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INTERNATIONAL JOURNAL OF ENVIRONMENTAL SCIENCES Volume 4, No 2, 2013

© Copyright by the authors - Licensee IPA- Under Creative Commons license 3.0

Research article ISSN 0976 – 4402

Received on May 2013 Published on September 2013 193

Phytoremediation: Emerging and green technology for the uptake of

cadmium from the contaminated soil by plant species Subhashini V1⃰, Swamy A.V.V.S1, Hema Krishna. R2

1- Department of Environmental Sciences, Acharya Nagarjuna University,

Nagarjuna Nager-522510, Guntur (Dist.) Andhra Pradesh, India.

2- Department of chemistry, University of Toronto, Ontario, Canada. M5S 3H6.

[email protected]

doi:10.6088/ijes. 2013040200008

ABSTRACT

The environmental cleanup of toxic metals such as cadmium was paramount important and it

was a potential environmental hazard. A novel, cost-effective and eco-friendly technologies

are needed to remove cadmium from the contaminated soil environment. The present

research study was designed to assess the naturally enhanced phytoextraction potential of

different plant species from the Cadmium contaminated soil. Uptake of cadmium by plant

species in a metal contaminated soil was studied in pot culture experiment. The pots were

filed with 5 kg of garden soil. Weed plants were grown in pots and were irrigated with known

heavy metal solutions in the concentration of 5ppm heavy metal (Cd) solution was added to

the pots alternate days up to 60 days. In controls normal water was used. The plants were

grown for a period of 60 days. The initial soil heavy metal concentration was analyzed. After

20, 40 and 60 days soil heavy metal concentrations were also analyzed. Analysis of heavy

metal was done in HNO3/HClO4 digested samples by Atomic Absorption spectrophotometer.

The plant species showed relatively good response to the higher level of heavy metal

concentration in the roots, stem and leafs suggested that these plant species were good metal

accumulator with the possibility of extracting Cd from contaminated soils.

Key words: Phytoremediation, Contaminated soil, Pot culture experiment, Cadmium, and

Plant species.

1. Introduction

Chemical pollution in the soil and water bodies has become a major source of concern and

has posed a serious health problem in many countries. Phytotechnologies are a set of

technologies using plants to remediate or contain contaminants in soil, groundwater, surface

water, or sediments. Some of these technologies have become attractive alternatives to

conventional cleanup technologies due to relatively low costs and the inherently aesthetic

nature of planted sites. The concept of using plants to clean up contaminated environments is

not new. About 300 years ago, plants were proposed for use in the treatment of wastewater

(Hartman 1975). At the end of the 19th century, Thlaspi caerulescens and Viola calaminaria

were the first plant species documented to accumulate high levels of metals in leaves

(Baumann1885). In 1935, Byers reported that plants of the genus Astragalus were capable of

accumulating up to 0.6 % selenium in dry shoot biomass. One decade later, (Minguzzi and

Vergnano 1948) identified plants able to accumulate up to 1% Ni in shoots. More recently,

(Rascio 1977) reported tolerance and high Zn accumulation in shoots of Thlaspi caerulescens.

Despite subsequent reports claiming identification of Co, Cu, and Mn hyperaccumulators, the

Phytoremediation: Emerging and green technology for the uptake of cadmium from the contaminated soil by

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Subhashini V et al

International Journal of Environmental Sciences Volume 4 No.2, 2013 194

existence of plants hyperaccumulating metals other than Cd, Ni, Se and Zn has been

questioned and requires additional confirmation (Salt et al., 1995). The idea of using plants to

extract metals from contaminated soil was reintroduced and developed by (Utsunamyia 1980)

and (Chaney 1983) and the first field trial on Zn and Cd phytoextraction was conducted in

(Baker et al .,1991). In the last decade, extensive research has been conducted to investigate

the biology of metal phytoextraction. Despite significant success, our understanding of the

plant mechanisms that allow metal extraction is still emerging. In addition, relevant applied

aspects, such as the effect of agronomic practices on metal removal by plants are largely

unknown. It is conceivable that maturation of phytoextraction into a commercial technology

will ultimately depend on the elucidation of plant mechanisms and application of adequate

agronomic practices. Natural occurrence of plant species capable of accumulating

extraordinarily high metal levels makes the investigation of this process particularly

interesting.

