heavy metals is restored back to theoriginal site. The removed pollutants are then transported tosome other site for dumping.42 Although, this approach is lesstime consuming and can be performed under controlledconditions, due to dumping and off-site burial of the removedcontaminants at the time of treatment, it can act as anotherthreat to the environment at another location.44



Fig. 3 Mechanisms of phytoremediation.


Publ

ishe

d on

12

Nov

embe

r 20

13. D

ownl

oade

d by

RSC

Int

erna

l on

23/1

2/20

13 1

0:52

:49.

View Article Online

The in situ method on the other hand is a technology whichremoves the heavy metals from contaminated soil, water and airwithout performing excavation and transport of contaminants.The treatment regimes are carried out at the same site thatprecludes off-site burial of the removed pollutants and thusprevents contamination of the clean soil.42 This method causesless ecological disturbance and is also economically viablewhich makes it a better alternative than ex situ technology. Thismethod is further divided into different categories to removetoxic metals from soil and water: phytoextraction, phytoltra-tion, phytostabilization, phytovolatilization, phytodegradation,and rhizodegradation (Fig. 2 and 3).

Phytoextraction

Also known as phytoaccumulation, this process includes theextraction of toxic metals from soil and water without disturb-ing its integrity. The absorption and uptake of heavy metals isperformed by plant roots followed by their translocation andnally accumulation and concentration above ground in thebiomass (shoots).43 Generally, this method is favored for thesites that are discreetly or supercially polluted.

The underlying mechanism behind phytoextration ishyperaccumulation. It is the process in which the plants likePteris vittata L, Thlaspi rotundifolium (L.) Gaudin, Fagopyrumesculentum Moench, and Betula papyrifera Marsh can accumu-late toxic metals at relatively higher concentrations. Hyper-accumulation is widely favored on metalliferous soils (soilsaffected with high concentrations of transition metals) and theplants which grow on this type of soil are referred to as metal-lophytes.45 The plants used for phytoextraction should havehigh hyper-accumulating capacity and should be capable ofgrowing on highly toxic soils (and water) in order to make the


extraction and translocation process of toxic metals in theshoots effective.46

In this process, regular cropping of the hyperaccumulatorplant is required until the metal concentration reduces down tothe desired level at the concerned site. Aer that the contami-nated plant biomass is either burned and converted into ash oris used in various industries (e.g. wood, cardboard, etc.). Thisprocess can also be put into commercial use such as phyto-mining which essentially involves extraction of biomass in theform of bio-ore (extracts of saleable heavy metals obtained bythe plant biomass ash).47

The phenomenon of hyperaccumulation is of two types:natural hyperaccumulation and chemically-enhanced hyper-accumulation. Natural hyperaccumulation utilizes specickinds of hyperaccumulators capable of absorbing the toxicmetals in the roots followed by its translocation in shoots.Finally, storage of these translocated heavy metals occurs the inaerial portion of the plant in a nontoxic form. These plants havehigh tolerance capacity and are credited with the ability of hightranslocation mobility of metals by secreting metal chelatingcompounds (phytosiderophores) and organic acids.48 Chemi-cally-enhanced hyperaccumulation is applied in case of somemetals like lead and gold that are immobile in soil and cannotbe absorbed readily. For this purpose some chemical inducerslike chelating agents (EDTA, NTA, malate etc.) or acidifyingagents are used which enhance their mobility in the soil byincreasing the bioavailability of the metals in soil which ulti-mately boosts their uptake.49

Kaur et al. reported that the chemically-enhanced phytoex-traction showed better accumulation capacity as compared tothe natural phytoextraction process. They also found thatBrassica juncea arawali can act as an excellent chemically-enhanced hyperaccumulator.50 A recent study showed theuptake of heavy metals from municipal solid waste by chelate-assisted Festuca arundinacea. It was also reported that thenitrilotriacetic acid signicantly enhanced the metal accumu-lation capacity of Festuca arundinacea in contrast to itsabsence.51 In a phytoextraction analysis the effect of inducerslike EDTA on texturally different soil was assessed. It wasrevealed that the induction of EDTA considerably enhanced thelead accumulation capacity of wheat shoots in loamy sand thanthat of the sandy clay loamy soil.52 In a recent study, assessmentof natural plants in Turkish serpentine soil was carried out foranalysis of its Ni accumulation capacity. Scientists tried toestablish the possible relationship between amount of phytoa-vailable Ni in the soil and the Ni content of potential accumu-lator plants. It was found that susceptibility and Ni requirementof a plant was species specic. They presented Isati spinnatilobaas potential Ni hyperaccumulator species.53

Phytoextraction can be performed by three means i.e. phy-toextraction by trees, by crops and by grasses. Each has its ownadvantages and disadvantages. Phytoextraction by treesproduces high biomass but due to shedding of leaves on tosurface the metal again becomes transported into the soil.Phytoextraction by grasses has high metal accumulationcapacity but has low biomass production as well as slow growthrate. Phytoextraction by crops has both the above mentioned




Publ

ishe

d on

12

Nov

embe

r 20

13. D

ownl

oade

d by

RSC

Int

erna

l on

23/1

2/20

13 1

0:52

:49.

View Article Online

advantages. However, they pose a threat due to the ingestion ofcrops by herbivores and thus entry of metals into the foodchain.54

There are four major steps involved in metal hyper-accumulation in plants:55

1. Bio-activation of trace metals in the rhizosphere.2. Root adsorption and compartmentation with the help of

transporters and chelators.3. Metal uptake by shoots.4. Distribution, detoxication, and sequestration of metal

ions.Hyperaccumulators need to have the facility of metal

homeostasis while growing in an impure surrounding. TheThlaspi family are hyperaccumulating plants among whichtwenty three species hyperaccumulate nickel, ten specieshyperaccumulate zinc, three species (T. caerulescens, T. praecoxand T. goesingense) hyperaccumulate cadmium and one specieshyperaccumulates lead.56 T. caerulescens is regarded as one ofthe nest and well-known hyperaccumulators.57 Interestingly,this plant is able to grow in serpentine soils, which containelevated levels of heavy metals including Zn, Co, Pb, Cr, Cd andNi, being capable of up taking up to 30 000 and 1000 mg kg�1

Zn and Cd, respectively in their shoots, while its developmentremains unaffected.58

Moreover, due to the advancement in genetic engineering,the genes which help in remediation of heavy metals have beenisolated and then inserted into the large biomass producingnon-accumulating plants.59

Phytofiltration

This process can be carried out in both terrestrial and aquaticenvironments, though mainly it is carried out to purify groundwater or other water bodies.60 This process enables the plantroots to absorb or adsorb, concentrate and precipitate the heavymetals from a polluted effluent source (industrial discharge,agricultural runoff, acid-mine drainage etc.). If plants use rootsfor remediation purposes then it is known as rhizoltration.Such kind of plants shows rapid growth of roots and removaltime of toxic metals is minimal.61

This process involves absorption of heavy metals by plantsroots followed by accumulation and transportation in the stemor leaves. The contaminants are removed through harvesting atan appropriate time.62 Development of feeder layer fertilization(suspending several layers of soil above a polluted stream ofwater through which the plant obtains the nutrients andsimultaneously removes heavy metals) system led to a boost inrhizofertilization technology. Extensive root network develop-ment can be carried out by regular application of concentratedfertilizers to this feeder layer.63

