Int. J. Aquat. Biol. (2014) 2(4): 206-214 E-ISSN: 2322-5270; P-ISSN: 2383-0956 Journal homepage: www.ij-aquaticbiology.com © 2014 Iranian Society of Ichthyology Original Article Phytoremediation efficiency of pondweed (Potamogeton crispus) in removing heavy metals (Cu, Cr, Pb, As and Cd) from water of Anzali wetland Hajar Norouznia1, Amir Hossein Hamidian*21 1Department of Environment, Faculty of Natural Resources and Earth Science, Kashan University, P.O. Box 8731751167, Kashan, Iran. 2Department of Environment, Faculty of Natural Resources, University of Tehran, P.O. Box 31585-4314, Karaj, Iran. Article history: Received 1 June 2014 Accepted 22 July 2014 Available online 2 5 August 2014 Keywords: Phytoremediation Heavy metal Potamogeton crispus Bioconcentration Translocation Abstract: Plant-based remediation (i.e. phytoremediation) is one of the most significant eco- sustainable techniques to cope with devastating consequences of pollutants. In the present study, the potential of a wetland macrophyt (i.e. Potamogeton crispus) for the phytoremediation of heavy metals (i.e. Cu, Cr, Pb, As and Cd) in the Anzali wetland was evaluated. The results showed that P. crispus tends to accumulate notable amounts of Cu, Cr, Pb, As and Cd according to their assayed concentrations as follows: 8.2 µg g-1 dw, 0.97 µg g-1 dw, 6.04 µg g-1 dw, 2.52 µg g-1 dw and 0.34 µg g-1 dw, respectively. Further accurate perception of the phytoremediation efficiency were conducted using both bioconcentration factor and translocation factor. The average of the highest bioconcentration factors was presented in a descending order as: 2.9×103, 1.9×103, 1.17×103, 0.68×103 and 0.46×103 for the Cu, Cr, Pb, Cd and As, respectively. Based on the results, P. crispus presents high potential to absorb all the alluded metals except for As and partly Cd. Correspondingly, the mean values of translocation factor were reported in the range of 0.41 to 2.24. Eventually, relying on the observed findings, the results support the idea that P. crispus species would be employed as the prospective candidate for the phytoremediation processes in Anzali wetland. Introduction Heavy metals pollution has been posed as a growing predicament worldwide. Hence, the natural environment quality levels tends to be worsened due to poor problem-solving skills and inadequate eco- suited techniques to diminish the detrimental demeanor of pollutants. Unlike organic pollutants, non-biodegradability property of heavy metals displays the immediate cause of bioaccumulation and its harmful subsequences on food chains (Khan et al., 2010; Hamidian et al., 2014). Therefore an acute health risk occurs to the environment and its living organisms. Thus, removing heavy metals from natural medium requires necessary action (Ali et al., 2013). The heavy metals are divided broadly into essential and non-essential. Essential heavy metals such as Zn, * Corresponding author: Amir Hossein Hamidian E-mail address: a.hamidian@ut.ac.ir Cu, Mn, Fe, Cr and Ni are necessary for functions in trace quantities in biological operations (Cempel and Nikel, 2006). In contrast, Hg, Cd, As and Pb known as non-essential heavy metals which are not only unnecessary for organism, but also have toxic and deleterious effects (Dabonne et al., 2010). Major origin of contaminants such as heavy metals are anthropological activities including industrial and municipal effluents, mining, untreated wastewater and agrochemicals and also, natural-driven origins such as volcanic eruption, ore weathering and mineral deposition (Kabata-Pendias and Pendias, 1989; Rai et al., 2008). Concern about marked release of heavy metal pollutants and cleaning up of contaminated areas has been increasing as the major controversial and disputed issues in the contemporary time. Therefore, 207 Norouznia and Hamidian/ Phytoremediation efficiency of pondweed in removing heavy metals many physical, chemical and biological approaches have been applied to reduce the impacts of pollutants especially heavy metals (Sheoran et al., 2011). Most of these methods appear to cost high and disruptive to the natural properties of environment and ended to soil erosion rapidly, and may also cause multiplied environmental problems (Ali et al., 2013). The most innovative and effective alternative method to omit the heavy metals