Heavy metals are conventionally defined as elements with metallic properties (Ductility,

conductivity, stability as cations, ligand specificity, etc.) and atomic number >20. The most

common heavy metal contaminants were: Cd, Cr, Cu, Hg, Pb, and Zn. Metals are natural

components in soil. Contamination, however, has resulted from industrial activities, such as

mining and smelting of metalliferous ores, electroplating, gas exhaust, energy and fuel

production, fertilizer and pesticide application, and generation of municipal waste (Kabata-

Pendias and Pendias,1989). High levels of metals in soil can be phytotoxic. Poor plant growth

and soil cover caused by metal toxicity can lead to metal mobilization in runoff water and

subsequent deposition into nearby bodies of water. Furthermore, bare soil is more susceptible

to wind erosion and spreading of contamination by airborne dust. In such situations, the

immediate goal of remediation is to reclaim the site by establishing a vegetative cover to

minimize soil erosion and pollution spread.Soil remediation is needed to eliminate risk to

humans or the environment from toxic metals. Human disease has resulted from Cd (Nogawa

et al., 1987; Kobayashi 1978; Cai et al., 1990). Majority of studies on phytoremediation was

based on pot experiments and hydroponic culture, and only a few reports evaluated the

phytoextraction potential of hyperaccumulators or high biomass crops under field conditions

(McGrath et al., 2006; Zhuang et al., 2007). Only a few attempts have been made to evaluate

the possibility of metal removal in response to modifications of agronomic practices

(Marchiol et al., 2007). Some weeds of the grass family have been experimented to be

suitable for phytoremediation because of their multiple ramified root systems. Keeping in

view the heavy metals exposure directly to the soil environments, this study was planned

under green house conditions with the objectives to evaluate the efficacy of different plant

species in the uptake and translocation of Cadmium from the contaminated soils.

2. Materials and methods

2.1 Selection of plant species

In this study ten plant varieties were selected for their ability to absorb heavy metal Cd from

the contaminated soil. Seedlings of the selected weed plant species, grass species and

flowering plants were collected from uncontaminated soils. The experimental plants were

four weed species, three flowering plants, three grass varieties. The used plant species were 1.

Weed species were; Acalypha indica, Abutilon indicum, Physalis minima and Cleome

viscose .2. Flowering plants were; Catharanthus roseus, Ruallia tuberose, and Canna indica.

3. Grass species were; Perotis indica, Echinocloa colona and Cyperus rotundus .


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Figure 1: Images showing various plants used for phytoremediation

2.2 Plant Sampling: Fresh plant samples were collected in the morning by pulling carefully

from the soil to avoid damage to the roots. Soil samples were collected from the surface to

subsurface portion of the soil around the plant roots at a range interval of 20 to 30 square

meters apart.


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2.3 Washing of plant samples: The collected samples were washed with distilled water

remove dust particles. The samples were then cut to separate the roots, stems and leaves. The

different parts (roots, stems and leaves) were air dried and then placed in a de hydrator for 2-3

days and then dried in an oven at 1000C. Dried samples of different parts of the plants were

ground into a fine powder using pestel and mortar and stored in polyethylene bags, until used

for acid digestion. Before the experiment soil parameters were done and for every 20 days the

soil samples were also collected and analysed for heavy metals by AAS.

2.4 Preparation of plant samples by wet digestion: Samples (0.5g) of each part (leaves,

stems and roots) of the plant were weighed in digestion flasks and treated with 5 ml of

concentrated HNO3. A blank sample was prepared applying 5ml of HNO3 into empty digestion

flask. The flasks were heated for 2 hours on an electric hot plate at 80-900C at which the

samples were made to boil and digestion continued until a clean solution was obtained. After

cooling, the solution was filtered with whatman No.42 filter paper. It was then transferred

quantitatively to a-25ml volumetric flask by adding distilled water. The filtrate was analyzed

for metal content using AAS (GBC 932 AA)

2.5 Preparation of standards and analysis of samples: Standard solution cadmium (Cd), was

prepared from the stock standard solution containing 1000 ppm of element in 2N nitric acid.

Calibration and measurements of elements were done on atomic absorption spectrophotometer

(a Analyst 300, perking Elmer Norkwalk, Conn, USA). The calibration curves were prepared

for this element individually. A blank reading was also taken and necessary correction was

made during the calculation of concentration of various elements.

