Effects of high stability iron-complexes on the kinetics of iron accumulation and excretion in Mytilus edvlis (L.)

The effects of small molecular weight complexes with a high affinity for iron on the uptake, accumulation and excretion of iron by the common mussel, Mytilus edulis (L.) have been investigated. Fe(III) complexes of citrate, EDTA and 1,10-phenanthroline increased both the rate of uptake and the total amount accumulated when compared to those for particulate ferric hydroxide in sea water, whereas ferrichrome b and the Fe(III) complexes of aceto- and benzo-hydroxamic acids give a decrease. Increased uptake is, however, compensated for by an increased rate of excretion resulting in an almost constant residence time for this metal. The iron is accumulated principally in the viscera with a smaller but significant proportion in the gills. The effects produced by prior complexation of the iron cannot be correlated with either the strength of binding of the complex to the iron or the exchangeability of the iron with other ligands but may be concerned with the endocytosis, the mechanism of uptake for iron previously shown to occur in Mytilus.     

Le fer dans la branchie des crustacés décapodes Carcinus maenas (L.) et Homarus americanus Milne Edw.: étude quantitative et histochimique

Carcinus maenas (L.) gills accumulate iron. This iron occurs in a ferric and inorganic state. Up to 99.9 % of this metal is included in the chitinous-cellular debris fraction as isolated by centrifugation. The remaining iron belongs to the soluble protein and mitochondrial fractions. At the site of the iron polymucosaccharic acids and polysaccharides are also observed. The iron content of Homarus americanus Milne Edw. gills is smaller than that of Carcinus maenas gills. In Homarus americanus the iron is accumulated as scattered ‘tablets’ along the branchial tubes, while in Carcinus maenas it coats the branchial lamellae. It seems that the accumulation of iron in Carcinus maenas is related to the rôle of filtration or adsorption by the chitin against the particulate iron of the sea water. The differences observed between Carcinus maenas and Homarus americanus are probably related to morphological and structural differences.     

Cadmium-induced alterations in essential trace element homoeostasis in the tissues of scallop Mizuhopecten yessoensis

Studies were performed regarding the effect of cadmium accumulation on the levels of essential elements (copper, zinc and iron) in the tissues of a marine bivalve mollusc, Mizuhopecten yessoensis, exposed to cadmium at 250 ppb during 2 weeks. It was found that the concentration of cadmium in the tissues increased in the order gonads < gills < hepatopancreas < kidney during exposure time. However, the highest value of concentration factor was recorded in the gills. Our data demonstrate that cadmium accumulation in all mollusc tissues is followed by the alterations in copper, zinc and iron concentration, but that the pattern of these changes varies with each tissue. Cadmium had the most pronounced effects on essential trace element homoeostasis in the kidney. The present study suggests that levels of the essential metals in a particular tissue can be modified depending on the level of cadmium accumulation. The possible mechanisms of the effects of cadmium on the essential trace elements are discussed.     

金鱼(Carassius auratus)鳃对颗粒吸附态铅的吸收

研究了金鱼（Ｃａｒａｓｓｉｕｓ ａｕｒａｔｕｓ）鳃在悬浮水铝矿吸附态Ｐｂ暴露实验中对Ｐｂ吸收过程,结果表明,在保持水中游离态Ｐｂ浓度不变（０．１ｍｇ．Ｌ＾－１）、吸附态Ｐｂ浓度增加的条件下,鳃中Ｐｂ,Ａｌ浓度均随暴露浓度增加而增加,血液中Ｐｂ浓度随鳃中Ｐｂ浓度的增加率（直线斜率０．１７１）低于溶解态Ｐｂ、暴露实验的增加率（直线斜率０．３４３）,附着鳃上的部分Ｐｂ未移转至血液中,鳃中Ｐｂ／Ａｌ比较水     

The influence of phosphorus,zinc and manganese on absorption and translocation of iron in watercress

Summary Absorption and translocation of iron by intact watercress plants (Rorippa nasturtium-aquaticum (L) Hayek) was studied in short period uptake experiments utilising ⁵⁹Fe labelled ferric chloride. Total translocation of iron was inhibited by increasing levels of phosphorus, zinc and manganese in the nutrient medium; the elevated phosphorus and zinc concentrations enhanced iron absorption into roots, but increased retention of absorbed iron in translocating portions of the plant. High levels of manganese in the medium reduced the initial absorption of iron into the root system.     

