Protective effects of zinc on cadmium toxicity in rodents

A study of acute and subacute toxicity of cadmium ions [Cd(II)] was carried out on male Swiss mice and Sprague-Dawley rats with and without previous administration of zinc chloride. The LD₅₀ of Cd(II) as cadmium sulfate (ip) was lower in animals previously given 10 mg/kg of zinc(II) chloride (sc). Factors such as animal weight variations, biochemical parameters, and accumulation patterns of Cd(II) and Zn(II) were taken into consideration when the subacute toxicity was evaluated. Alteration of the activities of glutamic pyruvic transaminase (GPT) and of glutamic oxaloacetic transaminase (GOT) was observed in short-term-exposure (<6 h) cases. These alterations reverted to normal after 1 wk. The activity of alkaline phosphatase (ALP) and the concentrations of cholesterol and triglycerides in serum are also changed, especially so in the groups given CdSO₄ alone. In the experimental groups treated with ZnCl₂ prior to administration of cadmium, proteinuria was detected 5 wk after the treatment. Also at 5 wk, both Zn-treated and nontreated groups showed an abnormally low liver mass with respect to total body mass. Both Cd and Zn are retained preferentially in the liver but show also in the kidneys. If CdSO₄ and ZnCl₂ are given simultaneously, especially after 1 wk of treatment, Cd is accumulated in greater amounts in these organs when compared to the groups given only cadmium sulfate.     

Chronic Cadmium Exposure Lead to Inhibition of Serum and Hepatic Alkaline Phosphatase Activity in Wistar Rats

Alkaline phosphatase (ALP) activity in the serum and liver from rats administered with cadmium (Cd) in drinking water was studied. After metal administration, Cd showed a time‐dependent accumulation in the liver, meanwhile metallothionein had a maximum increase at 1 month, remaining in this level until the end of the study. On the other hand, serum and liver ALP activity was decreased after 3 months exposure. To determine if Cd produced an inhibition on enzyme, apo‐ALP prepared from both nonexposed and exposed rats was reactivated with Zn, showing 60% more activity as compared with the enzyme isolated from nonexposed rats. In vitro assays showed that Cd‐ALP was partially reactivated with Zn; however, in the presence of cadmium, Zn‐ALP was completely inhibited. Kinetic studies indicate a noncompetitive inhibition by Cd; these results suggest that Cd can substitute Zn, and/or Cd can interact with nucleophilic ligands essential for the enzymatic activity.     

Activité phosphatase alcaline dans le labyrinthe du placenta de chatte,et problème de la localisation de l'ATPase de transport: étude ultrastructurale

Summary The ultrastructural localizations of alkaline phosphatase (ALPase) and of Na⁺–K⁺-dependent adenosine triphosphatase (Na⁺–K⁺-ATPase) were studied in the placental labyrinth of the cat during the last days of gestation. ALPase activity could be detected in the syncytiotrophoblast but was absent from maternal tissues. Enzyme activity was observed only along plasma membranes of microvilli and absorption tubules on the maternal surface of the syncytium and also on the podocytes-like cytoplasmic processes of the fetal face. The localization of the Na⁺–K⁺-ATPase activity as obtained with the method of Ernst was identical with that of ALPase. This activity was not very ouabaine sensitive or K⁺ dependent, but was almost completely inhibited by levamisole. The strong ALPase activity of the syncytiotrophoblast does not allow a specific detection of Na⁺–K⁺-ATPase. However, the localization of these enzymes activities on syncytiotrophoblast surfaces directly related to fetal and maternal capillaries could suggest that these surfaces are associated with transport mechanisms of the trophoblast.

