Metals (Ag+, Cd2+, Cr6+) affect ATPase activity in the gill,kidney, and muscle of freshwater fish Oreochromis niloticus following acute and chronic exposures

Freshwater fish Oreochromis niloticus were individually acutely exposed to different concentrations (0, 0.1, 0.5, 1.0, and 1.5 μg/mL) of Cd²⁺, Cr⁶⁺, and Ag⁺ for 96 h and 0.05 μg/mL concentration of the same metals for different periods (0, 5, 10, 20, and 30 days) chronically. Following each experimental protocol, Na⁺/K⁺‐ATPase, Mg²⁺‐ATPase, and Ca²⁺‐ATPase activities were measured in the gill, kidney, and muscle of O. niloticus. In vitro experiments were also performed to determine the direct effects of metal ions (0, 0.1, 0.5, 1.0, and 1.5 μg/mL) on ATPases. Except Ag⁺, none of the metals caused fish mortality within 30 days. Silver killed all the fishes within 16 days. Metal exposures generally decreased Na⁺/K⁺‐ATPase and Ca²⁺‐ATPase activities in the tissues of O. niloticus, although there were some fluctuations in Mg²⁺‐ATPase activity. Ag⁺ and Cd²⁺ were found to be more toxic to ATPase activities than Cr⁶⁺. It was also observed that metal efficiency was higher in the gill than in the other tissues. Results indicated that the response of ATPases varied depending on metals, exposure types, and tissues. Because ATPases are sensitive to metal toxicity, their activity can give valuable data about fish physiology. Therefore, they may be used as a sensitive biomarker in environmental monitoring in contaminated waters. © 2011 Wiley Periodicals, Inc. Environ Toxicol 28: 707–717, 2013.     

Metal sensitivity of the embryonic development of the ramshorn snail <Emphasis Type="Italic">Marisa cornuarietis</Emphasis> (Prosobranchia)

We investigated the effects of metal ions on the embryonic development of the ramshorn snail, Marisa cornuarietis, by exposing embryos to varying concentrations of copper (0, 50, 100, and 250 μg Cu²⁺/L), lead (0, 5, 10, and 15 mg Pb²⁺/L), lithium (0, 1, 2.5, and 3 mg Li⁺/L), or palladium (0, 50, 100, and 500 μg Pd²⁺/L). Effects of these metals were examined by recording mortality, the rate of tentacles and eyes formation, heart rate, hatching success, and weight after hatching. Compared to the control, we found a significant delay in the formation of tentacles and eyes after treatment with 100 μg Cu²⁺/L, 15 mg Pb²⁺/L, 2.5 mg Li⁺/L or 500 μg Pd²⁺/L. The heart rate decreased significantly at 500 μg Pd²⁺/L. At 10 mg Pb²⁺/L, 2.5 mg Li⁺/L, or 500 μg Pd²⁺/L, hatching was delayed significantly; 50 μg Cu²⁺/L induced a significantly earlier hatching, and reduced body weight. The LC₅₀ values were calculated to be about 50 μg Cu²⁺/L, 500 μg Pd²⁺/L, 2500 μg Li⁺/L, and 10000 μg Pb²⁺/L. These results show that the embryonic development of M. cornuarietis is about as sensitive to copper and lithium, compared to the most sensitive fishes used in embryo toxicity testing. Even though the MariETT is a laboratory-based assay focusing on toxicological endpoints of a selected model species, future application is envisaged to include testing of “natural” samples such as stream water or sediment interstitial water.     