DNA damage (comet assay) and 8-oxodGuo (HPLC-EC) in relation to oxidative stress in the freshwater bivalve Unio tumidus

The relationships between DNA damage and oxidative stress in the digestive gland, gills and haemocytes of the freshwater bivalve Unio tumidus were investigated. Two markers of genotoxicity were measured: DNA breaks by means of the comet assay, and oxidative DNA lesions by means of 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxodGuo) measured using high-performance liquid chromatography (HPLC) coupled to electrochemical detection. Lipid peroxidation was evaluated by measuring malondialdehyde (MDA) tissue levels. Effects were studied after exposure of bivalves for 6 days to benzo[a]pyrene (B[a]P) (50 and 100 μg?l^?1) and ferric iron (20 and 40 mg?l^?1), applied alone or in combination. Lipid peroxidation in the digestive gland and gills resulted from exposure to Fe³⁺ or B[a]P whatever the concentrations tested. DNA oxidatively formed lesions were induced in the two tissues at a higher level after B[a]P exposure than after Fe³⁺ treatment. No significant dose–response relationship was found with the two compounds and no synergistic effect was observed between Fe³⁺ and B[a]P. The gills appeared less sensitive than the digestive gland to DNA lesions expressed as 8-oxodGuo and comet results. Good correlations were noted between 8-oxodGuo and comet. MDA and DNA damage did not correlate as well, although it was stronger in the digestive gland than in the gills. Production of mucus by the gills likely served to prevent lesions by reducing the bioavailability of the chemicals tested, which could explain that dose–effect relationships and synergistic effects were not observed.     

Active Transport of Iron in Bacillus megaterium: Role of Secondary Hydroxamic Acids

Kinetics of radioactive iron transport were examined in three strains of Bacillus megaterium. In strain ATCC 19213, which secretes the ferric-chelating secondary hydroxamic acid schizokinen, ⁵⁹Fe³⁺ uptake from ⁵⁹FeCl₃ or the ferric hydroxamate Desferal-⁵⁹Fe³⁺ was rapid and reached saturation within 3 min. In strain SK11, which does not secrete schizokinen, transport from ⁵⁹FeCl₃ was markedly reduced; the two ferric hydroxamates Desferal-⁵⁹Fe³⁺ or schizokinen-⁵⁹Fe³⁺ increased both total ⁵⁹Fe³⁺ uptake and the ⁵⁹Fe³⁺ appearing in a cellular trichloroacetic acid-insoluble fraction, although 10 min was required to reach saturation. Certain characteristics of transport from both ferric hydroxamates and FeCl₃ suggest that iron uptake was an active process. The growth-inhibitory effect of aluminum on strain SK11 was probably due to the formation of nonutilizable iron-aluminum complexes which blocked uptake from ⁵⁹FeCl₃. Desferal or schizokinen prevented this blockage. A strain (ARD-1) resistant to the ferric hydroxamate antibiotic A22765 was isolated from strain SK11. Strain ARD-1 failed to grow with Desferal-Fe³⁺ as an iron source, and it was unable to incorporate ⁵⁹Fe³⁺ from this source. Growth and iron uptake in strain ARD-1 were similar to strain SK11 with schizokinen-Fe³⁺ or the iron salt as sources. It is suggested that the ferric hydroxamates, or the iron they chelate, may be transported by a special system which might be selective for certain ferric hydroxamates. Strain ARD-1 may be unable to recognize both the antibiotic A22765 and the structurally similar chelate Desferal-Fe³⁺, while retaining its capacity to utilize schizokinen-Fe³⁺.     

Chemotaxis of Pathogenic Vibrio Strains towards Mucus Surfaces of Gilt-Head Sea Bream (Sparus aurata L.)

