Temporal and spatial patterns of soil water following wildfire-induced changes in plant communities in the Great Basin in Nevada,USA

Infection of tall fescue (Festuca arundinacea Schreb.) with its endemicNeotyphodium coenophialum-endophyte (Morgan-Jones and Gams) Glenn, Bacon and Hanlin appears to reduce copper (Cu) concentrations in forage and serum of grazing animals, contributing to a range of immune-related disorders. A greenhouse experiment was conducted to identify effects of novel endophyte strains on Cu acquisition by tall fescue (Festuca arundinacea Schreb.) varieties Grasslands Flecha and Jesup infected with a novel, non ergot producing endophyte strain AR542, and two perennial ryegrass (Lolium perenne L.) varieties Aries and Quartet infected with a novel, non lolitrem B producing strain AR1, and their noninfected (E−) forms. Individual endophyte/grass associations were cultivated in nutrient solutions at 1.0 (P+) and 0.0 mM (P−) phosphorus concentrations. The Cu²⁺-binding activity of extracellular root exudates, and concentrations of Cu and other heavy metals in roots and shoots were measured. Extracellular root exudates of AR542-infected vs. E− tall fescue had higher Cu²⁺-binding activity only in P− nutrient solution as shown by lower concentration of free Cu²⁺ (0.096 vs. 0.188 mmol Cu²⁺ g⁻¹ root DM, respectively). The Cu²⁺-binding activity by root exudates of perennial ryegrass was not affected by endophyte infection, but was higher (i.e., lower concentration of free Cu²⁺) in P− vs. P+ nutrient solution (0.068 vs. 0.114 mmol Cu²⁺ g⁻¹ root DM). In this hydroponic experiment, Cu concentrations in shoots of both grasses were not a function of Cu²⁺-binding activity and endophyte effects on heavy metal concentrations in shoots and roots were specific for each variety. The Cu²⁺-binding activity of extracellular root exudates may affect Cu accumulation by field-grown, endophyte-infected tall fescue under P-limiting growth conditions and warrants verification by more specific methods.     

Human serum albumin coordinates Cu(II) at its N-terminal binding site with 1 pM affinity

The conditional stability constant at pH 7.4 for Cu(II) binding at the N-terminal site (NTS) of human serum albumin (HSA) was determined directly by competitive UV–vis spectroscopy titrations using nitrilotriacetic acid (NTA) as the competitor in 100 mM NaCl and 100 mM N-(2-hydroxyethyl)piperazine-N′-ethanesulfonic acid (Hepes). The log K _NTS^c value of 12.0 ± 0.1 was determined for HSA dissolved in 100 mM NaCl. A false log log K _NTS^c value of 11.4 ± 0.1 was obtained in the 100 mM Hepes buffer, owing to the formation of a ternary Cu(NTA)(Hepes) complex. The impact of the picomolar affinity of HSA for Cu(II) on the availability of these ions in neurodegenerative disorders is briefly discussed.     

Interaction between Cu toxicity and P deficiency in soil-grown cowpea (Vigna unguiculata (L.) Walp.)

Anthropogenic contamination with Cu is an important issue and it is necessary to understand how Cu toxicity influences the uptake/acquisition of nutrients by plants. An experiment was conducted with soil-grown cowpea (Vigna unguiculata (L.) Walp.) to investigate the interaction between Cu toxicity and P deficiency. Plant performance was related to the activity of Cu²⁺ at the outer surface of the root plasma membrane, {Cu²⁺} ₀ ^o , which was calculated from properties of the soil solution. The addition of Cu to the soil was found to reduce growth of plant shoots by inducing Cu toxicity, which was associated with a reduction in the shoot tissue Fe concentration. The critical value (50% reduction in shoot growth) determined for {Cu²⁺} ₀ ^o in this soil-based experiment (3.8 μM) corresponds well to values determined previously. Importantly, regression analyses indicated that although the alleviation of P deficiency improved overall growth, the P-status of the plant did not influence the apparent toxicity of the Cu. This result was unexpected, given that Cu inhibits the growth of roots hairs; these being important for the uptake of immobile nutrients such as P. This study advances our understanding of Cu toxicity and its impact upon nutrient uptake.     

