Earlier studies confirmed that the damage to plant root system and the decrease in transpiration strength has caused by excess of lead. Lead causes disorder in the composition of both the lipid membrane and the protein fraction, enabling its permeation into cells. Same results were obtained by some other studies at the calculated lead concentration: root, shoot and leaf growth, fresh and dry biomass were critically reduced in Pisum Sativum , 23 Zeamays, 15 Paspalum distichum and Cynodondactylon , 24 in Lycopersicon esculentum, 25 Ipomoea aquatic , 26 Phaseolus vulgaris and Lens culinaris.
In seeds, the testa avoids entry of lead into the internal tissues until it is ruptured by the developing radicle. Once the testa is ruptured, lead is taken up very rapidly, with distinguished exceptions occurring in the meristematic regions of the radicle and hypocotyls.
In cotyledons, lead moves through the vascular tissues and tends to accumulate in discrete areas in the distal parts. Like different heavy metals, lead treatment also affects the activity of a wide range of enzymes of different metabolic pathways. Chloramphenicol acetyl tranferase CAT is oxidoreductase that decomposes H 2 O 2 to water and molecular oxygen, and it is one of the important enzymes involved in the removal of toxic peroxides. CAT activities in cuttings and seedlings significantly increased at lower lead concentrations, while at higher lead concentrations, it decreased.
Reduced CAT activity at higher concentration of lead might be attributed to inactivation of enzyme by ROS, decrease in synthesis of enzyme, or change in assembly of its subunits. The process of photosynthesis is unfavorably affected by lead toxicity. Plants exposed to lead ions show a decline in photosynthetic rate which results from partial chloroplast ultra-structure, restrained synthesis of chlorophyll, plastoquinone and carotenoids, obstructed electron transport, inhibited activities of Calvin cycle enzymes, as well as deficiency of CO 2 as a result of stomatal closure.
Lead treatment also changes the lipid composition of thylakoid membranes. It harms the photosynthetic apparatus due to its affinity for protein N-and S-ligands.
When plants are exposed to lead, even at micromolar levels, adverse effects on germination and growth can occur. At low concentrations, lead inhibits the growth of roots and aerial plant parts. Arias et al. Plant biomass can also be restricted by high doses of lead exposure. In conclusion, lead is non essential element for plant although, it accumulates in different parts of plant and negatively affects various physiological processes.
Such physiological processes include photosynthesis, respiration, mineral nutrition, membrane structure and properties and gene expression. Therefore, this is the responsibility of Government and various environmental agencies to controlling heavy metals pollution especially lead. The authors wish to acknowledge the contribution of Dr.
This is an open access article distributed under the terms of the, which permits unrestricted use, distribution, and build upon your work non-commercially. Withdrawal Guidlines. Publication Ethics. Withdrawal Policies Publication Ethics. Review Article Volume 3 Issue 4. Keywords : lead, plant, growth, biochemical parameters. Sources of lead Human exposure to lead and its compounds occurs mostly in lead related occupations with various sources like leaded gasoline, industrial processes such as smelting of lead and its combustion, pottery, boat building, lead based painting, lead containing pipes, battery recycling, grids, arm industry, pigments, printing of books, etc.
Adverse effects of lead The visual general symptoms of lead toxicity are fast inhibition of root growth, underdeveloped growth of the plant, blackening of root system and chlorosis.
Effect of lead on photosynthesis The process of photosynthesis is unfavorably affected by lead toxicity. Cadmium: Toxicity and tolerance in plants. J Environ Biol. Weast RC.
The physiology of metal toxicity in plants. Ann Rev Plant Physiol J. Cadmium toxicity in plants. Braz J Plant Physiol.
Studies on cadmium toxicity in plants. Environ Pollut J. Hall JL. Cellular mechanisms for heavy metal detoxification and tolerance. J Exp Bot. Van Asshe F, Clijsters H.
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Lead toxicity in plants. To gain deep insights into the roles of NRAMP genes transporter in heavy metals uptake and homeostasis in plants, a more systematic characterization of the different members of the NRAMP family is further required.
The key feature of the ZIP family is that these proteins can transport heavy metals from the extracellular space or from organelles lumen into the cytoplasm. In Arabidopsis , 15 genes viz. Rest of the ZIP family members has been studied for their membrane localization and heavy metals they transport into or outside of a specific organelle Milner et al. Apart from the abovementioned transporters, recently another transporter arsenate reductase ACR has been characterized in yeast Saccharomyces cerevisiae , a model system for As resistance.
