Craig Musante

Concurrent plant uptake of heavy metals and persistent organic pollutants from soil

The extent of anthropogenic environmental pollution—in the United States (Black, 1999; Glass, 1999), in the European Union (Chaudhry et al., 2002), and in the third world is well documented. For example, the Food and Agriculture Organization of the United Nations has estimated that at a minimum 10 t of unwanted pesticides are in ‘‘storage’’ in undeveloped countries, with at least 2 10 t in African countries (Chaudhry et al., 2002). The potential for negative impacts of these stocks on humans and the environment is of major concern. In situ remediation techniques, such as phytoremediation— the attenuation of pollution through the use of plants—which impose minimal environmental disturbance, offer economic, agronomic, and societal benefits to all countries. Up to the present time phytoremediation of soilborne heavy metals and of organic contaminants has been pursued as two distinct disciplines. This compartmentalized approach applies to fundamental studies of the mechanisms of action, as well as to the development of remediation technologies. Based on data from the authors’ laboratories over the preceding several years we propose that far more convergence as opposed to divergence in the underlying plant physiology and soil science impacts the soil/vegetation microcosm to attenuate both soil-borne heavy metals and organic pollutants. For example, our published research has established that zucchini (Cucurbita pepo L.) and spinach (Spinacia oleracea) bioaccumulate soil-bound persistent organic pollutants (POPs) (Mattina et al., 2002). Other published reports have shown that spinach bioaccumulates heavy metals from soil (Romer and Keller, 2001). The data which are presented here demonstrate that these two plants simultaneously bioconcentrate and translocate both categories of weathered, soil-bound pollutant. Such simultaneous uptake and translocation of heavy metal and organic pollutants, if confirmed and optimized, could have enormous implications for plant/soil interaction mechanisms, and impact on practical remediation approaches, and ultimately on risk to human health.

Toxicity of silver and copper to Cucurbita pepo: Differential effects of nano and bulk‐size particles

The phytotoxicity of bulk and nanoparticle Cu and Ag was directly compared. NP Ag reduced biomass and transpiration by 66–84% when compared with bulk Ag. The Ag ion concentration was 4.4–10‐times greater in NP than bulk particle solutions. The Cu ion concentration was 1.4–4.4‐times greater in bulk than NP amended solutions. Humic acid (50 mg/L) decreased the ion content of bulk Cu solution by 38–42% but increased ion Cu content of NP solutions by 1.4–2.9 times. Bulk and NP Cu were highly phytotoxic; growth and transpiration were reduced by 60–70% relative to untreated controls. NP Cu phytotoxicity was unaffected by solution type, but humic acid (50 mg/L) completely alleviated phytotoxicity caused by bulk Cu. The data demonstrate differential toxicity of Ag NP relative to bulk Ag. The finding that humic acid and solution chemistry differentially impact bulk and NP behavior highlights the importance of evaluating nanoparticles under environmentally relevant conditions.

Analysis of Silver Nanoparticles in Antimicrobial Products Using Surface-Enhanced Raman Spectroscopy (SERS)

Silver nanoparticles (AgNPs) are the most commonly used nanoparticles in consumer products. Concerns over human exposure to and risk from these particles have resulted in increased interest in novel strategies to detect AgNPs. This study investigated the feasibility of surface-enhanced Raman spectroscopy (SERS) as a method for the detection and quantification of AgNPs in antimicrobial products. By using ferbam (ferric dimethyl-dithiocarbamate) as an indicator molecule that binds strongly onto the nanoparticles, AgNPs detection and discrimination were achieved based on the signature SERS response of AgNPs-ferbam complexes. SERS response with ferbam was distinct for silver ions, silver chloride, silver bulk particles, and AgNPs. Two types of AgNPs with different coatings, citrate and polyvinylpirrolidone (PVP), both showed strong interactions with ferbam and induced strong SERS signals. SERS was effectively applicable for detecting Ag particles ranging from 20 to 200 nm, with the highest signal intensity in the 60-100 nm range. A linear relationship (R(2) = 0.9804) between Raman intensity and citrate-AgNPs concentrations (60 nm; 0-20 mg/L) indicates the potential for particle quantification. We also evaluated SERS detection of AgNPs in four commercially available antimicrobial products. Combined with ICP-MS and TEM data, the results indicated that the SERS response is primarily dependent on size, but also affected by AgNPs concentration. The findings demonstrate that SERS is a promising analytical platform for studying environmentally relevant levels of AgNPs in consumer products and related matrices.

Accumulation and phytotoxicity of engineered nanoparticles to Cucurbita pepo.

