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Research Article - (2016) Volume 1, Issue 2

Physical, Chemical and Phytoremediation Technique for Removal of Heavy Metals

Sharma S1, Rana S2, Thakkar A1, Baldi A1, Murthy RSR1 and Sharma RK3*

1Indo Soviet Friendship College of Pharmacy, Moga, Punjab, India

2Division of CBRN Defence, Institute of Nuclear Medicine and Allied Sciences, Brig S.K Mazumdar Marg, Delhi, India

3Defence Food Research Laboratory, Siddartha Nagar, Mysuru , India

*Corresponding Author:

Sharma RK
Defence Food Research Laboratory,
Siddartha Nagar, Mysuru 570 011,
Tel: 0821-2473783, 09449651632;
Fax: 0821-2473468;

Received date: June 14, 2016; Accepted date: July 18, 2016; Published date: July 20, 2016

Citation: Sharma S, Rana S, Thakkar A, et al. Physical, Chemical and Phytoremediation Technique for Removal of Heavy Metals. J Heavy Met Toxicity Dis. 2016, 1:2.

Copyright: © 2016 Sharma S, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

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Mankind has been using plants and natural products since time immemorial for fighting the menace of heavy metal toxicity both in humans as well as in environment surrounding them. Nearly thirty five metals have been reported to cause occupational or accidental exposure to humans. Amongst these, twenty three are heavy metals. The increasing use of such heavy metals including radionuclides constitutes deleterious health issues. Presence of heavy metals in environment and their subsequent effects on humans down the food chain creates potential health hazard. Therefore removal of heavy metal has been a subject of paramount importance. Results of an exhaustive literature survey of natural and plant based compounds against heavy metal pollution including patents, books and scientific data from globally accepted scientific databases and search engines (Pubmed, Scopus and Web of Science, Sci Finder and Google Scholar), is systematically reviewed. It is conceived that a number of phytochemical agents as well microorganism can act as heavy metal removing agent both from human beings and the environment surrounding. Microbes which are used for the removal of heavy metals from the water bodies include bacteria, fungi, algae and yeast. Some important antioxidants such as flavonoids, pectin and phytic acid are also used for the elimination of the heavy metals from the human body. The present article is an extensive review that will offer a number of strategies and possible mechanisms for the heavy metals removal both from environment as well as from human body.


Heavy metals; Chelation; Adsorption; Absorption; Bio-sorption; Phytoremediation


Heavy metals are the chemical elements having density greater than 5. Some of these elements called trace elements are a part of our normal diet and are essential for good health and present in human and animal tissue in very low concentration. These trace elements may be essential or nonessential. The important essential elements along with their concentration in blood includes iron (0.06-0.26 mg/l), zinc (4-8 mg/l), cobalt (20 μg/l), copper (0.08-0.45 μg/l), chromium (0.08-0.5 μg/l), manganese (6.7-10.4 μg/l) and molybdenum (5-157 μg/l) [1,2]. Other elements called ultra-trace elements normally comprise less than 1 μg/g of a given organism. Their concentration in blood includes cadmium (0.1-2 μg/l), lead (40-290 μg/l), lithium (0.52-0.64 μg/l), nickel (1.1-4 μg/l), tin (120-140 μg/l) and vanadium (0.1-0.9 μg/l) [3,4]. However chronic exposure of toxic dose of these metals in humans results in various complications in nervous system, respiratory system, renal system, hepatic system as well as reproductive system. Metals are also reported to cause allergies and repeated long-term contact with some metals or their compounds may even prove carcinogenic. Most of the heavy metals are well known toxic and carcinogenic agents and represents a serious threat to the human population and the fauna and flora of the receiving water bodies as they are persistent and non-biodegradable.

Various agencies around the world take care of diminished quality of life and potential threat to environment associated with exposure to hazardous substances. Agency for Toxic Substances and Disease Registry (ATSDR), a part of U.S. department of health and human services, is the main agency which has compiled a priority list for hazardous substances. Canadian Environment Assessment Agency in Canada is the federal body performing high quality environmental assessment so that potential environmental effect of elements can be prevented. Likewise in India, Ministry of Environment and Forest (MoEF) is the agency for planning, promotion, coordination and overseeing of India’s environmental and forest policy and programs. This agency has promulgated “Manufacture, Storage, and Import of Hazardous Chemicals (MSIHC) rules, 1989” under Environment (Protection) Act, 1986 which classify toxic industrial chemicals as high, medium and low risk chemicals.

Heavy metals become toxic when they are not metabolized by the body and accumulate in the soft tissues inside the body. The source of heavy metals includes food, water, air, absorption through skin etc. The most common route of exposure for children is ingestion [5]. Less common routes of exposure are during a radiological procedure, from inappropriate dosing or monitoring during intravenous nutrition, from a broken thermometer [6] or from a suicide or homicide attempt. Likewise the source of heavy metals in environment is solid discharge from industry, nuclear power plants, smelting process of various metals, by-product from various process in chemical industry, volcanic eruption, combustion of fossil fuels, pesticides/insecticides etc. [7-9]. The presence of metals in environment is a potential source of toxicity owing to their transport down the food chain and their subsequent bio-magnification. They cannot be destroyed biologically and get transformed into different oxidation states or different organic complex [10,11]. Thus it is pertinent to explore remedy for removal of these toxic heavy metals both from environment as well as from human beings.

Removal of Heavy Metals: Strategies and Mechanisms

Heavy metal removal may be accomplished by different mechanism. Figure 1 summarizes different possible mechanisms involved in heavy metal removal.


Figure 1: An overview of different mechanisms involved in removal of heavy metals.