It was reported that a 4. Later on, Landrieu et al. Similarly, Ellis et al. Recent studies on A. Chao et al. However, in anaerobic paddy fields, As mainly Arsenite uptake is regulated by transporters of Si, namely Lsi1 low silicon 1; influx transporter and Lsi2 low silicon 2; efflux transporters Ma et al. Apart from these transporters, there are some other transporters as well that transfer arsenate and arsenite. Likewise, a transporter from P.
Investigations on the basic mechanisms of heavy metal tolerance and adaptation are the area of great scientific interest and an intensive research. Various stressors induce an expression of a set of genes in plants Nakashima et al. At molecular level, the regulation of gene expression is very important for the biological processes, which determines the fate of plant development as well as tolerance to heavy metal stress.
Stressors trigger large number of genes and several proteins in order to link the signaling pathways that confer stress tolerance Umezawa et al. These genes are classified into two groups: the regulatory genes and the functional genes Tran et al. However, the genes of functional group encode metabolic compounds such as amines, alcohols, and sugars, which play a crucial role in heavy metal stress tolerance.
The TFs, which are reported to be master regulators, control an expression of gene clusters and usually members of multigene families.
Studies reveal that a single TF can control the expression of many target genes via specific binding of the TF to the cis-acting element in the promoters of its target genes Wray et al. Most of the TFs contain a DNA-binding domain that interacts with cis-regulatory elements in the promoters of its target genes and via a protein—protein interaction domain that helps in oligomerization of TFs with other regulators Wray et al. LeDuc et al.
Liang et al. In addition to this, there are several other subgroups of bHLH family viz. Later, several researchers proposed that AtbHLH38 or AtbHLH39 interacts with FIT and forms heterodimers and directly activates transcription factors for ferric chelate reductase and ferrous transporters, which are the two major genes regulating Fe uptake under deficient condition Varotto et al.
In Arabidopsis , IRT1 has been reported to be the most essential ferrous transporter. A recent study by Wu et al. Transcriptome analysis in A. Similar induction of TFs has been reported in A. Differential expression of ERF factors under Cd indicates toward their responses to various levels of Cd stress. A study by Nakashima and Yamaguchi-Shinozaki reported down-regulation of dehydration-responsive element-binding protein DREB transcription factor involved in cold and osmotic stress responsive genes in roots of A.
Therefore, acquiring a deep knowledge of the interrelated mechanisms, which regulate the expression of these genes, is a crucial issue in plant biology and necessary to generate genetically improved crop plants for extreme environments like heavy metal stress Umezawa et al. Summary of an involvement of TFs in conferring heavy metal and other abiotic stresses tolerance is given in Table 2.
Table 2. Summary of transcription factors TFs whose overexpression in plants confers heavy metal stress tolerance. MAPK cascade are activated in response by plants when exposed to heavy metal stresses. This cascade has its significance in activation of signal transduction pathway used in hormone synthesis Jonak et al.
This cascade involves three kinases viz. The finally formed phosphorylated MAPK cascade phosphorylates substrates in cells including transcription factors in nucleus. Therefore, MAPK regulates the transduction of information downstream. Jonak et al. All these pathways finally lead to regulation of transcription factors that in turn activate genes for activation of metal transporters, biosynthesis of chelating compounds, and other defensing compounds. Proteomics is a well-established technique in the post-genomic era Liu et al.
Proteomics deals with the study of large-scale expression of proteins in an organism encoded by its genome Anderson and Anderson, Proteomics not only serves as a powerful tool for describing complete protein changes in any organisms but it can also be used to compare variation in protein profiles at organ, tissue, cell and organelle levels under various stress conditions including heavy metal stress Ahsan et al.
Although genomic analysis has enhanced our understanding regarding plants' response to heavy metal toxicity, transcriptomic changes in the genome are not always reflected at protein level Gygi et al. For instance, putative Zn and Mg transporter protein MHX was more abundant in Arabidopsis even though its corresponding transcript level was not different Elbaz et al. This suggests that transcription of any gene is not a guaranty that gene would be translated into a functional protein.