from environment is phytoremediation that is a green, safe, solar-oriented and cost effective method using the capability of plant species to absorb high levels of metals in specific tissues like roots and leaves especially in the aquatic substratum (Carranza-Άlvarez., 2008; Sigh and Prasade, 2011). The inception of phytoremediation idea was owing to Chaney (1983) and came to apply eagerly during the recent two decades as a better way to solve contamination problems particularly by aquatic plants for removal of pollutants from contaminated surface water (i.e. phytofilteration) (Mukhopadhyay and Maiti, 2010). Based on Ali et al. (2013), proper plant species for phytoremediation needs some distinctive features such as to spread widely, easy to grow and mainly high potential to accumulate heavy metals. The aquatic macrophyts have innate ability to uptake heavy metals from polluted water medium and wastewater (Hamidian et al., 2014). The aquatic plants can absorb more Cu, As, Br, Cr, Cd, Sr, V and Pb than terrestrial plants (Sood et al., 2012). Phytoremediation and metal removal using aquatic macrophyts can be enormously reinforced by preferring appropriate plants species (Fritioff and Greger, 2003; Hamidian et al., 2014). Identification of proper plants for phytoremediation in a contaminated area is a full-scale model to survey its mechanisms of heavy metals purification and accumulation (Carranza-Alvarez et al., 2008; Hamidian et al., 2014). Therefore, this study aimed to assess the phytoremediation potential of a native and dominant floating plant in Anzali wetland, Potamogeton crispus (Pondweed), and evaluate its innate capability to accumulate heavy metals as hyper-accumulators to meet the phytoremediation purposes. Potamogeton crispus is a widespread floating species in wetland ecosystems especially in Anzali wetland (Pajevic et al., 2008). This species is a floating plant and its metal-accumulation aspect is related to its surrounding water metal concentrations (Favas and Pratas, 2013). Hence, this study intends to determine the extent to which the above- mentioned species is well-suited for phytoremediation and whether could be used as a proper accumulator species to uptake specific heavy metals including Cu, Cr, Pb, As and Cd based on (1) to investigate the concentration of metals in water and plant body, (2) to determine the removal potential of Cu, Cr, Pb, As and Cd using P. crispus, (3) to estimate the metal transportability according to bioconcentration factor (BCF) for water and plant tissues; Moreover, transfer factor (TF) for shoot and root and (4) to confirm the positive correlation Figure 1. Location of Anzali wetland in Iran 208 Int. J. Aquat. Biol. (2014) 2(4): 206-214 between metal concentration in water and plant in the eastern part of Anzali wetland in Iran. Material and methods Description of study area: Anzali wetland is an outstanding coastal lagoon that is located in the southwest of Caspian Sea including an area of about 200 km2 located between 37°28´N and 49°25´E. It is an excellent natural aquatic ecosystems that enhances a widely heterogeneous floristic composition and wildlife refuges. The wetland is covering 1% of bird wintering immigrant community of middle-east region (Jafari, 2009). Anzali wetland similar to other coastal ecosystems is likely to be affected by anthropocentric activities. Hence, the contaminants deriving from industrial and urban effluents, agrochemicals and untreated wastewater can cause irretrievable adverse effects on this natural ecosystem (Fig. 1). Preparation of plant samples: Sampling was performed in October 2013. The water samples were removed from five stations and prepared by filtration and addition of 2% HNO3 subsequently. Also, twenty plant samples were collected from the same site. The body samples were washed with tap and distilled water to wipe out any adhering substances. The root and body of plants were homogenized and then dried in oven (60°C for 24 hrs). Then, the dried samples were pulverized and obtained powder was sieved (0.15 mm) (Kalra, 1998). According to the digestion protocol, 0.5 gram samples of plant tissues put into the digestion tube and then 10 ml of nitric acid was added and stored overnight. Furthermore, samples were heated (120°C) for 4 hrs. Finally, the samples were transferred into 25 