Metabolism-independent binding of toxic metals by Ulva lactuca: cadmium binds to oxygen-containing groups, as determined by NMR

The metabolism-independent metal binding characteristics of Ulva lactuca were investigated using both freeze-dried thalli and cell walls stripped of intracellular material by incubation in Triton-X followed by methanol. Biosorption of Cd, Zn, Cu and Co by freeze-dried thallus was concentration-dependent and followed Freundlich and Langmuir isotherms. The Freundlich plot suggested that freeze-dried U. lactuca had the greatest binding affinity for Cu compared with Cd, Zn and Co. The BET (Brunauer–Emmett–Teller) plot, which indicates a more complex form of adsorption, and the Scatchard plot were not adequate models for Cu adsorption. The Scatchard plot of Cd suggested that two Cd binding sites were available on the freeze-dried thallus, with the second, lower affinity site only becoming available at Cd loading capacities greater than 4.9mmol g dry wt. Cd nuclear magnetic resonance (NMR) studies confirmed that two binding sites were available for Cd on the freeze-dried algal powder, though only one was available on the cell wall, and that the affinity of the binding sites was greater for Cu than for Cd. The results of the NMR experiments suggested that Cd binds to oxygen-containing functional groups in the algal powder and on the cell wall. It is proposed that sulphate or hydroxyl groups attached to polysaccharide subunits are possible sites.     

Ulva lactuca as bioindicator of metal contamination in intertidal waters in Hong Kong

The use of Ulva lactuca L. as an indicator of metal contamination was assessed by analysing the levels of Mn, Fe, Ni, Cu, Zn, Cd and Pb in the alga collected from 24 intertidal sites around the Island of Hong Kong. Twelve of the sites are in the rural southern parts of the Island where the coastal waters are relatively clean. The remaining 12 sites are located in the north and within Victoria Harbour which receives, apart from industrial effluents, untreated domestic sewage from a population of some 3.5 million. The mean levels of Mn, Fe, Ni, Cu, Zn and Pb in Ulva from the urban sites were respectively 4.0, 4.6, 1.8, 2.3, 2.4 and 4.6 folds those from the rural sites. However, similar levels of Cd were found in the alga amongst all the sites. Locations of high levels of metal contamination, particularly to the eastern end of the Harbour, have been identified. Preliminary results indicate that Ulva is a good indicator of Mn, Fe, Cu, Zn and Pb contaminations.     

Ecological studies on Ulva lactuca L. from Veraval,India

Ulva lactuca L., at Veraval (20° 54N and 70° 22E) on the western coast of India, grows in the intertidal belt from June to late February. During the summer months of March, April and May, the species dries up, leaving rhizoidal fragments for perennation. High values of density, phytomass, frequency, weight, and volume have been recorded during September and January. Accumulation of phytomass appears to be chiefly density dependent. High dissolved oxygen content and low temperatures of the surface sea water favoured enhanced growth and, consequently, more phytomass.     

Chemical and physical-chemical characteristics of dietary fibres from Ulva lactuca (L.) Thuret and Enteromorpha compressa (L.) Grev.

Dietary fibres from Ulva lactuca (L.) Thuret (sea lettuce) and Enteromorpha compressa (L.) Grev. (A.O. nori) were measured according to a ‘standard’ method and a ‘physiological’ protocol simulating the gastric and intestinal environments. U. lactuca contained 15.8–8.0% soluble and 24.2–32.6% insoluble fibres according to the ‘standard’ and ‘physiological’ methods, respectively. For E. compressa, these values were 14.9–15.9 and 21.6–28.7%, respectively. For both algae, the composition suggests that the soluble fibres were xylorhamnoglycuronans sulphates and insoluble fibres were essentially composed of glucans. No marked chemical compositional variation was observed between soluble fractions extracted under the simulated gastric and intestinal conditions. Fibres in both algae are hydrophilic but the water holding capacities were higher after extraction of soluble fibres (5.5–9.5 g g⁻¹ for the dry algae; 14.0–16.0 g g⁻¹ for the standard insoluble fibres). Water soluble fibres demonstrated low intrinsic viscosities at 37 °C in buffers, particularly those from E. compressa (36.0–36.5 ml g⁻¹), and was affected by pH for those of U. lactuca (147.5 ml g⁻¹ at pH 3.0 and 175.0 ml g⁻¹ at pH 7.3).     