Purification and characterization of two cold-adapted extracellular tannin acyl hydrolases from an Antarctic strain <Emphasis Type="Italic">Verticillium</Emphasis> sp. P9

Two extracellular tannin acyl hydrolases (TAH I and TAH II) produced by an Antarctic filamentous fungus Verticillium sp. P9 were purified to homogeneity (7.9- and 10.5-fold with a yield of 1.6 and 0.9%, respectively) and characterized. TAH I and TAH II are multimeric (each consisting of approximately 40 and 46 kDa sub-units) glycoproteins containing 11 and 26% carbohydrates, respectively, and their molecular mass is approximately 155 kDa. TAH I and TAH II are optimally active at pH of 5.5 and 25 and 20°C, respectively. Both the enzymes were activated by Mg²⁺and Br⁻ ions and 0.5–2.0 M urea and inhibited by other metal ions (Zn²⁺, Cu²⁺, K⁺, Cd²⁺, Ag⁺, Fe³⁺, Mn²⁺, Co²⁺, Hg²⁺, Pb²⁺ and Sn²⁺), anions, Tween 20, Tween 60, Tween 80, Triton X-100, sodium dodecyl sulphate, β-mercaptoethanol, α-glutathione and 4-chloromercuribenzoate. Both tannases more efficiently hydrolyzed tannic acid than methyl gallate. E _a of these reactions and temperature dependence (at 0–30°C) of k _cat, k _cat/K _m, ΔG*, ΔH* and ΔS* for both the enzymes and substrates were determined. The k _cat and k _cat/K _m values (for both the substrates) were considerably higher for the combined preparation of TAH I and TAH II.     

Toxic effects of Pb2+ on the growth and mineral nutrition of signal grass (Brachiaria decumbens) and Rhodes grass (Chloris gayana)

Although grasses are commonly used to revegetate sites contaminated with lead (Pb), little is known regarding the Pb-tolerance of many of these species. Using dilute solution culture to mimic the soil solution, the growth of signal grass (Brachiaria decumbens Stapf cv. Basilisk) and Rhodes grass (Chloris gayana Kunth cv. Pioneer) was related to the mean activity of Pb²⁺ {Pb²⁺} in solution. There was a 50% reduction in fresh mass of signal grass shoots at 5 μM {Pb²⁺} and at 3 μM {Pb²⁺} for the roots. Rhodes grass was considerably more sensitive to Pb in solution, with shoot and root fresh mass being reduced by 50% at 0.5 μM {Pb²⁺}. The higher tolerance of signal grass to Pb appeared to result from the internal detoxification of Pb, rather than from the exclusion of Pb from the root. At toxic {Pb²⁺}, an interveinal chlorosis developed in the shoots of signal grass (possibly a Pb-induced Mn deficiency), whilst in Rhodes grass, Pb²⁺ caused a bending of the root tips and the formation of a swelling immediately behind some of the root apices. Root hair growth did not appear to be reduced by Pb²⁺ in solution, being prolific at all {Pb²⁺} in both species.     

Re-evaluating the effects of organic ligands on copper toxicity to barley root elongation in culture solution

Abstract

In the presence of weak ligands, both free ion activity and organic complexes of Cu should b considered when predicting Cu toxicity in aquatic and soil-plant systems. However, there is littl information about the quantitative contribution of Cu that is organically complexed to Cu toxicity. In thi study, a bioassay using barley root elongation in culture solution was used to investigate the effects o organic ligands with different conditional stability constants on Cu toxicity and the quantitativ contribution of the organically complexed Cu to the Cu toxicity. The results indicated that a significan decrease (p<0.05) in Cu toxicity, assessed by barley root elongation, was observed in response to th addition of organic ligands. The decrease differed, to some extent, with different organic ligands o disodium ethylenediaminetetraacetic acid (EDTA), nitrilotriacetic acid (NTA), oxalate and malate at low and constant free Cu²⁺ activity. Addition of EDTA or NTA resulted in strong reduction of Cu toxicity while modest reduction of Cu toxicity was observed for the addition of malate as the relatively wea ligand. Furthermore, the results of the present study revealed that the CuNTA^? and CuEDTA^2? complexes were not toxic, while the Cu–malate complexes were mildly toxic to barley root elongation More importantly, it was found that the toxicity of Cu–malate complexes were nearly 0.5-fold less than that of free Cu²⁺ ions.     