mL volumetric flasks and after addition of 3 mL diluted hydrochloric acid, filled with distilled water. Inductivity coupled Plasma-Optical Emission Spectrometry (ICP-OES) (PerkinElmer, USA) was used to measure the concentration of Cu, Cr, Pb and Cd. The detection limits for the analytical instrument were 2.5, 0.5, 0.5, 5 ppb for Cu, Cd, Cr and Pb, respectively. As concentrations were assayed by HG-FAAS (hydrogen generation flame atomic absorption spectroscopy) with detection limit of 1 ppb. The analysis procedure was confirmed by analyzing 2 blanks and 2 spiked specimens for each twenty plant samples considering all laboratory conditions being equal. Statistical Analysis: All statistical analysis were performed using Origin 8.0 software. Each metal was analyzed individually. Data was analyzed for normalization using Kolmogorov-smirnov test. ANOVA and Duncan tests were applied to parallel the mean values of metals in different sampling sites. Pearson correlation coefficients were used for estimating the correlation among metal concentrations in the water and plant tissues. Also, comparisons between metal concentrations in water and plant tissues were performed using t-test. When concentrations were not definite (e.g. below the detection limit), they were considered as half of the Element Sample (Mean ± SD, n=20) BCF TF Water (µg L-1) Shoot (µg g-1) Root (µg g-1) Cu 2.82 ± 0.14 4.35 ± 0.12 8.2 ± 0.17 2900 0.530 Cr 0.51 ± 0.03 0.471 ± 0.024 0.97 ± 0.05 1900 0.485 Pb 5.14 ± 1.43 6.04 ± 0.2 2.686 ± 0.134 1170 2.25 As 5.42 ± 0.65 1.037 ± 0.052 2.525 ± 0.127 460 0.411 Cd 0.50 ± 0.01 0.235 ± 0.012 0.344 ± 0.02 688 0.683 Bioconcentration factor (BCF) = Concentration in plant/Concentration in water Translocation factor (TF) = Concentration in shoot / Concentration in root Table 1. Heavy metals in Potamogeton crispus species tissues (µg-1, dry weight) and in Fresh water (μg lit-1). 209 Norouznia and Hamidian/ Phytoremediation efficiency of pondweed in removing heavy metals detection limit. Data were reported based on mean values of at least triplicates with standard deviation (SD). Result and Discussion Variation of heavy metals in water and plant tissues: No significant difference was observed in heavy metal concentrations of different sampling sites, in the same time (Table 1). Thus, it can be inferred that the metal ions were equally distributed throughout the wetland. However, the concentration of heavy metals in station 1 (Pirbazar river outfall to the wetland) and station 3 (Hendekhaleh river outfall to the wetland), which are located in the east and southeast of Anzali wetland, respectively, were little higher than those of other stations. Meanwhile, based on geographical interpretation of the sampling sites, it was obvious that heavy metal effluents outfall were participated in the contaminating of the water substratum (Pajevic et al., 2008; Pratas et al., 2012). The results showed a significant differences between the heavy metals concentration in plant tissues and water (P<0.05). The concentrations of heavy metals in water are shown in Figure 2. The heavy metals concentration in plant tissues are depicted in Figure 3. Based on the results, a range between 0.12 µg g-1 dw and 14.05 µg g-1 dw in plant and a range between 0.5 µg lit-1 and 6.91 µg lit-1 in water were recorded. This results show a significant differences in the different tissues of plant and water. Copper: Cu not only is a necessary nutrient for plants, but also a toxic element at extra concentration. It seems having high level of potency to accumulate in the lower tissues of plants (Kabata- Pendias and Pendias, 2001). The results revealed that the concentration of Cu in the water, P. crispus roots and shoots tissues were 2.82 µg lit-1, 8.20 µg g-1 dw and 4.01 µg g-1 dw, respectively. The highest registered BCF was calculated in P. crispus with the average of 2.9×103. Similarly, the TF were calculated in the range of 0.17 and 0.56. Moreover a significant (P<0.001) positive correlation between the Cu concentration in the plant and water was found. This result shows the positive aspects of P. crispus as a reliable accumulator species to omit Cu-oriented contaminants loaded to Anzali wetland. Chromium: Contents of Cr in plants have appealed warning notice not only due to its main function as an essential metal, but