Vibrio anguillarum and Vibrio alginolyticus exhibited significant adhesion to and chemotactic abilities towards mucus collected from the skin, gills, and intestine of gilt-head sea bream. Quadratic polynomial models for chemotaxis designed to estimate the influence of temperature demonstrated a differential bacterial chemotaxis depending of the source of the mucus, with the chemotaxis towards intestinal mucus being the least influenced.     

Isolation and determination of the ferric iron chelate of ethylenediamine di(o-hydroxyphenylacetic acid) in plant tissues

Summary A new technique for the extraction and quantitative determination of the ferric iron chelate of ethylenediamine di(o-hydroxyphenylacetic acid) from aqueous plant extracts is described. The metal chelate is salted out with ammonium sulfate and then is extracted into a small volume of n-amyl alcohol. Spectrophotometric data indicate that the extracted chelate has an absorption maximum corresponding to the iron chelate in aqueous solution and adheres to Beer's law in the range of 0 to 150 micrograms of the metal chelate. The chelate, being anionic in nature, can be absorbed and desorbed from an anion-exchange resin. Isolation and determination of the iron chelate in tomato tissues also is described. Recovery of the iron chelate from these tissues was 94 to 97 per cent.Paper No. 758, University of Arizona, Agr. Exp. Sta., Tucson, Arizona.     

Characterization of Phloem iron and its possible role in the regulation of fe-efficiency reactions

`Fe-efficiency reactions' are induced in the roots of dicotyledonous plants as a response to Fe deficiency. The role of phloem Fe in the regulation of these reactions was investigated. Iron travels in the phloem of Ricinus communis L. as a complex with an estimated molecular weight of 2400, as determined by gel exclusion chromatography. The complex is predominantly in the ferric form, but because of the presence of reducing compounds in the phloem sap, there must be a fast turnover in situ between ferric and ferrous (k ≈ 1 min⁻¹). Iron concentrations in R. communis phloem were determined colorimetrically or after addition of ⁵⁹Fe to the nutrient solution. The iron content of the phloem in Fe-deficient plants was lower (7 micromolar) than in Fe-sufficient plants (20 micromolar). Administration of Fe-EDTA to leaves of Phaseolus vulgaris L. increased the iron content of the roots within 2 days, and decreased proton extrusion and ferric chelate reduction. The increase in iron content of the roots was about the same as the difference between iron contents of roots grown on two iron levels with a concomitantly different expression of Fe-efficiency reactions. We conclude that the iron content of the leaves is reflected by the iron content of the phloem sap, and that the capacity of the phloem to carry iron to the roots is sufficient to influence the development of Fe-efficiency reactions. This does not preclude other ways for the shoot to influence these reactions.     

Selectivity of Ferric Enterobactin Binding and Cooperativity of Transport in Gram-Negative Bacteria

The ligand-gated outer membrane porin FepA serves Escherichia coli as the receptor for the siderophore ferric enterobactin. We characterized the ability of seven analogs of enterobactin to supply iron via FepA by quantitatively measuring the binding and transport of their ⁵⁹Fe complexes. The experiments refuted the idea that chirality of the iron complex affects its recognition by FepA and demonstrated the necessity of an unsubstituted catecholate coordination center for binding to the outer membrane protein. Among the compounds we tested, only ferric enantioenterobactin, the synthetic, left-handed isomer of natural enterobactin, and ferric TRENCAM, which substitutes a tertiary amine for the macrocyclic lactone ring of ferric enterobactin but maintains an unsubstituted catecholate iron complex, were recognized by FepA (K_d ≈ 20 nM). Ferric complexes of other analogs (TRENCAM-3,2-HOPO; TREN-Me-3,2-HOPO; MeMEEtTAM; MeME-Me-3,2-HOPO; K₃MECAMS; agrobactin A) with alterations to the chelating groups and different net charge on the iron center neither adsorbed to nor transported through FepA. We also compared the binding and uptake of ferric enterobactin by homologs of FepA from Bordetella bronchisepticus, Pseudomonas aeruginosa, and Salmonella typhimurium in the native organisms and as plasmid-mediated clones expressed in E. coli. All the transport proteins bound ferric enterobactin with high affinity (K_d ≤ 100 nM) and transported it at comparable rates (≥50 pmol/min/10⁹ cells) in their own particular membrane environments. However, the FepA and IroN proteins of S. typhimurium failed to efficiently function in E. coli. For E. coli, S. typhimurium, and P. aeruginosa, the rate of ferric enterobactin uptake was a sigmoidal function of its concentration, indicating a cooperative transport reaction involving multiple interacting binding sites on FepA.     