A resin buffered method for controlling metal speciation in nutrient solutions for plant toxicity tests

Aims

Root elongation tests are sensitive bioassays for testing metal toxicity in nutrient solutions. The metal speciation and, hence, metal exposure conditions are little controlled in the traditional set-up. A resin buffered solution system was developed to overcome this issue.

Methods

Barley (Hordeum vulgare L.) root elongation was tested in aerated 140 mL solution batch systems supplied with 3.3 g Dowex resin for two plants. Copper toxicity was measured in presence or absence of the resin (+R/?R) and in presence or absence of a metal complexing ligand (+NTA; nitrilotriacetic acid/?NTA). In addition, the toxicity in the traditional set without resin and with daily solution replacement was included as a reference.

Results

Metal desorption from the resin is fast in these systems (k?=?0.82 h^?1). Total dissolved Cu roughly halved during 4 days in ?R/?NTA systems due to uptake, while it increased by 30 % in the +R/?NTA, probably due to complexation reactions by root-derived molecules. The toxicity (50 % reduction in root length, EC50) of the initial free Cu²⁺ was equal in all resin or chelate buffered systems and in the solutions with daily replacement, whereas this threshold was significantly larger in the ?R/?NTA due to Cu²⁺ uptake and complexation reactions.

Conclusion

The resin method is a convenient system for high throughput screening of metal toxicity and avoids uncertainties in metal speciation inherent to chelator buffered systems. Details are given how to prepare the resin to obtain a target metal ion activity.     

Optimization of flow rate,initial metal ion concentration and biomass density for maximum removal of Cu2+ by immobilized Microcystis

The potential of alginate-immobilized Microcystis packed in a column for maximum removal of Cu²⁺ at different flow rates, biomass, and initial metal ion concentration was assessed in a continuous flow system. Although Cu²⁺ removal did occur at all the flow rates tested, it was maximum (54%) at 0.75-ml min⁻¹ flow rate, 30 μg ml⁻¹ initial metal ion concentration and 0.016 g biomass. Cu²⁺ removal was influenced by inlet metal ion concentration and biomass density. An increase in the biomass concentration from 0.016 to 0.128 g resulted in an apparent increase in percentage removal but the Cu²⁺ adsorbed per unit dry wt. declined. When the flow rate (0.75 ml min⁻¹) and biomass density (0.064 g) were kept constant and the inlet metal ion concentration was varied from 10 to 150 μg ml⁻¹, a 68% removal of Cu²⁺ was obtained at 50 μg ml⁻¹ initial concentration in a time duration of 15 min. The metal-laden columns were efficiently desorbed and regenerated following elution with double distilled water (DDW) (pH 2) (89%). This was followed by 1 mm EDTA > 1 mm NTA > 0.1 mm EDTA > 1 mm HCl > 1 mm HNO₃ > 5 mm CaCl₂ > DDW (pH 7.0) > 1 mm NaHCO₃ > 1 mm CaCl₂. Of the total (2.83 mg) adsorbed Cu²⁺, 1.89 mg (67%) was desorbed by DDW (pH 2) within the first 20 min of elution time. Thereafter the desorption rate slowed down and only 22% (0.632 mg) desorption was obtained in the last 20 min. In contrast to water pH 2, the desorption of Cu²⁺ by 1 mm EDTA was very slow, the maximum being 8% after 40 min of elution. This revised version was published online in July 2006 with corrections to the Cover Date.     