Influence of amitriptyline,chlorpromazine, and imipramine on the activity of phosphohydrolases

We have studied the effects of the antidepressants amitriptyline and imipramine and the neuroleptic chlorpromazine on the enzymatic function of Ca²⁺-activating-Mg²⁺-dependent ATPase of sarcoplasmic reticulum (SR), Na⁺, K⁺-dependent ATPase of endoplasmic reticulum (ER), and phosphodiesterase of cyclic adenosine monophosphate (cAMP). It is shown that interference with the enzymatic function (decrease of [Ca²⁺]/[ATP]) or inhibition of active transport of calcium ions by Ca²⁺, Mg²⁺-ATPase of SR by the psychotropic drugs leads to impairment of Ca²⁺ transport into the SR vesicule, which increases the force of muscle contraction. This effect decreases with increasing concentrations of amitriptyline and imipramine and with decreasing concentration of chlorpromazine, which is evidence for different mechanisms of action of Ca²⁺, Mg²⁺-ATPase of SR. Amitriptyline was a noncompetitive reversible inhibitor of Ca²⁺, Mg²⁺-ATPase of SR with an ATP hydrolysis inhibition constant K_i = 1.8 × 10⁻⁴ M and an active calcium transport inhibition constant K_i = 0.7 × 10⁻⁴ M. The goal of the investigation was to study the molecular mechanisms of action of amitriptyline, imipramine, and chlorpromazine. Translated from Khimiko-Farmatsevticheskii Zhurnal, Vol. 42, No. 9, pp. 3–10, September, 2008.     

The combined effect of copper and low pH on antioxidant defenses and biochemical parameters in neotropical fish pacu, <Emphasis Type="Italic">Piaractus mesopotamicus</Emphasis> (Holmberg, 1887)

Copper sulfate is widely used in aquaculture. Exposure to this compound can be harmful to fish, resulting in oxidative metabolism alterations and gill tissue damage. Pacu, Piaractus mesopotamicus, (wt = 43.4 ± 3.35 g) were distributed in experimental tanks (n = 10; 180 l) and exposed for 48 h to control (without copper addition), 0.4Cu (0.4 mg l⁻¹), 0CupH (without copper addition, pH = 5.0) and 0.4CupH (0.4 mg l⁻¹, pH = 5.0). In liver and red muscle, the superoxide dismutase (SOD) was responsive to the increases in the aquatic copper. The plasmatic intermediary metabolites and hematological variables in the fish of group 0.4Cu were similar to those of the control group. Conversely, the exposure to 0.4CupH caused an increase in the plasmatic lactate, number of red blood cells (RBC) and hemoglobin (Hb). Plasmatic copper concentration [Cu_p] increased in group 0.4Cu and 0.4CupH, which is higher in group 0.4CupH, suggests an effect of water pH on the absorbed copper. Exposure to 0.4Cu and 0.4CupH resulted in a reduction in the Na⁺/K⁺-ATPase activity and an increase in metallothionein (MT) in the gills. Exposure to 0CupH caused a decrease in glucose and pyruvate concentrations and an increase in RBC, Hb, and the branchial Na⁺/K⁺-ATPase activity. These responses suggest that the fish triggered mechanisms to revert the blood acidosis, save energy and increase the oxygen uptake. MT was an effective biomarker, responding to copper in different pHs and dissolved oxygen. Combined-factors caused more significant disturbance in the biomarkers than single-factors.     

<Emphasis Type="Italic">In vitro</Emphasis> Na<Superscript>+</Superscript>/K<Superscript>+</Superscript>-ATPase inhibitory activity and antimicrobial activity of sesquiterpenes isolated from <Emphasis Type="Italic">Thujopsis dolabrata</Emphasis>

A series of sesquiterpenes and hinokitiol-related compounds (1–15) was isolated from the essential oil of Thujopsis dolabrata Sieb. et Zucc. var. hondai Makino, and their structures were determined by combined spectroscopic analyses. The inhibitory effects of these compounds on microbial cell growth and Na⁺/K⁺-ATPase were evaluated in vitro. It was found that (−)-elema-1,3,11(13)-trien-12-ol (5), α,β,γ-costol (8), and chamigrenol (11) inhibit the activities of Na⁺/K⁺-ATPase, with IC₅₀ values of 11.2 ± 0.11, 12.2 ± 0.09, and 15.9 ± 0.54 μg/mL, respectively. Thujopsene (1), cedrol (9), γ-cuparenol (10), and chamigrenol (11) showed potent antibacterial activity, with MIC values in the range of 25–50 μg/mL, and β-thujaplicin (12) exhibited a broad spectrum of antibacterial and antifungal activity. These results indicate that these isolated compounds are promising candidates for the development of potent Na⁺/K⁺ ATPase inhibitors and antimicrobial agents.     