also because of its carcinogenic impacts. Chromium is slightly available to plants, thus it is accumulated eminently in roots. Chromium concentration vary between 0.4 and 3.2 mg kg-1 in rooted emergent plants species (Kabata-Pendias, 2011). Nonetheless, chromium concentrations more than 10 mg kg-1 have phytotoxic (Pais and Jones, 2000). In the current investigation, the mean values of Cr in the water and P. crispus roots and shoots were calculated to be 0.51 µg lit-1, 0.97 µg g-1 dw and 0.47 µg g-1 dw, respectively. The Cr values in the P. crispus tissues were measured in a descending order from root to shoot. Although, Cr concentration of P. crispus was shown positive correlation with its concentration in water. Low transport of Cr from root to aerial parts in Figure 2. Concentration of heavy metals in water (µg.litˉ1). Figure 3. Concentration of heavy metals in Potamogeton Crispus tissues (root (first column) and shoot (second column)) (µg g-1 dw). 210 Int. J. Aquat. Biol. (2014) 2(4): 206-214 Potamogeton sp. can be described due to being as a non-essential element for plant growth (Shewry and Peterson, 1974). On the other hand, the high accumulation of Cr in some wetland plants depends on the plant ability to decrease toxic Cr(VI) to the non-toxic Cr(III) form in roots and then trasmittancy of Cr(III) to the shoots (Lytle et al., 1998). The highest BCF was calculated with the average of 1.9×103. Also, the translocation factor values ranged between 0.47 and 0.6 in P. crispus. Based on TFs, Cr transfer ability of P. crispus from root to shoot, implying low transfer rate of Chromium due to incompetence in plant transfer system. According to the results a significant (<0.05) positive correlation among Cr concentrations in tissues and water was detected. Owing to the results, P. crispus seems to act as a proper alternative to reduce negative effects of pollution especially in the Cr-based contaminated aquatic environments. Arsenic: As is a prevalent metalloid that found in water, atmosphere, minerals and living organisms (Adriano, 2001). Concentration of As in unpolluted surface water differs from 1 to 10 μg lit-1. In freshwater, the As concentrations were reported in the range of 0.15–0.45 and 2 μg lit-1 (Sharma and Sohn, 2009; Smedley and Kinniburgh, 2002). Although, aquatic macrophyts have a significant effect on the As uptake, but the presence of As in plants is calculated more than 1 mg kg-1 (Sasmaz and Obek, 2009). García et al. (2010) noticed that As can hardly is removed via direct absorption by plants. In addition, Heung Lee (2013) mentioned that 0.5–1% of the total As input was accumulated in plant tissues. In the current investigation, the mean concentration of As in water, P. crispus roots and shoots tissues were measured as 5.4 µg lit-1, 2.52 µg g-1 dw and 1.03 µg g-1 dw, respectively. Besides, the assayed metal concentrations in plant were more considerable than those of water. The highest BCF was 0.46×103 and TFs values were varied between 0.28 and 0.42. TFs quantities pronounced that As were not transferred from roots to shoots efficiently. Moreover, a significant (P<0.001) positive correlation between As concentrations in the P. crispus and water was recognized. According to results, P. crispus does not appear to be a proper aquatic species to omit As. Lead: Pb is a major pollutant of the environment and absorbed by aerial tissues of plant passively and tightly is bound in root (Kabata- Pendias, 2011). The Pb transfer from roots to aerial parts is limited, whereas Raskin (1996) described, only 3% of the Pb in roots was transferred to the upper tissues. Pb is a gradually phytoavailable metal thus, hard to be phytofilter (Kabata-Pendias, 2011). In spite of above mentioned reports, in present study, the mean concentration of lead in water, P. crispus roots and shoots tissues were detected as 5.13 µg lit-1, 2.68 µg g-1 dw and 6.04 µg g-1 dw, respectively. The most concentration of Pb was found in the shoots of P. crispus (6.04 µg g-1 dw). This value were correlated with the Pb value of water. As a result, P. crispus was able to meet strong Pb concentrations overstepped that of the surrounding substratum. The highest BCFs of Pb in the P. crispus was 1.17×103. Also, the TFs were in the range of 1.77-2.24. Based on the results, the concentration of Pb in aerial