In vivo and in vitro effects of cadmium on selected enzymes in different organs of the fish Barbus conchonius ham. (Rosy Barb)

1. Enzyme modulation by cadmium in selected organs of the fish, Barbus conchonius (rosy barb), was investigated in vivo (48 hr exposure to 12.6 mg/1 cadmium chloride) and in vitro (10⁻⁶M cadmium chloride).2. The acetylcholinesterase (AchE) activity was depressed in the gills but stimulated in the skeletal muscles and brain in vivo. The hepatic, branchial, and renal acid phosphatase (AcP) activity decreased marginally in vivo but it was significantly increased in the gut and ovary. In vitro, except for the liver, the AcP activity was depressed in the selected organs. Collaterally, gut alkaline phosphatase (A1P) was significantly inhibited but a pronounced stimulation was noted in the kidneys and ovary in vivo. In vitro, the AIP activity was conspicuously elevated in the kidneys and gut, and moderately in the gills.3. Cadmium inhibited the glutamate-oxaloacetate and glutamate-pyruvate transaminases (GOT and OPT) in the liver, gills and kidneys in vivo. In vitro, the GOT and GPT activities were decreased in the liver, gills and kidneys. The lactic dehydrogenase (LDH) was significantly stimulated by Cd in the heart in vivo but in vitro the metal inhibited the enzyme in the gills.4. Enzymes in the liver, followed by those in the kidneys and gills seem to be most seriously affected by Cd poisoning in this fish.     

Comparison of injectable iron complexes in their ability to iron load tissues and to induce oxidative stress

Iron and copper homeostasis have been studied in various tissues after iron-loading with the polynuclear ferric hydroxide carbohydrate complexes, iron dextran, iron polymaltose, iron sucrose and iron gluconate for four weeks. There were significant increases in the iron content of the different rat tissues compared to controls, with the exception of the brain, which showed no change in its iron content following iron loading. However, the level of iron loading in the different tissues varied according to the preparation administered and only iron dextran was able to significantly increase the iron content of both broncho-alveolar macrophages and heart. The hepatic copper content decreased with iron loading, although this did not reach significance. However the copper content did not alter in the iron loaded broncho-alveolar macrophages. Despite such increases in hepatic iron content, there was little evidence of changes in oxidative stress, the activities of cytosolic (apart from iron dextran) or mitochondrial hepatic superoxide dismutase, SOD, were similar to that of the control rats, confirming the fact that the low reduction potential of these compounds prevents the reduction of the ferric moiety. It was not necessary for macrophages to significantly increase their iron content to initiate changes in NO^. release. Iron gluconate and iron sucrose increased NO^. release, while iron polymaltose and iron dextran decreased NO^. release although only the latter iron preparation significantly increased their iron content. It may be that the speciation of iron within the macrophage is an important determinant in changes in NO^. release after ex vivo stimulation. We conclude that tissues loaded with iron by such polynuclear iron complexes have variable loading despite the comparable iron dose. However, there was little evidence for participation of the accumulated iron in free radical reactions although there was some evidence for alteration in immune function of broncho-alveolar macrophages.     

