anzhen Li

Toxicity of zinc oxide nanoparticles in the earthworm, Eisenia fetida and subcellular fractionation of Zn

The extensive use of nanoparticles (NPs) in a variety of applications has raised great concerns about their environmental fate and biological effects. This study examined the impact of dissolved organic matter (DOM) and salts on ZnO NP dispersion/solubility and toxicity to the earthworm Eisenia fetida. To be able to better evaluate the toxicity of NPs, exposure in agar and on filter paper was proposed for enabling a comparison of the importance of different uptake routes. A dose-related increase in mortality was observed in earthworms exposed in agar with almost 100% mortality after 96 h exposure to the highest concentration (1000 mg ZnO/kg agar). Scanning electron microscopy (SEM) showed that the addition of salts enhanced the aggregation of ZnO NPs in agar and consequently affected the dissolution behavior and biological availability of the particles. On filter paper, mortality was the highest at the lowest exposure concentration (50 mg ZnO/L) and seemed to decrease with increasing exposure levels. TEM images of ZnO showed that the solubility and morphology of NPs were changed dramatically upon the addition of humic acids (HA). The subcellular distribution pattern of Zn in earthworms after 96 h exposure in agar and on filter paper showed that the Zn taken up via dietary ZnO particles (from agar) was mainly found in organelles and the cytosol while the Zn accumulated as soluble Zn from filter paper was mainly distributed in cell membranes and tissues. Antioxidant enzymatic activities (SOD, CAT, and GSH-px) were investigated in the worms surviving the toxicity tests. A slight increase of SOD activities was observed at the lowest exposure dose of ZnO (50mg/kg), followed by a decrease at 100mg/kg in the agar cubes. Activities of both CAT and GSH-Px enzymes were not significantly influenced in the worms exposed to agar, although a slight decrease at 500 and 1000 mg ZnO/kg agar was observed. A similar change trend of SOD activities was observed for the earthworms on filter paper, but a significant decrease began at a higher ZnO NP concentration of 500 mg ZnO/L. The use of soil extracts instead of deionized water (DW) to simulate a realistic exposure system significantly reduced the toxicity of the ZnO NPs on filter paper, which increases the predictive power of filter paper toxicity tests for the environmental risk assessment of NPs.

Metabolic responses in gills of Manila clam Ruditapes philippinarum exposed to copper using NMR-based metabolomics

Copper is an important heavy metal contaminant with high ecological risk in the Bohai Sea. In this study, the metabolic responses in the bioindicator, Manila clam (Ruditapes philippinarum), to the environmentally relevant copper exposures were characterized using NMR-based metabolomics. The significant metabolic changes corresponding to copper exposures were related to osmolytes, intermediates of the Krebs cycle and amino acids, such as the increase in homarine, branched chain amino acids and decrease in succinate, alanine and dimethylamine in the copper-exposed clam gills during 96 h exposure period. Overall, Cu may lead to the disturbances in osmotic regulation and energy metabolism in clams during 96 h experimental period. These results demonstrate that NMR-based metabolomics is applicable for the discovery of metabolic biomarkers which could be used to elucidate the toxicological mechanisms of marine heavy metal contaminants.

Benzo(a)pyrene-induced metabolic responses in Manila clam Ruditapes philippinarum by proton nuclear magnetic resonance ( 1 H NMR) based metabolomics

Benzo(a)pyrene is an important polycyclic aromatic hydrocarbon (PAH) which causes carcinogenic, teratogenic and mutagenic effects in various species and the level of contamination of this toxic agent in the marine environment is of great concern. In this study, metabolic responses induced by two doses (0.02 and 0.2μM) of BaP were characterized in the gill tissues of Manila clam Ruditapes philippinarum after exposure for 24, 48 and 96h. The high dose (0.2μM) of BaP induced the disturbances in energy metabolism and osmotic regulation based on the metabolic biomarkers such as succinate, alanine, glucose, glycogen, branched chain amino acids, betaine, taurine, homarine, and dimethylamine in clam gills after 24h of exposure. In addition, hormesis induced by BaP was found in clams exposed to both doses of BaP. Overall, our results demonstrated the applicability of metabolomics for the elucidation of toxicological effects of marine environmental contaminants in a selected bioindicator species such as the Manila clam.