parts is higher than that of roots, therefore Pb transferring from root to aerial part is feasible and furthermore, a well-developed root-rhizome form was not detected in plant species (Demirzen and Aksoy, 2004; Aksoy et al., 2005). A highly significant (P<0.001) positive correlation between Pb in P. crispus and water was found. Based on the findings, P. crispus seems to be an appropriate species to omit Pb from wastewater (Favas et al., 2012). Cadmium: Cadmium is noted as one of the main eco- toxic metals that reveals catastrophic effects on the plants and entire physiological processes of living organisms (Kabata-Pendias, 2011). Although Cd is a non-essential element for metabolic processes, it is easily absorbed by both root and leaf. There are proofs that acceptable values of Cd is absorbed passively by root and leaf (Kabata-Pendias, 2011). Hence, the linear relationship between Cd in plant and water medium was reported (Kabata-Pendias, 2011). Cd values in natural water are normally lower 211 Norouznia and Hamidian/ Phytoremediation efficiency of pondweed in removing heavy metals than 0.001 μg mL-1 and can be reached up to 1.9 µg g-1 in stems and leaves of aquatic plants (Friberg et al., 1986; Kabata-Pendias, 2004). The growth of plants in higher concentrations of Cd usually is stopped (Wang and Zhou, 2005). According to the results, the mean values of Cd in the water and P. crispus roots and shoots were recorded as 0.50 µg lit-1, 0.34 µg g-1 dw and 0.23 µg g-1 dw, respectively. The highest Cd concentrations in the roots of P. crispus (0.34 µg g-1 dw) indicates that this species is capable to accumulate a high Cd concentration. Therefore, the highest BCFs were estimated 0.68×103 for the root of P. crispus, indicating its moderate ability to accumulate Cd. Similarly, the TF recorded between 0.56 and 1.52. TFs reveal that Cd was not transferred from roots to leaves efficiently but was absorbed greater than Cu, Cr and As. A significant (P<0.001) positive correlation between Cd concentration and plant and water was observed, therefore P. crispus seems to be an appropriate species to remove Cd-derived pollutants from surrounding water effectively. Quantification of phytoremediation possibility: Evaluation of phytoremediation efficiency is determined using Bioconcentration Factor (BCFs) and Translocation Factor (TFs) (Pratas et al., 2012; Ali et al., 2013). Both factors (BCFs and TFs) would help trial to establish a greater degree of accuracy on phytoremediation efficiency. The both factor values are presented in Table 1. Bioconcentration factor: BCFs demonstrate the capability of plant to uptake heavy metals from surrounding medium, indicating that plants are able to accumulate heavy metals and therefore more appropriate for phytoremediation (Pratas et al., 2012). The bio-concentration factor is defined as metal concentration in dry mass in relation to its concentration in external substratum (Favas and Pratas, 2012). It is calculated as follows (Zhuang et al., 2007). Bioconcentration Factor (BCF) = 𝐶 ℎ𝑎𝑟𝑣𝑒𝑠𝑡𝑒𝑑 𝑡𝑖𝑠𝑠𝑢𝑒𝑠 𝐶 𝑤𝑎𝑡𝑒𝑟(𝑠𝑢𝑏𝑠𝑡𝑟𝑎𝑡𝑢𝑚) Where C harvested tissues is the concentration of the metal in the plant harvested tissue and C water(substratum) is the concentration of the same metal in water (substratum). A BCF value of > 1000 indicates a considerable hyper-accumulation potency of plant (Boonyapookana et al., 2002). In present study, the highest BCF value was calculated in P. crispus for Copper (2.9×103), showing that P. crispus exhibits acceptable efficiency to decline Cu in water. Similarly, the highest Chromium BCFs with the average of 1.9×103 was reported in this plant species. Also, the highest BCFs for Pb, Cd and As were as 1.17×103, 0.68×103 and 0.46×103, respectively. Consequently, P. crispus appears to be a noticeable accumulator in Anzali wetland for all mentioned metals except for As. Translocation Factor: Translocation factor explains the capacity of plant in transporting the concentrated metals from root to aerial parts (Ali et al., 2013). This factor is defined as the metal concentration in plant shoot in relation to its concentration in plant root (Pratas et al., 2012). It is calculated as follows (Padmavathiamma and Li, 2007): Translocation Factor (TF) = 𝐶 𝑠ℎ𝑜𝑜𝑡 𝐶 𝑟𝑜𝑜𝑡 Where C shoot is the concentration of metal in plant shoots and C root is its