Physical methods of heavy metal removal

Adsorption method: Adsorption is a physicochemical treatment processes which help in effective removal of heavy metals from metals contaminated wastewater and is one of the most preferred and efficient method. Its major advantage includes effectiveness at both high/low contaminant concentrations, selectivity by employing tailored adsorbents, regeneration ability of used adsorbents and a comparatively low cost. The various adsorbents and corresponding heavy metals adsorbed by them are described in Table 1.

Adsorbent Modifying agent(s) Heavy Metal Qmax
(mg g-1 )
Rice husk Water washed
Sodium hydroxide
Sodium bicarbonate
Cd(II) 8.58
Sawdust (C.deodarwood)
Sawdust (S. robusta)
Sawdust (Poplar tree)
Sawdust (Dalbergiasissoo)
Sawdust(Poplar tree) Sawdust (Pinussylvestris)
Sod. Hydroxide
Sulfuric acid
Sod. Hydroxide
Sod. Hydroxide Formaldehyde in Sulfuric acid
Ni (II)
Cd (II)
[84] [85]
Groundnut husk Sulfuric acid followed by silver
Cr (VI) 11.4 [86]
Wheat bran Sulfuric acid
Sulfuric acid
Cu (II)
Cd (II)
Banana pith Nitric acid Cu (II) 13.46 [89]
Cork powder Calcium Chloride, Sodium Chloride,
Sodium Hydroxide
Cu (II) 15.6
Corncorb Nitric acid
Citric acid
Cd (II) 19.3
(aquatic fern)
Hydrogen peroxide–Magnesium chloride Pb (II)
Cu (II)
Sugarcane bagasse Sodium bicarbonate Cu (II)
Pb (II)
Cd (II)
Sugarbeetpulp   Hydrochloric acid Cu (II)
Zn (II)
Coirpith ZnCl2 Cr(VI)
Cd (II)
NA [95]

Table 1: Summary of modified plant wastes as adsorbents for the removal of heavy metal ions from aqueous solution containing metals.

Majority of these adsorbents are chemically modified plant waste substances. The aqueous solution mentioned includes wastes water from different chemical industries and other synthetically made metal solutions for the purpose of study. Chemical pre-treatment of adsorbent results in higher adsorption capacity with respect to unmodified form. This is because pre-treatment causes higher number of active binding sites, better ion exchange properties and formation of new functional groups which have higher capacity of metal uptake. Chemical pre-treatment can be done by numerous chemicals which include mineral and organic acids, bases, oxidizing agent, organic compounds, etc. In one of the work by Gaballah and co-workers bark was studied for its removal efficiency from synthetic solution containing copper [12]. Bark was chemically pre-treated with alkali, acid and organic compound which lead to partial depolymerization of tannins. Pretreatment helped in efficient removal ability as tannins if present would have increase the biological oxygen demand of the solution and turned the solution brown. A retention of 43 mg of cu/g of dry modified bark was achieved.

Apart from activated/modified plant products, a plethora of other compounds can also act as adsorbents. Some of them include natural zeolite clinoptilolite, montmorillonite clay, activated carbons, sepiolite and kaolin. Natural zeolite clinoptilolite holds great potential for removing heavy metal cation from aqueous solution. In a study by Erdem and coworkers the adsoption behavior of clinoptilolite for Co2+, Cu2+, Mn2+ and Zn2+ was investigated [13]. The batch method was employed using metal concentrations in solution ranging from 100 to 400 mg/l. The adsorption phenomena depend on charge density and hydrated ion diameter and selectivity sequence for adsorption observed was Co2+>Cu2+> Zn2+> Mn2+. Thus natural zeolites can be suitably exploited to remove cationic heavy metal species from industrial wastewater. Lin and Juang in one of their batch expeeriment reported that montmorillonite suitably modified by anionic surfactant sodium dodecyl sulfate can also be used for removal of Cu2+ and Zn2+ from aqueous solutions. The removal capacity was relatively higher with respect to raw clay [14]. Sepiolite, a natural fibrous clay mineral, is also used for effective removal of various metal ions from polluted water. In a study by Lazarević and coworkers on natural and acid treated sepiolite, the divalent cation were retained on sepiolite in the order Pb2+>Cd2+>Sr2+. Batch experiments were performed using solutions of Pb(NO3)2, Cd(NO3)2 and Sr(NO3)2 with a concentration of 0.01 mol/dm3, at a ratio sepiolite to electrolyte solution of 0.05 g:25 cm3. It was observed that retention of Pb2+ and Cd2+ occurred by adsorption and Mg2+ ion exchange from sepiolite structure while electrostatic forces were main cause for retention of Sr2+ ions onto the surface of sepiolites [15]. Jiang and coworkers studied kaolinite clay obtained from Longyan, China for Pb(II), Cd(II), Ni(II) and Cu(II) uptake from wastewater. The results were impressive with maximum adsorption being observed within 30 minutes [16]. Thus adsorption method offers good option for removal of heavy metal in waste water from various industries. However selectivity does exist with regard to adsorbent and metal adsorbed by them. Careful pre-treatment and screening should be done for targeting the metal of choice.

Biosorption method: Various techniques have been employed for the treatment of metal bearing industrial effluents like precipitation, ion exchange, membrane and electrochemical technologies etc. However these techniques are expensive, not environment friendly and generally depends on the concentration of the waste. The search for an efficient, eco-friendly, cost effective and biological method for wastewater treatment culminates at biosorption method. The most striking advantage of biosorption method of heavy metal removal is the treatment of large volumes of effluents with low concentrations of biosorbent and no production of toxic secondary compounds. Other advantage includes short operation time. Biosorption essentially involves the passive uptake of metal ions by dead/inactive biological materials or by materials derived from biological sources. It consists of a solid phase (biosorbent) and a liquid phase (solvent, usually water) which contains dissolved species to be sorbed. The basic mechanism involves attraction of sorbent for the sorbate which are subsequently removed by different mechanisms. The biosorption process is affected by factors like status of biomass whether living or dead, type of biomaterial, pH, initial metal ion concentration etc. Biosorption can be attributed to a number of metabolism-independent processes that essentially take place in the cell wall. Important mechanisms involved are complexation, chelation, coordination, ion exchange, precipitation, reduction etc. Temperature does not have any significant effect on biosorption process in the range of 20-35ºC [17]. However pH, presence of other metal ion and biomass concentration greatly influences biosorption process. The most prominent effect is of pH as it influences solution chemistry of metal, activity of functional group in the biomass and the competition of metallic ions [18]. A vast array of biological materials, especially bacteria, algae, yeasts and fungi have received increasing attention for heavy metal removal and recovery due to their good performance, low cost and large available quantities. Biosorbents are cheaper, more effective alternatives for the removal of metallic elements, especially heavy metals from aqueous solution.