A novel three-stage seaweed (Ulva lactuca) biofilter design for integrated mariculture

Seaweed biofilters have proven their usefulness in the treatment of fishpond effluents. However, their performance poses a dilemma: TAN (Total Ammonia N) uptake rate – and with it seaweed yield and protein content – is inversely proportional to TAN uptake efficiency. The ideal for a seaweed biofilter performance would be a high uptake rate together with high uptake efficiency. The novel three-stage seaweed biofilter design described here has solved this dilemma. The design used the finding that the performance of seaweed ponds depended on the flux of TAN through them, and that therefore effluents with reduced TAN concentration could provide the seaweed with a high TAN flux if the water flow increased proportionally. Effluents from a seabream fishpond were passed through a series of three successively smaller (25, 12.5 and 6.25 m², respectively) air-agitated Ulva lactuca ponds. The diminished inflow TAN concentrations to the second and third ponds of the biofilter system were compensated for by the increased water exchange rates, inversely proportional to their sizes. The biofilter performance was evaluated under several TAN loads. TAN was efficiently removed (85–90%), at a high areal rate (up to 2.9 g N m^-2 d^-1) while producing high protein U. lactuca (up to 44% dw) in all three stages, although with mediocre yields (up to 189 g fresh m^-2 d^-1). Performance of each seaweed biofilter pond correlated not with TAN concentration, but with areal TAN loads. The novel three-stage design provides significant functional and economic improvements in seaweed biofiltration of intensive fishpond water.     

Mineral element content in Ulva lactuca L. with reference to eutrophication in Hong Kong coastal waters

The amounts of tissue nitrogen, phosphorus, potassium, sodium, calcium and iron were estimated in the green alga Ulva lactuca L. collected from 9 rural and 14 urban littoral sites in the waters around Hong Kong Island during 1978 and 1979. The mean levels of tissue nitrogen and phosphorus were respectively 65% and 87% more in urban sites than in rural ones. Very significant correlation (r = 0.920) was found between the logarithmic concentration of seawater inorganic nitrogen and that of tissue nitrogen. The same applied to soluble reactive phosphorus in seawater and tissue phosphorus (r = 0.886). The levels of potassium, sodium and calcium in the alga were relatively uniform. However considerable variation in the level of iron was detected. The potential use of Ulva as an indicator species for eutrophication is discussed.     

The concentration of cadmium,lead, copper and zinc inChironomus gr.thummi larvae (Diptera,Chironomidae) with deformedversus normal menta

The concentrations of Cd, Pb, Cu and Zn inChironomus gr.thummi were determined for 4th instar larvae from the polluted Dyle River, tributary of the Scheldt River (Belgium). Comparison was made between larvae with deformed and normal menta. Deformed larvae showed higher overall metal concentrations than normal larvae. Especially Pb and Cu had higher concentrations in deformed larvae (16.22 mg kg^–1 dry weight and 39.66 respectively) than in normal larvae (12.80 mg kg^–1 dry weight and 35.70 respectively). No significant differences were found in the concentrations of Cd and Zn (mean [Cd] = 0.81 mg kg^–1 dry weight and mean [Zn] = 313.12 mg kg^–1 dry weight). There was no difference between the two larval groups as far as total length, dry weight and developmental stage of the imaginal discs are concerned.     

Cell wall structure of the agarophytes Gracilaria tikvahiae and G. cornea (Rhodophyta) and penetration by the epiphyte Ulva lactuca (Chlorophyta)

Use of light, transmission, and scanning electronmicroscopes revealed that the epidermal cell wall ofthe red algal agarophytes Gracilaria tikvahiaeMcLachlan and G. cornea J. Agardh consists of adecklamelle and outer and inner wall layers. The twospecies differed, with G. cornea having asignificantly thicker outer wall and a more diffusedecklamelle. After induction, the zooids of Ulvalactuca would attach to glass slides and the twospecies of Gracilaria via an adhesion pad. Within a few days, 3–5 celled germlings penetrated thedecklamelle and outer wall layer of both basiphytes. By the time the epiphyte germlings reached the 15celled stage, they had penetrated the inner walllayer. The differences in epidermal cell wallconstruction between the two basiphytes may play arole in the ability of zooids of U. lactuca toattach in nature where epiphytization of G.cornea is infrequent.     