concentration in plant roots. The TF with higher valuse of 1, imply the high potency of plant metal transport systems (Zhao, 2002). Also, Kabata-Pendias and Pendias (2000) reported that the translocation factor in the range of 0.01-1 indicates the moderate bioavailiblity and accumulation of metals in aerial tissues of plants. In the present study, the mean values of TF in the P. crispus were calculated lower than 1 for Cu, As, Cr due to dysfunction in metal transmittancy operations (Sasmaz et al., 2008). The mean TFs in the plant evaluated higher than1 for Pb and relatively Cd. The highest TFs refer to P. crispus (2.24) for Pb, due to high transmittancy of Pb from roots to leaves actively. The mean TFs were presented in a descending order as Pb, Cd, Cu, Cr and As. This results confirmed that above-mentioned heavy metals were not transferred from roots to aerial parts efficaciously except for Pb and partly Cd. General conception: Many works have been 212 Int. J. Aquat. Biol. (2014) 2(4): 206-214 conducted to assess the bioaccumulation of heavy metals using aquatic macrophyts (Ye et al., 1997; Robinson et al., 2003; Pajevic et al., 2008; Carranza- Alvarez et al., 2008; Sasmaz et al., 2008; Alonso- Castro et al., 2009; Bonanno and Giudice, 2010; Pratas et al., 2012; Favas et al., 2012; Hamidian et al., 2014). Also, Rai (2008) has been emphasized to introduce P. crispus as proper metal accumulator. Ali et al. (1999) noticed the prominence of P. crispus to phytoremediate Cu, Cr, Pb and Zn. Demirzen and Aksoy (2004) investigated remediation potential of Potamogeton sp. to accumulate Cd, Pb, Cr, Ni, Zn and Cu in wetlands. Furthermore, Fritioff and Greger (2006) recognized P. crispus ability to participate on the heavy metals phytoremediation process. Moreover, Pajevic et al. (2008) pronounced that Potamogeton sp. can be used as reliable accumulators for heavy metals (Fe, Mn and Cd) pollution. Numerous factors may influence on the concentration-dependent variation of heavy metals in the plant species such as total concentration of metals in aquatic environment, the wide scope of metal species, various metal mechanisms and movability, and also plant–water interface. In addition, several physiochemical factors and physiological features such as water depth, water overflow, natural attributes of heavy metals, water pH, organic compound volume and biological characteristic of each plant species are used to determine whether a particular heavy metal is likely to be accumulated (Caranz-Alvarez et al., 2008). Conclusion Macrophyts are the key elements of wetland ecosystems. They not only uptake heavy metals, but also exhibit inherent strength to clean up surrounding water by absorbing of increasing pollutants particularly heavy metals (Jenssen et al., 1993). In addition, the highlighted functions of wetland macrophyts on phytoremediation and bioaccumulation assist to achieve a profound binary role (Rai et al., 2008). Eventually, this study showed a concentration-bound accumulation of Cu, Cr, Pb, As and Cd in P. crispus tissues. Namely, the metals concentrations in plant species was increased in a linear model along with increase in the water. The inspected positive correlation between metal concentrations in P. crispus and water, revealed its valuable role to remove metallic pollutants. The root of P. crispus is more adapted to concentrate metals than aerial parts, due to confined metal transporting system of plant. By returning to the objectives, it is now possible to state that P. crispus is an important qualified representative to meet the phytoremediation needs in the polluted substratum of Anzali wetland by Cu, Cr, Cd, Pb and As, respectively. Acknowledgment The authors are sincerely grateful to the anonymous reviewers for their profitable comments on the manuscript. References Adriano D. (2001). Trace elements in terrestrial environments. Biogeochemistry, bioavailability and risks of metals. New York, Springer, 45: 365-373. Aksoy A., Demirezen D., Duman F. (2005). Bioaccumulation, detection and analyses of heavy metal pollution in Sultan Marsh and its environment. Water Air and Soil Pollution, 164: 241-255. Ali H., Khan E., Sajad M.A. (2013). Phytoremediation of heavy metals - Concepts and applications. Chemosphere, 91: 869-881. Ali M.B., Tripathi R.D., Rai U.N., Pal A., Singh