Effects of cadmium alone and in combination with low molecular weight chitosan on metallothionein,glutathione‐S‐transferase,acid phosphatase,and ATPase of freshwater crab Sinopotamon yangtsekiense

Cadmium (Cd) is an environmental contaminant showing a variety of deleterious effects, including the potential threat for the ecological environment and human health via food chains. Low molecular weight chitosan (LMWC) has been demonstrated to be an effective antioxidant. Metallothionein (MT) mRNA levels and activities of glutathione‐S‐transferase (GST), superoxide dismutase (SOD), acid phosphatase (ACP), Na⁺,K⁺‐ATPase, and Ca²⁺‐ATPase as well as malondialdehyde (MDA) contents in the gills of the freshwater crab Sinopotamon yangtsekiense were analyzed in vivo in order to determine the injury of Cd exposure on the gill tissues as well as the protective effect of LMWC against this injury. The results showed that there was an apparent accumulation of Cd in the gills, which was lessened by the presence of LMWC. Moreover, Cd²⁺ significantly increased the gill MT mRNA levels, ACP activity and MDA content while decreasing the activities of SOD, GST, Na⁺,K⁺‐ATPase, and Ca²⁺‐ATPase in the crabs relative to the control. Cotreatment with LMWC reduced the levels of MT mRNA and ACP but raised the activities of GST, Na⁺,K⁺‐ATPase, and Ca²⁺‐ATPase in gill tissues compared with the crabs exposed to Cd²⁺ alone. These results suggest that LMWC may exert its protective effect through chelating Cd²⁺ to form LMWC‐Cd²⁺ complex, elevating the antioxidative activities of GST, Na⁺,K⁺‐ATPase, and Ca²⁺‐ATPase as well as alleviating the stress pressure on MT and ACP, consequently protecting the cell from the adverse effects of Cd. © 2012 Wiley Periodicals, Inc. Environ Toxicol 29: 298–309, 2014.     

Dietary exposure to titanium dioxide nanoparticles in rainbow trout, (Oncorhynchus mykiss): no effect on growth, but subtle biochemical disturbances in the brain

Our laboratory recently reported gut pathology following incidental ingestion of titanium dioxide nanoparticles (TiO₂ NPs) during aqueous exposures in trout, but there are almost no data on dietary exposure to TiO₂ NPs in fish. The aim of this experiment was to observe the sub-lethal effects of dietary exposure to TiO₂ NPs in juvenile rainbow trout (Oncorhynchus mykiss). Stock solutions of dispersed TiO₂ NPs were prepared by sonication without the use of solvents and applied to a commercial trout diet. Fish were exposed in triplicate to either, control (no added TiO₂), 10, or 100 mg kg⁻¹ TiO₂ NPs diets for 8 weeks followed by a 2 week recovery period where all fish were fed the control diet. TiO₂ NPs had no impact on growth or nutritional performance, and no major disturbances were observed in red or white blood cell counts, haematocrits, whole blood haemoglobin, or plasma Na⁺. Ti accumulation occurred in the gill, gut, liver, brain and spleen during dietary TiO₂ exposure. Notably, some of these organs, especially the brain, did not clear Ti after exposure. The brain also showed disturbances to Cu and Zn levels (statistically significant at weeks 4 and 6; ANOVA or Kruskal–Wallis, P < 0.05) and a 50% inhibition of Na⁺K⁺-ATPase activity during TiO₂ NP exposure. Na⁺K⁺-ATPase activity was unaffected in the gills and intestine. Total glutathione in the gills, intestine, liver and brain were not affected by dietary TiO₂ NPs, but thiobarbituric acid reactive substances (TBARS) showed up to 50% decreases in the gill and intestine. We conclude that TiO₂ NPs behave like other toxic dietary metals where growth rate and haematology can be protected during sub-lethal exposures, but in the case of TiO₂ NPs this may be at the expense of critical organs such as the brain and the spleen.     