Effect of cations on copper toxicity to wheat root: Implications for the biotic ligand model

The extent to which calcium, magnesium, sodium, potassium and hydrogen ions independently mitigate Cu rhizotoxicity to wheat (Triticumaestivum) in nutrient solutions was examined. Increasing activities of Ca2+ and Mg2+ but not Na+, K+ and H+ linearly increased the 2 d EC50 (as Cu2+ activity), supporting the concept that some cations can compete with Cu2+ for binding the active sites at the terrestrial organism-solution interface (i.e., the biotic ligand, BL). According to the biotic ligand model (BLM) concept, the conditional stability constants for the binding of Cu2+, Ca2+ and Mg2+ to the BL were derived from the toxicity data. They were 6.28, 2.43 and 3.34 for logK(CuBL), logK(CaBL) and logK(MgBL), respectively. It was calculated that on average 43.6% of BL sites need to be occupied by Cu2+ to induce 50% root growth inhibition. Using the estimated parameters, a BLM was successfully developed to predict Cu toxicity for wheat as a function of solution characteristics.

Cell Membrane Surface Potential (ψ0) Plays a Dominant Role in the Phytotoxicity of Copper and Arsenate

Negative charges at cell membrane surfaces (CMS) create a surface electrical potential (ψ0) that affects ion concentrations at the CMS and consequently affects the phytotoxicity of metallic cations and metalloid anions in different ways. The ζ potentials of root protoplasts of wheat (Triticum aestivum), as affected by the ionic environment of the solution, were measured and compared with the values of ψ0 calculated with a Gouy-Chapman-Stern model. The mechanisms for the effects of cations (H+, Ca2+, Mg2+, Na+, and K+) on the acute toxicity of Cu2+ and As(V) to wheat were studied in terms of ψ0. The order of effectiveness of the ions in reducing the negativity of ψ0 was H+ > Ca2+ ≈ Mg2+ > Na+ ≈ K+. The calculated values of ψ0 were proportional to the measured ζ potentials (r2 = 0.93). Increasing Ca2+ or Mg2+ activities in bulk-phase media resulted in decreased CMS activities of Cu2+ ({Cu2+}0) and increased CMS activities of As(V) ({As(V)}0). The 48-h EA50{Cu2+}b ({Cu2+} in bulk-phase media accounting for 50% inhibition of root elongation over 48 h) increased initially and then declined, whereas the 48-h EA50{As(V)}b decreased linearly. However, the intrinsic toxicity of Cu2+ (toxicity expressed in terms of {Cu2+}0) appeared to be enhanced as ψ0 became less negative and the intrinsic toxicity of As(V) appeared to be reduced. The ψ0 effects, rather than site-specific competitions among ions at the CMS (invoked by the biotic ligand model), may play the dominant role in the phytotoxicities of Cu2+ and As(V) to wheat.

Pathways of cadmium fluxes in the root of the halophyte Suaeda salsa

Halophyte plants offer a greater potential for phytoremediation research for reducing the levels of toxic metals from saline soils than salt sensitive plants. Using the scanning ion-selective electrode technique, we analyzed the pattern and rate of Cd(2+) fluxes at different regions of the root apex of Suaeda salsa. The Cd(2+) influx in the rhizosphere was greatest near the root tip (within 150μm of the tip). The results indicated that Cd(2+) influx into roots was significantly suppressed by the pre-treatment or in the presence of two kinds of Ca(2+) channel blockers; LaCl(3) and verapamil. The Cd(2+) influx was also reduced by N-ethylmaleimide, a thiol blocker. Cd content determination and labeling of Cd using fluorescent dye support our conclusion. The results of this study provide a more stable theoretical basis for the phytoremediation of Cd contamination in saline soils of coastal zones.

A metabolomic investigation on arsenic-induced toxicological effects in the clam Ruditapes philippinarum under different salinities

Arsenic is an important contaminant in the Bohai marine ecosystem due to the anthropogenic activities. In this work, we investigated the toxicological effects of arsenic in Ruditapes philippinarum under different seawater salinities using NMR-based metabolomics. Under normal salinity (31.1 ppt), arsenic decreased the levels of amino acids (glutamate, β-alanine, etc.), and increased the levels of betaine and fumarate. The metabolic biomarkers including decreased threonine, histidine, ATP and fumarate were found in the muscles of arsenic-treated clams under medium salinity (23.3 ppt). However, only elevated ATP and depleted succinate were detected in the arsenic-exposed clam samples under low salinity (15.6 ppt). These differential metabolic biomarkers indicated that arsenic could induce osmotic stress and disturbance in energy metabolism in clam under normal and medium salinities. However, arsenic caused only disturbance in energy metabolism in clam under low salinity. Overall, our results demonstrated that seawater salinity could influence the toxicological effects of arsenic.

