Anti-oxidative Responses on Hepatic Tissue of Zebrafish (<Emphasis Type="Italic">Danio rerio</Emphasis>) in a Short Duration of Sub-lethal Concentrations of Cadmium Exposure

The aim of the present study was to identify whether the responses of oxidative stress in zebrafish liver are similar to those in mammalians upon low doses of Cd²⁺ exposure in short durations. Fish were exposed to 1.78 μM Cd²⁺ (treatment) and 0.0 μM Cd²⁺ (control) for 0, 1, 3, and 6 h. The reactive oxygen species (ROS) and lipid peroxidation (LPO) of hepatic tissues significantly increased after 3 and 6 h of Cd²⁺ exposure, respectively. Antioxidants glutathione peroxidase (gpx1a), superoxide dismutase (sod), and catalase (cat) were up regulated after 1–3 h, and metallothionein isoforms (smtB and mt2) increased after 3–6 h of Cd²⁺ exposure. The caspase-3 and p53 mRNA expressions significantly increased threefolds after 1 h of Cd²⁺ exposure. Results confirmed that oxidative stress in the hepatic tissue was induced by Cd²⁺ within 3 h. However, anti-oxidative functions immediately up regulated, causing cell apoptosis levels to decrease after 6 h of Cd²⁺ exposure.     

Effects of Copper Sulfate on Ion Balance and Growth in Tilapia Larvae (<Emphasis Type="Italic">Oreochromis mossambicus</Emphasis>)

Newly hatched tilapia larvae were exposed to sublethal concentrations of Cu²⁺ (0, 30, 50, and 100 g/L) and lethal concentrations of Cu²⁺ (200 and 400 g/L) for 24–96 h. The interaction of the exposure dose and time was related to the Cu²⁺ accumulation rate, which showed a higher accumulation rate with sublethal concentrations of Cu²⁺ within 24 h compared to the other treatments. Furthermore, Cu²⁺ contents in the whole body of larvae significantly increased following Cu²⁺ exposure times up to 96 h. Cu²⁺ in the medium produced a dose-response effect on Na⁺ and K⁺ contents in larvae after 96 h of exposure time. Changes in Ca²⁺ contents statistically significantly decreased and were shown to be dose-responsive for larval exposure times exceeding 72 h. Changes of Ca²⁺ contents were more sensitive than those of Na⁺ and K⁺ with Cu²⁺ treatment of early larvae. Notably Na⁺ and K⁺ contents showed significant increases of 17–23% in larvae exposed to low concentrations of Cu²⁺ (30–50 g/L) for 24–72 h as compared to control larvae. Cu²⁺ caused no significant effect on body Cl^– content or osmolality except at 100 g/L Cu²⁺ for 24 h in tilapia larvae as compared to the control. However, there was a restoration phenomenon in larvae exposed to 100 g/L Cu²⁺ for longer than 72 h. The water content of larvae exposed to Cu²⁺ for 96 h significantly decreased. The yolk absorption rate of tilapia larvae was significantly suppressed when they were exposed to Cu²⁺ medium containing 30, 50, 100, 200, or 400 Cu²⁺ g/L from 72 h post transfer. These results obviously show that larvae are sensitive to Cu²⁺ during early development.     

DNA Damage Induced by Copper on Erythrocytes of Gilthead Sea Bream <Emphasis Type="Italic">Sparus aurata</Emphasis> and Mollusk <Emphasis Type="Italic">Scapharca inaequivalvis</Emphasis>

Heavy metal pollution in rivers and its impact on aquatic ecosystems is a dynamic process. Fish are ideal indicators of heavy metal contamination in aquatic systems because they occupy different trophic levels and are of different sizes and ages. In particular, copper is an essential trace metal for living organisms and it is present in all natural waters and sediments. In this paper, we report data on the effect of copper on DNA erythrocytes from the teleost gilthead sea bream Sparus aurata and the bivalve mollusk Scapharca inaequivalvis. In particular, the effect of 0.1 ppm of Cu²⁺ on the nucleated erythrocytes was analyzed using the comet assay. This test is a promising tool for estimation of DNA damage at the single cell level. The data obtained show that the in vivo treatment with 0.1 ppm of copper increased the susceptibility of DNA to be damaged. Exposure to Cu²⁺ produces a more evident effect on Sparus aurata, as all three comet parameters significantly increased (tail length, tail intensity, and tail moment). The higher comet parameters measured in Scapharca inaequivalvis compared to Sparus aurata could be due to the difference in stability of the respective hemoglobins. The comet assay could represent a useful test to evaluate the biological consequences of environmental contamination by metals on marine organisms.     