Biosorption by algae: Biosorption by algae requires high metal uptake and selectivity by substrate and suitable mechanical properties. Of all the algae brown algae have been proven to be the most effective and promising. It is their basic biochemical constitution that is responsible for this enhanced performance. More specifically, it is the properties of their cell wall constituents which are chiefly responsible for heavy metal uptake. Biosorption of the metallic cations to the algal cell wall component is essentially a surface process. Carboxyl, amino, sulfhydryl, and sulfonate are the main chemical groups which are involved in metallic cation biosorption. These groups are part of the algal cell wall structural polymers namely, polysaccharides (alginic acid, sulfated polysaccharides), proteins, and peptidoglycans. Ion exchange is one of the main biosorption mechanisms for heavy metal uptake by algae. However other binding mechanisms like micro-precipitation and complexation are also involved in the process of heavy metal uptake [19]. Table 2 explains some of the main algae used for this purpose and the metal ion biosorbed by them. Results of different batch experiments show the biosorption capacity of algae to various metals. Therefore biosorption using algae presents an innovative depurative process employing biomaterials which are abundantly present in nature and can be used as a valuable option for treatment of industrial waste water and other heavy metal contaminated water.

Algae Metal Sorbed Results Reference
Spirogyra Chromium (IV) Batch experiments at 5 mg/l of initial metal concemtration showed removal of 14.7×103 mg metal/kg of dry weight biomass atpH 2.0 in 120 minutes. [96]
Sargassam sp. (chromophyta) Copper Batch experiments using aqueous solution containing copper showed a high metal uptake capacity of 1.48 mmol/g biomass. Experiments were performed using 100 mg of dried biomass added to 25 ml of copper solution in 500 ml polypropylene flasks. [97]
Lyngbeaputealis Chromium(VI) 82% biosorption of chromium at pH 2-3 and 45°C at initial chromium concentration of 50–60 mg/l of solution [98]
Sargassumfluitans Uranium Uranium sorption capacity observed was 560 mg/g, 330 mg/g and 150 mg/g at pH 4.0, 3.2 and 2.6 respectively. [99]

Table 2: Biosorption by different algae and the corresponding metal sorbed.

Biosorption by fungi: The cell wall of fungus can make up 30% or more of its dry and is made mostly of polysaccharides, which constitute about 80% of the dry weight. Fungi can act as efficient bio-sorbent owing to their high percentage of cell wall material, which shows excellent metal binding properties [20]. Fungi have large amounts of chitin, chitosan, glucan and mannan as well as small amount of glycoprotein in their cell walls. These polymers are abundant sources of metal binding ligands like carboxyl, amine, hydroxyl and phosphate groups [21]. Fungal mycelium, the vegetative part of fungus consisting of thread like hyphae, has also been reported for its Zn2+ metal ion biosorption [22]. Use of fungus for biosorption process has many advantages which includes its ease to cultivate at large scale owing to its short multiplication cycle and high yield of biomass. It can be easily grown using unsophisticated fermentation techniques and inexpensive growth media. Fungal biomass is also very easily available as industrial waste products and certainly provides an economic advantage as compared to other biosorbents. Most importantly major fungi used as biosorbent are non-pathogenic and can be easily exploited without any safety concerns.

Biosorption by fungi is affected by many factors each functioning independently which should be taken into consideration in order to exploit their full potential. Some of the important factors include initial solute concentration, type and nature of biomass, biomass concentration (biosorbent dose/solution volume) in solution and physicochemical factors like pH, temperature and ionic strength. Fungi can be used in many forms as free/immobilized, living/dead, raw/pretreated, lab culture/waste industrial biomass etc. Several studies been done so far have shown excellent potential of fungi as biosorbent particularly for treating industrial waste water full of toxic heavy metals. In one of the finding by Velkova and colleagues, biosorption of Cu (II) onto chemically modified waste mycelium of Aspergillus awamori was studied [23]. Maximum biosorption capacity was reached by sodium hydroxide pre-treated waste fungal mycelium at pH 5.0. Table 3 enlists various fungi that have been used in different biosorption experiments. The results discussed of various batch experiments and laboratory investigations proves the potential of fungus for treating metal contaminated waste waters from different sources by selectively using the most optimum biosorbent.

