Heavy Metals in Different Environmental Matrices

The problem of such heavy metals and metalloids as cadmium, mercury, nickel, lead or chromium accumulation in various ecosystems is not new and is increasing significantly because of the improvement of the level of life, population expansion and heavy industry development. High natural levels of the pollutants in environmental matrices are the result of agricultural and semi-industrial activities, energy supply, mining or waste disposal. Heavy metal ions are ubiquitous in water (ground or surface water), plants, animals, soil as sediments and sewage sludge. They are present in all environmental matrices but in diverse concentration levels, depending on the metal, the matrix and the distance from pollutant sources, which - considering their toxicity and easy translocation in living organisms - causes the real threats.

Heavy metal ions are able to modify many vital mechanisms, to inhibit enzymatic processes by binding to bio-molecules and in consequence to destroy organisms (animals, plants). Recognition of the heavy metal concentration level in environmental matrices is of particular importance due to their significant effects on vegetation followed by human health risk (entering and accumulating in the food chain - fruits, vegetables, crops, plant tissue, etc). Uptake of heavy metal ions by plants and/or animals is one of the main causes of their presence in food. In environmental matrices there are several heavy metals essential in animal nutrition (As, Co, Cr, Cu, Fe, Mn, Mo, Ni, Se, Sn, V and Zn) or in plant growth (B, Cu, Fe, Mo, Mn, Ni, Zn), although a few of them (Cd, Hg and Pb) are not recognized as essential in living organisms. Owing to the acute toxicity of the above-listed elements, they and also Cu, Ni and Zn are included in the US Environmental Protection Agency's (EPA) list as priority pollutants and also in the lists of the American Agency for Toxic Substances and Disease Registry (ATSDR).

The presence of metals and metalloids in matrices influences almost the whole ecosystem, which necessitates environmental monitoring followed by application of proper, continually improved and modernized remediation technology. The heavy metal removal efficiency depends on the method, concentration level and kind of pollutants in the matrix and the presence of other pollutants (interaction, change of redox potential and pH). In contrast to organic compounds (possible to be decomposed or mineralized with final CO2 and H2 O products), heavy metals are not decomposable to a simpler form and will never be completely removed from polluted matrices but can be immobilized (technical methods) or taken up by plants (phytoremediation) only.

The uptake of heavy metals by plants and their accumulation in tissue has been of great interest recently, not just because of their negative impact on ecosystems and human health. The improved resistance of some plant species to toxic elements has been found as a promising tool in phytoremediation techniques, with efficient extraction of heavy metals from contaminated soil, sewage and water followed by their accumulation in plant tissue (Kumar et al. 1995). Plants have developed numerous mechanisms to compensate for toxicity of heavy metals including avoidance, detoxification and non-specific resistance (Zenk 1996). However, when their concentration in the environment exceeds the species-specific phytotoxicity threshold, the excessive accumulation of heavy metals within cells leads to numerous toxic effects, such as rapid inhibition of overall growth, leaf chlorosis and premature leaf senescence due to the reduction of photosynthesis rate and transpiration, reduction of root elongation and inhibition of seed germination (Borowiak et al. 2011; G^secka et al. 2011; Obrouchova et al. 1998). Accumulation of heavy metals in leaves via roots and/or stomata causes a reduction in the size of guard cells as the effect of enhanced biosynthesis of abscisic acid and further decline in the size of the photosyn-thetic area. Heavy metals interfere with active sites of many enzymes (binding directly with thiol groups of the active centre or with carboxyl groups stabilizing the secondary structure of the enzyme protein), including phosphatase, ATPase and enzymatic antioxidants (catalase, glutathi-one reductase, ascorbate peroxidase, superoxide dismutase), which results in inhibition of enzyme activity (van Assche and Clijsters 1990; Verma and Dubey 2003). Heavy metals also inhibit the biosynthesis of chlorophyll, cause structural changes in chloroplasts depressing the photosynthesis rate, bind to nucleic acids inducing the aggregation and condensation of chromatin and inhibit replication and transcription, reduce cellular respiration, and lead to macro- and microelement deficits (iron, potassium, magnesium and chlorine) (Borowiak et al. 2011; Seregin et al. 2004; Stroinski and Koziowska 1997).