Effects of Ca<Superscript>2+</Superscript> or Na<Superscript>+</Superscript> on Metallothionein Expression in Tilapia Larvae (<Emphasis Type="Italic">Oreochromis mossambicus</Emphasis>) Exposed to Cadmium or Copper

The objectives of this study were to try to determine the reasons of the external Ca²⁺ and Na⁺ enhancement of Cd²⁺ and Cu²⁺ resistance in fish. Tilapia larvae at 3 days posthatch were exposed to (A) 0 (control), 40 μg/L Cd²⁺, 40 μg/L Cd²⁺ + 2 mM Ca²⁺ (Cd/hyper-Ca), and 2 mM Ca²⁺ or (B) 0 (control), 75 μg/L Cu²⁺, 75 μg/L Cu²⁺ + 0.52 mM Na⁺ (Cu/hyper-Na), and 0.52 mM Na⁺. After 48 hours, results indicated that (1) Cd/hyper-Ca and Cu/hyper-Na treatments showed decreased growth inhibition induced by the metals; (2) metal accumulation in Cd/hyper-Ca–treated larvae was lower compared with those exposed only to Cd; and (3) metallothionein (MT) expression was significantly higher in Cu/hyper-Na–treated larvae than in the group treated with Cu only. Taking all of this into account, either supplementary Ca²⁺ or Na⁺ in ambient water may help fish to maintain Ca²⁺ or Na⁺ homeostasis, which could decrease metal accumulation and its detrimental effects. Consequently, the fish increase MT expression and retard the growth inhibition caused by metals.     

Antioxidant Modulation in Response to Metal-Induced Oxidative Stress in Algal Chloroplasts

To investigate adaptive responses to metal stress at the subcellular level, the oxidative balance in isolated chloroplasts was evaluated for the first time in the unicellular alga Gonyaulax polyedra exposed to the toxic metals Hg²⁺, Cd²⁺, Pb²⁺, and Cu²⁺. Different antioxidant responses were verified according to the metal and model of stress applied. Cells chronically exposed to metals exhibited high activity of the antioxidant enzymes superoxide dismutase and ascorbate peroxidase, high glutathione content, and decrease of peridinin levels, whereas no significant changes were detected for β-carotene levels. In contrast, cells subjected to acute metal stress displayed twice as much β-carotene but only a slight increase in superoxide dismutase and ascorbate peroxidase activities. The correlation of acute metal treatment and oxidative stress was inferred from the higher oxygen uptake and decreased reduced glutathione pool found in treated cells. In addition, increased oxidative damage to proteins and lipids occurred mainly in cells under acute stress. Pb²⁺ was the most damaging toxicant, causing protein oxidation and lipid peroxidation even at chronic treatment. These results indicate that heavy metals are able to induce oxidative stress in chloroplasts of G. polyedra, particularly under acute conditions. Nevertheless, the maintenance of a high antioxidant capacity within chloroplasts seems to be an important strategy during acclimation of G. polyedra to chronic metal stress. By acting at the subcellular site, where oxidative stress is triggered, induction of such chloroplast antioxidants might be crucial for cell survival during exposure to heavy metals. Received: 4 April 2000/Accepted: 9 July 2000     

Influence of divalent metal ions on E2-induced ER pathway in goldfish (Carassius auratus) hepatocytes