2.6 Pot culture experiment: Uptake of heavy metals by plants in a metal contaminated and

normal soil was studied in pot culture experiment. Weed plants, flowering plants and grass

species were grown in pots and were irrigated with known heavy metal solutions in the

concentration of 5ppm heavy metal solution (Cd) was added to the pots alternate days up to 60

days. In controls normal water was used. The plants were grown for a period of two months

(60days). The contaminated soil received the metals Cd as Cd (NO3)2 .4H2OThe initial soil

heavy metal concentration was analysed. After 20, 40 and 60 days soil heavy metal

concentrations was analyzed. Every 20 days the plant samples from each pot were collected

and washed thoroughly under running tap water and distilled water so that no soil particles

remained. Analysis of heavy metals was done in HNO3/HClO4 digested samples (Tappi Test

Method, 1989) and analyzed by Atomic Absorption spectrophotometer.(GBC 932). The

experiment was carried out in the garden by using heavy metal solutions. The plant sample was

uniform in size and which were free of disease symptoms. The pots were filed with 5 kg of

garden soil. The plants were watered once every day and care was taken to avoid leaching of

water from the pots. After 20 days from each pot plant samples were collected. The plants were

harvested after 20 days, 40 days and 60 days for heavy metal analysis.

2.7 Statistical analysis: Average data uptake, transfer coefficient were submitted for statistical

analysis. The uptake and mobilization of PTM in plant tissues were assessed by

bioaccumulation coefficient (BC). Differences in heavy metal concentrations among different

parts of the weeds were detected using One-way ANOVA, followed by multiple comparisons

using Turkey tests. A significance level of (p < 0.05) was used throughout the study. All

statistical analyses were performed using STATISTICA (Version 7), MSEXCEL (Microsoft

office 2012) and GrphPad Prism (Version 5).


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3. Results and discussion

3.1 Soil physico-chemical properties

The results obtained from the soil analysis were shown in Table 1. Soil particle size was

determined using Bouyoucos hydrometer method. All reagents used were of analytical grade

(BDH Laboratory supplies, Poole, England) The soil pH was determined in a mixture of soil

and deionized water (1:2, w/v) with a glass electrode (McLean E.O 1882). Total organic

carbon content was determined using the Walkey-Black wet oxidation (Nelson. D.W et al.,

1982), Total Nitrogen was determined using the Kjeldhal method( Bremner. J.M, and

Mulvaney C.S, 1982). Cation exchange capacity (CEC) and amounts of exchangeable Ca and

Mg were determined using the ammonium acetate method (Rhoades J.D 1982). Total

phosphorus was determined calorimetrically. Total background cadmium concentration was

determined using Atomic Absorption Spectrophotometer (Buck scientific, 210 VGP). The

electrical conductivity (EC) was measured by using digital meters (Elico, Model LI-120)

with a combination of 1-cm platinum conductivity cell.

Table 1: Soil physico-chemical properties

CEC: Cation exchange capacity; EC: Electrical conductivity

The taxonomic classification of the experimental soil (Table 1) was sandy loam with pH of

6.94, EC of 455 mS/cm. The high pH level of the soil is generally within the range for soil in

the region; soil pH plays an important role in the sorption of heavy metals, it controls the

solubility and hydrolysis of metal hydroxides, carbonates and phosphates and also influences

ion-pair formation, solubility of organic matter, as well as surface charge of Fe, Mn and Al-

oxides, organic matter and clay edges (Tokalioglu et al., 2006).

3.2 Experimental procedure

The experimental plants were intended to grow for 2 months (60 days) in the soil. The pots

used for the experiment contained 5kg of soil. The seedlings of the experimental plants were

collected from the uncontaminated soils. In each pot eight seedlings were planted. Grass

species were also used for heavy metal accumulation. The plant species used for heavy metal

accumulation are tabulated in table-1.the total experimental period 60 days was divided into 3

S.No Parameter Result

1 PH 6.94

2 Organic matter 1.20

3 Total N (%) 0.85

4 Total (P) % 1.24

5 Sand (%) 18

6 Silt (%) 16

7 Clay (%) 42

8 CEC(mol/g soil) 10.99

9 EC(mS/cm) 455.00

10 Moisture content (%) 35.00

11 Cadmium mg/kg 0.366


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stages. The first 20 days period was regarded as I stage. The next 40 days period regarded as

II stage. The total 60 days period regarded as III stage. Cadmium total accumulation values

in selected plants were shown in Table.2.

Table 2: Cadmium total accumulations in selected plants

growing. After 20 days two plants were collected from each pot and soil samples were also

collected. The collected plant samples were separated into roots, stem and leaves. These are

air dried first for two days, and then dried in hot air oven at 101 for 12 hours. After the

plant parts are powdered using mortar and pistle. Then 2g of the root leaves and stem

powders are weighed and stored in polyethylene small bags for heavy metal analysis by

Atomic Absorption Spectrometer.