Rhizofiltration by symbiotic fungi and bacteria associated with the root system and the exudation of chelating agents into the rhizosphere, i.e. hydrocarbons, organic acids, amino acids and glycoproteins, are the first mechanisms of heavy metal avoidance (Marchner et al. 1996). In the root apoplast, heavy metal transport up to aerial plant organs is restricted due to their binding with carboxyl groups of galacturonic and glucuronic acids present in the structure of the cell wall. Simultaneously, root endodermis prevents radial transport of heavy metals into vascular tissue and further to upper parts of the plant. Among heavy metals, lead is a hardly mobile element, and 70-95% of its content in plants is associated within roots (Piechalak et al. 2002). Heavy metals transported through vascular tissue to upper plant parts accumulate in leaves. However, atmospheric deposition of particulate matter - taken up through stomata or adsorbed by epicuticular wax - is a significant source of toxic elements in photosyn-thetic tissue (Wozny 1995). At low concentrations, heavy metals are immobilized within the apoplastic space, bound to cell wall polysaccha-rides. However, higher concentrations perpetuate their transport via the cell membrane to the cell interior (passive - simple or facilitated diffusion; active - with trans-membrane proteins, and endo-cytosis) (Samardakiewicz and Wozny 2000). Plant resistance to heavy metals is mostly dependent on the efficiency of the detoxification mechanisms in intercellular liquid, among which chelation and further subcellular compartmenta-tion in vacuoles is the most abundant one (Clemens 2001; Piechalak et al. 2002). Phytochelatins (PCs), a family of cysteine-rich peptides (glutathione-derived oligomers) are metal-chelating molecules that are crucial in plant tolerance versus metal ions (Cobbett and Goldsbrough 2002; Yruela 2005). PC synthesis via glutathione transpeptidation is catalyzed by phytochelatin synthase induced by heavy metal ions bound to the thiol group of glutathione (Chen et al. 1997) ; Metal ions complexed with PCs are further transported to the vacuole, where after being released by hydrogen ions they react with organic acids, polyphenols and glucosides, and as a result form less or non-toxic complexes (Cobbett 2000; Zenk 1996). However, Landberg and Greger (2004) did not detect PCs in willow - a plant considered in application of phytoextraction - during long-term treatment of different Salix genotypes with a wide range of heavy metals. The mechanism of willow tolerance may be determined by other factors, perhaps metal complexation with low-molecular carboxylic organic acids in the cytoplasm (Clemens 2001; G^secka et al. 2011).

Furthermore, plants employ non-specific resistance mechanisms such as enhanced amino acid biosynthesis, mainly proline which chelates heavy metal ions and serves as a free radical scavenger, thus preventing cell membrane lipids from being oxidized (Alia et al. 2001; Mehta and Gaur 1999). Heavy metals induce chemical changes in cell wall composition, i.e. enhanced formation of callose and suberin, accumulation of reactive oxygen species, induction of the anti-oxidant system in the apoplast and cell interior, as well as biosynthesis of signaling compounds, phytohormones and other regulatory compounds in a controlled response to heavy metal stress (Malecka et al. 2001; Zenk 1996).

5 Biological Methods of Environment Decontamination: Significance and Improvement of Phytoremediation

There is no doubt that increasing consumption to some extent is a sign of the times indicating an increasing and improved level of life, which in consequence results in deterioration of the environment by pollution (rubbish, refuse and debris). The large and growing number and range of pollutants and xenobiotics present in the soil, sediments, sewage and surface water makes it necessary to develop new, more efficient methods of environment cleaning. Elaborated methods are divided into different groups depending on the pollutant specificity, site and cost of the method used. The economic dimension is especially significant when reclamation is performed on a large polluted area. The new environment cleaning methods, including application of nanomaterials to environment protection (mesoporous and siliceous materials usually used for water but also for polluted soil), are still too expensive for the industrial scale.