A second generation technology which uses plant seedlingsfor the removal of heavy metals from contaminated water isknown as blastoltration.64 The seedlings have the potential toabsorb or adsorb a high percentage of toxic heavy metals.Special kind of seedling cultures are prepared througheconomic means using seeds, water and along with appropriateexposure to light and darkness.65 This process shows


advantages over rhizoltration because of the fact that seedscan grow independent of environmental conditions and absorbhigher amounts of heavy metal during initial phase of theirlife cycle.66

Removal of lead from wastewater using Carexpendula wasachieved by Yadav et al. using the rhizoltration technique.They carried out pot and simulation experiments and foundthat Carexpendula accumulated a signicant amount of leadespecially in root biomass as compared to shoot.67 In anotherstudy, rhizoltration of cadmium and lead was performed byusing four different macrophytes (Pistiastratiotes L., Salviniaauriculata Aubl, Salvinia minima Baker and AzollaliculoidesLam). It was seen that Pistiastratiotes L. had extensive bio-accumulation efficiency of removing lead and cadmium. Theaccumulation of these two heavy metals in the roots was 10-foldhigher than that of the leaves.68 Vesely et al. reported the effi-ciency of organic acids to enhance the mobility of heavy metalsthrough rhizoltration. They studied the bioaccumulationpotential of Pistiastratiotes L. against the removal of Cd, Pb andZn. They found that the organic acid substantially increased themobility of all heavy metals. However, translocation of heavymetals decreased in the plant in a time-dependent manner.69

Rhizoltration has an advantage of absorbing metals readilybut this method works only in water and not in soil. Moreover,metals get accumulated in the plant biomass which must bedisposed of regularly to reduce the risk of contamination.70

Phytostabilization

Phytostabilization is a process which involves absorption andprecipitation of contaminants like heavy metals by plantsthrough immobilization. The process aims at the stabilizationof the heavy metal at the contaminated site instead of itsremoval. This prevents movement of these contaminants viaground water and wind.71 The underlying mechanisms thatdetermine the phenomenon of phytostabilization are asfollows:72,73

(a) Phytostabilization in the root zone: In this the root getsexudated (converting contaminants into less bioavailable form)in the rhizosphere so as to immobilize the heavy metal in theroot zone itself.

(b) Phytostabilization of the root membrane: This step leadsto the binding of the heavy metals to the root surfaces whichprevents their entry inside the plant.

(c) Phytostabilization in the root cells: This step furtherprevents the translocation of heavymetals by sequestering theminto the cell vacuole.

For effective phytostabilization, the plants should have richroot (to absorb large quantity of water) and shoot systems but apoor translocation mechanism so as to prevent entry of heavymetals into the shoots. Dense coverings of shoots tend toincrease transpiration which prevents precipitation of heavymetals into the groundwater. Moreover, upward ow can bemaintained by fast transpiration by plants which preventsdownward leaching.62

Cambrolle et al. investigated the capability of two Spartinaspecies in terms of phytostabilization and bioaccumulation of




Publ

ishe

d on

12

Nov

embe

r 20

13. D

ownl

oade

d by

RSC

Int

erna

l on

23/1

2/20

13 1

0:52

:49.

View Article Online

heavy metals like Co, Cr and Ni in two marshes with differentlevels of contamination. They reported that in all the sitesamples, the concentration of these heavy metals in bothspecies were higher in below-ground tissues as compared to theabove-ground tissues. Both species of Spartina showed goodphytostabilization capacity towards Co in the contaminatedsoil.74 Varun et al. reported the phytostabilization potential ofTyphalatifolia L. in industrial sludge. Their ndings showedthat Typhalatifolia L. had higher potential for immobilization ofZn, Mn, Cr and As but had less phytostabilisation potentialtowards Ni, Cd and Co.75 Recently, the phytostabilizationcapacity of six different plant species was assessed for theimmobilization of the lead in mine tailing using eld and potexperiments. It was found that A. mangium had the best abilityfor phytostabilization towards lead mine tailing out of six plantspecies. A. mangium stored a higher concentration of lead in theroots which aerward could be used in the timber industry orpaper industry etc.76 In another study, a Sorghum species wasused by Soudek et al. for the immobilization of heavy metals(Zn, Cd) in soil deposited due to industrial activities. Theyfound that initially the root had a higher concentration of heavymetal, but as the concentration of zinc and cadmium in thesolution increased they were transferred into the shoots whichultimately caused toxicity to leaves. The toxicity affected the Chla/b ratio in the leaves.77

This technology is highly cost-effective in nature and doesnot require the disposal of soil and contaminants aer treat-ment. However, this technology is not feasible for all sites(restricted only to water) and also requires containment ofcontaminants for an indenite period as they remain inside thesoil for a long time.78

Phytovolatilization

This process uses plants for the uptake of contaminants fromsoil and water followed by subsequent degradation into lesstoxic forms which are then transpired into the environment.79

Plants can volatilize both organic and inorganic contaminantsprovided that the inorganic contaminants should not formmethyl and hydride derivatives.80 Contaminants which havehigh Henry's constant (KH is characteristic of particular solute,solvent and temperature)81 i.e. KH> 10 atm-m3 water per (m3 air)are applicable for the phytovolatilization mechanism.82

The mechanism includes open stomata of the leaves todiffuse volatile contaminants in the environment in less toxicforms. The plants used in this process shows high levels of uxof the pollutant towards the atmosphere through the transpi-ration process.80 This method not only removes the pollutantsfrom contaminated site in a volatile form but the removal isdone in safer forms of that particular pollutant.

Sakakibara et al. reported the eradication of As through aremediation process by using Pterisvittata. They found thatPterisvettata had a good efficiency of volatilizing As (90%) fromarsenic-polluted soil. However, secondary arsenic pollution wassaid to be caused if a large amount of arsenic is released into theenvironment.83 In another study, the effect of ethylene glycol onthe phytovolatilization of 1,4-dioxane was estimated. DN34


poplar trees were used for this study and it was seen that when10 g l�1 of ethylene glycol was present in ground water itreduced the growth rate of plants to 28%. Similarly the effect ofethylene glycol on Arabidopsis was also observed and it wasunderstood that ethylene glycol had an inhibitory effect on itsgrowth and under hydroponic conditions it inhibited the phy-tovolatilization of 1,4-dioxane.84 Carvalho et al. carried outstudies on four aquatic plants (Typha domingensis, Lemnaobscura, Hydrilla verticillata Royle and Crinum americanum) forthe removal of aqueous selenium. Initially, they found that theplants accumulated the selenium in their tissues. But, later itwas concluded that the main mechanism behind seleniumaccumulation was phytovolatilization. In this process, plantsconverted the inorganic form of the selenium into the organicform which is less toxic and was then transpired.85