Fungi Metal Sorbed Important Results Reference
Penicillinochrochloron Copper Culture studies at pH 2-8 and at copper concentration 5000 ppm in solution showed metal uptake of upto 4.0 × 105 µg/g dry weight of biomass after 1 day. Experiments with lake water containing metal showed removal and recovery of metal. [100]
Penicillinchrysogenum Radium Culture experiments done using radium at a concentration of 1000pCi/Lshowed 5 × 104nCi/g radium being biosorbed at pH 7 by the biomass. [101]
Rhizopusarrhizus Uranium, Thorium Rhizopusarrhizus at pH 4 and a maximum metal concentration of 5.5 mg/liter of solution in laboratory experiments exhibited the uranium and thorium biosorptive uptake capacity in excess of 180 mg/g. [102]
Agaricusmacrosporus Cadmium, Mercury, Copper Agaricusmacrosporus effectively extracted cadmium and mercury from the contaminated substrate. Different experiments at cadmium (10 mg per kg dw) and multisubstrate experiments at cadmium, mercury and leadeach at 10 mg kg-1 showed that fungi biomass efetively extracted metal from the substrate solution containing metals. [103]
Termitomycesclypeatus Chromium Biosorption of chromium from effluents coming from tannery industries was studied using live fungi biomass. The sorption of hexavalent chromium was best obtained at pH 3 and showed prominent reduction in level of metal from the solution. [104]
Aspergillus parasiticus Lead Batch experiments using contaminated lead solution showed biosorption capacity of the fungal biosorbent at 4.02 × 10-4mol g-1 at pH 5.0 and 20°C in 70 minutes. Regeneration cycles also showed no significant loss of sorption performance during four biosorption-desorption cycles. [105]
Aspergilusniger Zinc Experiments were done both batch wise and at column mode. Results showed that biosorption was function of pH (increasing with increasing pH between 1 to 9), biomass concentration (decreasing at high biomass concentartion) and zinc concentration. Pretreatment of biomass with NaOH further increased its biosorption capacity from contamianted metal solution. [22]
Aspergillus awamori Copper Sodium hydroxide and DMSO pre-treatments increased Cu (II) uptake capacity of fungal biomass by 48.20% and 20.05%, respectively. Biosorption experiments were done in 250 ml Erlenmeyer flasks by adding 0.1 g biosorbent to 100 mL metal solution at 20°C. [23]

Table 3: Biosorption of metals by different fungi species.

Biosorption by bacteria: Potent metal biosorbents under the class of bacteria include genre of Bacillus, Pseudomonas and Streptomyces. The bacterial cell wall consists of many functional groups like carboxyl, phosphonate, amine and hydroxyl groups [24,25]. Amongst them, carboxyl groups are abundantly available, negatively charged and actively participate in binding to metal cations. The amine group is also very effective for removing metal ions as it chelates cationic metal ions as well as adsorbs anionic metal species through electrostatic interaction or hydrogen bonding [26,27]. Bacteria are classified into gram positive and gram negative depending on its cell wall composition. Anionic functional groups found in the peptidoglycan, teichoic acids and teichuronic acids of Gram-positive bacteria, and the phospholipids, peptidoglycan and lipopolysaccharides of Gram-negative bacteria are the components primarily responsible for the anionic character and metal-binding capability of their cell wall. Using potentiometric titrations, metal uptake capacity can be correlated with amount of acidic groups [28]. FT-IR analyses can help to detect nature of binding sites and their involvement during biosorption [29,30]. Table 4 provides basic information to evaluate the possibility of using bacterial biomass for the uptake of metal ions from waste water. This model can be used for employing bacteria for waste water treatment systems. The extent of biosorption depends upon type of metal ions and the bacterial genus as different genus has variable cellular contents. Using bacteria in fine powder form in various batch process helps in quick achievement of equilibrium and improved biosorption capacity due to nonexistence of mass transfer resistances. The solution chemistry affects bacterial surface chemistry and metal speciation in the solution. Therefore optimum conditions for biosorption and careful pretreatment of biomass need to be fully understood before full exploitation of bacterial biosoprtion potential.

Bacteria Metal M=Biomass dosage, teq=Equilibrium time Uptake (mg/g) Reference
Bacillus coagulans Chromium (VI) M=2 g/l, teq=1 h 39.9 (E) at pH 2.5 [106]
Bacillus licheniformis Chromium (VI) M=1 g/l, teq=2 h 69.4 (L) at pH 2.5 [107]
Bacillus megaterium Chromium (VI) M=2 g/l, teq=1 h 30.7 (E) at pH 2.5 [106]
Bacillus thuringiensis Chromium (VI) M=1 g/l 83.3 (L) at pH 2.0 [108]
Chryseomonasluteola Chromium (VI) M=1 g/l, teq=1 h 3.0 (L) at pH 4.0 [109]
Pseudomonas cepacia Copper NA 65.3 (L) at pH 7 [110]
Pseudomonas putida Copper NA 6.6 (L) at pH 6.0 [111]
SpHaerotilusnatans Copper M=3 g/l; teq=0.5 h 60 (E) at pH 6.0 [112]
Streptomyces coelicolor Copper M=1 g/l; teq=8 h 66.7 (L) at pH 5.0 [113]
Bacillus circulans Cadmium M=0.5 g/l; teq=2 h 26.5 (E) at pH 7.0 [114]
Pseudomonas putida Cadmium NA 8.0 (L) at pH 6.0 [111]
Streptomyces rimosus Cadmium M=3 g/l 64.9 (L) at pH 8.0 [115]
Corynebacterium glutamicum Lead M=5 g/l, teq=2 h 567.7 (E) at pH 5.0 [116]
Pseudomonas putida Lead M=1 g/l, teq=24 h 270.4 (L) at pH 5.5 [117]
Streptomyces rimosus Lead M=3 g/l; teq=3 h 135.0 (L) [118]
Streptoverticilliumcinnamoneum Lead M=2 g/l, teq=0.5 h 57.7 (E) at pH 4.0 [119]
Lactobacilliusbulgaricus Lead M=4.5 g/l. 42.6 mg/gm at pH 6.0 [120]
Bacillus thuringiensis Nickel M=1 g/l, teq=8 h 45.9 (L) at pH 6.0 [121]
Streptomyces rimosus Nickel M=3 g/l, teq=2 h 32.6 (L) at pH 6.5 [122]
Arthrobacternicotianae IAM 12342 Thorium M=0.15 g/l, teq=1 h 75.9 (E) at pH 3.5 [123]
Bacillus licheniformis IAM 111054 Thorium M=0.15 g/l, teq=1 h 66.1 (E) at pH 3.5 [123]
Bacillus megaterium IAM 1166 Thorium M=0.15 g/l, teq=1 h 74.0 (E) at pH 3.5 [123]
Bacillus subtilis IAM 1026 Thorium M=0.15 g/l, teq=1 h 71.9 (E) at pH 3.5 [123]
Corynebacterium equi IAM 1038 Thorium M=0.15 g/l, teq=1 h 46.9 (E) at pH 3.5 [123]
Pseudomonas sp. (strain MTCC 3087) Thorium, uranium M=2g/l, teq=12 h Uptake of 43–54% of cell dry weight at pH 4-5 [124]
Citrobacterfreudii Uranium M=6g/L Uptake of 94.68% at pH 6.0 [125]
Arthrobacternicotianae IAM 12342 Uranium M=0.15 g/l, teq=1 h 68.8 (E) at pH 3.5 [123]
Bacillus licheniformis IAM 111054 Uranium M=0.15 g/l, teq=1 h 45.9 (E) at pH 3.5 [123]
Bacillus megaterium IAM 1166 Uranium M=0.15 g/l, teq=1 h 37.8 (E) at pH 3.5 [123]
Bacillus subtilis IAM 1026 Uranium M=0.15 g/l, teq=1 h 52.4 (E) at pH 3.5 [123]
Zoogloearamigera IAM 12136 Uranium M=0.15 g/l, teq=1 h 49.7 (E) at pH 3.5 [123]