Metal ions existing in the environment could influence the estrogen pathway in aquatic animal, but the detailed mechanism is still delusive. We here showed that in male Carassius auratus hepatocytes, copper (Cu) or cadmium (Cd), did not directly induce vitellogenin (VTG) expression. Interestingly, co-exposure with Cd²⁺ (or Cu²⁺) and 17-β-estradiol (E2) greatly increased the VTG level, comparing with single treatment of E2. Meanwhile, Cd²⁺ or Cu²⁺ (but not E2) triggers HSP70 expression. But, mixture of Cd²⁺ or Cu²⁺ with E2 did not obviously raise HSP70 level. E2 also had no obvious effect on reactive oxygen species. Co-treatment of Cd²⁺ and E2 showed no obvious increase compared to single treatment with Cd²⁺. We further assume that Cd²⁺-involved oxidative stress generates misfolded proteins, resulting in the competition of HSP70 proteins from a heterocomplex (with estrogen receptor). Thus, dissociation of the heterocomplex actives the receptor-ligand binding activity and promotes the E2-induced VTG expression.     

Acrosome Reaction of Sperm in the Mud Crab Scylla serrata as a Sensitive Toxicity Test for Metal Exposures

In order to test the sensitivity of the sperm cell of the mud crab Scylla serrata to heavy metals, the toxic effects of Ag⁺, Cd²⁺, Cu²⁺, and Zn²⁺ on the acrosome reaction (AR) were studied by artificially inducing the AR of sperm exposed to heavy metals, counting the AR rates by light microscopy, and observing structural changes in sperm by transmission electron microscopy. The AR in S. serrata occurs at two stages. The first stage (ARI) is the eversion of the subacrosomal material. The second stage (ARII) is the ejection of the acrosomal filament. The results showed the EC₅₀ values of the AR based on (ARI + ARII)% for Ag⁺, Cd²⁺, Cu²⁺, and Zn²⁺ were 10.02, 2.14, 13.69, and 2.21 μg/L, and the EC₅₀ values based on ARII % of Ag⁺, Cd²⁺, Cu²⁺, and Zn²⁺ were 1.96, 0.20, 1.46, and 0.34 μg/L. The order of toxicity is Cd²⁺ > Zn²⁺ > Cu²⁺ > Ag⁺ based on the percentage of reacted sperm at the second stage. Sperm cells exposed to heavy metals showed an increased rate of swelling, shape irregularities, and the acrosomal filament of some sperm cells was, crooked, ruptured, and even dissolved. The AR of the sperm cell from S. serrata is more sensitive to the tested heavy metals compared to sea urchin sperm cell toxicity tests.     

Antioxidant responses of rice seedling to Ce⁴+ under hydroponic cultures

Since the 1980s, rare earth elements have been commonly used in China because of their enriched fertilizers. To understand the potential benefits or damages of Ce⁴⁺ on rice, the antioxidant responses (superoxide dismutase, ascorbate peroxidase, catalase activities, and ascorbate and glutathione contents) of rice seedling to Ce⁴⁺ under hydroponic cultures were investigated. The results showed that Ce⁴⁺ induced H₂O₂ and O₂⁻ production of rice seedling. The inhibition studies with diphenylene iodonium suggested that the key enzyme responsible for oxidative bursts was primarily NADPH oxidase. Ce⁴⁺ (0.02 mM) increased the antioxidant capacity of reduced ascorbate and glutathione and the levels of superoxide dismutase, ascorbate peroxidase, and catalase. However, antioxidant enzymes and antioxidant capacity of rice seedling were decreased by 0.2 mM Ce⁴⁺ treatment, indicating that higher content of Ce⁴⁺ damaged the mechanism of defense responses and emerged the peroxidation of membrane lipids. These results will help us to understand the mechanism of Ce⁴⁺ on rice and concern about its environmental impact in agricultures.     

Effects of [C2mim][OAc] (1-Ethyl-3-methyl-imidazolium acetate) on the Growth of Wheat Seedlings Under Cd2+ Stress

The joint effect on the growth of wheat seedlings under Cd²⁺ stress (0, 0.5 mmol L^?1) and [C₂mim][OAc] with a series of concentrations (0, 100, 200, 300, 400, 500 mg L^?1) was investigated. Physiological characteristics including superoxide dismutases, peroxidases, catalase and malondialdehyde were studied in hydroponic and soil cultures. Results indicated that, compared to samples treated under Cd²⁺ stress alone, joint treatment groups showed Cd²⁺ stress was mitigated by ionic liquid at appropriate concentrations (≤400 mg L^?1 in hydroponic treatments and ≤300 mg L^?1 in soil culture treatments). Use of inductively coupled plasma mass spectroscopy showed less Cd²⁺ in organelles in the joint treatment group. This phenomenon could be due to [C₂mim][OAc] forming metal complexes with Cd²⁺, thus reducing the amount of free Cd²⁺ available for absorption by the seedlings.     