This may occur due to the potential impact of post-transcriptional and translational modifications, protein folding, stability and localization, protein—protein interactions, which are considered important determinants of a protein function Dalcorso et al.
Therefore, depth analyses of proteomics offer a new platform for identifying target proteins, which take part in heavy metal detoxification, and in studying complex biological processes and interactions among the possible pathways that involve a network of proteins Ahsan et al. Furthermore, it is known that proteins directly take part in plant stress responses, and plant adaptations to heavy metal stress are always accompanied with deep proteomic changes.
Therefore, technique of proteomics can be exploited for deciphering the possible relationships between proteins abundance and plant stress adaptation. Studies have revealed that an abundance of defense proteins was increased for scavenging of ROS, and molecular chaperones play a role in re-establishing the conformation of a functional protein that contributes in helping heavy metal stressed plants to maintain the redox homeostasis Zhao et al.
Under heavy metal stress, modulations of various metabolic pathways occur such as photosynthesis, respiration, nitrogen metabolism, sulfur metabolism, etc. The cellular mechanism of stress sensing and further transduction of signals into the cell appear to be the first reactions in the plant cell against heavy metal.
Furthermore, an intracellular communication of stress signals plays a fundamental role in signal transduction pathways under stress, which ultimately activate defense-related genes and thus signaling cascades Hossain et al.
Therefore, to decipher an underlying molecular mechanism of alterations in the protein signature of a plant cell in order to withstand stress, a deep study on the cellular as well as organelle proteomics would be of great importance in developing heavy metal-tolerant crops. Alterations in protein profile under heavy metal stress, which could be utilized for developing heavy metal-tolerant plants, are given in Table 3. Table 3. Summary of heavy metal-induced changes in protein expressions and their potential uses in developing heavy metal tolerant plants.
In the following section, we will discuss about the roles of peptides in heavy metal tolerance. Phytochelatins PCs have been the best-characterized chelators in plant systems. It has been reported that PCS were activated under heavy metal exposure Rauser, ; Cobbett, , and similar PC synthase activity has been observed in several other crops Klapheck et al. Loeffler et al. Haag-Kerwer et al.
Due to the presence of thiol group, they have the capability of chelating metals and forming complexes Cobbett, , which are then sequestrated in the vacuole. From the preceding discussion, it is clear that chelation by PCs is not a simple process but involves a complex molecular mechanism, where firstly, the PCS gets activated by metal ion and biosynthesis of PCs takes place; secondly, formation of complexes and sequestration in vacuole; thirdly, more complexation with the sulfides or organic acids in the vacuole, and finally detoxified.
Besides detoxifying heavy metals, PCs also play a major role in metal ion homeostasis and thus regulating the metal ion availability in plant cells Guo et al. Like PCs, metallothioneins MTs are synthesized and activated under heavy metal toxicity.
They belong to a family of low molecular weight protein having cysteine-rich metal binding peptide. Due to the presence of mercaptides, they have the ability of binding metal ions. In addition, Zhou and Goldsbrough reported restoration in Cu tolerance ability of MT-deficient yeast strains, when provided with the Arabidopsis MTs. Similar to this, Zhigang et al. Moreover, comparative study of mutant and wild-type A.
In terms of transcript amount, expression of MT genes varies during different developmental stages of plant as well as under varying environmental condition Rauser, Beside, chelating metal ions MTs can also catalyze antioxidant protection mechanism as well as plasma membrane repair Hamer, Ferritins are other multimeric proteins that could accumulate iron atom Harrison and Arosio, However, animal ferritins have been reported to store other metals like Cu, Zn, Cd, etc.
These are synthesized in plants when there is excess Fe in the surroundings and thus represents first-line defense against Fe-induced oxidative stress Ravet et al. These are not only involved in storing or releasing Fe but also involved in scavenging free reactive iron Ravet et al.
In spite of five classical plant hormones, i. Furthermore, it is also expected that some more growth hormones are yet to be discovered in future. In laboratory as well as filed studies, two strategies have been used for plant hormone-mediated increase in stress tolerance as well as crop yield.
These strategies include exogenous application of plant hormones and genetic manipulation of their endogenous contents. Both approaches have given promising results for increasing crop yield and enhancing stress tolerance in a variety of crop species Vriet et al. Although SA and GAs both are cost effective and can easily be availed for their exogenous application in crop fields under stress conditions, high cost of synthetic BRs and the variability of the results have discouraged the use of exogenous BRs in agriculture and horticulture Khripach et al.