Table 4: Detailed list of metal biosorption by various bacteria along with their metal uptake capacity. Note: (E)=Experimental uptake, (L)=Uptake predicted by the Langmuir model. NA means not available.

Biosorption by yeast: Biosorption by yeast biomass have been studied extensively because of the ease of availability of large amount of waste fungal biomass from various fermentation industries and its amenability to genetic and morphological manipulations. Of all the fungi, bisorption potential of fungi like Rhizopus, Aspergillus, Streptoverticillum, Phanerochaete and Saccharomyces has been studied the most. Yeast such as Saccharomyces cerevisiae is widely used in food and beverage production and is easily cultivated using cheap media. Various batch and culture experiments using yeast have proved the bisorption potential of yeast and its ability to remove metals form contaminated waste waters. Experimental parameters affecting biosorption process are pH, biosorbent dose, initial metal concentration, contact time and particle size as is the case with other biosorbents. Treatment of biomass with mineral acids causes desorption and help in regenerate ability of biomass. Yeast can accumulate inordinate amount of metals due to production of extracellular yeast glycoproteins. The biosorption mechanisms have been related to different cell wall constituents [31-36]. {Murray, 1975 #85}The major functional groups involved for biosorption are carbonyl, amino groups and methyl groups present in biomass cell surface [37]. Table 5 enlists different experiments using yeast for biosorption of metals and important inference.

Yeast Metal sorbed Results Reference
Phanerochaetecrysosporium Chromium (VI) Batch experiments (shake flask condition) using chromium containing wastewater was studied for biosorption capacity of yeast. Maximum biosorption of63.72% was obtained at pH 2 for acid-treated biomass type at initial concentration of 100 ppm. [126]
Saccharomyces cerevisiae subsp. uvarum Mercury Batch experiments using aqueous solution (20 ml) containing 0.5 mmol/Lof Hg2+was incubated with magnetically modified yeast cells at a pH of 7.0 The maximum Hg2+ biosorption capacity was 114.6 mg/g at 35ºC. [127]
Candida tropicalis CBL-1 Cadmium Lab experiments on metal solution at a concentration of 100 mg/l of Cd(II) showed that Candida tropicalis CBL-1 reduced Cd(II) 59%, 64% and 70% from the medium after 48, 96 and 144 h, respectively. Moreover the yeast was also able to remove Cd(II) 46% and 60% fromthe wastewater containing Cd(II)after 6 and 12 days, respectively. [128]
Candida tropicalis Copper Experiments carried out in culture flasks at different concentration of copper solution showed decrease in uptake capacity with increase in biomass concentration at optimalpHrange of 5 to 7. Uptake was reported to be dependent on cell age. Cells at stationary growth phase had highest uptake capacity. [129]
Saccharomyces cerevisiae Lead Entrapment of the biomass in a sol–gel matrix was observed. The yeast cells were homogeneously distributed into the solid matrix and could take up hazardous heavy metals from aqueous solution.
Using inactive biomass, the maximum metal ions uptake at optimum biosorption temperature of 25°C were found to be 270.3, 46.3 and 32.6 mg g-1, respectively for Pb(II), Ni(II) and Cr(VI).
[130]   [131]
Copper Saccharomyces cerevisiae immobilized on sepiolite was able to retain metal when metal solution was passed through the column at pH 8 clearly indicating metal binding capacity of yeast for metal.
Metal uptake capacity of 8.0-8.1 mg/gm for copper from aqueous solution was obtained with formaldehyde cross-linked Saccharomyces cerevisiae in column bioreactors.
[132]    [133]
Cadmium The adsorption process was pseudo-second-order with respect to metal ion concentration and occurred in four distinct steps.
Biosorption experiments using artificial aqueous solution and pretreated yeast biomass showed maximum metal uptake values (qmax, mg g-1) at31.75.
[134]   [135]
Mercury Yeast cells were succesfully used to separate methyl mercury from Hg2+using biosorption.Binding of methyl mercury to yeast was independent of solution pH, temperature, incubation time, amount of biomass etc. [136]
Nickel Biosorption experiments using inactive yeast showed maximum Ni2+ ions uptake of46.3 mg g-1at 25°C. [131]
Chromium (VI) Sorption was exhibited by both intact cell and dehydrated cells with the latter having greater potential at 30°C or 45°C. [137]
Uranium Batch experiments at contact time of 1 h, pH=6.5 and 10-1 M UO2(CH3COO)2 solution as uranyl source showed the maximum degree of bioaccumulation at 8.75 mmol UO22+/g yeast.
Batch experiments using non-living yeast as biosorbent showed an optimum uranium uptake at pH 5 and 100 µm particle size of biomass at adsorbent dose of 10
g/l and initial metal concentration of 100mg/l. Maximum uptake was observed after contact time of 75 minutes.
[138]   [37]

Table 5: Biosorption by different yeast species and important inferences.