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Activité phosphatase alcaline dans le labyrinthe du placenta de chatte, et problème de la localisation de l'ATPase de transport: étude ultrastructurale

Résumé Les localisations de la phosphatase alcaline (ALPase) et de l'adénosine triphosphatase Na⁺–K⁺-dependante (Na⁺–K⁺-ATPase) ont été recherchées dans le labyrinthe du placenta de chatte en fin de gestation. L'activité ALPase est localisée de manière discontinue le long de la membrane plasmique du syncytiotrophoblaste: microvillosités et tubules d'absorption de la face maternelle, digitations cytoplasmiques à allure podocytique de la face foetale. La localisation de l'activité Na⁺–K⁺-ATPase par la méthode de Ernst est identique à celle de l'ALPase. Cette activité, très peu sensible à la ouabaïne et à l'absence de K⁺, est presque totalement inhibée par le tétramisole. Cette présence importante d'activité ALPase empêche la mise en évidence spécifique de la Na⁺–K⁺-ATPase. La localisation de ces enzymes dans des zones en rapport avec les capillaires foetaux et maternels, permet de soutenir la thèse d'une spécialisation de ces zones dans les mécanismes de transport.

     

Cadmium accumulation and interactions with zinc, copper, and manganese, analysed by ICP-MS in a long-term Caco-2 TC7 cell model

The influence of long-term exposure to cadmium (Cd) on essential minerals was investigated using a Caco-2 TC7 cells and a multi-analytical tool: microwave digestion and inductively coupled plasma mass spectrometry. Intracellular levels, effects on cadmium accumulation, distribution, and reference concentration ranges of the following elements were determined: Na, Mg, Ca, Cr, Fe, Mn, Co, Ni, Cu, Zn, Mo, and Cd. Results showed that Caco-2 TC7 cells incubated long-term with cadmium concentrations ranging from 0 to 10 μmol Cd/l for 5 weeks exhibited a significant increase in cadmium accumulation. Furthermore, this accumulation was more marked in cells exposed long-term to cadmium compared with controls, and that this exposure resulted in a significant accumulation of copper and zinc but not of the other elements measured. Interactions of Cd with three elements: zinc, copper, and manganese were particularly studied. Exposed to 30 μmol/l of the element, manganese showed the highest inhibition and copper the lowest on cadmium intracellular accumulation but Zn, Cu, and Mn behave differently in terms of their mutual competition with Cd. Indeed, increasing cadmium in the culture medium resulted in a gradual and significant increase in the accumulation of zinc. There was a significant decrease in manganese from 5 μmol Cd/l exposure, and no variation was observed with copper.     

Subcellular localisation of Cd and Zn in the leaves of a Cd-hyperaccumulating ecotype of<Emphasis Type="Italic"> Thlaspi caerulescens</Emphasis>

Thlaspi caerulescens (Ganges ecotype) is able to accumulate large concentrations of cadmium (Cd) and zinc (Zn) in the leaves without showing any toxicity, suggesting a strong internal detoxification. The distribution of Cd and Zn in the leaves was investigated in the present study. Although the Cd and Zn concentrations in the epidermal tissues were 2-fold higher than those of mesophyll tissues, 65–70% of total leaf Cd and Zn were distributed in the mesophyll tissues, suggesting that mesophyll is a major storage site of the two metals in the leaves. To examine the subcellular localisation of Cd and Zn in mesophyll tissues, protoplasts and vacuoles were isolated from plants exposed to 50 M Cd and Zn hydroponically. Pure protoplasts and vacuoles were obtained based on light-microscopic observation and the activities of marker enzymes of cytosol and vacuoles. Of the total Cd and Zn in the mesophyll tissues, 91% and 77%, respectively, were present in the protoplast, and all Cd and 91% Zn in the protoplast were localised in the vacuoles. Furthermore, about 70% and 86% of total Cd and Zn, respectively, in the leaves were extracted in the cell sap, suggesting that most Cd and Zn in the leaves is present in soluble form. These results indicate that internal detoxification of Cd and Zn in Thlaspi caerulescens leaves is achieved by vacuolar compartmentalisation.     

Bioaccumulation du Cd et du Zn chez les plants de tomates (Lycopersicon esculentum L.)