Removing heavy metals from synthetic effluents using “kamikaze” Saccharomyces cerevisiae cells

One key step of the bioremediation processes designed to clean up heavy metal contaminated environments is growing resistant cells that accumulate the heavy metals to ensure better removal through a combination of biosorption and continuous metabolic uptake after physical adsorption. Saccharomyces cerevisiae cells can easily act as cation biosorbents, but isolation of mutants that are both hyperaccumulating and tolerant to heavy metals proved extremely difficult. Instead, mutants that are hypersensitive to heavy metals due to increased and continuous uptake from the environment were considered, aiming to use such mutants to reduce the heavy metal content of contaminated waters. In this study, the heavy metal hypersensitive yeast strain pmr1∆ was investigated for the ability to remove Mn²⁺, Cu²⁺, Co²⁺, or Cd²⁺ from synthetic effluents. Due to increased metal accumulation, the mutant strain was more efficient than the wild-type in removing Mn²⁺, Cu²⁺, or Co²⁺ from synthetic effluents containing 1–2 mM cations, with a selectivity $ {\text{Mn}}^{{{\text{2}} + }} > {\text{Co}}^{{{\text{2}} + }} ~ > {\text{Cu}}^{{{\text{2}} + }} $ {\text{Mn}}^{{{\text{2}} + }} > {\text{Co}}^{{{\text{2}} + }} ~ > {\text{Cu}}^{{{\text{2}} + }} and also in removing Mn²⁺ and Cd²⁺ from synthetic effluents containing 20–50 μM cations, with a selectivity Mn²⁺ > Cd²⁺.     

A quantitative appraisal of the ambivalent metal ion binding properties of cytidine in aqueous solution and an estimation of the<Emphasis Type="Italic"> anti</Emphasis>–<Emphasis Type="Italic">syn</Emphasis> energy barrier of cytidine derivatives

The recently defined versus straight-line plots for L = pyridine-type (PyN) and ortho-aminopyridine-type (oPyN) ligands now allow the evaluation in a quantitative manner of the stability of the 1:1 complexes formed between cytidine (Cyd) and Ca²⁺, Mg²⁺, Mn²⁺, Co²⁺, Ni²⁺, Cu²⁺, Zn²⁺ or Cd²⁺ (M²⁺); the corresponding stability constants, , including the acidity constant, , for the deprotonation of the (N3)H⁺ site had been determined previously under exactly the same conditions as the mentioned plots. Since the stabilities of the M(PyN)²⁺ and M(oPyN)²⁺ complexes of Ca²⁺ and Mg²⁺ are practically identical, it is concluded that complex formation occurs in an outer-sphere manner, and this is in accord with the fact that in the pK_a range 3–7 metal ion binding is independent of or . Ca(Cyd)²⁺ and Mg(Cyd)²⁺ are more stable than the corresponding (outer-sphere) M(PyN)²⁺ complexes and this means that the C2 carbonyl group of Cyd must participate, next to N3 which is most likely outer-sphere, in metal ion binding, leading thus to chelates; these have formation degrees of about 50% and 35%, respectively. Co(Cyd)²⁺ and Ni(Cyd)²⁺ show no increased stability based on the hence, the (C2)O group does not participate in metal ion binding, but the inner-sphere coordination to N3 is strongly inhibited by the (C4)NH₂ group. In the M(Cyd)²⁺ complexes of Mn²⁺, Cu²⁺, Zn²⁺ and Cd²⁺, this inhibiting effect on M²⁺ binding at N3 is partially compensated by participation of the (C2)O group in complex formation and the corresponding chelates have formation degrees between about 30% (Zn²⁺) and 83% (Cu²⁺). The different structures of the mentioned chelates are discussed in relation to available crystal structure analyses. (1) There is evidence (crystal structure studies: Cu²⁺, Zn²⁺, Cd²⁺) that four-membered rings form, i.e. there is a strong M²⁺ bond to N3 and a weak one to (C2)O. (2) By hydrogen bond formation to (C2)O of a metal ion-bound water molecule, six-membered rings, so-called semichelates, may form. (3) For Ca²⁺ and Mg²⁺, and possibly Mn²⁺, and their Cyd complexes, six-membered chelates are also likely with (C2)O being inner-sphere (crystal structure) and N3 outer-sphere. (4) Finally, for these metal ions also complexes with a sole outer-sphere interaction may occur. All these types of chelates are expected to be in equilibrium with each other in solution, but, depending on the metal ion, either the one or the other form will dominate. Clearly, the cytidine residue is an ambivalent binding site which adjusts well to the requirements of the metal ion to be bound and this observation is of relevance for single-stranded nucleic acids and their interactions with metal ions. In addition, the anti–syn energy barrier has been estimated as being in the order of 6–7.5 kJ/mol for cytidine derivatives in aqueous solution at 25 °C.Abbreviations ADP^3– adenosine 5-diphosphate - AMP^2– adenosine 5-monophosphate - ATP^4– adenosine 5-triphosphate - CDP^3– cytidine 5-diphosphate - cl closed - CMP^2– cytidine 5-monophosphate - 3-CMP^2– cytidine 3-monophosphate - CTP^4– cytidine 5-triphosphate - Cyd cytidine - DNA deoxyribonucleic acid - I ionic strength - K_a acidity constant - K_I intramolecular equilibrium constant - L general ligand - M²⁺ general divalent metal ion - NTP^4– nucleoside 5-triphosphate - op open - oPyN ortho-aminopyridine-type ligand - PyN pyridine-type ligand - t-RNA transfer ribonucleic acid - Tu tubercidin (7-deazaadenosine)In honor of Professor Liang-Nian Ji on the occasion of his 70th birthday in friendship and with best wishes.     