The effect of heavy metals on nicotinamideN-methyltransferase activityin vitro relating to Parkinson’s disease

Objective The aims of this study were to determine the effects of heavy metals such as manganese on nicotinamideN-methyltransferase (EC 2.1.1.1) (NNMT) activity and to consider the possibility of involvement of NNMT activation in the pathogenesis of heavy metal induced Parkinson’s disease. Methods NNMT activity in supernatants separated from brain, liver and kidney homogenates of 5 elderly male Wistar rats by centrifugation were measured by high performance liquid chromatography system with fluorescence. NNMT activity under the conditon of 0.5 or 5.0 mM Mn²⁺, Fe²⁺, Cu²⁺ or Cd²⁺ was compared with control (no metal ion existence). Results NNMT activities in rat brain, liver and kidneys were significantly decreased by Cu²⁺, and those in the liver and kidneys were significantly decreased by Cd²⁺. Mn²⁺ reduced NNMT activity only in the liver. Fe²⁺ had no effect on NNMT activity. Conclusions No metal increased NNMT activity in this study, contrary to our hypothesis. Further study is needed to clarify the reason why the effects of Mn²⁺ and Fe²⁺ which have a high relevance to Parkinson’s disease on NNMT activity differ from those of Cu²⁺ and Cd²⁺     

Toxicity of heavy metals and salts to eurasian watermilfoil (Myriophyllum spicatum L.)

The toxicity of various heavy metals and salts to Eurasian watermilfoil (Myriophyllum spicatum L.) was determined under controlled growth conditions. Toxicants were added to water or to soil in systems with and without woods earth in the substrate.Fifty precent inhibition of root weight occurred with concentrations of 0.25 ppm Cu⁺², 1.9 ppm Cr₂O₇ ^–2, 3.4 ppm Hg⁺², 2.9 ppm AsO₂ ^–1, 7.4 ppm Cd⁺², 2.5 ppm Al⁺³, 9.9 ppm Cr⁺³, 41.2 ppm Ba⁺², 21.6 ppm Zn⁺², 13.3 ppm NH₄ ⁺¹, 22.4 ppm CN^–1, 143 ppm B₄O₇ ^–2, 363 ppm Pb⁺², 10,228 ppm Na₂SO₄, and 8,183 ppm NaCl. Soil increased toxicity of Cr⁺² and Ba⁺² but decreased toxicity of Cr₂O₇ ^–2, Cu⁺², Cd⁺², Al⁺³, and Hg⁺². In distilled water, CaCl₂ increased toxicity of Cr⁺³ but not Cr₂O₇ ^–2.For most toxicants there was a consistent relationship between inhibition of length and inhibition of weight and between inhibition of roots and inhibition of shoots. However, Cr₂O₇ ^–2 disproportionately decreased dry weight, and Hg⁺² and Na₂SO₄ disproportionately decreased stem length growth. With Cd⁺² and Cu⁺² stem length was greater relative to other measures of growth. Toxicity of Na₂SO₄ and NaCl was the same when concentrations were calculated as osmotic pressure but not when calculated as Na atoms or as total molecules/L.     