The advantage of this technology is that it does not requiredisposal of any contaminant thereby circumventing any sitedisturbance and erosion. This process is restricted only forabolition of volatile compounds and cannot be applied for theremoval of nonvolatile heavy metals. However, the maindisadvantage of phytovolatilization is that the heavy metals arestill toxic to some level even when they are volatilized. The rateof their migration and translocation cannot be predicted in thepolluted area.62

Phytodegradation

This process exploits the capability of plants that possesscertain specialized enzymes (dehalogenase, reductase and oxy-genase) or cofactors for the degradation of contaminants fromsoil and groundwater.86 This method is limited only to organicpollutants because these are biodegradable in nature. Phyto-degradation differs from rhizodegradation mainly because ofthe fact that the former encompasses the breakdown ofcontaminants with the help of microorganisms present in therhizosphere and is a relatively slower mechanism. Flavonoidsand carbohydrates secreted by plants facilitating phytode-gradation further enhance the microbial activity. Properties likesolubility, polarity, hydrophobicity and partitioning coefficient(Kd) of organic contaminants directly interferes with their entryinto plant through the root membrane.87 For the removal ofheavy metals some genetically modied plants have beendeveloped such as transgenic poplars.88

Farias et al. worked on petroleum-contaminated soil andstudied the tolerance and phytodegradation potential of Eryth-rina crista-galli L. in three different conditions: non-contami-nated soil, vegetation-contaminated soil and non-vegetationcontaminated soil. They found that the growth of Erythrinacrista-galli L. in vegetation-contaminated soil was reduced ascompared to non-contaminated soil. On the other hand thedegradation of petroleum in vegetation-contaminated soil washigher as compared to non-contaminated soil.89 Recently, astudy has been done on a transgenic tobacco plant whichexpresses bacterial organophosphorus hydrolase (an enzymethat degrades organophosphorus pesticides). Aer 14 days ofgrowth it was found that the tobacco plant degraded more than92% of methyl parathion and gives more root and shoot




Publ

ishe

d on

12

Nov

embe

r 20

13. D

ownl

oade

d by

RSC

Int

erna

l on

23/1

2/20

13 1

0:52

:49.

View Article Online

biomass as compared to the wild tobacco plant. This researchholds importance for the removal of organophosphoruscompounds from the environment.90

Biosorption

Biosorption is a process which uses biological materials for theremoval of contaminants through different mechanisms likeadsorption, absorption, surface complexation, precipitationand ion exchange. It depends on numerous factors likesubstance to be sorbed, environmental issues, biosorbent used,presence and absence of metabolic process (in living organ-isms).91 The two terms absorption (process in which onesubstance gets incorporated into another of different state) andadsorption (physical phenomenon in which adherence andbinding of ions or molecule occur on the surface of anothermolecule) comes under sorption process. In the case ofadsorption, the adsorbate is the substance which gets adsorbedon a solid surface and the adsorbent is the soil surface.92 If theadsorption phenomenon results in the formation of a stablemolecular phase at the interface, it is described as a surfacecomplex which can be of two types: inner and outer spheresurface complexes. In the former one, the adsorbent gets boundto at least onemolecule of the hydration sphere of the adsorbatebut in the latter one without any hydration sphere the moleculegets directly bound to the adsorbent.93,94

The contaminants that can be removed by biosorption couldbe organic and inorganic or soluble and insoluble. Metals (K+,Mg+) that are highly mobile and accordingly do not get accu-mulated with biomass during phytoremediation can be easilyremoved through biosorption.95 Heavy metals (lead, arsenic,cadmium, uranium, mercury) along with dyes, phenoliccompounds and pesticides are receiving a lot of attention fortheir eradication through this process.96

Fig. 4 Mechanisms of biosorption can be categorised on the basis ofcellular metabolism and location of biosorption. Further, cellularmetabolism based biosorption is divided into metabolism-dependentand non-metabolism-dependent. Metabolism-dependent includestransport through cell membrane and precipitation. Non-metabolism-dependent includes ion exchange, precipitation, complexation andsurface adsorption. On the basis of location, biosorption is classified asextra cellular accumulation/precipitation, cell surface sorption/precipitation, intracellular accumulation and further they are divided inthe same fashion as metabolism-dependent and non-metabolism-dependent processes.

Types of biosorbent

Primarily biosorbents fall into the following categories: livingbiomass and non-living biomass. Living biomass includesbacteria (gram-positive bacteria, gram-negative bacteria andcyanobacteria), fungi (mould, mushroom and yeast), algae(micro-algae, macro-algae, brown seaweeds and red seaweeds).While non-living biomass includes industrial waste (fermenta-tion wastes, food/beverages waste, activated sludges, anaerobicsludges), agricultural waste (fruit/vegetable waste, rice straws,wheat bran, soybean hull etc.), natural residues (plant residues,sawdust, tree bark, weeds etc.) and other biomaterials (chitosan-based materials, cellulose-based materials etc.).91,97

Many studies show that non-living biomass has gainedmore preference over living biomass for the biosorptionprocess because it does not require any maintenance andnutrient supply.98 Moreover, the biomass can be easilyobtained from industrial waste which adds the ease of avail-ability and makes the process economic.99 Whereas, livingbiomass demands proper maintenance of healthy microbialculture coupled with sustained environmental conditions.Even by providing these conditions, recovery of heavy metals


cannot be done efficiently from living biomass as metals bindintracellularly.98,100

Peptidoglycan carboxyl groups and phosphate groupsprovide the metal binding sites in gram-positive and gram-negative bacteria respectively.101 In addition, the proteinaceousS-layer and sheath also contribute to metal binding which aremade up of proteins and polysaccharides. The metal bindingcomponent of many cyanobacterial cell walls contains pepti-doglycan and some of them contain a sheath as well as extra-cellular polymeric substances.102 Pseudomurein is another cellwall component which resembles peptidoglycans present inarchaea bacteria along with sulfonated polysaccharides andglycoproteins which provides the anionic sites (carboxyl andsulphate groups).103 Amongst all the components present in thealgal cell wall, cellulose is common in all algal diversity whichalong with other components (depending on the presence) likepolysaccharides (mannan, alginic acid, xylans) and proteinsprovides the binding sites (phosphate, sulphate, hydroxyl,amine groups) for metal attachment.104

Chitins, glucans, mannans and proteins are the componentsof fungal cell walls. Apart from these it also contains otherpolysaccharides, lipids and pigments (melanin) which facilitatebinding of many metal ions.105 An important structuralcomponent of the fungal cell wall is chitin which is cheaper ascompared to activated carbon and acts as an efficient bio-sorbent for metals as well as radionuclides.106 Like chitin, chi-tosan (derived from deacetylation of chitin) and otherderivatives of chitin also have effective biosorption capacity.107

Carboxyl, phenolic, hydroxyl, carbonyl and methoxyl groups are




Publ

ishe

d on

12

Nov

embe

r 20

13. D

ownl

oade

d by

RSC

Int

erna

l on

23/1

2/20

13 1

0:52

:49.

View Article Online

important as they bind to the oxygen binding sites which arepresent in the phenolic polymers and melanins of the fungalcell wall.93 Due to the availability of fungal biomass and rapidgrowth rate these are receiving considerable attention as bio-sorbents among living cells.