In this context, modulation of endogenous BRs levels by genetic engineering has emerged an efficient strategy for enhancing crop yield under normal as well as adverse growth conditions Divi and Krishna, Herein, we have summarized recent advances made in enhancing heavy metal tolerance as well as achieving high yield with desired agronomic traits by using salicylic acid SA , brassinosteroids BRs , and gibberellins GA. In recent years, SA has gained much scientific attention due to its function as an endogenous signaling molecule conveying local and systemic plant—pathogen defense responses.
Besides this, it has been reported that SA also plays a role in plant response against abiotic stresses such as heavy metal toxicities, chilling, drought, osmotic stress, and heat. Being well characterized and studied role of SA in pathogen resistance, an exogenous application of SA could also provide protection against several types of abiotic stresses such as heavy metals, high or low temperature, salinity, radiation, etc.
Since under stress condition, reduced plant growth could result from an altered hormonal status, and thus, an exogenous application of plant hormones like SA has been an attractive approach to attenuate heavy metal stress. Studies carried out so far demonstrated that SA treatment to plants evoke acclimatization effect, which causes an enhanced tolerance toward heavy metal stress primarily due to the adjustment of metabolic processes such as enhanced antioxidative capacity.
In one of the first works, it was demonstrated that SA may induce protective effects against Cu toxicity in tobacco and cucumber Strobel and Kuc, Later, an increasing numbers of studies have demonstrated SA-mediated amelioration of toxicities produced by various heavy metals.
Zhou et al. It indicates a role of ROS signaling in such an amelioration process. In maize plant, Cd declined the growth by inhibiting chlorophyll synthesis, ribulose 1,5-bisphosphate carboxylase and phosphoenolpyruvate carboxylase, and enhancing oxidative damage such as lipid peroxidation and electrolyte leakage, whereas SA pretreatment of seeds reversed these toxic effects Krantev et al.
In cucumber, an exogenous application of SA has also been reported to enhance Mn tolerance by modulating nutrients' statuses and antioxidant defense system Shi and Zhu, Similarly, in pea seedlings, Cd toxicity caused decline in growth due to an inhabited photosynthetic process and enhanced oxidative damage, whereas SA pretreatment alleviated damaging consequences of Cd on growth and photosynthesis Popova et al. Moreover, Guo et al.
Conversely, Metwally et al. For instance, SA has been shown to potentiate generation of ROS in photosynthetic tissue under abiotic stresses and thus causes tissue damage Borsani et al. Therefore, it can be concluded that the concentration of SA appears to be important in regulating stress responses.
The SA-mediated alterations in genes that are involved in mediating stress tolerance are listed in Table 4. Table 4. Summary of plant hormone-mediated alterations in genes and their relation with an increased heavy metal stress tolerance. It is known that SA also involves in the regulation of oxidative stress caused by various stress factors Yang et al.
An enhanced level of SA under heavy metal stress suggests a connection between the extent of plant tolerance to heavy metal, which is mediated by the SA signal and the redox balance Metwally et al.
Moreover, Cui et al. In spite of considerable progress in the understanding of SA signaling, molecular events, which are involved in the SA signaling in order to alleviate heavy metal stress, are still poorly known Figure 4.
Figure 4. Schematic representation of plant hormone-mediated alleviation of heavy metal toxicity in plants. Heavy metals' signals are perceived by receptors, and receptors transduce signals via cAMP, pH, etc.
ROS cause damage to macromolecules and thus create oxidative stress inside the cell. These factors, in turn, initiate expression of the nuclear genes encoding defense proteins, transcription factors TFs , heat shock proteins HSP , and metal transporter proteins MTs.
MTs protect electron transport chains against heavy metals by regulating their uptake. Defense proteins protect plant against ROS under heavy metal stress Numbers 1 and 2 designated to chloroplast and mitochondria show the sources of ROS in cell. Brassinosteroids are group of hormones having ability of regulating ion uptake in plant cells and very effectively reducing the heavy metal accumulation in plants.