Electrospray ionization mass spectrometry of zinc, cadmium, and copper metallothioneins: evidence for metal-binding cooperativity

Electrospray ionization (ESI) mass spectra of both well-characterized and novel metallothioneins (MTs) from various species were recorded to explore their metal-ion-binding modes and stoichiometries. The ESI mass spectra of the zinc- and cadmium-binding MTs showed a single main peak corresponding to metal-to-protein ratios of 4, 6, or 7. These findings combined with data obtained by other methods suggest that these MTs bind zinc or cadmium in a single predominant form and are consistent with the presence of three- and four-metal clusters. An unstable copper-specific MT isoform from Roman snails (Helix pomatia) could be isolated intact and was shown to preferentially bind 12 copper ions. To obtain additional information on the formation and relative stability of metal-thiolate clusters in MTs, a mass spectrometric titration study was conducted. One to seven molar equivalents of zinc or of cadmium were added to metal-free human MT-2 at neutral pH, and the resulting complexes were measured by ESI mass spectrometry. These experiments revealed that the formation of the four-metal cluster and of the thermodynamically less stable three-metal cluster is sequential and largely cooperative for both zinc and cadmium. Minor intermediate forms between metal-free MT, Me4MT, and fully reconstituted Me7MT were also observed. The addition of increasing amounts of cadmium to metal-free blue crab MT-I resulted in prominent peaks whose masses were consistent with apoMT, Cd3MT, and Cd6MT, reflecting the known structure of this MT with two Me3Cys9 centers. In a similar reconstitution experiment performed with Caenorhabditis elegans MT-II, a series of signals corresponding to apoMT and Cd3MT to Cd6MT species were observed.     

Maternal and neonatal scalp hair concentrations of zinc, copper, cadmium, and lead

Postpartum scalp hair samples from 82 term-pregnancy mother/neonate pairs were analyzed for their concentration of zinc (Zn), copper (Cu), cadmium (Cd), and lead (Pb), using inductively coupled plasma-mass spectrometry. Maternal and neonatal Zn concentrations had geometric means (and 99% confidence intervals) of 122.5 μg/g (117.9–131.5 μg/g) and 146.9 μg (141.5–156.7 μg/g) respectively. Corresponding Cu values were 18.4 μg/g (17.6–23.8 μg/g) and 6.7 μg/g (6.3–7.6 μg/g). Those of Cd were 0.49 μg/g (0.47–0.69 μg/g) in the mothers and 0.57 μg/g (0.55–0.86 μg/g) in the neonates. For Pb, they were 7.95 μg/g (7.60–9.32 μg/g) and 4.56 μg/g (4.39–5.56 μg/g). Cigaret smoking, despite its relatively low prevalence (19.5%), was associated with lower Zn and higher Cd and Pb concentrations and in lower Zn/Cd and Zn/Pb molar concentration ratios. Smoking also altered interelemental relationships, particularly those of Zn with Cd and Pb and those between Cd and Pb. Smoking frequency appeared to show negative dose-response effects on maternal and neonatal Zn concentrations, Zn/Pb molar concentration ratios, and birth weight. Mothers with a history of oral contraceptive (OC) usage had significantly higher Cu concentrations and lower Zn/Cu molar concentration ratios than nonusers, with the highest Cu concentrations and lowest Zn/Cu values being associated with third-generation OCs. No similar effects were elicited in the respective neonatal Cu concentrations. Neither alcohol consumption nor prenatal supplementation with iron and/or folic acid had discernible effects on the maternal or neonatal elemental concentrations. The data from this study suggest that in a given population of term-pregnancy mothers and neonates, significant interindividual variations in hair trace element concentrations can occur, irrespective of commonality of general environment, and that lifestyle factors, including cigaret smoking and OC usage history, can be significant contributory factors to such variations. The data are discussed in relation to the effects of smoking-associated exposure to Cd and Pb exposure on Zn availability for placental transfer, as well as on the quantitative maternal Zn supply levels to the fetus resulting from the known tendency of smokers to have lower dietary intakes of Zn. The higher Cu concentrations in OC users are discussed in relation to altered Cu metabolism, characterized by increased synthesis of the Cu-binding protein, ceruloplasmin, as an acute-phase antioxidant response to altered lipid profile and increased lipid oxidation.     

