Peter M. Kopittke

Trace metal phytotoxicity in solution culture: a review

Solution culture has been used extensively to determine the phytotoxic effects of trace metals. A review of the literature from 1975 to 2009 was carried out to evaluate the effects of As(V), Cd(II), Co(II), Cu(II), Hg(II), Mn(II), Ni(II), Pb(II), and Zn(II) on plants grown in solution. A total of 119 studies was selected using criteria that allowed a valid comparison of the results; reported toxic concentrations varied by five orders of magnitude. Across a range of plant species and experimental conditions, the phytotoxicity of the trace metals followed the trend (from most to least toxic): Pb approximately Hg >Cu >Cd approximately As >Co approximately Ni approximately Zn >Mn, with median toxic concentrations of (muM): 0.30 Pb, 0.47 Hg, 2.0 Cu, 5.0 Cd, 9.0 As, 17 Co, 19 Ni, 25 Zn, and 46 Mn. For phytotoxicity studies in solution culture, we suggest (i) plants should be grown in a dilute solution which mimics the soil solution, or that, at a minimum, contains Ca and B, (ii) solution pH should be monitored and reported (as should the concentrations of the trace metal of interest), (iii) assessment should be made of the influence of pH on solution composition and ion speciation, and (iv) both the period of exposure to the trace metal and the plant variable measured should be appropriate. Observing these criteria will potentially lead to reliable data on the relationship between growth depression and the concentration of the toxic metal in solution.

Fate of ZnO Nanoparticles in Soils and Cowpea (Vigna unguiculata)

The increasing use of zinc oxide nanoparticles (ZnO-NPs) in various commercial products is prompting detailed investigation regarding the fate of these materials in the environment. There is, however, a lack of information comparing the transformation of ZnO-NPs with soluble Zn(2+) in both soils and plants. Synchrotron-based techniques were used to examine the uptake and transformation of Zn in various tissues of cowpea ( Vigna unguiculata (L.) Walp.) exposed to ZnO-NPs or ZnCl2 following growth in either solution or soil culture. In solution culture, soluble Zn (ZnCl2) was more toxic than the ZnO-NPs, although there was substantial accumulation of ZnO-NPs on the root surface. When grown in soil, however, there was no significant difference in plant growth and accumulation or speciation of Zn between soluble Zn and ZnO-NP treatments, indicating that the added ZnO-NPs underwent rapid dissolution following their entry into the soil. This was confirmed by an incubation experiment with two soils, in which ZnO-NPs could not be detected after incubation for 1 h. The speciation of Zn was similar in shoot tissues for both soluble Zn and ZnO-NPs treatments and no upward translocation of ZnO-NPs from roots to shoots was observed in either solution or soil culture. Under the current experimental conditions, the similarity in uptake and toxicity of Zn from ZnO-NPs and soluble Zn in soils indicates that the ZnO-NPs used in this study did not constitute nanospecific risks.

In Situ Distribution and Speciation of Toxic Copper, Nickel, and Zinc in Hydrated Roots of Cowpea

The phytotoxicity of trace metals is of global concern due to contamination of the landscape by human activities. Using synchrotron-based x-ray fluorescence microscopy and x-ray absorption spectroscopy, the distribution and speciation of copper (Cu), nickel (Ni), and zinc (Zn) was examined in situ using hydrated roots of cowpea (Vigna unguiculata) exposed to 1.5 μm Cu, 5 μm Ni, or 40 μm Zn for 1 to 24 h. After 24 h of exposure, most Cu was bound to polygalacturonic acid of the rhizodermis and outer cortex, suggesting that binding of Cu to walls of cells in the rhizodermis possibly contributes to the toxic effects of Cu. When exposed to Zn, cortical concentrations remained comparatively low with much of the Zn accumulating in the meristematic region and moving into the stele; approximately 60% to 85% of the total Zn stored as Zn phytate within 3 h of exposure. While Ni concentrations were high in both the cortex and meristem, concentrations in the stele were comparatively low. To our knowledge, this is the first report of the in situ distribution and speciation of Cu, Ni, and Zn in hydrated (and fresh) plant tissues, providing valuable information on the potential mechanisms by which they are toxic.