As discussed above, due to the abundance and lesser pro-cessing requirement, agricultural and industrial waste and theirby-products are very economic and have received good accep-tance. Adsorbents include leafs, bers, fruit peels, saw dust barketc. from agricultural and forest industries has been used inremoving metals from contaminated water. Due to their phys-ico-chemical characteristics and their availability they can beused as adsorbents.108

Vargas et al. worked on waste fruit cortex for the removal ofheavy metals from contaminated water. They checked the bio-sorption capacity of banana (Musa paradisiaca), lemon (Citruslimonum) and orange (Citrus sinensis) peel. They found thatlemon and orange cortex showed good biosorption potential forlead and copper as compared to banana. In the case ofcadmium, banana showed greater biosorption efficiency thanlemon and orange. They also studied the relationship betweenparticle size and surface area and found them to be inverselyrelated to each other.109 In another investigation, Amaranthushybridus stalk and Carica papaya were used for removal of Mnand Pb ions from wastewater. The study showed that amongboth substrates Mn had greater percentage removal than lead.The adsorptive capacity of Carica papaya in all cases was higheras compared to Amaranthus hybridus stalk.110 In a recent study,researchers worked on chitin and a-(1,3)-b-D-glucan (fromindustrial bio-waste exhausted from brewer's yeast) for theremoval of heavy metals from acid mine drainage (Merladet andFaith open-cast mines). They found that the Faith minedrainage was contaminated with U, Al, Cu, Mn, etc. and theMerladet mine drainage with Al, Mn, Zn, and Cu. They reportedthat Saccharomyces cerevisiae L. acted as an efficient biosorbentwhich eliminated heavy metals from polluted water.111 Suryanet al. used paper mill waste for the removal of heavy metals (Pb,Cd, Ni and Cu) from aqueous solution. They found thatadsorption process was affected by pH and adsorption rate incase of all metal ions was above 70% (pH 2 to5). They concludedthat the paper mill waste did not require any pre-treatment andrecommended this as an option for better utilization of waste.112

Biosorption mechanism(s)

The mechanism of biosorption is a highly complex processowing to the complexity of the biological structures involved.Functional groups like carboxyl, phosphate, hydroxyl, amino,thiol etc. are present on the structure of biomass which interactswith different heavy metals with variable degree which may beaffected by physico-chemical factors. There are number offactors on which the binding of sorbate and sorbent dependslike number of binding site in the biosorbent, binding strengthof pollutant and functional groups present on the biosorbent,availability and accessibility of sites.

There are various criteria on the basis of which mechanismsof biosorption can be divided. This includes cell metabolism


and the location of biosorption (Fig. 4). Cell metabolismmediated biosorption can further be divided into metabolism-dependent and non-metabolism-dependent processes:113

� Metabolism dependent: occurs only in viable cells andplays a vital role in the defence mechanism of microbesshowing reaction with toxic metals. Metabolism-dependentbiosorption may further be classied as:

» Transport across cell membrane: microorganisms sharesthe same mechanism for transport of heavy metals across themembrane as well as transport of metabolic ions like sodium,magnesium etc.88 It has no association with metabolic activityand comprises two steps:

1. Metabolism-independent binding where the metals bindto the cell walls.

2. Metabolism-dependent intracellular uptake which incl-udes transport of metal ions across the cell membrane.

» Precipitation: metabolism-dependent precipitation isoen related to the microbe's defense mechanism wheretheir reaction in the presence of toxic metals producescompounds favouring precipitation.114

� Non-metabolism-dependent: involves the interactionbetween metal and functional groups present on the microbialsurface. As already discussed, many functional groups(carboxyl, phosphate, sulphate etc.) are present on themicrobialsurface because the microbial cell wall is made up of poly-saccharides, proteins and lipids. It can further be categorised asfollows:

» Physical adsorption: a physical phenomenon involvingvan der Waal's and electrostatic forces. Even dead biomasses ofalgae fungi and yeasts have shown adsorption of heavy metalslike copper, uranium, cobalt, zinc and cadmium through elec-trostatic interactions.93

» Ion exchange: here, the polysaccharides of microbial cellwalls act as counter ions and facilitate the exchange of bivalentmetal ions. The marine algae alginates which occur as salts ofMg2+, K+, Na+, Ca2+ can adsorb heavy metals by exchange ofcounter ions like Co2+, Cu2+, Cd2+ and Zn2+.115

» Complexation: interaction of metals with active groupsmediates their removal from solution via the formation of cellsurface complexes. Organic acids produced by microbes alsoplay an important role in chelating toxic metals by their sol-ubilisation and leaching resulting in generation of metallo-organic molecules. Complexation and adsorption of metals isbrought about by carboxyl groups in microbial polysaccharidesand polymers.116

» Precipitation: metabolism-independent precipitationresults from the metal and cell surface interaction which is achemical phenomenon.

The location-dependent biosorption can be categorised onthe basis of location where removed metals from solutionaccumulate. It is of the following types:

� Extracellular accumulation/precipitation.� Cell surface sorption/precipitation.� Intracellular accumulation.These can further be categorised in exactly the same fashion

as that in the case of metabolism-dependent and independentbiosorption.114




Publ

ishe

d on

12

Nov

embe

r 20

13. D

ownl

oade

d by

RSC

Int

erna

l on

23/1

2/20

13 1

0:52

:49.

View Article Online

Merits and demerits of biosorption

Conventional methods like ion exchange, electrodialysis,reverse osmosis etc. are expensive and their efficiency ofremoving heavy metals is very low. For this reason, biosorptionis receiving much attention because it shows many advantagesover traditional methods stated as follows:88

3 It is cost effective, selective and shows efficient removal ofmetals even at low concentrations.

3Unlike conventional methods it does not produce any toxicsludge during the removal process which presents the oppor-tunity of metal recovery and recycling of biosorbent.

3 Low cost adsorbents like industrial solid waste, agricul-tural waste etc. have shown excellent heavy metal removalefficiency.

3 It has a great advantage of being used in situ without theneed of any industrial process in integration with other eco-friendly systems.

3 With dead biomass, concerns related to toxicity, microbialconstrains and media formulation is alleviated.

The only disadvantage of using biosorbents is there is nocontrol over the biological characteristic of the biosorbent andearly saturation while performing experiments.

Recent developments inbioremediation

Recently new strategies for the process of bioremediation havebeen uncovered. Scientists have shown the application ofnanoparticles, non-living biomass and genetically modiedplants for the removal of heavy metal toxicants from differentsources. These approaches are credited with having quick andhigh bioremediation capacity.