Evaluation of genotoxicity and effects on reproduction of nonylphenol in <Emphasis Type="Italic">Oreochromis niloticus</Emphasis> (Pisces: cichlidae)

Nonylphenol ethoxylate (NPE) is widely used as a component of detergents, paints, pesticides, and many other products. In the aquatic environment NPE breakdown to 4-nonylphenol (NP), which is more stable and persistent. NP is estrogenic in fish, avian, and mammals and is described as an environmental pollutant with endocrine disruptor characteristics. The genotoxicity of NP was evaluated through micronuclei assay and single cell gel electrophoresis (Comet assay) in peripheral erythrocytes of Oreochromis niloticus exposed in vivo. The study on reproductive development was also carried out in male and female gonads of O. niloticus. Lethal concentration (LC 50%) of 0.032 ml l⁻¹ was previously determined. We ran assays with O. niloticus exposed to concentrations of 1.0, 10.0, and 16.0 μl l⁻¹ of NP diluted in water. Our results showed that NP was not genotoxic. However, 3-day exposure to NP in concentrations of 1.0, 10.0, and 16.0 μl l⁻¹ of water increased the frequency of reproductive stages in males and females. The histology of the reproductive tract of the treated fish was significantly altered in females treated with 16.0 μl l⁻¹ of water when compared to controls. Analogous estrogenic effects were observed, such as accelerated maturation of oocytes and spermatogenesis. These results showed that the O. niloticus reproductive system is sensitive to NP estrogenicity.     

Chronic exposure to dichloroacetamide induces biochemical and histopathological changes in the gills of zebrafish

To evaluate the impact of DCAcAm on zebrafish gill, we measure the responses of antioxidant enzyme (superoxide dismutase, SOD), lipid peroxidation (malondialdehyde, MDA), ATPase (Na⁺/K⁺‐ATPase and Ca²⁺/Mg²⁺‐ATP) and histopathological changes of gill in adult zebrafish, after exposed to different concentrations of DCAcAm (0, 1, 10, 100, and 1000 μg L^?1) for 30 days. Results indicated that DCAcAm first increased and then decreased SOD activity, and DCAcAm also lowered the activities of Na⁺/K⁺‐ATPase and Ca²⁺/Mg²⁺‐ATPase. These results indicated that high affinity of DCAcAm probably be a main factor, which can damage the structures of enzymes, thereby inhibiting the SOD and ATPase activities. Besides, histopathological investigation results also manifested that chronic exposure to DCAcAm can damage the gill tissues, disrupting the normal function of gills. We conclude that chronic exposure to DCAcAm was harmful to organisms, not only influence gill function, but also further cause damage on the gill tissues.     

Purification and Characterisation of a Newly Isolated Stable Long-Life Tannase produced by F. subglutinans (Wollenweber and Reinking) Nelson et al.

A stable long-life tannase was synthesised by Fusarium subglutinans and the fermentation processing parameters were optimised. Maximum enzyme production (9.38 U/ml) was recorded after 96 h of incubation at 35°C, initial pH 5, in submerged culture (200 rpm) utilising 2% (w/v) tannic acid as a sole carbon source. The tannase produced was purified to electrophoretic homogeneity through two-step column chromatography and the purified form remained stable in a pH range of 6–8. Its midpoint of thermal inactivation (T _m) was recorded at 70°C after 60 min of exposure. Maximum tannase activity was enabled at pH 6 and 40°C. Ca²⁺, K⁺, Mg²⁺ and Mn⁺ showed a stimulatory effect while Ba²⁺, Co²⁺, Cu²⁺, Fe³⁺ and Zn⁺ showed a competitive inhibitory effect on enzyme activity. Values of K _m, V _max, K _cat and the molecular mass of the purified enzyme were 0.116 μM ml⁻¹ min⁻¹, 3.57 mM, 1.16 μM ml⁻¹ min⁻¹ and 150 kDa, respectively. The participation of the SH group and carbohydrates in the enzyme structure was also suggested by the results. The stability of the purified and partially purified enzyme at −15°C extended to 13 months.     