This work aims at evaluating the accumulation of cadmium (Cd) and zinc (Zn) (trace elements) in the organs of young tomato plants (Lycopersicon esculentum L. var. Rio Grande) and their effects on the rate of chlorophyll and enzyme activities involved in the antioxidant system: catalase (CAT), glutathion-S-transferase (GST) and peroxysase ascorbate (APX). Plants previously grown on a basic nutrient solution were undergoing treatment for 7 days, either by increasing concentrations of CdCl₂ or ZnSO₄ (0, 50, 100, 250, 500 μM) or by the combined concentrations of Cd and Zn (100/50, 100/100, 100/250, 100/500 μM). The results concerning the determination of metals in the various compartments of tomato plants as a function of increasing concentrations of Cd or Zn, suggest a greater accumulation of Cd and Zn in the roots compared to leaves. The combined treatment (Cd/Zn) interferes with the absorption of the two elements according to their concentrations in the culture medium. The presence of Zn at low concentrations (50 μM of Zn/100 μM Cd) has little influence on the accumulation of Cd in the roots and leaves, while the absorption of these two elements in the leaves increases and decreases in roots when their concentrations are equivalent (100/100 μM) compared to treatment alone. When the concentration of Zn is higher than that of Cd (500 μM of Zn/100 μM Cd) absorption of the latter is inhibited in the roots while increasing their translocation to the leaves. Meanwhile, the dosage of chlorophylls shows that they tend to decrease in a dose-dependent for both treatments (Cd or Cd/Zn), however, treatment with low concentrations of Zn (50 and 100 μM) stimulates chlorophyll synthesis. However, treatment with different concentrations of Cd seems to induce the activity of the enzymes studied (CAT, APX, GST). It is the same for treatment with different concentrations of Zn and this particularly for the highest concentrations. Finally, the combined treatment (Zn/Cd) also appears to cause enzyme inductions: CAT, APX and GST.     

水淹生境下秋华柳对Cd污染土壤微生物数量及酶活性的影响

三峡库区消落带面临水淹及Cd污染双重胁迫,为探究秋华柳(Salix variegata Franch.)在水淹条件下对Cd污染土壤的修复能力,以秋华柳扦插苗为试验材料,设置正常供水(CK)和水淹组(FL)两个水分处理方式,4个Cd浓度梯度:对照组(0mg/kg)、低浓度(0.5mg/kg)、中浓度(2mg/kg)及高浓度(10mg/kg),分别对处理60 d和120 d的土壤微生物数量及酶活性变化特征进行研究。试验结果表明:(1)Cd浓度处理均未显著影响土壤微生物数量(P0.05),水淹显著降低处理60 d土壤细菌数、真菌数及处理120 d的土壤放线菌数及真菌数(P0.05)。(2)种植秋华柳显著提高处理60 d土壤细菌数量(P0.05),对土壤放线菌、真菌数量也有一定提升。(3)Cd浓度处理显著影响处理60 d土壤磷酸酶活性及处理120 d脲酶活性(P0.05),水淹显著降低处理60 d土壤磷酸酶活性及处理120 d脲酶、蔗糖酶和磷酸酶活性(P0.05)。(4)正常供水及水淹条件下,种植秋华柳对土壤酶活均有一定改善作用。种植秋华柳显著提高了处理60 d土壤磷酸酶活性以及处理120 d脲酶和蔗糖酶活性(P0.05)。研究结果表明:水淹生境中,秋华柳对Cd污染土壤微生物数量及酶活性具有改善作用,在Cd污染土壤修复方面有一定应用前景。     

Activation of alkaline phosphatase with Mg2+ and Zn2+ in rat hepatoma cells. Accumulation of apoenzyme