Use of nanoparticles

Application of nanotechnology is widely being used for thedevelopment of resourceful, efficient and environment friendlynanomaterial systems in different spheres of biotechnologyincluding bioremediation. The physiochemical properties ofthe nanoscale particles vary signicantly from their largercounterparts. This is due to the very high surface to volume ratioof the nanomaterials which provides them with high adsorptioncapacity. Moreover, they have low cost and augmentedbioavailability which makes them excellent candidates forbioremediation.117

Lately, superparamagnetic iron oxide nanoparticles (SPION)have been used for the separation of contaminants fromwastewater because of their ultrane structure and highcompetence. In this technique, the carriers contain a polymericshell having functional groups and a magnetic core (FeO, Fe3O4

and Fe2O3) which provides a strong magnetic response.118 Shenet al., prepared and implemented Fe3O4 nanoparticles for thepurication of wastewater contaminated with heavy metals(Cd2+, Cr6+, Cu2+ and Ni2+). The nanoparticles prepared were ofdifferent size and were prepared by co-precipitation and polyolmethod. They found that particles of 8 nm size were veryeffective for the recovery and removal of metals from


wastewater. It was found that the adsorption capacity of Fe3O4

particles increased with decreasing the particle size orincreasing the surface area. Furthermore, maximum adsorptionwas seen to occur at pH 4.0 under room temperature (20 �C) andthe adsorption capacity of nanoparticles was found to be ashigh as 35.46 mg g�1 (7 times higher than that of the coarseparticles).119 In another nding, the bioaccumulation capacityof magnetic gel beads (prepared from Fe2O3 nanoparticle andgellan gum) as potent bioadsorbents was assessed. Theyconcluded that the magnetic gel beads were effective for theremoval of lead, manganese and chromium in the order ofPb2+ > Cr3+ > Mn2+. Additionally, it showed the high desorptioncapacity with sodium citrate which would prove be veryeconomical.120 Bezbaruah et al., used calcium alginate beads forthe entrapment of zero-valent iron nanoparticle (Feo) to removethe test contaminant present in ground water (nitrate). Zero-valent iron nanoparticles (nZVI) have been known to eradicatevarious groundwater contaminants like chlorinatedcompounds, pesticides and heavy metals. However, it suffersfrom higher mobility, agglomeration and settlement problemsby non-target compounds. They found that the overall removalefficiency of entrapped nZVI towards contaminants wascomparable to that of bare nZVI.121 Another investigationreported the formulation of magnetic chitosan nanoparticles bya one-step in situ co-precipitation method with an aim toexamine the sorption property for removing Cu(II) from aqueoussolution. The process was found to be highly competent and themaximum sorption capacity was calculated to be 35.5 mg g�1.122

Use of non-living biomass

Another favorable strategy that is used extensively for biore-mediation includes use of living microorganisms such asbacteria, fungi, algae etc. They provide a large surface area tovolume ratio because of the smaller size of microorganisms.However, they suffer from several disadvantages such ascausing redox reactions between cell and medium which leadsto an increase in pH of the system. Also, this method increasesthe biological oxygen demand and chemical oxygen demand asit requires nutrient uptake.123 In contrast to this, the applicationof dead biomass can be regarded as a suitable and favorablealternative. Dead biomass does not require growth media ornutrients and does not cause toxicity during the metal removalprocess. Besides, this method is very economical in comparisonto living biomass.124

A recent investigation showed the biosorption capability ofnonliving biomass of marine macrophytes for arsenic removal.Different phyla of alga were used to check their capability underdifferent pH conditions. Considerable adsorption was exhibitedby all the species. The highest observed value was by red algaCeramium (1.3 � 0.1 mg g�1) and seagrass Zostera, comparableto other low-cost adsorbents like activated carbon. It was alsoobserved that there were many factors on which biosorptionrested like composition and structure of outer layer of macro-phytes, availability of functional groups present on arsenic,different pH levels and counter ionic interaction with arsen-iate.125 Ekmekyapar et al., studied the biosorption capacity of




Publ

ishe

d on

12

Nov

embe

r 20

13. D

ownl

oade

d by

RSC

Int

erna

l on

23/1

2/20

13 1

0:52

:49.

View Article Online

Cladonia rangiformis (a non-living lichen) for the removal of leadfrom aqueous solution. They found that the non-living lichen isa natural biomass, harmless and easily available and exhibited ahigh accumulation capacity towards lead which can be exploi-ted for the treatment of industrial effluents.126 Mohamad et al.,reported the capability of dead cells ofMesorhizobium amorphaeto act as a robust biosorbent for the removal of Cu2+ fromaqueous solutions.123 Biosorption capacity of non-livingbiomass of Spirulina sp. for the removal of lead and zinc wasalso reported in a work carried out by Goyal et al.127

Use of genetically modied plants (GMP)

The main aim of genetic engineering in the eld of phytor-emediation is to enhance the capacity of plants to tolerate,accumulate and absorb contaminants. Many genes fromdifferent organisms have been identied and characterized thatare involved in acquisition, allocation and decontamination ofmetals. The recombinant proteins produced by these transgenicplants play an important role in chelation (e.g. citrate, phy-tochelatins, metallothioneins, phytosiderophores and ferritin),assimilation and membrane transport of metals.128

Recently transgenic Arabidopsis thaliana was developed toincrease the tolerance and accumulation of arsenic andcadmium by simultaneous over-expression of AsPCS1 and YCF1genes. These genes are derived from garlic and baker's yeast.This work was based on chelation of metals and vacuolarcompartmentalization which are the main strategies for heavymetals/metalloids detoxication. It was found that the twogenes simultaneously increased the accumulation capacity ascompared to use of a single gene.129 Zhang et al., carried out astudy on a transgenic alfalfa plant with a motive to enhance itsresistance capability towards the heavy metals and organicpollutants. This transgenic plant expressed both humanCYP2E1 and glutathione S-transferase which were producedfrom hypocotyl segments by the use of Agrobacterium-mediatedtransformation. They found that the transgenic alfalfa plantwhich expressed both genes simultaneously had a remarkablepotential to remove mixed contaminants as compared to thewild type and transgenic plant expressing the single gene.130

Another GMP, Nicotiana tabacum carrying a yeast metal-lothionein gene was shown to accumulate cadmium in the rootof the transgenic plant.131

Conclusion

Levels of heavy metals are increasing day by day due toincreased industrial usage causing their accumulation in livingbeings. Their exposure can cause fatal consequences to organsystems through several mechanisms (primarily due to gener-ation of oxidative stress). Oxidative stress leads to the produc-tion of free radicals followed by the decrease in the level ofantioxidants and nally leading to cell death.

Presently, conventional remediation methods like ionexchange, chemical precipitation, reverse osmosis, landllingand bio-piles are used for the removal of heavy metal contam-inants. Although they have several advantages like ease of metal


recovery, pure effluent production and high productivity, theyhave severe disadvantages which includes their high cost,production of toxic sludge and incomplete removal of metals.

So, as a potential alternative to these methods, bioremedia-tion is a promising upcoming technology which uses plants,microbes and their enzymes for the removal of heavy metals inan eco-friendly manner. Bioremediation involves twoapproaches i.e. phytoremediation and biosorption. Phytor-emediation makes use of plants which have the capability toaccumulate, degrade and/or volatilize the heavy metals, hydro-carbons and organic solvents leading to improvement in thequality of soil and water. Depending upon the conditions for e.g.on the basis of the site (soil or water) and property of contam-inants (organic or inorganic), phytoremediating plants imple-ment different mechanisms which include phytoextraction,phytoltration, phytostabilization, phytovolatilization andphytodegradation.