Quantifying the adsorption and uptake of CuO nanoparticles by wheat root based on chemical extractions.

Extensive application of metal nanoparticles is attracting more attention because of their potential environmental risks. Many studies have focused on the uptake of metal nanoparticles (NPs) by plant, but the adsorption of nanoparticles on root surface is often mistakenly regarded as their uptake. This study optimized the methods to distinguish the adsorption and uptake of CuO-NPs on the wheat root by applying different metal competing ions (Na+, Mg2+, and La3+), surfactant (i.e., sodium dodecyl benzene sulfonate, SDBS), or complexing agents like NaOAc and Na4EDTA, as well as ultrasonic technique. The results indicated that some CuO-NPs is strongly adsorbed on the plant root surface, and part of them by mechanical adhesion. Competing ions could not desorb the CuO-NPs from the root surface, while NaOAc and Na4EDTA well dissolved the adsorbed CuO-NPs. In addition, the uptake and adsorption of CuO-NPs increased with increasing exposure concentrations of CuO-NPs in the range of 5-200 mg/L. The amount of CuO-NPs adsorption is always lower than that of their uptake.

Subcellular distribution of Cd and Pb in earthworm Eisenia fetida as affected by Ca2+ ions and Cd-Pb interaction.

A semi-static solution culture method was used to study the effects of Ca(2+) supply and interaction of Cd-Pb on the subcellular distribution of Cd and Pb in earthworm Eisenia fetida. The subcellular distribution of Cd and Pb was shown to be metal specific. About 80% of the Cd was distributed in the cytosol (fraction G), and only about 20% of total Cd was found in the tissue and cell membrane (fraction E) and the microsomes (fraction F). Nearly 50% of the Pb was rich in the tissue and cell membrane (fraction E). The supply of Ca(2+) ions significantly decreased Cd concentration in the cytosol (fraction G) and the whole tissue of the E. fetida. At the subcellular level, the addition of Pb(2+) ions significantly decreased the Cd percentage associated with fraction G from 83.7% to 58.4% and increased fraction E from 10.7% to 34.0%, respectively.

Evaluating mechanisms for plant-ion (Ca2+, Cu2+, Cd2+ or Ni2+) interactions and their effectiveness on rhizotoxicity

Electrical properties of plant cell membranes (CMs) provide a new avenue for exploring the mechanisms of plant-ion interactions and the biotic effects of ions. The modulating effect of Ca2+ on rhizotoxicity of metallic ions was studied to evaluate the mechanisms of plant-ion interaction in terms of the electrical potential at the CM exterior surface CMSe (ψ0o). Adding Ca2+ to root bathing media (BM) reduced the negativity of ψ0o at the CMSe. This reduction caused decreases in the activities of toxic metal ions at the CMSe and hence alleviated the toxicity (denoted as Mechanism I). Calcium is an essential element for growth and adding Ca2+ could also restore metal-displaced Ca2+ at the CMSe and alleviated Ca2+ deficiency (Mechanism II). The reduced surface negativity increased the surface-to-surface transmembrane potential difference (Em,surf), thus increasing the electrical driving force for transport of toxic metallic ions across the CM (Mechanism III). The Mechanism III would increase toxicity, but did not offset the alleviation by Mechanisms I and II. Regression analysis of relative root elongation in appropriate nonlinear equations incorporating the effects of above mechanisms provided evidence that under the current experimental conditions, Mechanism I is the most important mechanism for plant-ion interactions. In addition, the intracellular concentration of metals in root was a better predictor of toxicity than the free metal ion activity in the BM and could be predicted with an electrostatic uptake model developed previously. Based on the intracellular concentration and the EUM, the toxicity threshold (EC50, activities producing 50% growth inhibition) could be predicted generally within a factor of 2 of the observed values, indicting its potential utility in risk assessment of toxic metals.