Repeated removal of cadmium and zinc from an industrial effluent by waste biomass Sargassum sp.

Waste biomass Sargassum sp. biosorbed 100% of Cd²⁺ and 99.4% of Zn²⁺ from a 3 and 98 mg l^–1 solution (pH 4.5), respectively, at the end of four serial experiments. Of the five desorbents studied in consecutive adsorption/desorption cycles, CaCl₂ 0.05 M eluted nearly 40% of both metals and decreased the biosorption in only 8% and 17% of Cd²⁺ and Zn²⁺, respectively. Although NaOH desorbent improved the heavy metal uptake from the second cycle onwards, it did not elute metals from the pre-loaded biomass.     

Changes in selenium, zinc, copper and cadmium contents in human milk during the time when selenium has been supplemented to fertilizers in Finland

Sodium selenate has been supplemented to all agricultural fertilizers used in Finland since 1984. We followed the changes in selenium, cadmium, zinc and copper content in Finnish human milk between the years 1987 and 1993-1995. A total of 257 milk samples was collected, four weeks after delivery, in two areas: In Helsinki, an urban area, and in Kuopio, a rural area, where elevated copper concentrations have been found in the bedrock. Direct atomic absorption spectrophotometric methods without digestion were used for the analyses. The dependence of trace element content on study time, living area, smoking habits, fish eating frequency, and parity of mothers was studied by analysis of covariance. Inter-element correlations and correlations with mothers' age and fat content in milk were studied by partial correlation. Significant increases were observed in mean selenium (16.4 microg/l and 18.9 microg/l, p < 0.001) and in fat contents (3.4% and 4.0%, p < 0.001), whereas significant decreases were seen in mean zinc (3.00 mg/l and 1.47 mg/l, p < 0.001), copper (0.52 mg/l and 0.43 mg/l, p < 0.001) and cadmium contents (0.095 microg/l and 0.062 microg/l, p < 0.01). In 1987, zinc had a positive correlation with copper and fat. Copper correlated inversely with the mothers' age. In 1993-1995, selenium correlated positively with copper, and zinc correlated inversely with mothers' age. Mothers living area had an effect on copper content in milk. Our results confirm that selenium supplementation to fertilizers in Finland has increased the selenium level in human maternal milk and most likely it also has an effect on the zinc and copper concentrations in maternal milk.     

Valve closure responses of the Asiatic clam Corbicula fluminea exposed to cadmium and zinc

The valve movement patterns of immobilized Asiatic clams (Corbicula fluminea) were monitored during exposure to constant concentrations of cadmium (0.0, 0.1, 0.2, 0.3, and 0.4 mg 1^–1) or zinc (0.0, 0.1, 0.3, 0.5, and 0.9 mg 1^–1) for 24 h following a 24-h acclimation period. Data indicate that the duration of response was concentration dependent and toxicity related. Durations of periods with valves parted declined as the concentration of heavy metal increased. Behavior was consistent for both mean time to first closure following the initial exposure and mean time per valve parting episode over a 24-h exposure period. Mean time per valve parting episode during the 24-h exposure period ranged from 600 minutes for control trials to 36 and 69 minutes for the highest concentrations of cadmium and zinc tested, respectively. There was no association between durations of periods with valves sealed and solutions of cadmium; however, lengthening durations of periods with valves sealed coincided with exposure of clams to progressively more concentrated zinc solutions. In addition, Asiatic clams demonstrated a greater rate of response (decline in the duration of periods with valves parted) to progressively more concentrated solutions of cadmium than to comparable increases in the concentrations of zinc solutions.     