Hexavalent Chromium Reduction by <Emphasis Type="Italic">Bacillus</Emphasis> sp. Strain FM1 Isolated from Heavy-Metal Contaminated Soil

A Cr(VI) reducing bacterial strain FM1 was isolated from heavy metal contaminated agricultural soil irrigated with tannery effluents of Jajmau, Kanpur (India), and was identified as Bacillus sp. on the basis of biochemical methods and 16S rDNA gene sequence analysis. FM1 strain was found to be resistant to some toxic heavy metals (Cr(VI), Cr(III), Cu²⁺, Co²⁺, Cd²⁺, Ni²⁺ and Zn²⁺) up to several fold concentrations to the normal levels occurring in highly polluted region. FM1 was resistant to very high concentration of Cr(VI) (1,000 mg/L) and completely reduced 100 mg/L Cr(VI) within 48 h. Factors (pH, temperature, initial Cr(VI) concentration) affecting Cr(VI) reduction under culture condition were also evaluated. Reduction was optimum at 37°C and pH 8. Cr(VI) reduction was enhanced by addition of glucose. The presence of heavy metal cations, such as Cu²⁺, Co²⁺, Cd²⁺, Ni²⁺ and Zn²⁺ showed differential effect on reduction. Since strain FM1 could grow in the presence of significant concentrations of metals and due to high Cr(VI) reduction ability, this bacterium may be potentially applicable in Cr(VI) detoxification.     

Magnetization of Bauxite Residue to Enhance the Removal Efficiency Towards Heavy Metals

Bauxite residues are a mass of industrial wastes derived from aluminum metallurgy. This work provided a simple pyrolysis method to magnetize the bauxite residue to serve as a magnetic adsorbent towards heavy metals removal. The X-ray diffraction patterns and Mossbauer spectrum results confirmed the partial reduction of iron species with an obvious enhancement in magnetization. The magnetized bauxite residue exhibited excellent removal efficiencies for Cu²⁺, Cd²⁺ and Pb²⁺ with maximum adsorption capacities of 219.0 mg g^?1, 275.4 mg g^?1, and 100.4 mg g^?1, which could be quickly separated through a magnet. The adsorption equilibrium data were fitted to the Langmuir isotherm model, while the adsorption kinetics followed a pseudo-first-order model. According to the characterization results, chemical precipitation and sorption was the major mechanism for the removal of Cu²⁺, Pb²⁺, and Cd²⁺. Thus, the magnetized bauxite residue exhibited promising applications for heavy metals removal in wastewater.

     

Cadmium kinetics in freshwater clams. III. Effects of zinc on uptake and distribution of cadmium inAnodonta cygnea

Freshwater clams (Anodonta cygnea) were exposed to Cd²⁺ (25 g/L) or to Cd²⁺ (25 g/L) plus Zn²⁺ (2.5 mg/L). In the presence of zinc, uptake of cadmium in whole clam was halved. In organs such as the gills, mantle, and labial palps a still larger reduction of Cd accumulation occurred. By contrast, accumulation in midgut gland and kidney was hardly affected by the presence of zinc. It is shown that in the gills zinc competes with cadmium for metal binding sites both within the particulate fraction and the high-molecular weight fraction of the cytosol. Zinc probably did not induce an enlarged synthesis of specific metal-binding, metallothionein-like, proteins. In conclusion, zinc exerts antagonistic effects on uptake of cadmium by gills, and accelerates Cd transport from the gills towards the internal organs.     

Acute toxicities of selected heavy metals to the softshell clam,Mya arenaria

Summary Static acute toxicity bioassays with adult softshell clams and salts of copper, cadmium, zinc, lead, manganese, and nickel were conducted at 30 salinity and 22°C. Concentrations fatal to 50% in 168 hours, in mg/ (ppm) metal added at start, were 0.035 for Cu, 0.150 for Cd, 1.55 for Zn, 8.80 for Pb, 300.0 for Mn, and >50.0 for Ni. Additional tests were conducted with Zn²⁺ and Cu²⁺ at 30 during fall (17.5°C) and winter (4°C); clams displayed increasing survival with decreasing temperature. For Cu, LC-50 (336 hr) values at 17°C and 4°C were 0.086 and >3.00 mg/, respectively; for Zn these were 2.65 and >25.0, respectively.     