The effect of bulk and engineered nanoparticle (NP) Ag, Au, Cu, Si, and C at 250 and 750 mg/L on zucchini biomass, transpiration, and element content was determined. The pH of bulk and NP solutions prior to plant growth frequently differed. Nanoparticle Cu solution pH was significantly higher than bulk Cu, whereas for Ag and C, the NPs had significantly lower pH. Plants were unaffected by Au, regardless of particle size or concentration. NP Ag reduced plant biomass and transpiration by 49–91% compared to equivalent bulk Ag. NP Si at 750 mg/L reduced plant growth and transpiration by 30–51% relative to bulk Si. Bulk and NP Cu were phytotoxic but much of the effect was alleviated by humic acid. The shoot Ag and Cu content did not differ based on particle size or concentration. The accumulation of bulk Au was greater than the NP, but humic acid increased the accumulation of NP and bulk Au by 5.6-fold and 80%, respectively. The uptake of NP Si was 5.6–6.5-fold greater than observed with the bulk element. These findings show that the NPs may have unique phytotoxicity or accumulation patterns and that solution properties can significantly impact particle fate and effects.

Characterization of substances released from crumb rubber material used on artificial turf fields

Crumb rubber material (CRM) used as infill on artificial turf fields can be the source of a variety of substances released to the environment and to living organisms in the vicinity of the CRM. To assess potential risks of major volatilized and leached substances derived from CRM, methods were developed to identify organic compounds and elements, either in the vapor phase and/or the leachate from CRM. A qualitative method based on solid phase micro-extraction (SPME) coupled with gas chromatography/mass spectrometry (GC-MS) was developed to identify the major volatile and semi-volatile organic compounds out-gassing from CRM samples under defined laboratory conditions. Direct vapor phase injection into the GC-MS was applied for the quantitative analysis. Ten organic compounds were identified in the vapor phase by the SPME method. Volatile benzothiazole (BT) was detected at the highest level in all commercial CRM samples, in the range 8.2-69 ng g(-1) CRM. Other volatile PAHs and antioxidants were quantified in the vapor phase as well. A decrease of volatile compounds was noted in the headspace over CRM samples from 2-years-old fields when compared with the virgin CRM used at installation. An outdoor experiment under natural weathering conditions showed a significant reduction of out-gassing organic compounds from the CRM in the first 14 d; thereafter, values remained consistent up to 70 d of observation. Zinc was the most abundant element in the acidified leachate (220-13000 microg g(-1)), while leachable BT was detected at relatively low amounts.

Accumulation of zinc, copper, or cerium in carrot ( Daucus carota ) exposed to metal oxide nanoparticles and metal ions

The release of engineered nanoparticles (ENPs) into the environment has raised concerns about the potential risks to food safety and human health. There is a particular need to determine the extent of ENP uptake into plant foods. Belowground vegetables growing in direct contact with the growth substrate are likely to accumulate the highest concentration of ENPs. Carrot (Daucus carota) was grown in sand amended with ZnO, CuO, or CeO2 NPs or the same concentrations of Zn2+, Cu2+, or Ce4+. Treatment with ZnO or Zn2+ produced a concentration-dependent decrease in root and total biomass. Ionic Cu2+ and Ce4+ caused a greater reduction in shoot biomass as compared to the corresponding ENP treatments. Accumulation of Zn, Cu, or Ce in the taproot was restricted to the taproot periderm. Metal concentrations in the taproot periderm were higher for the ionic treatments than for the ENP treatments. Radial penetration of the metals into the taproot and subsequent translocation to shoots were also generally greater for plants receiving the ionic treatment than those receiving the ENP treatment. The distribution of the metals from the ENP treatments across the periderm, taproot, and shoots differed from that observed for the ionic treatments. Overall, the ENPs were no more toxic than the ionic treatments and showed reduced accumulation in the edible tissues of carrot. The results demonstrate that the understanding of ionic metal transport in plants may not accurately predict ENP transport and that an additional comparative study is needed for this and other crop plants.

Uptake and Accumulation of Bulk and Nanosized Cerium Oxide Particles and Ionic Cerium by Radish (Raphanus sativus L.)

The potential toxicity and accumulation of engineered nanomaterials (ENMs) in agricultural crops has become an area of great concern and intense investigation. Interestingly, although below-ground vegetables are most likely to accumulate the highest concentrations of ENMs, little work has been done investigating the potential uptake and accumulation of ENMs for this plant group. The overall objective of this study was to evaluate how different forms of cerium (bulk cerium oxide, cerium oxide nanoparticles, and the cerium ion) affected the growth of radish (Raphanus sativus L.) and accumulation of cerium in radish tissues. Ionic cerium (Ce(3+)) had a negative effect on radish growth at 10 mg CeCl3/L, whereas bulk cerium oxide (CeO2) enhanced plant biomass at the same concentration. Treatment with 10 mg/L cerium oxide nanoparticles (CeO2 NPs) had no significant effect on radish growth. Exposure to all forms of cerium resulted in the accumulation of this element in radish tissues, including the edible storage root. However, the accumulation patterns and their effect on plant growth and physiological processes varied with the characteristics of cerium. This study provides a critical frame of reference on the effects of CeO2 NPs versus their bulk and ionic counterparts on radish growth.