BRs can also impart plant stress tolerance against variety of biotic and abiotic stresses such as heavy metal, salinity, drought, low and high temperatures, and pathogen attack Bajguz and Hayat, ; Hao et al. An increasing numbers of studies have shown that an exogenous application of BRs is widely used in order to improve crop yield as well as stress tolerance in various plant species Divi and Krishna, ; Peleg and Blumwald, ; Li et al.
However, it has been reported that Cd-induced toxicity can be lowered with BR. For instance, Janeckzo et al. Hayat et al. In Vigna radiata L. Wilczek, Al stress caused a reduction in length, fresh and dry mass of root and shoot; activity of carbonic anhydrase; water use efficiency; relative water content; chlorophyll content; and the rate of photosynthesis, whereas addition of BR reversed these toxic effects and protected the plants via elevated level of proline in an association with an antioxidant defense system which at least in part was responsible for the amelioration of Al stress Ali et al.
The Cr, a known toxic metal, reduced the growth performance of Raphanus sativus L. Micronutrients such as Cu and Ni are essential for growth and development, but in excess, they cause severe toxic effects. Cu, which has increasingly attained interest due to its use in fungicides, fertilizers, and pesticides, is also highly toxic to plants, but when seeds of B.
Similar protective responses of exogenous BR on B. Besides higher plants, BR has also been found to be effective in alleviating heavy metals such as Cu, Pb, and Cd toxicities in algae, Chlorella vulgaris , through the regulation of antioxidant defense system Bajguz, The BR-mediated alterations in the gene expressions and their roles in stress tolerance are listed in Table 2.
The gibberellins GAs are a large family of tetracyclic diterpenoid plant growth hormone associated with the plant growth and developmental processes Matsuoka, Of these, GA has been a focus of plant scientists Hisamatsu et al. Several studies revealed that GA alleviates various abiotic stresses including heavy metal toxicity. It is reported that an exogenous addition of GA reprograms the growth of soybean under stress conditions by enhancing the levels of daidzein and genistein contents, suggesting protective role of GA in mitigating adverse consequences of stressors Hamayun et al.
In wheat seedlings, Ni 50 mM has been shown to decline growth, chlorophyll content, and carbonic anhydrase activity by enhancing oxidative stress, whereas an addition of GA ameliorates Ni-induced toxic effects Siddiqui et al. Gangwar et al. It has been observed that Pb and Zn affect seed germination in Cicer arietinum cv.
Aziziye by altering hormonal balance, and an exogenous application of GA reverses the toxic effect of heavy metals Atici et al. Furthermore, Sharaf et al.
These studies clearly indicate that GA plays an important role in protecting plant metabolism against various stresses; however, this may occur via various routes suggesting complex GA signaling during plant acclimation against stresses. The GA-mediated alterations in genes and their relation with stress tolerance are summarized in Table 4.
Around 3. It is expected that world population will be about 10 billion by the middle of the twenty-first century, and we will witness serious food shortages Smith et al. Furthermore, the situation will likely to be severe due to increased anthropogenic activities that have resulted into unwanted changes in the environment such as soil, air, and water pollution with various factors including heavy metal.
These situations pollution and population are posing a continuously increasing burden on global crop productivity, and hence, there are demands for crop varieties that should be adaptive and resistant to various stresses. In contrast to biotic stress, which is under the control of monogenic trait, abiotic stress tolerance is a genetically complex process that involves many components of signaling pathways, multigenic in nature, and thus, comparatively more difficult to control and engineer Vinocur and Altman, Therefore, plant-engineering strategies for heavy metal tolerance depend on the expression of gene s whose product s are involved either in signaling and regulatory pathways or in the synthesis of functional and structural proteins and metabolites that confer heavy metal stress tolerance.
Recently, several efforts are being made to improve heavy metal stress tolerance capacity through genetic engineering with several achievements; however, the genetically complex mechanisms of heavy metal stress tolerance and transfer of technology to field conditions make it difficult. Furthermore, genetic engineering of heavy metal-responsive genes particularly TFs , metabolites, and proteins has shown surprising results but its full potential remains to be exploited.
The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. We are also very grateful to Department of Botany, University of Allahabad for providing necessary lab facilities and also to University Grant Commission for providing financial support to PP and RS as research fellow to carry out this work. Abul Kashem, M.