II- stage: After 20 days 5ppm/1000ml of heavy metal (Cd) solutions was added alternate days

to the all pots. The plants were growing. In these days the plants growth and physical changes

were noted. After 40 days two plants from each pot were collected and soil sample was taken

for heavy metal analysis. The collected plant samples plant parts were separated and dried in

hot air oven at 101 for 12 hours. After the plant parts are powdered using mortar and pastel,

then 2g of the root stem and leaves powders are weighed and stored in polyethylene bags for

heavy metal analysis.

III-stage: After 40 days the heavy metal concentrations was increased. 5ppm/1000ml of

heavy metal solutions such Cd solution was added alternate days to the all plots. The plants

were growing. In all these days growth, development and physical characters were recorded.

After 60 days two plants from each pot was collected for heavy metal analysis. The soil

samples were also collected from each pot for heavy metal analysis. The collected plant

samples the plant parts were separated into root, stem and leaves. These are air dried24hours

and then dried in the hot air oven at 101 for 12 hours. After the plant parts are powered

using mortar and pistle.2g of the plant material was weighed and stored in small polyethylene

bags for heavy metal analysis by AAS. In the third stage the collected soil samples were also

measured for heavy metal analysis. All these stages all the plants were grown in identical

environmental conditions. The experimental plants were four weed species, three flowering

plants, three grass varieties. Generally grass species does not having stem and leaves except

roots, hence we considered root analysis in grass varieties. Cadmium total accumulation

values in selected plants were shown in figure 1.


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Figure 1: Cadmium total accumulation in selected plants

3.3 Metal transfer coefficients

3.3.1. Accumulation Factor (AF)

(Baker 1981) divides plants in three category i.e. accumulator, excluder and indicator. In

accumulator plants the concentration ratio of the element in the plant to that in the soil is >1.

In excluder plant metal concentrations in aerial parts are maintained low (<<1) and constant

over a wide range of soil concentrations. In indicator plants the uptake and transport of metals

were regulated in such a way that the ratio of the concentration of element in the plant to that

in the soil is near 1.As total heavy metal concentration of soils is poor indicator of metal

availability for plant uptake, accumulation factor was calculated based on metal availability

and its uptake by a particular plant were shown in Table 3. Accumulation Factor for plants

was calculated as:

Table 3: Cadmium total accumulation standard deviation of selected plants

Name of the plant Cadmium mg/kg

20TH DAY 40TH DAY 60TH DAY

Acalypha indica 5.8932±0.0740 6.8241±0.1568 19.838±0.1190

Abutilon indicum 6.2076±0.0821 7.7793±0.1211 25.312±0.0813

Physalisminima 7.1715±0.0485 12.0761±0.0222 29.1225±0.0185

Cleome viscosa 1.3284±0.1609 3.1399±0.0222 23.9365±0.01918

Catharanthus roseus 3.6076±0.0847 6.5859±0.2326 27.4451±0.0360

Ruellia tuberosa 4.1713±0.0779 6.9153±0.1648 7.8289±0.1856

Canna indica 3.2688±0.1180 6.4778±0.1317 34.6278±0.0846

Perotis indica 1.949±0.0236 7.2003±0.0057 8.5223±0.0408

Echinocloa colona 1.6252±0.0473 8.0523±0.0073 12.9059±0.0230

Cyperus rotundus 2.5709±0.0620 13.1129±0.0865 16.3502±0.0806


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3.3.2 Translocation factor (TF)

Translocation factor is a measure of the ability of plants to transfer accumulated metals from

the roots to the shoots. It is given by the ratio of concentration of metal in the shoot to that in

the roots (Cui et al., 2007).

Table 4: Cadmium Translocation Factor of selected plants

TFs were also varied under different Cd concentrations in the growth media and a significant

difference (p≤0.05) observed among treatments in TFs. This index was in the range of 0.2674

to 0.7984 shown in Table.4. It was observed that TFs increased with increase in the applied

Cd concentration in the growth media. However, the TFs under different Cd levels were <1

except in Catharanthus roseus, indicating that all plant species were less able to tanslocate

Cd from the roots to the shoots efficiently thus labeling these plants as trace metal excluders

where as Catharanthusroseus only tanslocate Cd from the roots to the shoots efficiently

hence it is considered as accumulator plant (Cd levels was >1 ) . The translocation of Cd is

often restricted due to the ability of this element to create Cd-phytochelatin complex by

sequestration in the vacuole (Lux et al., 2011). Cd move-ment from root to shoots probably

occurs within the xylem. The levels of free Cd in the symplast can be influenced highly by

cellular sequestration of Cd and, therefore, it can affect the movement of Cd throughout the

plants (Niu et al., 2007).