In vitro and in vivo effects of cadmium on cholinesterases in Nile tilapia fingerlings: implications for biomonitoring aquatic pollution

Effects of cadmium on in vitro and in vivo cholinesterase (ChE) activities of brain and muscle tissues of Oreochromis niloticus fingerlings were evaluated, considering its potential use in biomonitoring tropical water pollution. Results show that in vitro ChE activities were depressed significantly by millimolar concentration ranges of Cd²⁺. The IC₅₀ values of Cd²⁺ on in vitro ChE activity in brain and muscle tissues were 1.56 and 4.31 mM, respectively. Exposure of fish to environmentally relevant concentrations of Cd²⁺ (5–30 μg l^−l) for 28 days evoked only a transient inhibition (21–34%) of in vivo ChE activities. Prior exposure and co-exposure of fish to 15 μg l⁻¹ of Cd²⁺ enhanced the extent of inhibition of ChE levels induced by the organophosphorous insecticide chlorpyrifos. As high concentrations of cadmium have the potential to depress ChE activities, monitoring of metal levels in water bodies with suspected high levels of metal inputs is necessary to accurately interpret the fish ChE inhibition data in relation to insecticide contaminations.     

The inhibition by di(2-ethylhexyl)-phthalate of <Emphasis Type="Italic">erg</Emphasis>-mediated K<Superscript>+</Superscript> current in pituitary tumor (GH<Subscript>3</Subscript>) cells

DEHP (bis(2-ethylhexyl)-phthalate) known to be an endocrine-disrupting chemical is a widely used phthalate. Little information regarding the effects of phthalate esters on ion currents is available. In this study, the effects of DEHP and other phthalate esters (DBEP: di(2-butoxyethyl)-phthalate and DMGP: di(2-methylglycol)-phthalate) on ion currents were investigated in pituitary GH₃ cells. Hyperpolarization-elicited K⁺ currents in GH₃ cells bathed in high-K⁺, Ca²⁺-free solution were examined to evaluate the effects of DEHP, DBEP, and DMGP on the ether-à-go–go-related-gene (erg) K⁺ current (I _K(erg)). Addition of DEHP to GH₃ cells suppressed the amplitude of I _K(erg) in a concentration-dependent manner with an IC₅₀ value of 16.3 μM. With a two-pulse protocol, addition of DEHP shifted the activation curve of I _K(erg) to a depolarized potential by approximately 10 mV with no change in the rate of I _K(erg) deactivation. This compound did not have any effects on delayed rectifier K⁺ current in GH₃ cells, while 4-aminopyridine-3-methanol (100 μM) suppressed this current significantly. DBEP (30 μM) had little or no effect on I _K(erg), while DMGP (30 μM) slightly reduced it. In inside-out configuration, DEHP (30 μM) applied to the bath slightly reduced the activity of large-conductance Ca²⁺-activated K⁺ channels. DEHP (30 μM) increased the frequency of spontaneous action potentials (APs); however, this compound at the same concentration had no effect on AP firing in KCNH2 siRNA-transfected GH₃ cells. The effects described herein can contribute to their actions on functional activity of endocrine or neuroendocrine cells if similar results are found in vivo.     

Influence of kaempferol,a flavonoid compound,on membrane-bound ATPases in streptozotocin-induced diabetic rats

Abstract

Context: Kaempferol is a flavonoid found in many edible plants (e.g. tea, cabbage, beans, tomato, strawberries, and grapes) and in plants or botanical products commonly used in traditional medicine. Numerous preclinical studies have shown that kaempferol have a wide range of pharmacological activities, including antioxidant, anti-inflammatory, anticancer, cardioprotective, neuroprotective, and antidiabetic activities.