Another mode of bioremediation exists called biosorptionwhich uses low cost adsorbents like industrial waste, agricul-tural waste, microbial biomass and their derivatives for thetreatment of aqueous waste. Different mechanisms are used bythe adsorbent on the basis of location of biosorption andcellular metabolism. Further, they are divided into ionexchange, precipitation, complexation and physical adsorption.The heavy metal adsorption depends upon the type of adsor-bent used, surface area, particle size, shape of the adsorbentand experimental conditions. Biosorption is a promisingapproach and is not only cost effective but also shows selectivityand high efficiency towards the removal of heavy metals.Moreover, it does not produce any toxic sludge.

The latest addition to this technology is the application ofnanoparticles, non-living biomass and transgenic crops. Thesenovel approaches carrying out bioremediation have given highlyencouraging results and in addition of being efficient they arealso economical and give rapid results.

Thus, bioremediation has bright prospects for the abolitionof heavy metals from polluted sites. Also, its applicability couldbe further enhanced by identifying and implementing morenovel plants and biosorbents that can offer better scope forheavy metal removal without the need for any substrate modi-cation. Consequently, the processing cost for modication ofadsorbents could be saved. Conclusively, it can be said that thebioremediation technology has given us a platform that coulddirect us towards the elimination of heavy metal pollution in aneco-friendly manner.

Competing interest

The authors declare that they have no conict of interest.

Abbreviations

ROS
Reactive Oxygen Species BBB Blood Brain Barrier EDTA Ethylenediaminetetraacetic acid



Publ

ishe

d on

12

Nov

embe

r 20

13. D

ownl

oade

d by

RSC

Int

erna

l on

23/1

2/20

13 1

0:52

:49.

View Article Online

CaNa2EDTA

Environ. Sci.: Pr

Calcium disodium ethylenediaminetetraaceticacid

DPA
D-Penicillamine BAL Dimercaprol NTA Nitrilotriacetic acid Chl Chlorophyll KH Henry constant Kd Partitioning coefficient.
Acknowledgements

I heartily acknowledge Ms. Batul Diwan (School of Biotech-nology, Rajiv Gandhi Proudyogiki Vishwavidyalaya, Bhopal,India) who played a major role in correcting and editing of themanuscript.

References

1 V. Marcano, P. Benitez and E. Palacios-Pru, Planet. SpaceSci., 2003, 51, 159–166.

2 I. G. Dubus, J. M. Hollis and C. D. Brown, Environ. Pollut.,2000, 110, 331–344.

3 R. Singh, N. Gautam, A. Mishra and R. Gupta, Indian J.Pharmacol., 2011, 43, 246–253.

4 S. J. Flora, M. Mittal and A. Mehta, Indian J. Med. Res., 2008,128, 501–523.

5 V. Vinod and R. Sashidhar, Indian J. Biotechnol., 2011, 10,113–120.

6 N. Singh and R. Gadi, J. Environ. Chem. Ecotoxicol., 2012, 4,137–142.

7 S. N. Singh and R. R. D. Tripathi, Environmentalbioremediation technologies, Springer-Verlag, BerlinHeidelberg, 2007.

8 A. N. Glazer and H. Nikaido, Microbial Biotechnology:Fundamentals of Applied Microbiology, W.H. Freeman, 1995.

9 D. Di Baccio, R. Tognetti, L. Sebastiani and C. Vitagliano,New Phytol., 2003, 159, 443–452.

10 J. Kelly, M. Haggblom and R. Tate, III, Biol. Fertil. Soils,2003, 38, 65–71.

11 R. A. Wuana and F. E. Okieimen, ISRN Ecol., 2011, 402647.12 J. Buekers, Ph.D. thesis, Katholieke Universiteit Lueven,

Dissertationes De Agricultura, Doctoraatsprooefschri nr,2007.

13 J. Shiowatana, R. G. McLaren, N. Chanmekha andA. Samphao, J. Environ. Qual., 2001, 30, 1940–1949.

14 K. Jomova, D. Vondrakova, M. Lawson and M. Valko, Mol.Cell. Biochem., 2010, 345, 91–104.

15 K. A. Al-Ghanim, Afr. J. Biotechnol., 2011, 10, 13860–13866.16 R. R. Schroeder, Thesis for the Master of Environmental

Study Degree, The Evergreen State College, 2010.17 G. Flora, D. Gupta and A. Tiwari, Interdiscip. Toxicol., 2012,

5, 47–58.18 S. Mahjoub and A. H. Moghaddam, Iranian Journal of Health

and Physical Activity, 2011, 2, 1–5.19 J. Kasten-Jolly, Y. Heo and D. A. Lawrence, Toxicol. Appl.

Pharmacol., 2010, 247, 105–115.

ocesses Impacts

20 N. Fretellier, N. Bouzian, N. Parmentier, P. Bruneval,G. Jestin, C. Factor, C. Mandet, F. Daubine, F. Massicotand O. Laprevote, Toxicol. Sci., 2013, 131, 259–270.

21 B. Anilkumar, A. G. Reddy, P. Ravikumar, Y. R. Reddy andC. Haritha, Inventi Rapid: Molecular Pharmacology, 2012,Inventi:pmp/167/12.

22 T. O. Llewellyn, Cadmium (materials Flow), Department ofthe Interior. Bureau of Mines, United States, 1994.

23 V. Arroyo, K. Flores, L. Ortiz, L. Gomez-Quiroz andM. Gutierrez-Ruiz, J. Drug Metab. Toxicol., 2012, 5, 2.

24 H. Baba, K. Tsuneyama, M. Yazaki, K. Nagata,T. Minamisaka, T. Tsuda, K. Nomoto, S. Hayashi, S. Miwaand T. Nakajima, Mod. Pathol., 2013, 1228–1234.

25 M. Filipic, T. Fatur andM. Vudrag,Hum. Exp. Toxicol., 2006,25, 67–77.

26 G. J. Flora, Curr. Trends Biotechnol.Pharm., 2012, 6, 280–289.27 S. Chanda, B. Ganguli, U. B. Dasgupta and D. G. Mazumder,

J. Toxicol. Environ. Health Sci., 2011, 3, 171–175.28 D. GuhaMazumder and U. Dasgupta, Kaohsiung J. Med. Sci.,

2011, 27, 360–370.29 K. Jomova, Z. Jenisova, M. Feszterova, S. Baros, J. Liska,

D. Hudecova, C. Rhodes and M. Valko, J. Appl. Toxicol.,2011, 31, 95–107.

30 V. Pachauri, A. Mehta, D. Mishra and S. J. Flora,Neurotoxicology, 2013, 137–145.

31 L. Patrick, Alternative Med. Rev., 2002, 7, 456–471.32 J.-D. Park and W. Zheng, Journal of Preventive Medicine and

Public Health, 2012, 45, 344–352.33 M. Korbas, J. L. O'Donoghue, G. E. Watson, I. J. Pickering,

S. P. Singh, G. J. Myers, T. W. Clarkson and G. N. George,ACS Chem. Neurosci., 2010, 1, 810–818.

34 C. Tschirhart, P. Handschumacher, D. Laffly andE. Benece, Hum. Ecol., 2012, 40, 511–523.

35 M. Valko, H. Morris and M. T. Cronin, Curr. Med. Chem.,2005, 12, 1161–1208.

36 A. P. Society and H. Press, Cell Physiology, AmericanPhysiological Society, 1992.

37 A. Erakhrumen, Educational Research and Review, 2007, 2,151–156.

38 N. Adams, D. Carroll, K. Madalinski, S. Rock, T. Wilson andP. Pivetz, Introduction to phytoremediation, National RiskManagement Research Laboratory, Cicinnati, Ohio, USA,2000.