Nanotechnology: A New Opportunity in Plant Sciences

The agronomic application of nanotechnology in plants (phytonanotechnology) has the potential to alter conventional plant production systems, allowing for the controlled release of agrochemicals (e.g., fertilizers, pesticides, and herbicides) and target-specific delivery of biomolecules (e.g., nucleotides, proteins, and activators). An improved understanding of the interactions between nanoparticles (NPs) and plant responses, including their uptake, localization, and activity, could revolutionize crop production through increased disease resistance, nutrient utilization, and crop yield. Herewith, we review potential applications of phytonanotechnology and the key processes involved in the delivery of NPs to plants. To ensure both the safe use and social acceptance of phytonanotechnology, the adverse effects, including the risks associated with the transfer of NPs through the food chain, are discussed.

Plasma Membrane Surface Potential: Dual Effects upon Ion Uptake and Toxicity

Electrical properties of plasma membranes (PMs), partially controlled by the ionic composition of the exposure medium, play significant roles in the distribution of ions at the exterior surface of PMs and in the transport of ions across PMs. The effects of coexisting cations (commonly Al3+, Ca2+, Mg2+, H+, and Na+) on the uptake and toxicity of these and other ions (such as Cu2+, Zn2+, Ni2+, Cd2+, and H2AsO4−) to plants were studied in terms of the electrical properties of PMs. Increased concentrations of cations or decreased pH in rooting media, whether in solution culture or in soils, reduced the negativity of the electrical potential at the PM exterior surface (ψ0o). This reduction decreased the activities of metal cations at the PM surface and increased the activities of anions such as H2AsO4−. Furthermore, the reduced ψ0o negativity increased the surface-to-surface transmembrane potential difference, thus increasing the electrical driving force for cation uptake and decreasing the driving force for anion uptake across PMs. Analysis of measured uptake and toxicity of ions using electrostatic models provides evidence that uptake and toxicity are functions of the dual effects of ψ0o (i.e. altered PM surface ion activity and surface-to-surface transmembrane potential difference gradient). This study provides novel insights into the mechanisms of plant-ion interactions and extends current theory to evaluate ion bioavailability and toxicity, indicating its potential utility in risk assessment of metal(loid)s in natural waters and soils.

Fast X-Ray Fluorescence Microtomography of Hydrated Biological Samples

Metals and metalloids play a key role in plant and other biological systems as some of them are essential to living organisms and all can be toxic at high concentrations. It is therefore important to understand how they are accumulated, complexed and transported within plants. In situ imaging of metal distribution at physiological relevant concentrations in highly hydrated biological systems is technically challenging. In the case of roots, this is mainly due to the possibility of artifacts arising during sample preparation such as cross sectioning. Synchrotron x-ray fluorescence microtomography has been used to obtain virtual cross sections of elemental distributions. However, traditionally this technique requires long data acquisition times. This has prohibited its application to highly hydrated biological samples which suffer both radiation damage and dehydration during extended analysis. However, recent advances in fast detectors coupled with powerful data acquisition approaches and suitable sample preparation methods can circumvent this problem. We demonstrate the heightened potential of this technique by imaging the distribution of nickel and zinc in hydrated plant roots. Although 3D tomography was still impeded by radiation damage, we successfully collected 2D tomograms of hydrated plant roots exposed to environmentally relevant metal concentrations for short periods of time. To our knowledge, this is the first published example of the possibilities offered by a new generation of fast fluorescence detectors to investigate metal and metalloid distribution in radiation-sensitive, biological samples.

Alleviation of Cu and Pb Rhizotoxicities in Cowpea (Vigna unguiculata) as Related to Ion Activities at Root-Cell Plasma Membrane Surface