Objective: The present study investigates the effect of kaempferol on membrane-bound ATPases in erythrocytes and in liver, kidney, and heart of streptozotocin (STZ)-induced diabetic rats.

Materials and methods: Diabetes was induced into adult male albino rats of the Wistar strain, by intraperitoneal administration of STZ (40?mg/kg body weight (BW)). Kaempferol (100?mg/kg BW) or glibenclamide (600?µg/kg BW) was administered orally once daily for 45?d to normal and STZ-induced diabetic rats. The effects of kaempferol on membrane-bound ATPases (total ATPase, Na⁺/K⁺-ATPase, Ca²⁺-ATPase, and Mg²⁺-ATPase) activity in erythrocytes and in liver, kidney, and heart were determined.

Results: In our study, diabetic rats had significantly (p?<?0.05) decreased activities of total ATPases, Na⁺/K⁺-ATPase, Ca²⁺-ATPase, and Mg²⁺-ATPase in erythrocytes and tissues. Oral administration of kaempferol (100?mg/kg BW) or glibenclamide (600?µg/kg BW) for a period of 45?d resulted in significant (p?<?0.05) reversal of these enzymes' activities to near normal in erythrocytes and tissues when compared with diabetic control rats.

Discussion and conclusion: Thus, obtained results indicate that administration of kaempferol has the potential to restore deranged activity of membrane-bound ATPases in STZ-induced diabetic rats. Further detailed investigation is necessary to discover kaempferol’s action mechanism.     

Effects of 4-nonylphenol,fish predation and food availability on survival and life history traits of <Emphasis Type="Italic">Daphnia magna</Emphasis> straus

This study aimed to investigate the compound effect of environmentally relevant 4-nonylphenol (NP) concentrations and natural stressors—namely fish predation and food availability—on Daphnia magna, which were exposed to four NP concentrations (0, 1, 5 and 10 μg l⁻¹) under optimum or low food concentrations (1.00 and 0.075 mg C l⁻¹, respectively) in water (un)conditioned by a fish predator (Alburnus alburnus). A(n) “environmentally relevant” and “no observable effect” concentration (NOEC) of NP (10 μg l⁻¹) resulted in a significant reduction (P < 0.01**) in daphnids’ survival when it was encountered concurrently with conditions of low food availability and presence of fish predation. The significance of the results lies in the observation that not only environmentally relevant concentrations of NP but also NP concentrations reported to have no observable effect on daphnids may in reality have unexpected critical effects on D. magna survival under conditions more parallel to natural ecosystems. The deterioration of the life-history traits—namely, NP-induced delay in the age at first reproduction (P < 0.001***) and fish kairomone-induced reduction in the size at first reproduction (P < 0.001***)—of the D. magna individuals is also crucial, as such alterations could significantly influence future generations and result in ultimate adverse effects at the community level because large-bodied daphnids are key-stone species in freshwater ecosystems. The results of this study demonstrate the importance of taking into account environmentally realistic conditions while investigating the effects of NOEC levels of toxicants on non-target aquatic species.     

EFFECTS OF SELECTED ORGANIC SOLVENTS ON THE ASTROCYTE MEMBRANE ATPase IN VITRO

1. The present study deals with astrocyte cultures as a model for studying the membrane-mediated central nervous system-depressing effect of organic solvents. 2. The primary astrocyte cultures were prepared from neonatal rat cerebella. The cells were cultured in modified essential medium. The astrocyte membranes isolated from the cultures were exposed to solvents in incubation mixture at different dose levels (3, 6 and 9 mmol/L) for 1h. The physiologically important integral proteins Na⁺, K⁺-ATPase and Mg²⁺-ATPase were studied. 3. The aromatic hydrocarbons (benzene, toluene, styrene, xylene and ethylbenzene) inhibited the ATPase activities according to their lipid solubilities. n-Hexane and cyclohexane clearly had less effect than aromatic hydrocarbons, despite their greater lipid solubilities. 4. Astrocytes were shown to be sensitive targets to the effects of organic solvents, measured as the inhibition of the integral enzymes Na⁺, K⁺-ATPase and Mg²⁺-ATPase.     