Study on the In Vitro Effects of the Mixtures of Polycyclic Aromatic Hydrocarbons (PAHs) and Heavy Metals on Ethoxyresorufin-O-Deethylase (EROD) Activity in Mossambica tilapia Liver

This paper reports in vitro effects of individual heavy metals (Cd²⁺, Cu²⁺ and Hg²⁺), and PAHs, including benzo[a]pyrene(BaP), indeno[1,2,3-cd]pyrene (IP) and fluoranthene (FL), and their mixtures on ethoxyresorufin-O-deethylase (EROD) activities using a plate-reader method. The results showed that all three metals inhibited EROD activity, while BaP/IP significantly induced the enzyme. However, FL alone decreased EROD activity. Moreover, co-treatment with BaP/IP and heavy metals inhibited PAH-induced EROD activities, while combined exposure to FL and heavy metals induced FL-inhibited EROD activities.     

Binding of cadmium by cyanobacterial growth media: Free ion concentration as a toxicity index to the cyanobacteriumNostoc UAM 208

A quantitative study of cadmium binding to three different growth media for nitrogen-fixing cyanobacteria was done with the aid of a solid state ion-specific electrode. Kratz and Myers modified medium and Arnon medium bound large amounts of Cd²⁺, BG11₀ medium had less binding capacity. Of the media components, phosphate ion showed the greatest ability to bind Cd²⁺. Different pHs, the size of cell inoculum and two buffers (Tricine and HEPES, 25 mM) also changed the availability of free cadmium ion in solution.The effect of free Cd²⁺ ion towards the cyanobacteriumNostoc UAM 208, isolated from a heavy metal polluted environment, also was tested. The effective concentration affecting 50% of population (EC₅₀), at 120 h of exposure, was less for nitrogenase activity (0.26 g/mL) than for growth (0.55 g/mL), suggesting that this enzyme activity is more sensitive to cadmium than growth. Furthermore, cadmium toxicity was influenced by the addition of buffers to the growth medium. In the presence of buffer, Tricine (25 mM), growth and nitrogenase activity was reduced by 50% at a total cadmium concentration of about 115 g/mL, although no free ion was detected in this case.These results suggest that although generally cadmium toxicity is a function of free metal ion concentration, this can also vary in the presence of complexing agents.     

Low-molecular-weight-chitosan ameliorates cadmium-induced toxicity in the freshwater crab, Sinopotamon yangtsekiense

Cadmium (Cd) has been shown to induce oxidative stress. Low-molecular-weight-chitosan (LMWC) has been demonstrated to exhibit potent antioxidant effects. We investigated the regulation role in Cd²⁺-induced oxidative damage in the hepatopancreas of the freshwater crab Sinopotamon yangtsekiense and the protective effect of LMWC. The results showed that Cd²⁺ significantly increased the hepatopancreatic metallothionein (MT) mRNA levels and protein kinase C (PKC) activity while decreasing the activities of Na⁺,K⁺-ATPase and Ca²⁺-ATPase in crabs relative to the control group. Co-treatment with LMWC suppressed the levels of MT and PKC but raised the activities of Na⁺,K+-ATPase and Ca²⁺-ATPase in hepatopancreatic tissues compared with the crabs exposed to Cd²⁺ alone. We postulate that LMWC may exert its protective effect through regulating the expressions of MT, PKC, Na⁺,K⁺-ATPase and Ca²⁺-ATPase, thereby enhancing antioxidant defense. These observations suggest that LMWC may be beneficial because of its ability to alleviate the Cd²⁺-induced damages to the crabs.     

Effects of Pb and Ni stress on oxidative stress parameters in three moss species

Antioxidative responses of the mosses Hypnum plumaeforme, Thuidium cymbifolium, and Brachythecium piligerum to short-term Pb and Ni stress were investigated. Both Pb and Ni treatment increased the formation of reactive oxygen species and lipid peroxidation, decreased superoxide dismutase (SOD), and catalase (CAT) activities in H. plumaeforme and T. cymbifolium. However, SOD activity in B. piligerum was increased under 10 mM Pb stress and Ni increased CAT activity in B. piligerum under 1 mM Ni stress. Peroxidase (POX) activity in the three mosses was increased by Pb and Ni exposure, indicating that POX plays an important role in preventing heavy metal-induced oxidative stress. The accumulation of O₂⁻ and H₂O₂ in mosses is related to the decline in SOD and CAT activities. B. piligerum is the most sensitive and T. cymbifolium is the most tolerant species to Pb and Ni stress among the three bryophytes.