Permeability of Plant Young Root Endodermis to Cu Ions and Cu-Citrate Complexes in Corn and Soybean

The non-selective apoplastic passage of Cu and Cu-citrate complexes into the root stele of monocotyledonous corn and dicotyledonous soybean was investigated using an inorganic-salt-precipitation technique. Either Cu ions or Cu-citrate complexes were drawn into root through the apoplast from the root growth medium, and K₄[Fe(CN)₆] was subsequently perfused through xylem vessels or the entire root cross section. Based on microscopic identification of the reddish-brown precipitates of copper ferrocyanide in the cell walls of the xylem of corn and soybean roots, Cu²⁺ passed through the endodermal barrier into the xylem of both species. When the solution containing 200 μM CuSO₄ and 400 μM sodium citrate (containing 199.98 μM Cu-citrate, 0.02 μM Cu²⁺) was drawn via differential pressure gradients into the root xylem while being perfused with K₄[Fe(CN)₆] through the entire root cross-section, reddish-brown precipitates were observed in the walls of the stele of soybean, but not corn root. However, when a CuSO₄ solution containing 0.02 or 0.2 μM free Cu²⁺ was used, no reddish-brown precipitates were detected in the stele of either of the two plants. Results indicated that endodermis was permeable to Cu-citrate complexes in primary roots of soybean, but not corn. The permeability of the endodermal barrier to the Cu-citrate complex may vary between dicotyledonous and monocotyledonous plants, which has considerable implications for chelant-enhanced phytoextraction.     

Development of a Multi-Species Biotic Ligand Model Predicting the Toxicity of Trivalent Chromium to Barley Root Elongation in Solution Culture

Little knowledge is available about the influence of cation competition and metal speciation on trivalent chromium (Cr(III)) toxicity. In the present study, the effects of pH and selected cations on the toxicity of trivalent chromium (Cr(III)) to barley (Hordeum vulgare) root elongation were investigated to develop an appropriate biotic ligand model (BLM). Results showed that the toxicity of Cr(III) decreased with increasing activity of Ca²⁺ and Mg²⁺ but not with K⁺ and Na⁺. The effect of pH on Cr(III) toxicity to barley root elongation could be explained by H⁺ competition with Cr³⁺ bound to a biotic ligand (BL) as well as by the concomitant toxicity of CrOH²⁺ in solution culture. Stability constants were obtained for the binding of Cr³⁺, CrOH²⁺, Ca²⁺, Mg²⁺ and H⁺ with binding ligand: log K_CrBL 7.34, log K_CrOHBL 5.35, log K_CaBL 2.64, log K_MgBL 2.98, and log K_HBL 4.74. On the basis of those estimated parameters, a BLM was successfully developed to predict Cr(III) toxicity to barley root elongation as a function of solution characteristics.     