Chemical method of removal deals with chelation of heavy metal with suitable natural compound. These natural compounds can be of animal or plant origin such as alginates, citrates, flavonoids and phytic acid.

Alginates: Alginate is an anionic polysaccharide found in cell walls of brown algae. Chemically, it is a linear copolymer with homopolymeric blocks of (1-4)-linked β-D-mannuronate (M) and its C-5 epimer α-L-guluronate (G) residues, respectively. These two units are covalently linked together in different sequences or blocks. Alginates are obtained from sea weeds and bacteria. Seaweeds include the giant kelp Macrocystis pyrifera, Ascophyllum nodosum and various types of Laminaria whereas bacterial source of alginates are Pseudomonas and Azotobacter genera. Alginates from different sources, collected at different seasons from plants of different age have variable physical property, chemical property and yield [38]. In one of the study by Tanaka and coworkers, partially degraded alginates yields products which can better prevent strontium absorption in the body compared to parent seaweeds [39]. Degraded alginates form relatively non-viscous solutions and are relatively easier to administer with food or in drinking water. Alginates have the capability of reducing body burden of radiostrontium and acts by binding to radioisotope [40]. Binding of radioisotope with alginate is through divalent bonding. Thus alginates are potential candidates for removing or inhibiting heavy metals uptake from the body when taken internally. The ‘egg box’ model explains the binding of alginates with divalent metal ions [41]. Figure 2 shows a schematic representation of calcium induced gelation of alginate in accordance with ‘egg-box’ model. The divalent calcium cations (Ca2+) binds to guluronate blocks of the alginate chains as they have high affinity to divalent cation. The guluronate blocks of one polymer then form junctions with the guluronate blocks of adjacent polymer via ionic bridges formed between ionized carboxyl groups of the adjacent alginate. Sodium Alginate at a concentration range of 1.4%, 12% and 24% reduces Sr-89 uptake in a constant proportion. In a study reported by Harrison and coworkers, oral use of commercial jelly containing 1.5 gms of sodium alginate caused two fold reduction of strontium absorption in the body [42]. Such properties of alginates can be attributed to its complexation ability. The binding strength of alkaline earth metals to both polymannuronate and polyguluronate was found to decrease in the order Ba2+2+2+2+. They also interpreted the preferential binding of heavier ions to stereochemical effects [43].


Figure 2: ‘Egg box model’ showing calcium induced gelation of alginates. Here Ca2+ is the calcium ion being held in ionic bridges formed between calcium ions and the ionised carboxyl groups of adjacent alginate chain.

Citrus pectin: Citrus pectin is complex polysaccharide derived from the inner peel white pulp of citrus fruit. Its principal monosaccharide is D-galacturonic acid along with some amount of neutral sugars. It has the ability to bind toxic heavy metals and excrete them from the body. Essential minerals are not disturbed during this process [44]. Pectin can bind with Pb, Cu, Co, Ni, Zn, Cd as well as with Ba, Zn, Sr, Mn, Mg [45]. Citrus pectin is used for effective treatment of lead poisoning in children. Clinical study by Zhao and his colleagues on oral administration of modified citrus pectin (MCP) showed effective lowering of lead level in the blood of children between the ages of 5 and 12 years [46]. Another study by Li and co-workers showed that modified pectin cross linked with adipic acid has a rough, porous phase covered with carboxy groups and shows high adsorption capacity. The saturated loading capacity for Pb2+, Cu2+ and Zn2+ reached 1.82 mmol/g, 1.794 mmol/g and 0.964 mmol/g, respectively [47]. In another study by Schiewer and Patil on removal of heavy metals from food processing and agriculture industry, it was observed that pectin-rich fruit can be effectively used for removal of dangerous heavy metal cadmium form waste water. They observed that metal uptake got increased with pH, with uptake capacities ranging between 0.5 and 0.9 meq/g of dry peel [48]. Sorption followed second order model. Owing to their low cost and high physical stability, pectin rich fruit can be suitably explored. The modified form of citrus pectin (MCP) consists of approximately 10% rhamnogalacturonan II and is widely known metal cation chelator [49,50]. Oral administration of modified citrus pectin (MCP) increases total urinary excretion of arsenic and cadmium from body [44]. Pectin has same mechanism of binding as alginate (i.e. ‘egg-box’ mechanism). According to this, long fiber chains in solution stack together in groups and leads to formation of pockets. Pockets serve as the point where metal cations can form complex with the fibers. This model has been confirmed by X-ray diffraction [51] and NMR spectroscopic analyses [52].

Flavonoids: They are a class of plant secondary metabolites that perform a variety of functions such as plant pigment, UV filters and physiological regulator. Different studies by Bukhari and co-workers and Zhou and co-workers found that the antioxidant activity of flavonoids depends mainly on number as well as position of hydroxyl group in the flavonoid structure [53,54]. A number of flavonoids have been shown to have chelation with heavy metal ions particularly with copper and iron. Studies have shown that complex formation can take place between metal (copper and iron) and flavonoids having the stochiometries of metal:flavanoids 1:1, 1:2, 1:3, 2:2, 2:3 [55]. Optimal pH for complex formation is around 6.0. Flavonoids can also be used as active antidote for heavy metal poisoning In vivo [56]. Quercetin forms complex with Al (III) thereby reducing its overload in diet [57]. Fluorescence spectroscopy has shown that quercetin can form 3:1 complex with eight rare earth metals [54]. It is the 3-or 5- hydroxypyran-4-one group in B flavonoid ring which plays a principal role in chelating action. Morin forms Pd (II) and Pt (II)-complexes as is shown in Figure 3 [58].