Different iron sources to study the physiology and biochemistry of iron metabolism in marine micro-algae

We compared ferric EDTA, ferric citrate and ferrous ascorbate as iron sources to study iron metabolism in Ostreococcus tauri, Phaeodactlylum tricornutum and Emiliania huxleyi. Ferric EDTA was a better iron source than ferric citrate for growth and chlorophyll levels. Direct and indirect experiments showed that iron was much more available to the cells when provided as ferric citrate as compared to ferric EDTA. As a consequence, growth media with iron concentration in the range 1–100 nM were rapidly iron-depleted when ferric citrate—but not ferric EDTA was the iron source. When cultured together, P. tricornutum cells overgrew the two other species in iron-sufficient conditions, but E. huxleyi was able to compete other species in iron-deficient conditions, and when iron was provided as ferric citrate instead of ferric EDTA, which points out the critical influence of the chemical form of iron on the blooms of some phytoplankton species. The use of ferric citrate and ferrous ascorbate allowed us to unravel a kind of regulation of iron uptake that was dependent on the day/night cycles and to evidence independent uptake systems for ferrous and ferric iron, which can be regulated independently and be copper-dependent or independent. The same iron sources also allowed one to identify molecular components involved in iron uptake and storage in marine micro-algae. Characterizing the mechanisms of iron metabolism in the phytoplankton constitutes a big challenge; we show here that the use of iron sources more readily available to the cells than ferric EDTA is critical for this task.     

Mucus as a diffusion barrier to oxygen: Possible role in O2 uptake at low pH in carp (Cyprinus carpio) gills

• 1.1. The standard V_o2 of carp (Cyprinus carpio) is reduced at low pH, concurrently with the formation of mucus on the gills.
• 2.2. The rate of diffusion of oxygen through mucus produced by carp in response to low pH was measured in a diffusion chamber and found to be 2.60 × 10⁻⁵ cm²/min per atm. The diffusion constant for water measured with the same apparatus was 3.60 × 10⁻⁵cm₂/min per atm.
• 3.3. The inhibition of the diffusion of oxygen into the gill bloodstream, and the inhibition of the flow of respiratory water between the secondary lamellae, are calculated for a gill with assumed dimensions. At thicknesses of mucus on the gills up to 5 μm the effects are similar, but at greater thicknesses the effect of slowing the water flow predominates. We conclude that mucus on the gills contributes to the hypoxia observed in fish subjected to high acidities.

     

Multilocus isozyme systems in African lungfish,Protopterus annectens: distribution,differential expression and variation in dipnoans

Distribution of ADH, ALP, FBALD, GAPDH, G3PDH, G6PDH, GPI, LDH, MDH, PGM, and SOD was identified in retina, heart, muscle, liver, kidney, gills, brain, gut, lung and ovary of the African lungfish. Data are compared with patterns previously described in dipnoans and other vertebrates. The number of loci expressed for all enzymes was found to be similar to those of diploid Actinopterygii. Differences in the number of loci expressed in Amphibia were found for ALP, sG3PDH, GPI, LDH, MDH and SOD. Differences in tissue distribution were noted in ALP due to the absence of an intestinal-specific form typical of teleostean fish, amphibians, reptiles and birds, and in GPI and MDH, due to the tissue expression, as in primitive fish. There were also differences in LDH, where a third locus (LDH-C*) was expressed in the gills of Protopterus annectens and not in the retina or liver tissues, as in teleosts. LDH-A₄ was most common in all the tissues. Major differences were noted in the tissue patterns of protein expression in the three dipnoans compared. As expected, the least divergence was found between the two species belonging to the same family (Lepidosirenidae). The highest index of divergence was observed between Neoceratodus forsteri and Lepidosiren paradoxa, belonging to the families Ceratontidae and Lepidosirenidae, respectively. The divergence is revealed by changes at the enzyme and morphological levels. These results suggest that P. annectens occupies an interesting systematic position, its biochemical characteristics distinguishing it from N. forsteri, L. paradoxa, the advanced fish and amphibians.     