Cations, such as Ca and Mg, are generally thought to alleviate toxicities of trace metals through site-specific competition (as incorporated in the biotic ligand model, BLM). Short-term experiments were conducted with cowpea (Vigna unguiculata L. Walp.) seedlings in simple nutrient solutions to examine the alleviation of Cu and Pb toxicities by Al, Ca, H, Mg, and Na. For Cu, the cations depolarized the plasma membrane (PM) and reduced the negativity of ψ(0)(o) (electrical potential at the outer surface of the PM) and thereby decreased {Cu(2+)}(0)(o) (activity of Cu(2+) at the outer surface of the PM). For Pb, root elongation was generally better correlated to the activity of Pb(2+) in the bulk solution than to {Pb(2+)}(0)(o). However, we propose that the addition of cations resulted in a decrease in {Pb(2+)}(0)(o) but a simultaneous increase in the rate of Pb uptake (due to an increase in the negativity of E(m,surf), the difference in potential between the inner and outer surfaces of the PM) thus offsetting the decrease in {Pb(2+)}(0)(o). In addition, Ca was found to alleviate Pb toxicity through a specific effect. Although our data do not preclude site-specific competition (as incorporated in the BLM), we suggest that electrostatic effects have an important role.

Laterally resolved speciation of arsenic in roots of wheat and rice using fluorescence-XANES imaging

• Accumulation of arsenic (As) within plant tissues represents a human health risk, but there remains much to learn regarding the speciation of As within plants. • We developed synchrotron-based fluorescence-X-ray absorption near-edge spectroscopy (fluorescence-XANES) imaging in hydrated and fresh plant tissues to provide laterally resolved data on the in situ speciation of As in roots of wheat (Triticum aestivum) and rice (Oryza sativa) exposed to 2 μM As(V) or As(III). • When exposed to As(V), the As was rapidly reduced to As(III) within the root, with As(V) calculated to be present only in the rhizodermis. However, no uncomplexed As(III) was detected in any root tissues, because of the efficient formation of the As(III)-thiol complex - this As species was calculated to account for all of the As in the cortex and stele. The observation that uncomplexed As(III) was below the detection limit in all root tissues explains why the transport of As to the shoots is low, given that uncomplexed As(III) is the major As species transported within the xylem and phloem. • Using fluorescence-XANES imaging, we have provided in situ data showing the accumulation and transformation of As within hydrated and fresh root tissues.

Toxic effects of Cu2+ on growth, nutrition, root morphology, and distribution of Cu in roots of Sabi grass

Sabi grass (Urochloa mosambicensis (Hack.) Dandy) (a C4 species of Poaceae) is commonly used to revegetate disturbed sites in low-rainfall environments, but comparatively little is known regarding copper (Cu) toxicity in this species. A dilute nutrient solution culture experiment was conducted for 10 d to examine the effects of elevated Cu(2+) activities ({Cu(2+)}) on the growth of Sabi grass. Growth was inhibited by high Cu in solution, with a 50% reduction in the relative fresh mass occurring at 1.0 microM {Cu(2+)} for the roots and 1.2 microM {Cu(2+)} for the shoots. In solutions containing 1.2-1.9 microM {Cu(2+)}, many of the roots ruptured due to the tearing and separation of the rhizodermis and outer cortex from the underlying tissues. Transmission electron microscopy revealed that Cu-rich deposits were found to accumulate predominantly within vacuoles. Due to limited translocation of Cu from the roots to the shoots, phytotoxicity is likely to be more of a problem in remediation of Cu-toxic sites than is Cu toxicity of fauna consuming the above-ground biomass.

Metal-induced cell rupture in elongating roots is associated with metal ion binding strengths

Low concentrations of Al, Cu and La rapidly decrease root elongation and cause transverse ruptures to the rhizodermis and outer cortex, but it is not known if other trace metals have similar effects. Six trace metals, Ga, Gd, Hg, In, Ru, and Sc, decreased cowpea root growth and caused ruptures similar to those caused by Al, Cu and La. Calculated speciation of the metals showed that only Gd was almost exclusively present as the trivalent ion (Gd3+), but the other test solutions were dominated by Ga(OH)2+, HgCl20, either In3+, In(OH)2+, In(OH)2+, In(OH)30, or InCl2+, and Sc3+ or ScOH2+ (no thermodynamic constants were available for Ru). The results from this and other studies suggest that the ability of these trace metals (plus Al, Cu, and La) to cause ruptures is related to the strength to which the trace metals bind to the cell wall. Therefore, it is proposed that the toxic effects of trace metals results from (1) the strength of binding (either ionically or covalently), and (2) other toxic effects of the metals not dependent on cell wall interactions.