Uptake of lead,cadmium and zinc by the fairy shrimp,Branchinecta longiantenna (Crustacea: Anostraca)

Individuals of the fairy shrimp, Branchinecta longiantenna, were subjected to 5 concentrations (0.1 to 15 mg l^–1) of Pb in diluted habitat water at 13 °C. Lead concentrations (mg kg^–1 wet weight) in the animals were determined at 2-day intervals by digestion in nitric acid followed by atomic absorption analysis. The shrimp were also subjected to 0.1 mg l^–1 media of Cd and Zn, separately.Uptake rates by the fairy shrimp for the three metal ions at 0.1 mg l^–1 were: 0.111, 0.0885, and 0.0460 mg kg^–1 day^–1 for Zn, Pb, and Cd, respectively. After 2 days in 1.0 mg l^–1 Cd or Zn, the animals expired; but they surviced for 8 days in a 10 mg l^–1 Pb medium and for 2 days in 25 mg l^–1 Pb. Lead uptake demonstrated a linear dependence on the Pb concentration in the media.Shrimp survived at much higher tissue accumulations of Pb compared to Zn and Cd. Estimated lethal doses were 20, 1.2–2.4, and 0.4–1.4 mg kg^–1 wet weight for Pb, Zn, and Cd, respectively. Pb was found to be at much lower concentration than Cd or Zn in the natural pond water but between Cd and Zn levels in the sediment. Thus Cd and Zn probably present a greater threat to B. longiantenna than Pb, although Pb may be in higher concentration in the environment.Contribution 47, Laboratory of Ecology, The Claremont Colleges, Claremont, CA 91711, USA. Send reprint requests to Clyde Eriksen.     

Comparative strategies of heavy metal accumulation by crustaceans: zinc,copper and cadmium in a decapod,an amphipod and a barnacle

This study investigates the comparative strategies of accumulation under standardised laboratory conditions of the essential metals zinc and copper, and the non-essential metal cadmium by three crustaceans of different taxa; vizPalaemon elegans Rathke (Malacostraca: Eucarida: Decapoda),Echinogammarus pirloti (Sexton & Spooner) (Malacostraca: Peracarida: Amphipoda) and the barnacleElminius modestus Darwin (Cirripedia: Thoracica).The decapodP. elegans regulates body zinc concentrations to a constant level (ca. 79 µg Zn g^–1) over a wide range of dissolved metal availabilities until regulation breaks down at high Zn availabilities and net accumulation begins. The amphipodE. pirloti accumulates zinc at all dissolved zinc concentrations but at a low net rate such that the accumulation strategy approaches that of regulation. The barnacleE. modestus accumulates zinc to high body concentrations with no significant excretion of accumulated zinc. In the case of copper,P. elegans similarly regulates body copper concentrations to a constant level (ca. 129 µg Cu g^–1) over a range of dissolved copper availabilities until regulation breaks down at high copper concentrations. Both the amphipodE. pirloti and the barnacleE. modestus on the other hand accumulate copper at all dissolved copper exposures with no evidence of regulation. All three crustaceans accumulate the non-essential metal cadmium at all dissolved cadmium concentrations without regulation.Heavy metal accumulation strategies therefore vary between crustacean taxa and between metals. Uptake rates for zinc and cadmium have been estimated for the three crustaceans and can be interpreted in terms of cuticle permeability and way of life of each crustacean. Examination of these uptake rates provides an insight into possible reasons behind the adoption of particular metal accumulation strategies.     

Effect of chronic exposure to cadmium and copper on Asellus aquaticus (L.) (Crustacea,Isopoda)

The effects of chronic exposure to 5 g·1^–1 cadmium or copper on the crustacean Isopod Asellus aquaticus (L.) were studied by analyzing survival and body growth in the first stages of the life-cycle and by determining fecundity and survival of embryo-bearing females. Juveniles survival is differently affected by the two metals in that embryonic development is more sensitive to cadmium while juvenile development is more sensitive to copper. Juvenile body growth is stimulated by cadmium and depressed by copper. Embryo-bearing female survival and fecundity are significantly reduced by cadmium but are not affected by copper. The consequence of environmental contamination by a sublethal cadmium or copper concentration is discussed.     