Modulation of contractile enzyme activity by anticonvulsant drugs

Anticonvulsant drugs of the heterocyclic amide family inhibit in vitro enzymatic activity of Ca²⁺, Mg²⁺-ATPase of sarcoplasmic reticulum, the activated transmembrane calcium-ion transfer/ATP hydrolysis specific rate ratio being retained to provide lower concentrations of calcium ions in myofibrilla and relaxed muscles. Ethosuximide is a noncompetitive inhibitor with reversible action on basic enzyme catalyzed processes. The inhibition of the hydrolytic activity of the enzyme under the action of ethosuximide, phenytoin, and primidone was compared to their effect on other phosphatases (cAMP phosphodiesterase and Na⁺, K⁺-dependent ATPase). Translated from Khimiko-Farmatsevticheskii Zhurnal, Vol. 42, No. 9, pp. 6–8, September, 2008.     

Physiological response of Cu and Cu mine tailing remediation of <Emphasis Type="Italic">Paulownia fortunei</Emphasis> (Seem) Hemsl

The physiological responses and Cu accumulation of Paulownia fortunei (Seem) Hemsl. were studied under 15.7–157 μmol L⁻¹ Cu treatments in liquid culture for 14 days; the impacts of Cu concentration in the seedlings were evaluated under Cu mine tailing culture with acetic acid and EDTA treatment for 60 days. Results showed that the concentrations of Chl-a, Chl-b and Carotenoids significantly increased (p < 0.05) at 15.7–78.7 μmol L⁻¹Cu treatment and significantly decreased at 157 μmol L⁻¹ treatment after 14 days of Cu exposure. The activities of superoxide dismutase (SOD) and catalase (CAT) significantly increased as Cu levels were enhanced and the activities of both SOD and CAT under 157 μmol L⁻¹ Cu stress were 2.9 and 1.9 times higher than that of control, respectively. The concentrations of proline and soluble sugars in the leaves of P. fortunei significantly increased as the Cu concentrations were elevated. Cu concentrations in roots, stems and leaves of P. fortunei increased significantly as Cu levels increased and reached 1911, 101 and 93 μg g⁻¹ dry weights (DW) at 157 μmol L⁻¹ Cu treatment, respectively. The seedlings of P. fortunei cultivated in Cu tailing experienced unsuccessful growth and loss of leaves in all treatments due to poor nutrition of the Cu tailing. The dry weight of P. fortunei increased under all the treatments of acetic acid after 60 days exposure. However, dry weight significantly decreased under both levels of EDTA. The Cu concentrations increased significantly in roots and decreased in leaves when each was treated with both concentrations of acetic acid. The Cu concentrations in the roots, stems and leaves increased significantly, and the concentrations of Cu in the stems and leaves under the treatment of 2 μmol L⁻¹ EDTA reached 189.5 and 763.1 μg g⁻¹ DW, respectively. The result indicated that SOD, CAT, proline and soluble sugars played an important role in coping with the oxidative stress of copper. Acetic acid could promote growth and EDTA at the experimental levels, which could also enhance Cu absorption and translocation into the stems and leaves of P. fortune. Furthermore, acetic acid and EDTA could be rationally utilized in Cu-contaminated soil.     