Most frequently, technical (chemical and physical, such as excavation and burial of soil, reverse osmosis, ion exchange, microfiltration or fixation/inactivation) and biological (bioremedia-tion and phytoremediation) methods are applied (Juwarkar et al. 2010; Jabeen et al. 2009). Technical methods cause a lot of irreversible changes in properties of polluted matrices (e.g. soil destruction) and the final product is often completely deprived of the valuable elements necessary for plants' development. For this reason, biological methods are of prime concern, since they are almost non-invasive to the environment. In these methods plants and microorganisms active in natural processes and present in soil or water ecosystems are usually used.

Phytoremediation ("phyto" in Latin means plant and "remedium" means restore) is defined as a biological method using selected species/ varieties of plants for the effective accumulation of inorganic pollutants or degradation of organic ones (Vamerali et al. 2010). An important restriction is the long time required to achieve satisfactory effects of the process, but it also has a lot of benefits, which means that this effective method may support technical methods or may be used as an individual method of deteriorated environment cleaning. Additionally, the methods are gradually replacing technical methods, especially in the case of areas temporarily excluded from practical use or production.

The development of phytoremediation in the last 20 years is focused on the selection of new plants and/or modification of the process conditions to increase its effectiveness (Lone et al. 2008) . The weak point - at the beginning - was the lack of information on complex mechanisms of plant-soil interactions and their influence on transport (translocation) of pollutants to the above-ground plant tissues. Recently, a wide range of studies on the mechanisms of processes in physiology, biochemistry and botany, and with the support of genetic engineering, with emphasis on rapid development of phytoremediation, have been performed. In recent years, phytore-mediation as an interdisciplinary technology has been examined in terms of its practical effectiveness in cleaning post-industrial areas, characterized by high contents of various pollutants (organic and inorganic). The most spectacular progress is observed in phytoextraction and phy-tostabilization studies. The uptake, chelation, translocation or volatilization of heavy metals needs to be developed, since the expected results are very important in elucidation of pollutant transport pathways in the ecosystem, estimation of the risk of phytoremediation for natural processes in the environment, and also identification and introduction of genes in transgenic plants.

Changes in phytoremediation are observed mostly in terms of plant features. Hyperac-cumulators able to accumulate above average amounts of pollutants are of prime concern in the area (Memon and Schröder 2009). As the next step, a combination of phytoaccumulation with high and fast efficiency in plant biomass production is important to improve the process rationalization (simpler harvest of plants), and to increase the accumulation with the perspective of wood utilization in energy production. Results presented recently underline physiological and biochemical aspects of phytoremediation with the influence of environmental factors (plant-fungus or plant-soil interactions) with possibilities of more efficient implementation of this process. An interesting prospect in this scope may be the exploration of combined techniques (e.g. selected plants with phytoremediation abilities with such technical or semi-technical methods as electroki-netic remediation under constant voltage across the soil).

Heavy metals belong to an ecologically significant and toxicologically unique class of toxicants, because they are spread everywhere, particularly in industrialized areas. Effective phytoextraction requires the regular (not significantly inhibited) growth of plants in polluted areas followed by the activation of defence mechanisms. Plants with phytoremediation abilities have to meet several fundamental criteria: high effectiveness of phytoaccumulation/ phytodegradation, high biomass increase, well-developed root system, high resistance to pollutants, easy adaptation to different environmental conditions and simple environmental requirements (Vangronsveld et al. 2009). Plants used in phytoremediation should exhibit no or very small risk of metals' transport to higher trophic levels (reduced possibility to contaminate the food chain).

To date the following topics have been elucidated and are well known: removal of pollutants by aerial plant organs, transport of metals through the plasmalemma, and detoxification of pollutants in the cell (chaperones, phytochelatins, met-allothioneins, low-molecular-weight organic acids - LMWOAs) (Pal and Rai 2010). The above possibilities allow plants to defend themselves against viruses, microbes and fungi with effective phytoaccumulation/phytodegradation of pollutants present in the environment (Rascio and Navari-Izzo 2011).

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