Effects of aluminium on nitrate uptake and assimilation

A study was conducted to examine the hypothesis that the effects of external Al on NO₃^? uptake and assimilation depend upon the concentration of Al present. Young soybean seedlings [Glycine max (L.) Merrill, cv. Essex], growing under moderate acidity stress at pH 4-2, were exposed to a range of {A1³⁺} in solution for 3d, and to labelled 99 atom %¹⁵NO₃^? during the final hour of Al exposure. Uptake of ¹⁵NO₃^?g^?1 root dry weight was increased by about 28% in the presence of Al at {A1³⁺} below 10 mmolm^?3, and NO₃^? uptake was decreased by about 12% when the {A1³⁺} increased to 44mmoln^?3. The stimulation phase closely paralleled stimulation of root elongation. At higher {A1³⁺}, the inhibition of root elongation was much more severe than that of NO₃^? uptake. There was no indication of a separate effect of Al on root ¹⁵NO₃^? reduction in situ, as the accumulation of reduced ¹⁵N in the root remained a similar percentage of ¹⁵NO₃^? uptake at all {A1³⁺}. At higher {A1³⁺}, the atom %¹⁵N enrichment of the insoluble reduced-N (protein) fraction of root tips increased. This suggests that the Al inhibition of root elongation did not result from disruption of the N supply to the root apex.     

Plant cell responses to heavy metals: molecular and physiological aspects

The effect of lead, cadmium and cooper on protein pattern, free radicals and antioxidant enzymes in root of Lupinus luteus L. were investigated. Heavy metals inhibited growth of lupin roots, which was accompanied by increased synthesis and accumulation of a 16 kDa polypeptide (Przymusiński et al. 1991 Biochem. Physiol. Pflanzen., 187:51–57). This component has been earlier identified as immunologically related to Cu,Zn-superoxide dismutase (Przymusiński et al. 1995 Env.Exp.Bot., 35:485–495). However, more detailed study revealed that this stress-stimulated protein is composed of four to six polypeptides of different electrophoretic mobility. The most abundant polypeptides of the 16kDa region were found to be closely homologous to pathogen related proteins. The number and intensity of these polypeptides was highly variable in roots of individual seedlings, which suggests that they might represent separate allelic forms. Electron paramagnetic spectra revealed that at low lead concentrations the amplitude of the first derivative was similar to the control and distinctly increased at higher metal concentrations. On the other hand, at the lower lead concentrations the activity of antioxidant enzymes increased, whereas at higher metal doses the enzyme activities did not raise further (SOD) or even dropped (CAT, APOX). This implies that the responses of antioxidant system to lead is dose-dependent stimulated by low metal concentrations, whereas at the higher metal level the free radical emission is beyond the quenching capacity of antioxidant enzymes, which in turn might contribute to the reduced root growth. The effect of various heavy metals: Pb²⁺, Cd²⁺ and Cu²⁺ on phytochelatins and antioxidant enzymes depends on the kind of metal ion. Pb²⁺ and Cd²⁺ stimulated the PCs formation whereas Cu²⁺ was not effective. On the other hand, in root exposed to Cu the activity of catalase (CAT) was the highest as was the production of H₂O₂. The strong oxidative effect of Cu²⁺ ions which were not complexed by PCs suggests that these peptides might by involved in the cellular defense system by binding excessive heavy metal ions. On the basis of our results it can be concluded that in lupin roots exposed to heavy metals there is a complex defense system against metal phytotoxicity, which comprises of specific proteins, antioxidant enzymes and phytochelatins.     

Complexation of oxoanions and cationic metals by the biscatecholate siderophore azotochelin

Azotochelin is a biscatecholate siderophore produced by the nitrogen-fixing soil bacterium Azotobacter vinelandii. The complexation properties of azotochelin with a series of oxoanions [Mo(VI), W(VI) and V(V)] and divalent cations [Cu(II), Zn(II), Co(II) and Mn(II)] were investigated by potentiometry, UV–vis and X-ray spectroscopy. Azotochelin forms a strong 1:1 complex with molybdate (log K = 7.6 ± 0.4) and with tungstate and vanadate; the stability of the complexes increases in the order Mo < V < W (log K _app^Mo = 7.3 ± 0.4; log K _app^V = 8.8 ± 0.4 and log K _app^W = 9.0 ± 0.4 at pH 6.6). The Mo atom in the 1:1 Mo–azotochelin complex is bound to two oxo groups in a cis position and to the two catecholate groups of azotochelin, resulting in a slightly distorted octahedral configuration. Below pH 5, azotochelin appears to form polynuclear complexes with Mo in addition to the 1:1 complex. Azotochelin also forms strong complexes with divalent metals. Of the metals studied, Cu(II) binds most strongly to azotochelin , followed by Zn(II) , Mn(II) and Co(II) . Since very few organic ligands are known to bind strongly to oxoanions (and particularly molybdate) at circumneutral pH, the unusual properties of azotochelin may be used for the separation and concentration of oxoanions in the laboratory and in the field. In addition, azotochelin may prove useful for the investigation of the biogeochemistry of Mo, W and V in aquatic and terrestrial systems. Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users.     