Figure 3: Probable structure of M(II)-morin complex. M = Pt(II), Pd(II) and Zinc (II).

The structure is suggested due to the fixation of benzene ring caused by the effect of coordination after the complex was formed. Malesev and Kuntić showed that benzoyl moiety is basic site for metal chelation by their IR spectroscopy of Pd (II)-quercetin and UO2-rutin complexes. The complexes with rutin, morin or 3-hydroxyflavone are quiet stable with WO42- anion in the center with the ligand-metal interaction partly electrostatic [59].

Phytic Acid: Phytate (myo-inositol (1, 2, 3, 4, 5, 6) hexakisphosphate) is a natural compound formed during maturation of plant seeds and grains. Structurally phytic acid has 12 replaceable protons in its molecule which gives it tremendous potential of forming complex with positively charged multivalent cations and positively charged proteins [60]. Due to its structure, it can strongly interact with many metals and nonmetals, proteins and starch. The interaction is mainly electrostatic [61-63]. Rimbach and Pallauf reported that bioavailability and toxicity of cadmium was significantly reduced by phytic acid. Similar results were obtained for lead [64]. Phytic acid and iron form insoluble complexes that is not available for absorption under pH conditions of the small intestine [65].

Phytic acid forms variety of salts with metal ions easily and exist as phytate metal ion complex at a certain pH. A change in pH leads to formation of other complex having altered stability. Vohra and coworker reported the order of stability of phytate-metal complex as Cu2+>Zn2+>Ni2+>Co2+>Mn2+>Ca2+ at pH 7.4 [66]. Phytic acid immobilized on suitable surface such as poly 4-vinyl pyridine can act as very good adsorbent. The order of metal ion adsorption at pH 6.5 by PVP- Phytic acid complex was Ni2+>Zn2+>Cu2+>Co2+> Cd2+>Pb2+ (Tsao et al. 1997). According to International Union of Pure and Applied Chemistry (IUPAC), potentiometry and multinuclear NMR have been the main instrumental techniques used for the determination of stability constants of such complexes [67]. A study by Reinhold and coworkers on phytate rich diet found that phytate has inhibitory effect on Zn absorption [68]. Several subsequent single meal studies clearly showed a negative correlation between presence of phytate or ionositol phosphates and zinc absorption in humans [69-72]. In another study by Bohn and coworkers on 20 human volunteers (10 males and 10 females) it was found that fractional magnesium absorption from white bread was significantly impaired by addition of phytic acid. The effect was dose dependent as addition of 1.49 mmol of phytic acid lowered magnesium absorption from 32.5 ± 6.9% to 13.0 ± 6.9% and addition of 0.75 mmol phytic acid lowered magnesium absorption from 32.2 ± 12.0% to 24.0 ± 12.9% [73]. Phytic acid is also known to chelate uranium. A study by Cebrian and his colleagues found that the In vitro ability of phytic acid to chelate uranium was 2.0, 2.6 and 16 times higher than that observed for ethidronate, citric acid and diethylenetriaminepenta-acetic acid (DTPA) respectively [74].


The build-up of toxic pollutants such as metals, radionuclides and organic contaminants in soil, sludges, surface water and groundwater by various anthropogenic activities affects natural resources and causes a major strain on ecosystem. Phytoremediation, also referred to as botanical bioremediation, is the use of green plants for the treatment of soil, water and air pollution [75]. It is an effective in situ remediation technology that utilizes the inherent abilities of living plants to cleanse nature. It is an ecologically friendly and solar energy driven clean-up technology. Phytoremediation involves growing plant in a contaminated matrix for a required period of time to remove contaminants from the matrix or to facilitate the immobilization (binding/containment) or degradation of pollutants. The plant can be subsequently harvested, processed or even disposed. Plants have remarkable metabolic and absorption capabilities as well as transport system that can take up nutrients or contaminants selectively from growth matrix, soil or water. The uptake of contaminants in plants occurs primarily through the root system, in which the principal mechanisms for preventing contaminant toxicity are found. The root system owing to their enormous surface area causes absorption and accumulation of water and nutrients essential for growth as well as other nonessential contaminants. It is the genetic adaption by plants to handle the accumulated pollutants which results in effective contaminant uptake from soil and waste water. Phytoremediation takes advantage of natural plant processes and requires comparatively less equipment and labor than other methods since plants do most of the work. Also, the site can be cleaned up without digging up and hauling soil or pumping groundwater, which saves energy. The widespread plant cover help control soil erosion, reduce noise, and improve overall surrounding air quality. Other advantage includes its low cost, wide spectrum of action against different metals, generation of recyclable plant products and public acceptance. Phytoremediation of land contaminated with inorganic and/or organic pollutants has been a subject of considerable attention and research over the last decade [76-78]. The degradation by-products from plants may be mobilized in groundwater or bio-accumulated in animals. The depth of plant root in soil limits the treatment zone which in most cases is shallow. Climatic factors will also influence its effectiveness. The success of remediation depends on carefully selecting plant community. Introducing new plant species to an area may cause widespread ecological ramifications. Moreover the overall process time is too long taking several years to clean up a site. Phytoremediation can be achieved by different mechanisms that include phytoextraction, phytostabilization, phytotransformation, phytostimulation, phytovolatilization and rhizofiltration.