S.P. (1999). Physico-chemical characteristics and pollution level of Lake Nainital (U.P., India): role of macrophytes and phytoplankton in biomonitoring and phytoremediation of toxic metal ions. Chemosphere, 39(12): 2171-2182. Alonso-Castro A.J., Carranza-Álvarez C., Alfaro-De la Torre A., Chávez-Guerrero A., Ramo´n L., Garcıa- De la Cruz F. (2009). Removal and accumulation of cadmium and lead by Typha latifolia exposed to single and mixed metal solutions. Archives of Environmental Contamination and Toxicology, 57: 688-696. Bonanno G., Lo Giudice R. (2010). Heavy metal bioaccumulation by the organs of Phragmites australis (common reed) and their potential use as 213 Norouznia and Hamidian/ Phytoremediation efficiency of pondweed in removing heavy metals contamination indicators. Ecological Indicators, 10: 639-645. Boonyapookana B., Upatham E.S., Kruatrachue M., Pokethitiyook P., Singhakaew S. (2002). Phytoaccumulation and phytotoxicity of cadmium and chromium in Duckweed Wolffia globosa. International Journal of Phytoremediation, 4: 87-100. Carranza-Alvarez C., Alonso-Castro A.J., Alfaro-De La Torre M.C., Garcıa-De La Cruz R.F. (2008). Accumulation and distribution of heavy metals in Scirpus americanus and Typha latifolia from an artificial lagoon in San Luis Potos, Mexico. Water Air Soil Pollution, 188: 297-309. Cempel M., Nikel G. (2006). Nickel: a review of its sources and environmental toxicology. The Polish Journal of Environmental Studies, 15: 375-382. Chaney R.L. (1983). Plant uptake of inorganic waste constituents. In: Parr, J.F.E.A. (Ed.), Land Treatment of Hazardous Wastes. Noyes Data Corp. Park Ridge, NJ. 3: 50-76. Dabonne S., Koffi B., Kouadio E., Koffi A., Due E., Kouame L. (2010). Traditional utensils: Potential sources of poisoning by heavy metals. British Journal of Pharmacology and Toxicology, 1: 90-92. Demirezen D., Aksoy A. (2004). Accumulation of heavy metals in Typha angustifolia and Potamogeton pectinatus living in Sultan Marsh (Kayseri, Turkey). Chemosphere, 56: 685-693. Favas P.J.C., Pratas J., Prasad M.N.V. (2012). Accumulation of arsenic by aquatic plants in large- scale field conditions: Opportunities for phytoremediation and bioindication. Science of the Total Environment, 433: 390-397. Favas P.J.C., Pratas J. (2012). Uranium in soils, waters and plants of the abandoned uranium mine (Central Portugal). In: 12th International Multidisciplinary Scientific Geo Conference (SGEM 2012). Conference Proceedings 5:17-23, Albena, Bulgaria. Technology Ltd. 5: 1023-1028. Favas P.J.C., Pratas J. (2013). Uptake of uranium by native aquatic plants: potential for bioindication and phytoremediation. Published by EDP Sciences, E3S Web of conferences in Portugal 1, 13007: 674-677. Friberg L., Nordberg G.R., Vouk V.B. (1986). Handbook on the toxicology of metals. 2nd ed. NewYork, NY: Elsevier. 362 p. Fritioff A., Greger M. (2003). Aquatic and terrestrial plant species with potential to remove heavy metals from storm water. International Journal of Phytoremediation, 5: 211-224. Fritioff A., Greger M. (2006). Uptake and distribution of Zn, Cu, Cd, and Pb in an aquatic plant: Potamogeton natans. Chemosphere, 63: 220-227. García J., Rousseau D.P.L., Morató J., Lesage E., Matamoros V., Bayona J.M. (2010). Contaminant removal processes in subsurface-flow constructed wetlands: a review. Critical Reviews in Environmental Science and Technology, 40: 561-661. Hamidian A.H., Atashgahi M., Khorasani N. (2014). Phytoremediation of heavy metals (Cd, Pb and V) in gas refinery wastewater using common reed (Phragmites australis). International Journal of Aquatic Biology, 2(1): 29-35. Heung Lee J. (2013). An overview of phytoremediation as a potentially promising technology for environmental pollution control. Biotechnology and Bioprocess Engineering, 18: 431-439. Jafari N. (2009). Ecological integrity of wetlands, their functions and sustainable use. Journal of Ecology and Natural Environment, 1: 45-54. Jenssen P., Maehlum T., Krogstad T. (1993). Potential use of constructed wetlands for wastewater treatment in northern environments. Water Science and Technology, 28: 149-157. Kabata-Pendias A. (2004). Trace elements in soils and plants. CRC Press, Washington, D.C. 637 p. Kabata-Pendias A. (2011). Trace elements in soils and plants. CRC Press, Washington, D.C. 534 p. Kabata-Pendias A., Pendias H. (2000). Trace elements in soils and plants. CRC Press, Washington, D.C. 534 p. Kabata-Pendias A., Pendias H. (1989). Trace elements in soil and plants. Boca Raton, FL, CRC.564 p. Kabata-Pendias A., Pendias H. (2001). Trace elements in soils and plants. CRC Press, Washington, D.C. 696 p. Kalra A.P. (1998). Handbook of references methods for plant analysis .Soil and plant analysis council, Inc. CRC Press. Boca Raton Boston. London. New York. Washington, D.C. 281 p. Khan S., Hesham A.E.