Photooxidation of wetland and riverine dissolved organic matter: altered copper complexation and organic composition

In natural waters, the uptake of transition metals such as copper (Cu) by aquatic biota depends on the activity of the free cupric ion ({Cu²⁺}) rather than on total Cu concentration. Thus, an important ecological function of dissolved organic matter (DOM) in aquatic ecosystems is Cu–DOM complexation, which greatly decreases the {Cu²⁺}. However, Cu bioavailability is greatly modified by source and environmental history of DOM because DOM affinity for Cu varies by orders of magnitude among DOM sources; moreover, DOM is photochemically unstable. During 72-h irradiation experiments at intensities approximating sunlight with DOM from a palustrine wetland and a third-order river, we investigated photooxidative effects on DOM complexation of Cu as well as spectral and chemical changes in DOM that might explain altered Cu complexation. Irradiation decreased Cu complexation by riverine DOM, but unexpectedly increased Cu complexation by wetland DOM, resulting in 150% greater {Cu²⁺} in riverine DOM at the same dissolved organic carbon concentrations. The specific ultraviolet absorption (SUVa) and humic substances tracked photochemical changes in the conditional stability constants of Cu–DOM complexes, suggesting that the aromaticity of DOM influences its affinity for Cu. Carbonyl concentration in ¹³C nuclear magnetic resonance spectra (¹³C-NMR) covaried directly with Cu binding-site densities in DOM. However, no aspect of Cu–DOM complexation consistently covaried with fluorophores (i.e., the fluorescence index) or low molecular weight organic acids. Our results suggest that global increases in UV radiation will affect Cu–DOM complexation and subsequent Cu toxicity depending on light regime as well as DOM source. Handling editor: K. Martens     

Enhanced sorption of Cu2+ and Ni2+ by acid-pretreated Chlorella vulgaris from single and binary metal solutions

The influence of HCl pretreatment (0.1 mM) on sorption ofCu²⁺ and Ni²⁺ by Chlorella vulgariswas tested using single and binary metal solutions. The optimal initial pH forsorption was 3.5 for Cu²⁺ and 5.5 for Ni²⁺. Second orderrate kinetics described well sorption by untreated and acid-pretreated cells.The kinetic constant q_e (metal sorption at equilibrium) for sorptionof test metals from single and binary metal solutions was increased afterpretreatment of the biomass with HCl. The Langmuir adsorption isotherm wasdeveloped for describing the various results for metal sorption. In single metalsolution, acid pretreatment enhanced q_max for Cu²⁺ andNi²⁺ sorption by approximately 70% and 65%, respectively.Cu²⁺ and Ni²⁺ mutually interfered with sorption of theother metal in the binary system. The combined presence of Cu²⁺ andNi²⁺ led to their decreased sorption by untreated biomass by 19% and88%, respectively. However, acid-pretreated biomass decreased Cu²⁺and Ni²⁺ sorption by 15 and 22%, respectively, when both the metalswere present in the solution. The results suggest a reduced mutual interferencein sorption of Cu²⁺ and Ni²⁺ from the binary metal systemdue to the acid pretreatment. Acid-pretreated cells sorbed twice the amount ofCu²⁺ and ten times that of Ni²⁺ than the untreated biomassfrom the binary metal system. Acid pretreatment more effectively enhanced thesorption of Ni²⁺ form the binary metal solution. The total metalsorption by untreated and acid-pretreated biomass depended on theCu²⁺ : Ni²⁺ ratio in the binary metal system. Acidpretreatment of C. vulgaris could be an effective andinexpensive strategy for enhancing Cu²⁺ and Ni²⁺ sorptionfrom single and binary metal solutions.     