39 P. Ahmadpour, F. Ahmadpour, T. Mahmud, A. Abdu,M. Soleimani and F. H. Tayefeh, Afr. J. Biotechnol., 2012,11, 14036–14043.

40 Y. D. Jing, Z. L. He and X. E. Yang, J. Zhejiang Univ., Sci., B,2007, 8, 192–207.

41 M. I. Lone, Z. L. He, P. J. Stoffella and X. E. Yang, J. ZhejiangUniv., Sci., B, 2008, 9, 210–220.

42 M. Ghosh and S. Singh, Applied Ecology and EnvironmentalResearch, 2005, 3, 1–18.

43 S. Hayat and A. Ahmad, Brassinosteroids: A Class of PlantHormone, Springer London, Limited, 2011.

44 L. V. Pavel andM. Gavrilescu, Environ. Eng. Manage. J., 2008,7, 815–834.

45 N. Rascio and F. Navari-Izzo, Plant Sci., 2011, 180, 169–181.




Publ

ishe

d on

12

Nov

embe

r 20

13. D

ownl

oade

d by

RSC

Int

erna

l on

23/1

2/20

13 1

0:52

:49.

View Article Online

46 M. M. Lasat, The use of plants for the removal of toxic metalsfrom contaminated soils, American Association for theAdvancement of Science, Environmental Science andEngineering Fellow, Washington, D.C., USA, 2000, p. 33.

47 V. Sheoran, A. S. Sheoran and P. Poonia, Miner. Eng., 2009,22, 1007–1019.

48 T. Mahmood, Soil Environ., 2010, 29, 91–109.49 B. V. Tangahu, S. Abdullah, S. Rozaimah, H. Basri, M. Idris,

N. Anuar and M. Mukhlisin, Int. J. Chem. Eng., 2011, 2011,939161.

50 L. Kaur, K. Gadgil and S. Sharma, International Journal ofBioassays, 2013, 2, 352–357.

51 S. Zhao, L. Jia and L. Duo, Bioresour. Technol., 2013, 129,249–255.

52 Saifullah, M. H. Zia, E. Meers, A. Ghafoor, G. Murtaza,M. Sabir, M. Zia-ur-Rehman and F. M. G. Tack,Chemosphere, 2010, 79, 652–658.

53 H. Altinozlu, A. Karagoz, T. Polat and I. Unver, Turk. J. Bot.,2012, 36, 269–280.

54 W. A. Number, C. Ma, J. Kingscott and M. Evans, U.S.Environmental Protection Agency Office of Solid Waste andEmergency Response Technology Innovation Office,Washington D.C., 1997.

55 H. Patra and M. Mohanty, Development, 2013, 25, 27.56 S. Mukhopadhyay and S. Maiti, Global J. Environ. Res., 2010,

4, 135–150.57 C. Cosio, E. Martinoia and C. Keller, Plant Physiol., 2004,

134, 716–725.58 J. B. Wells, T. U. o. T. a. S. A. Earth and E. Science, Effect of

Infection with Glomus Mosseae on Cadmium Uptake in VetiverGrass [Chrysopogon Zizanioides (L.) Roberty], University ofTexas, San Antonio, 2008.

59 M. L. Lang, Y. X. Zhang and T. Y. Chai, Chin. J. Biotechnol.,2004, 20, 157–164.

60 M. S. Islam, Y. Ueno, M. T. Sikder and M. Kurasaki, Int. J.Phytorem., 2012, 1010–1021.

61 D. Sarkar, R. Datta and R. Hannigan, Concepts andApplications in Environmental Geochemistry, ElsevierScience, 2011.

62 N. A. Anjum, M. E. Pereira, I. Ahmad, A. C. Duarte andS. Umar, Phytotechnologies: Remediation of EnvironmentalContaminants, CRC Press Inc, 2012.

63 S. K. Ong, Natural Processes and Systems for HazardousWaste Treatment, American Society of Civil Engineers,2008.

64 P. D. M. Jumba, Genetically Modied Organisms The MysteryUnraveled, AEG Literary Publishing Services, Incorporated,2010.

65 I. Raskin, R. D. Smith and D. E. Salt, Curr. Opin. Biotechnol.,1997, 8, 221–226.

66 B. Thomas, D. J. Murphy and B. G. Murray, Encyclopedia ofApplied Plant Sciences, Three-Volume Set, Elsevier Science,2003.

67 B. K. Yadav, M. A. Siebel and J. J. A. van Bruggen, Clean: Soil,Air, Water, 2011, 39, 467–474.

68 T. Vesely, P. Tlustos and J. Szakova, Int. J. Phytorem., 2011,13, 859–872.


69 T. Vesely, P. Tlustos and J. Szakova, Int. J. Phytorem., 2012,14, 335–349.

70 C. S. K. Mishra, Environmental Biotechnology, APHPublishing Corporation, 2007.

71 R. Mohee and A. Mudhoo, Bioremediation andSustainability: Research and Applications, Wiley, 2012.

72 S. Shilev, M. Benlloch, R. Dios-Palomares and E. D. Sancho,in Predictive Modeling and Risk Assessment, Springer, 2009,pp. 225–242.

73 R. Costa and K. Kristbergsson, Predictive Modeling and RiskAssessment, Springer, USA, 2009.

74 J. Cambrolle, E. Mateos-Naranjo, S. Redondo-Gomez,T. Luque and M. Figueroa, Hydrobiologia, 2011, 671,95–103.

75 M. Varun, R. D'Souza, J. Pratas and M. Paul, Biotechnol.Bioinf. Bioeng., 2011, 1, 137–145.

76 W. Meeinkuirt, P. Pokethitiyook, M. Kruatrachue,P. Tanhan and R. Chaiyarat, Int. J. Phytorem., 2012, 14,925–938.

77 P. Soudek, S. Petrova and T. Vanek, InternationalProceedings of Chemical, Biological and EnvironmentalEngineering (IPCBEE), 2012, 46, 25–29.

78 L. K. Wang, N. K. Shammas and Y. T. Hung, WasteTreatment in the Metal Manufacturing, Forming, Coating,and Finishing Industries, Taylor & Francis, 2010.

79 S. Feroz, D. G. Rao, R. Senthilkumar and J. A. Byrne,Wastewater Treatment: Advanced Processes andTechnologies, Taylor & Francis Group, 2012.

80 B. P. Singh, Industrial Crops and Uses, CABI Publishing,2010.

81 D. L. Reger, S. R. Goode and D. W. Ball, Chemistry, Principlesand Practice, BROOKS COLE Publishing Company, 2009.

82 P. J. Alvarez and W. A. Illman, Bioremediation and NaturalAttenuation: Process Fundamentals and MathematicalModels, Wiley, 2005.