Two rice plasma membrane intrinsic proteins, OsPIP2;4 and OsPIP2;7, are involved in transport and providing tolerance to boron toxicity

Boron (B) toxicity is responsible for low cereal crop production in a number of regions worldwide. In this report, we characterized two rice genes, OsPIP2;4 and OsPIP2;7, for their involvement in B permeability and tolerance. Transcript analysis demonstrated that the expression of OsPIP2;4 and OsPIP2;7 were downregulated in shoots and strongly upregulated in rice roots by high B treatment. Expression of both OsPIP2;4 and OsPIP2;7 in yeast HD9 strain lacking Fps1, ACR3, and Ycf1 resulted in an increased B sensitivity. Furthermore, yeast HD9 strain expressing OsPIP2;4 and OsPIP2;7 accumulated significantly higher B as compared to empty vector control, which suggests their involvement in B transport. Overexpression of OsPIP2;4 and OsPIP2;7 in Arabidopsis imparted higher tolerance under B toxicity. Arabidopsis lines overexpressing OsPIP2;4 and OsPIP2;7 showed significantly higher biomass production and greater root length, however there was no difference in B accumulation in long term uptake assay. Short-term uptake assay using tracer B (10B) in shoots and roots demonstrated increased 10B accumulation in Arabidopsis lines expressing OsPIP2;4 and OsPIP2;7, compare to wild type control plants. Efflux assay of B in the roots showed that 10B was effluxed from the Arabidopsis transgenic plants overexpressing OsPIP2;4 or OsPIP2;7 during the initial 1-h of assay. These data indicate that OsPIP2;4 and OsPIP2;7 are involved in mediating B transport in rice and provide tolerance via efflux of excess B from roots and shoot tissues. These genes will be highly useful in developing B tolerant crops for enhanced yield in the areas affected by high B toxicity.

Enhanced Boron Tolerance in Plants Mediated by Bidirectional Transport Through Plasma Membrane Intrinsic Proteins

High boron (B) concentration is toxic to plants that limit plant productivity. Recent studies have shown the involvement of the members of major intrinsic protein (MIP) family in controlling B transport. Here, we have provided experimental evidences showing the bidirectional transport activity of rice OsPIP1;3 and OsPIP2;6. Boron transport ability of OsPIP1;3 and OsPIP2;6 were displayed in yeast HD9 mutant strain (∆fps1∆acr3∆ycf1) as a result of increased B sensitivity, influx and accumulation by OsPIP1;3, and rapid efflux activity by OsPIP2;6. RT-PCR analysis showed strong upregulation of OsPIP1;3 and OsPIP2;6 transcripts in roots by B toxicity. Transgenic Arabidopsis lines overexpressing OsPIP1;3 and OsPIP2;6 exhibited enhanced tolerance to B toxicity. Furthermore, B concentration was significantly increased after 2 and 3 hours of tracer boron (10B) treatment. Interestingly, a rapid efflux of 10B from the roots of the transgenic plants was observed within 1 h of 10B treatment. Boron tolerance in OsPIP1;3 and OsPIP2;6 lines was inhibited by aquaporin inhibitors, silver nitrate and sodium azide. Our data proved that OsPIP1;3 and OsPIP2;6 are indeed involved in both influx and efflux of boron transport. Manipulation of these PIPs could be highly useful in improving B tolerance in crops grown in high B containing soils.

Defense mechanisms and nutrient displacement in Arabidopsis thaliana upon exposure to CeO2 and In2O3 nanoparticles

Metal-based nanoparticles (NPs) can cause toxicity to terrestrial plants, however there is little understanding of plant defense mechanisms that may counteract nanotoxicity. In the present study, we investigated the defense mechanisms of Arabidopsis thaliana in response to 250 mg L−1 and 1000 mg L−1 cerium oxide (CeO2) and indium oxide (In2O3) NPs exposure. Excessive amounts of total reactive oxygen species (ROS) were measured upon exposure to both NPs, demonstrating clear oxidative stress in Arabidopsis. Analysis of ROS scavenger activity indicated that activities of superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), and peroxidase (POD) were significantly elevated upon exposure to CeO2 NPs, while these elevations were only evident for SOD and POD activities in the In2O3 NP treatments. In addition, the activities of glutathione S-transferase (GST) and glutathione reductase (GR) were increased by approximately 15% and 51% by 1000 mg L−1 CeO2 and In2O3 treatment, respectively. Furthermore, the activities of phenylanine ammonialyase (PAL) and polyphenol oxidase (PPO) were greatly induced in response to both types of NPs. Additionally, both NPs disrupted the uptake of elemental nutrients, as is evident from the significantly lower levels of Fe accumulation in Arabidopsis root tissues exposed to CeO2 and In2O3 NPs. These results were further supported by the differential regulation of three iron-regulating genes, including ferric chelate reductase (FRO), iron-regulated transporter (IRT) and ferritin (FER), at various time points. The findings provide useful mechanistic information for plant detoxification pathways following NP exposure.