The involvement of transition metal ions on iron-dependent lipid peroxidation

The metals iron (Fe) and copper (Cu) are considered trace elements, and the metals cobalt (Co) and nickel (Ni) are known as ultra-trace elements, considering their presence in low to very low quantity in humans. The biologic activity of these transition metals is associated with the presence of unpaired electrons that favor their participation in redox reactions. They are part of important enzymes involved in vital biologic processes. However, these transition metals become toxic to cells when they reach elevated tissue concentrations and produce cellular oxidative damage. Phospholipid liposomes (0.5 mg/ml, phosphatidylcholine (PC)/phosphatidylserine (PS), 60/40) were incubated for 60 min at 37°C with 25 μM of Fe²⁺ in the absence and in the presence of Cu²⁺, Co²⁺, and Ni²⁺ (0–100 μM) with and without the addition of hydrogen peroxide (H₂O₂, 5–50 μM). Iron-dependent lipid peroxidation in PC/PS liposomes was assessed by thiobarbituric acid-reactive substances (TBARS) production. Metal transition ions promoted lipid peroxidation by H₂O₂ decomposition and direct homolysis of endogenous hydroperoxides. The Fe²⁺-H₂O₂-mediated lipid peroxidation takes place by a pseudo-second order process, and the Cu²⁺-mediated process by a pseudo-first order reaction. Co²⁺ and Ni²⁺ alone do not induce lipid peroxidation. Nevertheless, when they are combined with Fe²⁺, Fe²⁺-H₂O₂-mediated lipid peroxidation was stimulated in the presence of Ni²⁺ and was inhibited in the presence of Co²⁺. The understanding of the effects of transition metal ions on phospholipids is relevant to the prevention of oxidative damage in biologic systems.     

Inhibition of ATPase activity in the freshwater fish Labeo rohita (Hamilton) exposed to sodium cyanide

Present study concerns the effect of sodium cyanide on the Indian major carp, Labeo rohita. Fishes were exposed to lethal (0.32?mg/L) and sublethal (0.064?mg/L) concentrations of sodium cyanide. The effect of intoxication was studied on Na⁺K⁺ATPase, Mg⁺²ATPase and Ca²⁺ATPase in various physiological tissues (gill, liver, and muscle) at the end of 1, 2, 3 and 4 days of lethal and 5, 10 and 15 days of sublethal exposure periods. Sodium cyanide induced significant inhibitory effects on the ATPase activity of the fish. Inhibition of the ATPase blocked the active transport system of the gill epithelial as well as chloride cells, and thus altered the osmo-regulatory mechanism of the fish. The value of the measured responses as an indicator of stress caused by water contamination discussed. The results confirm that ATPase levels significantly decreased in treated fish, indicating that ATPases could be used as sensitive and useful biomarkers for cyanide pollution.     

Enzymatic alterations and RNA/DNA ratio in intestine of rainbow trout, <Emphasis Type="Italic">Oncorhynchus mykiss</Emphasis>, induced by chronic exposure to carbamazepine

We investigated the effect of long-term exposure to carbamazepine (CBZ) on the enzymatic alterations and RNA/DNA ratio in intestine tissue of rainbow trout. Fish were exposed to sublethal concentrations of CBZ (1.0 μg/l, 0.2 or 2.0 mg/l) for 42 days. Digestive enzymes (proteolytic enzymes and amylase) and energy metabolic enzyme (Na⁺-K⁺-ATPase) and antioxidant enzymes (superoxide dismutase [SOD], catalase [CAT], glutathione peroxidase [GPx], and glutathione reductase [GR]) in fish intestine were measured. In addition, intestinal RNA/DNA ratio was determined after 42 days exposure. Carbamazepine exposure at 2.0 mg/l led to significantly inhibited (P < 0.05) activity of Na⁺-K⁺-ATPase. Activities of the antioxidant enzymes SOD, CAT, and GPx in CBZ-treated groups gradually increased at lower concentration of CBZ (1.0 μg/l and 0.2 mg/l), then significantly inhibited (P < 0.05) at 2.0 mg/l. After 42 days, the RNA/DNA ratio in fish intestine was significantly lower (P < 0.05) in groups exposed to CBZ at 2.0 mg/l than in other groups. However, there was no statistical significance (P > 0.05) in the activities of digestive enzymes (proteolytic enzyme and amylase) and GR in all groups. In short, prolonged exposure to CBZ resulted in different responses of various enzymes and significantly lower RNA/DNA ratio in fish intestine. Furthermore, molecular and genetic mechanisms of these physiological responses in fish are not clear, which need to be further studied.