83 M. Sakakibara, A. Watanabe, M. Inoue, S. Sano andT. Kaise, Proceedings of the Annual International Conferenceon Soils, Sediments, Water and Energy, 2010.

84 M. R. Edwards, M. F. Hetu, M. Columbus, A. Silva andD. D. Lefebvre, Int. J. Phytorem., 2011, 13, 702–716.

85 K. M. Carvalho and D. F. Martin, J. Aquat. Plant Manage.,2001, 39, 33–36.

86 P. A. Kulakow and V. V. Pidlisnyuk, Application ofPhytotechnologies for Cleanup of Industrial, Agricultural andWastewater Contamination, Springer, 2009.

87 D. Tsao and M. K. Banks, Phytoremediation, Springer, 2003.88 H. Fulekar, Bioremediation Technology: Recent Advances,

Springer, Netherlands, 2010.89 V. de Farias, L. T. Maranho, E. C. de Vasconcelos, M. A. da

Silva Carvalho Filho, L. G. Lacerda, J. A. M. Azevedo,A. Pandey and C. R. Soccol, Appl. Biochem. Biotechnol.,2009, 157, 10–22.

90 X. Wang, N. Wu, J. Guo, X. Chu, J. Tian, B. Yao and Y. Fan,Biochem. Biophys. Res. Commun., 2008, 365, 453–458.

91 G. M. Gadd, J. Chem. Technol. Biotechnol., 2009, 84, 13–28.92 W. Stumm and J. J. Morgan, Aquatic Chemistry: Chemical

Equilibria and Rates in Natural Waters, Wiley, 2012.




Publ

ishe

d on

12

Nov

embe

r 20

13. D

ownl

oade

d by

RSC

Int

erna

l on

23/1

2/20

13 1

0:52

:49.

View Article Online

93 G. Crini and P. M. Badot, Sorption processes and pollution:Conventional and non-conventional sorbents for pollutantremoval from wastemasters, Presses Universitaires deFranche-Comte, 2011.

94 G. Sposito, The Chemistry of Soils, Oxford University Press,USA, 2008.

95 Z. Aksu, Process Biochem., 2005, 40, 997–1026.96 G. M. Gadd, FEMS Microbiol. Lett., 1992, 100, 197–203.97 D. Park, Y.-S. Yun and J. Park, Biotechnol. Bioprocess Eng.,

2010, 15, 86–102.98 K. Vijayaraghavan and Y.-S. Yun, Biotechnol. Adv., 2008, 26,

266–291.99 R. H. Vieira and B. Volesky, Int. Microbiol., 2000, 3,

17–24.100 J. Modak and K. Natarajan,Miner. Metall. Process., 1995, 12,

189–196.101 P. Kotrba, M. Mackova and T. Macek, Microbial biosorption

of metals, Springer, Netherlands, 2011.102 G. M. Gadd, Microbiology, 2010, 156, 609–643.103 G. Seltmann and O. Holst, The Bacterial Cell Wall, Springer,

2002.104 T. A. Davis, B. Volesky and A. Mucci, Water Res., 2003, 37,

4311–4330.105 J.-P. Latge, Cell. Microbiol., 2010, 12, 863–872.106 J. Wang and C. Chen, Biotechnol. Adv., 2009, 27, 195–226.107 E. Lichtfouse, J. Schwarzbauer and D. Robert,

Environmental Chemistry for a Sustainable World:Remediation of air and water pollution, SpringerScience+Business Media B.V., 2011.

108 M. Rafatullah, O. Sulaiman, R. Hashim and A. Ahmad,J. Hazard. Mater., 2010, 177, 70–80.

109 K. Kelly-Vargas, M. Cerro-Lopez, S. Reyna-Tellez,E. R. Bandala and J. L. Sanchez-Salas, Phys. Chem. Earth,2012, 37–39, 26–29.

110 J. Egila, B. Dauda, Y. Iyaka and T. Jimoh, Int. J. Phys. Sci.,2011, 6, 2152–2157.

111 F. Ramırez-Paredes, T. Manzano-Munoz, J. Garcia-Prieto,G. Zhadan, V. Shnyrov, J. Kennedy and M. Roig,Biotechnol. Bioprocess Eng., 2011, 16, 1262–1272.

112 S. Suryan and S. Ahluwalia, International Journal ofEnvironmental Sciences, 2010, 1331–1343.


113 N. Ahalya, T. Ramachandra and R. Kanamadi, Res. J. Chem.Environ., 2003, 7, 71–79.

114 L. K. Wang and J. H. Tay, Environmental Bioengineering,Humana, 2010.

115 S. K. Sharma and A. Mudhoo, Green Chemistry forEnvironmental Sustainability, Taylor & Francis, 2010.

116 S. Srivastava and P. Goyal, Novel Biomaterials:Decontamination of Toxic Metals from Wastewater,Springer, Berlin Heidelberg, 2010.

117 S. Cameotra and S. Dhanjal, in Bioremediation Technology,ed. M. H. Fulekar, Springer, Netherlands, 2010, pp. 348–374.

118 V. V. Mody, R. Siwale, A. Singh and H. R. Mody, J. Pharm.BioAllied Sci., 2010, 2, 282.

119 Y. F. Shen, J. Tang, Z. H. Nie, Y. D. Wang, Y. Ren and L. Zuo,Sep. Purif. Technol., 2009, 68, 312–319.

120 X. Wang, C. Zhao, P. Zhao, P. Dou, Y. Ding and P. Xu,Bioresour. Technol., 2009, 100, 2301–2304.

121 A. N. Bezbaruah, S. Krajangpan, B. J. Chisholm, E. Khanand J. J. Elorza Bermudez, J. Hazard. Mater., 2009, 166,1339–1343.

122 C. Yuwei and W. Jianlong, Chem. Eng. J., 2011, 168, 286–292.

123 O. A. Mohamad, X. Hao, P. Xie, S. Hatab, Y. Lin and G. Wei,Microbes Environ., 2012, 27, 234–241.

124 C. M. Monteiro, P. M. Castro and F. X. Malcata, Biotechnol.Prog., 2012, 28, 299–311.

125 C. Pennesi, F. Veglio, C. Totti, T. Romagnoli andF. Beolchini, J. Appl. Phycol., 2012, 24, 1495–1502.

126 F. Ekmekyapar, A. Aslan, Y. Bayhan and A. Cakici, Int. J.Environ. Res., 2012, 6, 417–424.

127 R. K. Aneja, G. Chaudhary, S. S. Ahluwalia and D. Goyal,Indian J. Microbiol., 2010, 50, 438–442.

128 T. Vamerali, M. Bandiera and G. Mosca, Environ. Chem.Lett., 2010, 8, 1–17.

129 J. Guo, W. Xu and M. Ma, J. Hazard. Mater., 2012, 199–200,309–313.

130 Y. Zhang and J. Liu, J. Hazard. Mater., 2011, 189, 357–362.131 O. Krystofova, O. Zitka, S. Krizkova, D. Hynek,

V. Shestivska, V. Adam, J. Hubalek, M. Mackova, T. Macekand J. Zehnalek, Int. J. Electrochem. Sci., 2012, 7, 886–907.



Documents

Review Paper Nisha