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Soil adjacent to the industrial area contains the highest concentration of heavy metals. Among heavy metals, lead and cadmium are not essential elements for plants and are generally of low availability in soils,7,6 investigated the cadmium and lead uptake grown in three different textured soils and found Cd and Pb concentration in plant parts were highly correlated with Cd and Pb application, respectively.
Similarly, the essentiality of chromium for plants has not been demonstrated,8 whereas the importance of nickel has been documented by a few scientists.
There are no studies about the pollution of alzinc ursine second in Africa continental after South Africa of production the zinc metal, so far. Therefore, this study assesses the pollution effects of heavy metals on agricultural soils and plant alimentary at Ghazaouet. The objectives of the present study were i to characterise the fate and dispersal of Pb, Zn, Ni, Cu, Cd, Mn, Cr, Fe and As in soils around a former mining area, ii to measure Pb, Zn, Ni, Cu, Cd, Mn, Cr, Fe and As levels in the three plant alimentary artichoke, grape and pepper and iii determination the rate risk of health man et animals.
In summary, we address an initial strategy to waste dumpsite risk in this site that also takes into account the presence of significant concentrations of heavy metals. Soils and plants were sampled in the surrounding of the dumpsite around m2.
Sampling was collected out between June and July Shoots of several plant species were collected, as well as representative soil samples from the soil directly adjacent to the sampled plants cm, topsoil layer , obtaining a total of 3 soil samples and 3 plants samples.
Three regions were identified:. For better preparation against contamination during sampling, soil were collected with plastic spatulas and stored in polypropylene boxes. After collection, pebbles and twigs were removed. All soil samples were taken by mixing six sub-samples from three sites of each plot at cm depth. Each soil sample was air-dried and sieved to The methods of analysis used are presents in Table 1, vegetable and soil samples were taken randomly across the field during summer June , near from potential contamination Sources industrial plants, busy roads, residential areas, etc.
Plant samples collected from the field were washed under running tap water to remove adhered soils. The absorption wavelengths were The roots and sheets of the different plants were analyzed separately for heavy metal content. The interrelationship between these properties determines the capabilities and limitations of the soil for plant growth. Thus the soil acts as a reservoir for plant nutrients. Not all the nutrients are present in plant available form. Some are components of rock minerals or organic compounds that must be simplified before they can be utilized by the plant.
If all the nutrients are present in adequate amounts, the plant should exhibit good growth and vigor. Table 2. In the present study, it was found The pH of the soils ranged between moderately acidic 5. The amount of organic matter and total nitrogen is median, Soil reaction or pH is a simple and direct measure of the overall chemical condition of soils. Similarly it commonly recognized that at pH 6. The pH of soil in the study area ranges from 7. This pH value also favors the widest range of soil organisms and their activities.
Organic matter is needed for the chemical well being of the plant and soil fertility status because it is the source of nearly all the nitrogen and most of the phosphorus in some soils. This moderate amount of organic matter in present study is supported by the fact that pH will be higher where organic matter is well decomposed and incorporated into the surface mineral horizon.
The ratio between the nitrogen and organic matter is, thus, also rather constant14 states that organic matter contents in soil cannot be increased without simultaneously increasing its nitrogen contents and vice versa. The other macronutrients essential for plant growth, potassium, calcium and magnesium were present in adequate amount, Among them, calsium was present in slightly higher amount i. Heavy metals are chemical elements common to all types of soils, and their abundance ranks between percentage iron only and parts per million.
Hence heavy metals are synonyms to pollution and toxicity. Soil contamination by heavy metals is increasing nowadays,17 in the present study, the heavy metals Pb, Zn, Ni, Cu, Cd, Mn, Cr, Fe and As concentration with the exception of iron, decreased with the increase in distance from the road, i. The highest levels of Zn, Cd, Mn and Fe were found in the composite soil from adjacent of the waste dumpsite the alzinc industrial area indicating the waste as the source of soil pollution and contamination Table 3.
Table 3. Concentration of heavy metals in soils and plants collected from waste dumpsite of alzinc ursine. The high metal concentrations of the soil could have negative effects on microbial activities,20 provoking a low organic matter mineralization during the plant growth.
The apparent diminishing of heavy metals concentration away from the waste dumpsite of alzinc almost certainly confirms the waste as the potential source of soil contamination and their concentrations in plants. Samples of plant collected from the immediate environment of the waste were grossly contaminated with Zn, Mn and Cd.
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