Assessing the potential for zinc and cadmium phytoremediation with the hyperaccumulator Thlaspi caerulescens

Thlaspi caerulescens is a Zn and Cd hyperaccumulator, and has been tested for its phytoremediation potential. In this paper we examine the relationships between the concentrations of Zn and Cd in soil and in T. caerulescens shoots, and calculate the rates of Zn and Cd extraction from soil. Using published data from field surveys, field and pot experiments, we show that the concentrations of Zn and Cd in the shoots correlate with the concentrations of Zn and Cd in soils in a log-linear fashion over three orders of magnitude. There is little systematic difference between different populations of T. caerulescens in the relationship between soil and plant Zn concentrations. In contrast, populations from southern France are far superior to those from other regions in Cd accumulation. Bioaccumulation factors (plant to soil concentration ratio) for Zn and Cd decrease log-linearly with soil metal concentration. Model calculations show that phytoremediation using T. caerulescens is feasible when soil is only moderately contaminated with Zn and Cd, and the phytoremediation potential is better for Cd than for Zn if the populations from southern France are used. Recent progress in the understanding of the mechanisms of Zn and Cd uptake by T. caerulescens is also reviewed.     

Comparison of different extractive procedures for proteins from the edible seaweeds Ulva rigida and Ulva rotundata

Proteins have been extracted from the edible seaweeds Ulva rigida Agardh and Ulva rotundata Bliding using classical or enzymatic procedures. The protocols using NaOH under reductive conditions or a two-phase system (PEG/K₂CO₃) produced the best protein yields. The cleavage or the limitation of the linkages between proteins and polysaccharides caused by these experimental conditions probably explains the efficiency of these protocols. In SDS PAGE, the protein fraction obtained after NaOH extraction from U. rotundata is characterised by the presence of three major bands with apparent molecular weights of 45 600, 31 800 and 18 600. The protein fraction from U. rigida presents two specific bands with apparent molecular weights of about 27 000 and 12 000. These fractions are mainly rich in aspartic and glutamic acids, alanine, glycine and contain few hydroxyproline residues (0.91–2.44% total amino acid content). The use of cellulase does not significantly improve the extraction of algal proteins in comparison with the blank procedure (without enzymes). The weak accessibility of the substrates in the intact cell wall could explain these experimental data. The improvement of protein yield after the use of the polysaccharidase mixture (-glucanase, hemicellulase, cellulase) partially confirms this hypothesis.     

Adaptive and cross resistance to cadmium (II) and zinc (II) by Pseudomonas aeruginosa BC15

Cadmium and zinc appear in the combined forms and they are co-pollutants. Cd is the most hazardous metal ion for human beings and causes renal dysfunction, liver and lungs damage, bone degeneration and blood damage. Though Zn is an essential nutrient, excess of Zn is toxic. Biological process was more important because conventional methods fail to remediate these pollutants due to high costs and less affordability. The screening and understanding of the functioning of microorganism plays an important role in removal and recovery of metals from heavy-metal-polluted water and soil. In our study, the strain Pseudomonas aeruginosa BC15 was isolated from oil-mill-treated waste water and it showed to be highly resistant to 6 mM Cd and 20 mM Zn in the solid and liquid media. The growth studies of BC15 strain in the medium without induction exhibited high tolerable capacity when compared to other microbes. Pretreatment of P. aeruginosa BC15 with sub-lethal concentrations of Cd induced adaptive resistance to lethal doses of Cd. Cadmium-induced cells also showed cross resistance to lethal concentration of zinc. The organism had high resistance against Cd and Zn. This has been clearly proven through biosorption studies: Cd was absorbed up to 62% and Zn about 60% in single solution, whereas in binary solution Cd was biosorbed up to 82% and Zn 85%. In conclusion, this study reveals the significance of using the strain P. aeruginosa BC15 in the bioremediation of Cd and Zn from industrial waste water and contaminated soil.     

Accumulation, depuration and distribution of cadmium and zinc in the green-lipped mussel Perna viridis (Linnaeus) under laboratory conditions

Ecotoxicological tests were conducted in the green-lipped mussel Perna viridis under laboratory conditions. Different rates of accumulation and depuration in soft tissues are found and this might be due to different mechanisms of metal binding and regulation. At the end of depuration, Cd levels in soft tissues of P. viridis were 10–30 times higher than before exposure, while Zn levels in soft tissues were almost similar to levels before exposure. These results indicate that P. viridis is a good biomonitoring organism for Cd but Zn levels might be actively regulated. It remains uncertain whether P. viridis is a good biomonitoring organism of environmental Zn contamination. However, the positive patterns, although different rates, of accumulation and depuration for Cd and Zn support the use of P. viridis as a biomonitoring agent for such metals.