Phytoextraction involves the uptake of contaminants from contaminated soil or water by plant and their simultaneous translocation to harvestable parts of plant. This follows a complex series of events starting from dissolution of metal, its absorption transport and finally storage. Phytoextraction is an effective In situ technique for removing heavy metals from polluted soils and promote long term clean-up of soil or wastewater. Use of hyper accumulators is an important strategy for phytoextraction as they can accumulate inordinate amount of elements within their tissues. A large number of plants have been used for phytoextraction. Some of them are enlisted in Table 6.

Metal removed Plant Species Family Mechanism for Removal Reference
Cadmium Chamomillarecutita and Hypericumperforatum L Asteraceae Secondary metabolites of plant complexes with cadmium forming less toxic organo-metallic complexes. [139]
Zinc Brassica juncea Brassicaceae Due to production of high biomass of shoot. [140]
Nickel Psychotriadouarre Asteraceae Metal removal attributed to high concentrations of tannins in leaves which functions as a detoxicant for elevated cytoplasmic metal concentrations, in addition to providing defensive benefits. [141]
Uranium Brassica chinensis, Brassica juncea, Brassica narinosa, Amaranthus species Brassicaceae Chelation [142]
Thallium Iberis intermedia Brassicaceae Due to high amount of thallium accumulation in the leaves. [143]
Mercury Eichhorniacrassipes Potederiaceae Due to binding of Hg ionically to oxygen ligands in roots, most likely to carboxylate groups and by covalent binding to sulfur groups in shoots. [143]

Table 6: List of plants used in phytoextraction, their mechanisms and the target metal.

Plant root mediates dissolution by secreting phytosiderophores, organic acids, or carboxylates which helps in capturing metal in the rhizosphere and transports it over the cell wall. The transport of metal from root to shoot is regulated by various transporters. For hyperaccumulators, leaves in the shoot system stores maximum amount of heavy metals.

Phytostabilization involves the reduction of mobility of heavy metal in soil through absorption and accumulation by roots, adsorption onto roots or precipitation within the root zone of plant. The addition of soil amendments result in decrease of solubility of metals in soil and minimize its leaching to groundwater. Various soil amendments include phosphates minerals (hydroxyapatite, phosphoric acid), iron and manganese oxides, aluminosilicates (bentonite, montmorillonite, zeolites) etc. The net result is that pollutants become less bioavailable and thus human exposure is significantly reduced. In one of the experiments by Blaylock and his colleagues it was observed that phytostabilization may reduce metal leaching by converting metals from a soluble oxidation state to an insoluble oxidation state. This technology does not remove contaminant from its location and thus excludes the need for treatment of secondary waste and further adds to the fertility of soil. The plants which are used for phytostabilization be tolerant to metal and should not accumulate contaminants in above-ground parts which are liable to be consumed by humans or animals.

Phytotransformation is use of plants for transformation of contaminants in sediments, soil and natural water to environmentally more acceptable products. The process involves breakdown of contaminants taken up by plant through various metabolic processes occurring within the plant or breakdown of contaminants in the vicinity of plant by the effect of various compounds such as enzymes produced by the plant. The various complex molecules present as pollutants in the soil or water are degraded into simple molecules which are simultaneously incorporated into the plant tissue thereby promoting plant growth.

Phytostimulation, also called rhizodegradation, is the plant assisted breakdown of organic contaminants in the soil via enhanced microbial activity in the plant root zone or rhizosphere. The enhanced microbial activity can be attributed to various secretions like sugar, amino acids, carbohydrates and enzymes by the roots. The root system brings oxygen to the rhizosphere thereby ensuring aerobic transformations. Thus microbes help in digestion and breakdown of various pollutants present in the soil. The successful design of phytostimulation experiments requires dense root system and microbes which can degrade the contaminants. However this technique works at low level of pollutants in shallow areas and is a much slower process.

Phytovolatilization is the ability to take up contaminants in the transpiration stream and then transpire volatile contaminants. Thus plants may serve as effective pump-andtreat systems for mobile contaminants including volatile compounds like carbon tetrachloride (CCl4) and ethylene dibromide (EDB). Removal of tightly sorbed contaminants from micropores within the soil may be the rate-limiting step for their remediation. Dewatering increases the potential for gasphase diffusion within the soil. Thus phytovolatilization offers good option for effective removal of volatile contaminants particularly from the soil, surface and water.


The problem of heavy metal pollution is worsening day-byday due to human activities. Therefore, the removal of metals from human body and environment becomes a subject of paramount importance. Removal of heavy metals can be done by both physical as well as chemical means. For physical removal, adsorption and bio-sorption are employed mechanism while chemical removal mainly employs chelation. Adsorption of metal ions using plant waste products presents an effective as well as economical approach for heavy metal removal from aqueous effluent. The stability and adsorption capacity of adsorbent can be enhanced by suitable pretreatment with alkali or acids which causes an increase in active binding sites, better ion exchange properties or may even lead to formation of new functional group that may favor metal ion uptake. Biosorption is also an effective technique used for heavy metal removal from aqueous wastes. For algae biomass, ion exchange is shown to play an important role in metal sequestering mechanisms. For yeast and bacteria, biosorption can be attributed to different cell wall constituents. This technique has been extensively used for treating heavy metal contamination especially in the waste water coming from electroplating, mining and textile bath industries which can contaminate the environment. Another method of heavy metal removal is chelation. Chelation is exhibited by many active principles of plants and animals. Alginates, citrus pectin, phytic acid and flavonoids appear promising in this regard. Majority of these compounds can be used orally after minor modifications for treating heavy metals toxicity as they have the ability to effectively chelate the metal.

The use of plants in metal extraction has also appeared as a promising alternative for the heavy metal removal from water, sludges and soil. The various mechanisms employed include extraction, containment and immobilization, and volatilization etc. However commercial application of phytoremediation is relatively low in spite of large amount of research being done in this field. Selection of appropriate plant species and modification of cultivation condition can help in an enhanced removal of pollutants using plants. Biotechnological interventions can help in creation of new plant species having increased remediation ability.


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