3.3.3 Bio Concentration Factor (BCF)

Bioconcentration factor (BCF) is defined as the ratio of heavy metal concentration in plant

roots to that in soil (Malik et al., 2010). Bioconcentration factor (BCF) was used to evaluate

the efficiency of Cd phytoextraction and was calculated using the following Equation:

BCF = CCd-Roots /CCd-soil

where, CCd-roots (mg/kg) is the Cd concentration in roots part of the plant . CCd -soil

(mg/kg) is the Cd concentration in soil (Ghosh and Singh, 2005) and (Yoon et al., 2006).

Name of the plant Shoot Root Translocation factor

Acalypha indica 4.1857 15.6523 0.2674

Abutilon indicum 11.2378 14.0742 0.7984

Physalisminima 9.2134 19.9091 0.4627

Cleome viscosa 9.6954 14.2407 0.6808

Catharanthus roseus 24.1914 3.2537 7.435

Ruellia tuberosa 2.2484 5.5805 0.4029

Canna indica 8.6327 25.9951 0.332

Perotis indica 8.5223

Echinocloa colona 12.9059

Cyperus rotundus 16.3502


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Table 5: Cadmium- bio concentration factors of selected plants

The BCFs were varied under different Cd con-centrations in the soil and it was in the range

of 2.13904 to 9.46115 has shown in Table.5. There was a significant difference (p≤0.05)

among treatments in BCFs. The highest BCF (that is 9.46 ± 0.28) of Cd was found in Canna

indica as compared to other treatment levels. The BCFs were >1 under various treatment

levels. The BCF value of Cd usually ranged from 1 to 10 (Li et al., 2006). The BCFs

decreased with increase in the Cd concentration in the growth media, which may indicate the

restriction in soil-root transfer at higher Cd concentrations in the soil (Justin et al., 2011).

3.3.4 Relation between translocation factor and bio concentration factor

Phytostabilisation, metal-tolerant plants are used to reduce the mobility of metals, thus, the

metals are stabilized in the substrate (N.T. Abdel-Ghani, 2007) Plants with both

bioconcentration factor and translocation factor greater than one (TF and BCF> 1) have the

potential to be used in phytoextraction. Besides, plants with bioconcentration factor greater

than one and translocation factor less than one (BCF> 1 and TF< 1) have the potential for

phytostabilization (J. Yoon 2006). By comparing BCF and TF, the ability of different plants

in taking up metals from soils and translocating them to the shoots can be compared (J. Yoon

2006) .As shown in Tables 4 and 5 , among the sampled plants,except Catharanthus roseus

none of them were suitable for phytoextraction of Cd-metal but they were most suitable for

phytostabilization where as Catharanthus roseus have the potential to be used in

phytoextraction.

4. Conclusion

In our opinion the accumulated metal Cd concentration in roots, stems and leafs, growing

conditions, and their economic values were three major factors for selecting the plants. Based

on among the 10 sampled plant species, metal translocation into shoots appears to be very

restricted in the some plant species so that these plants will not be an effective source of

metal removal in this experiment. However, in the view of toxicology, this could be a

desirable property, as metals would not pass into the food chain via herbivores, and thus

avoid potential risk to the environment. Therefore, plants with low shoot accumulation should

be used in order to stabilize the metals and reduce the metal dispersion through grazing

animals or at leaf senescence. whereas some plant species like Catharanthusroseus have the

potential to be used in phytoextraction .Hence this study has been proved the possibility of

Name of the plant Total accumulation Bio concentration factor

Acalypha indica 19.838 5.42022

Abutilon indicum 25.312 6.95184

Physalisminima 29.1225 7.95696

Cleome viscosa 23.9365 6.54003

Catharanthus roseus 27.4451 7.49866

Ruellia tuberosa 7.8289 2.13904

Canna indica 34.6278 9.46115

Perotis indica 8.5223 2.32849

Echinocloa colona 12.9059 3.5262

Cyperus rotundus 16.3502 4.46726


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using 10 sampled plant species for phytoremediation of Cadmium and also possible for

phytoextration. These techniques reduce leaching, runoff and erosion via stabilization of soil.

Acknowledgement

The authors would like to thank to Prof. Z. Vishnuvarthan, Dean and Board of the Studies,

Department of environmental sciences Acharya Nagarjuna University, India, for his

cooperation and encouragement. Our special thanks also reserved for our family members

and friends who stand always by our side in all our endeavors.

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