Measurement of water drinking rates in marine Crustacea

Tests were made of γ-emitting compounds as potential non-absorbed reference markers for estimating water ingestion in the western rock lobster Panulirus longipus cygnus (George) and the penaeid prawn Penaeus latisulcatus (Kishinouye). The extent of any marker absorption into the tissues was measured in animals totally immersed and in rock lobsters with only the gills perfused. Distribution of marker within the body, transport into the gut, and excretion were examined following injection of the label into the blood of the rock lobster.¹²⁵I-sodium iothalamate, ⁵⁸Co-EDTA, and ⁵¹Cr-EDTA were all satisfactory reference markers with very low absorption, ⁵¹Cr-EDTA being the best. Most injected sodium iothalamate was excreted from the body over 24 h, probably via the urine, which suggests its further use in studies of antennal gland function.⁴⁶ScCl₃, ⁵¹CrCl₃, ⁵⁸CoCl₂, and ¹¹⁰AgCl were of some value for estimating drinking in penaeid prawns, although ¹¹⁰AgCl probably gave levels that were too high. ¹¹⁰AgCl was useless for the rock lobster, which absorbed much more marker than was retained in the gut.¹²⁵I-rose bengal was not practicable because it slowly precipitated in sea water.Requirements for reference markers appear to be more critical for animals with high permeability to major sea-water ions than those which extensively regulate all major ions.     

Toxicity and bioaccumulation of iron in soil microalgae

Microalgae are extensively used in the remediation of heavy metals like iron. However, factors like toxicity, bioavailability and iron speciation play a major role in its removal by microalgae. Thus, in this study, toxicity of three different iron salts (FeSO₄, FeCl₃ and Fe(NO₃)₃) was evaluated towards three soil microalgal isolates, Chlorella sp. MM3, Chlamydomonas sp. MM7 and Chlorococcum sp. MM11. Interestingly, all the three iron salts gave different EC50 concentrations; however, ferric nitrate was found to be significantly more toxic followed by ferrous sulphate and ferric chloride. The EC₅₀ analysis revealed that Chlorella sp. was significantly resistant to iron compared to other microalgae. However, almost 900 μg g^?1 iron was accumulated by Chlamydomonas sp. grown with 12 mg L^?1 ferric nitrate as an iron source when compared to other algae and iron salts. The time-course bioaccumulation confirmed that all the three microalgae adsorb the ferric salts such as ferric nitrate and ferric chloride more rapidly than ferrous salt, whereas intracellular accumulation was found to be rapid for ferrous salts. However, the amount of iron accumulated or adsorbed by algae, irrespective of species, from ferrous sulphate medium is comparatively lower than ferric chloride and ferric nitrate medium. The Fourier transform infrared spectroscopy (FTIR) analysis shows that the oxygen atom and P?=?O group of polysaccharides present in the cell wall of algae played a major role in the bioaccumulation of iron ions by algae.     

DNA damage (comet assay) and 8-oxodGuo (HPLC-EC) in relation to oxidative stress in the freshwater bivalve Unio tumidus.

The relationships between DNA damage and oxidative stress in the digestive gland, gills and haemocytes of the freshwater bivalve Unio tumidus were investigated. Two markers of genotoxicity were measured: DNA breaks by means of the comet assay, and oxidative DNA lesions by means of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo) measured using high-performance liquid chromatography (HPLC) coupled to electrochemical detection. Lipid peroxidation was evaluated by measuring malondialdehyde (MDA) tissue levels. Effects were studied after exposure of bivalves for 6 days to benzo[a]pyrene (B[a]P) (50 and 100 microg l(-1)) and ferric iron (20 and 40 mg l(-1)), applied alone or in combination. Lipid peroxidation in the digestive gland and gills resulted from exposure to Fe3+ or B[a]P whatever the concentrations tested. DNA oxidatively formed lesions were induced in the two tissues at a higher level after B[a]P exposure than after Fe3+ treatment. No significant dose-response relationship was found with the two compounds and no synergistic effect was observed between Fe3+ and B[a]P. The gills appeared less sensitive than the digestive gland to DNA lesions expressed as 8-oxodGuo and comet results. Good correlations were noted between 8-oxodGuo and comet. MDA and DNA damage did not correlate as well, although it was stronger in the digestive gland than in the gills. Production of mucus by the gills likely served to prevent lesions by reducing the bioavailability of the chemicals tested, which could explain that dose-effect relationships and synergistic effects were not observed.