-L., Qiao M., Rehman S., He J.-Z. (2010). Effects of Cd and Pb on soil microbial community structure and activities. Environmental Science and Pollution Research, 17: 288-296. Lytle C. M., Lytle F. W., Yang N., Quian J. H., Hansen D., Zayed A., Terry N. (1998). Reduction of Cr (VI) to Cr (III) by wetland plants: Potential for in situ heavy metal detoxification. Environmental Science and 214 Int. J. Aquat. Biol. (2014) 2(4): 206-214 Technology, 32: 3087-3093. Mukhopadhyay S., Maiti S.K. (2010). Phytoremediation of metal enriched mine waste: a review. Global Journal, 4: 365-377. Pais I., Jones J.B. (2000). The Handbook of Trace Elements. St. Luice Press, Florida. 387 p. Pajević S., Kevrešan Ž., Radulović S., Radnović D.,Vučković M., Matavulj M. (2003). The role of macrophytes in monitoring the impact of heavy metal effluents on the aquatic environment. Central European Journal of Occupational and Environmental Medicine, 9: 317-321. Padmavathiamma P.K., Li L.Y. (2007). Phytoremediation technology: hyperaccumulation metals in plants. Water, Air, and Soil Pollution, 184: 105-126. Perkin Elmer. (1994). Analytical methods for atomic absorption spectrometry. USA. Pratas J., Favas P J.C., Paulo C., Rodrigues N., Prasad M.N.V. (2012). Uranium accumulation by aquatic plants from uranium-contaminated water in central Portugal. International Journal of Phytoremediation, 14(3): 221-234. Rai P.K. (2008). Heavy metal pollution in aquatic ecosystems and its phytoremediation using wetland plants: An eco-sustainable approach, International Journal of Phytoremediation, 10(2): 133-160. Raskin I. (1996). Plant genetic engineering may help with environmental clean-up. Proceedings of the National Academy of Sciences of United State of America, 93(8): 3164-3166. Robinson B.H., Duwig C., Bolan N.S., Kannathasan M., Saravanan A. (2003). Uptake of arsenic by New Zealand watercress (Lepidium sativum). Science of the Total Environment, 301: 67-73. Sasmaz A., Obek E., Hasar H. (2008). The accumulation of heavy metals in Typha latifolia L. grown in a stream carrying secondary effluent. Ecological Engineering, 33: 278-284. Sasmaz A., Obek E. (2009). The accumulation of arsenic, uranium, and boron in Lemna gibba L. exposed to secondary effluents. Ecological Engineering, 35: 1564-7. Sharma VK., Sohn M. (2009) Aquatic arsenic: toxicity, speciation, transformations, and remediation. Environment International, 35: 743-59. Sheoran V., Sheoran A., Poonia P. (2011). Role of hyperaccumulators in phytoextraction of metals from contaminated mining sites: a review. Critical Reviews in Environmental Science and Technology, 41: 168- 214. Shewry P.R., Peterson P. J. (1974). The uptake and transport of chromium by Barley seedlings (Hordeum vulgare L.). Planta (Berl), 132: 209-214. Singh A., Prasad S.M. (2011). Reduction of heavy metal load in food chain: technology assessment. Reviews in Environmental Science and Biotechnology, 10: 199- 214. Smedley P.L., Kinniburgh D.G. (2002). A review of the source, behaviour and distribution of arsenic in natural waters. Applied Geochemistry, 17: 517-68. Sood A., Uniyal P.L., Prasanna R.S., Ahluwalia A. (2012). Phytoremediation potential of aquatic macrophyte, Azolla. Royal Swedish Academy of Sciences. AMBIO, 41: 122-137. Wang X. F., Zhou Q. X. (2005). Ecotoxicological effects of cadmium on three ornamental plants. Chemosphere, 60(1): 16-21. Ye Z.H., Baker A.J.M., Wong M.H., Willis A.J. (1997). Zinc, lead and cadmium tolerance, uptake and accumulation by Typha latifolia. New Phytologist, 136: 469-480. Zhao, F.J., Dunham, S.J., McGrath, S.P. (2002). Arsenic hyperaccumulation by different fern species. New Phytologist, 56: 27-31. Zhuang P., Yang Q., Wang H., Shu W. (2007). Phytoextraction of heavy metals by eight plant species in the field. Water, Air, and Soil Pollution, 184: 235- 242.