Purification and properties of <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis><Emphasis Type="Italic">S</Emphasis>-adenosylmethionine synthetase expressed in recombinant <Emphasis Type="Italic">Pichia pastoris</Emphasis>

An intracellular S-adenosylmethionine synthetase (SAM-s) was purified from the fermentation broth of Pichia pastoris GS115 by a sequence chromatography column. It was purified to apparent homogeneity by (NH₄)₂SO₄ fractionation (30–60%), anion exchange, hydrophobic interaction, anion exchange and gel filtration chromatography. HPLC showed the purity of purified SAM-s was 91.2%. The enzyme was purified up to 49.5-fold with a final yield of 20.3%. The molecular weight of the homogeneous enzyme was 43.6 KDa, as determined by electro-spray ionization mass spectrometry (ESI-MS). Its isoelectric point was approximately 4.7, indicating an acidic character. The optimum pH and temperature for the enzyme reaction were 8.5 and 35 °C, respectively. The enzyme was stable at pH 7.0–9.0 and was easy to inactivate in acid solution (pH ≤ 5.0). The temperature stability was up to 45 °C. Metal ions, such as, Mn²⁺ and K⁺ at the concentration of 5 mM had a slight activation effect on the enzyme activity and the Mg²⁺ activated the enzyme significantly. The enzyme activity was strongly inhibited by heavy metal ions (Cu²⁺ and Ag²⁺) and EDTA. The purified enzyme from the transformed Pichia pastoris synthesized S-adenosylmethionine (SAM) from ATP and l-methionine in vitro with a K _m of 120 and 330 μM and V _max of 8.1 and 23.2 μmol/mg/min for l-methionine and ATP, respectively.     

Effects of Metal Combinations on the Production of Phytochelatins and Glutathione by the Marine Diatom Phaeodactylum tricornutum

Copper, Cd and Zn can be found at elevated concentrations in contaminated estuarine and coastal waters and have potential toxic effects on phytoplankton species. In this study, the effects of these metals on the intracellular production of the polypeptides phytochelatin and glutathione by the marine diatom Phaeodactylum tricornutum were examined in laboratory cultures. Single additions of Cu and Cd (0.4 μM Cu² and 0.45 μM Cd²⁺) to the culture medium induced the production of short-chained phytochelatins ((γ-Glu-Cys)_n-Gly where n = 2–5), whereas a single addition of Zn (2.2 μM Zn²⁺) did not stimulate phytochelatin production. Combination of Zn with Cu resulted in a similar phytochelatin production compared with a single Cu addition. The simultaneous exposure to Zn and Cd led to an antagonistic effect on phytochelatin production, which was probably caused by metal competition for cellular binding sites. Glutathione concentrations were affected only upon exposure to Cd (85% increase) or the combination of Cd with Zn (65% decrease), relative to the control experiment. Ratios of phytochelatins to glutathione indicated a pronounced metal stress in response to exposures to Cu or Cd combined with Zn. This study indicates that variabilities in phytochelatin and glutathione production in the field can be explained in part by metal competition for cellular binding sites.     

Effect of Cu Toxicity on Growth of Cowpea (Vigna unguiculata)

Accurate determination of the rhizotoxicity of Cu in dilute nutrient solutions is hindered by the difficulty of maintaining constant, pre-determined concentrations of Cu (micromolar) in solution. The critical Cu²⁺ activity associated with a reduction in the growth of solution-grown cowpea (Vigna unguiculata (L.) Walp. cv. Caloona) was determined in a system in which Cu was maintained constant through the use of a cation exchange resin. The growth of roots and shoots was found to be reduced at solution Cu²⁺ activities ≥1.7 μM (corresponding to 90% maximum growth). Although root growth was most likely reduced due to a direct Cu²⁺ toxicity, it is considered that the shoot growth reduction is attributable to a decrease in tissue concentrations of K, Ca, Mg, and Fe and the formation of interveinal chlorosis. At high Cu²⁺ activities, roots were brown in color, short and thick, had bent root tips with cracking of the epidermis and outer cortex, and had local swellings behind the roots tips due to a reduction in cell elongation. Root hair growth was reduced at concentrations lower than that which caused a significant reduction in overall root fresh weight.