In Reuber rat hepatoma cells (R-Y121B), alkaline phosphatase activity increased without de novo enzyme synthesis (Sorimachi, K., and Yasumura, Y. (1986) Biochim. Biophys. Acta 885, 272-281). The enzyme was partially purified by butanol extraction from the particulate fractions. The incubation of the extracted alkaline phosphatase with the cytosol fraction induced a large increase in enzyme activity (5-10-fold of control). The dialyzed cytosol was more effective than the undialyzed cytosol during an early period of incubation at 37 degrees C. This difference between the dialyzed and the undialyzed cytosol fractions was due to endogenous Na+. For maximal activation of the enzyme, both Mg2+ above 1 mM and Zn2+ at low concentrations (below 0.01 mM) were needed, although Zn2+ at high concentrations (above 0.1 mM) showed an inhibitory effect. Zn2+ and Mg2+ alone slightly increased alkaline phosphatase activity. This activation of the enzyme was temperature dependent and was not observed at 0 or 4 degrees C. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate showed that the increase in alkaline phosphatase activity did not involve the fragmentation of the enzyme and that 65Zn2+ bound to it during enzyme activation with 65Zn2+ and Mg2+. The cytosol fraction not only supplied Zn2+ to the nascent enzyme but also increased the maximal enzyme activity more than did direct addition of metal ions. Ferritin and metallothionein contributed to the activation of alkaline phosphatase with the metal ions. Since the binding of Zn2+ and Mg2+ to the nascent alkaline phosphatase is disturbed in Reuber rat hepatoma cells (R-Y121B), the apoenzyme is accumulated inside the cells. The binding of Zn2+ and Mg2+ to the apoenzyme readily takes place in the cell homogenates accompanied by an increase in catalytic activity without new enzyme synthesis.     

Physiological responses to Cd and Zn in two Cd/Zn hyperaccumulating Thlaspi species

In a model hyperaccumulation study a Cd/Zn hyperaccumulator Thlaspi caerulescens accession Ganges and a recently reported Cd/Zn hyperaccumulator Thlaspi praecox grown in increasing Cd and Zn concentrations in the substrate and in field collected polluted soil were compared. Plant biomass, concentrations of Cd and Zn, total chlorophylls and anthocyanins, antioxidative stress parameters and activities of selected antioxidative enzymes were compared. Increasing Cd, but not Zn in the substrate resulted in the increase of biomass of roots and shoots of T. praecox and T. caerulescens. The two species hyperaccumulated Cd in the shoots to a similar extent, whereas T. caerulescens accumulated more Zn in the shoots than T. praecox. Cadmium amendment decreased total chlorophyll concentration and glutathione reductase activity, and increased non-protein thiols concentration only in T. praecox, suggesting that it is less tolerant to Cd than T. caerulescens. In the field-contaminated soil, T. caerulescens accumulated higher Cd concentrations; but as T. praecox produced higher biomass, both species have similar ability to extract Cd.     

Etude du transfert de quelques oligo-éléments dans les chaînes trophiques néritiques et estuariennes: Accumulation biologique chez les poissons omnivores et super-carnivores

Five species of fishes(Dicentrarchus labrax, Gobius microps, Stizostedion lucioperca, Gadus luscus, Merlangius merlangus) and their major prey organisms were collected monthly from two stations in the Loire estuary (France). The levels of several trace elements (Cd, Pb, Cu, Zn) in their tissues were determined by means of atomic absorption spectrophotometry. The concentrations of Cd, Pb and Cu were shown to decrease in the highest trophic levels: the relatively highest metal levels were determined in annelids, followed by crustaceans, the lowest levels being encountered in fishes. However, a preferential uptake of Cu was observed in crustaceans. There is no biomagnification for these three metals, the concentrations in preys being generally lower than in predators. For Zn, the highest concentrations were measured in worms and copepods but preys such as shrimps and mysids exhibit values of the same order of magnitude compared to predator fishes.

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Avec la collaboration technique de R. Ferre.     

Localisation ultrastructurale de la phosphatase acide et de la phosphatase alcaline dans les cloisons septales stériles de l'anthozoaire Pachycerianthus fimbriatus

Résumé La phosphatase acide et la phosphatase alcaline ont été localisées dans les cellules des cloisons septales stériles d'un Cérianthe. Chez les animaux à jeun, l'activité de la phosphatase acide a été observée dans les citernes de l'appareil de Golgi, les lysosomes et les phagosomes provenant d'un repas précédent. Chez les animaux nourris dans les 12 heures précédentes, le précipité est localisé dans les mêmes structures ainsi que dans des phagosomes récemment formés par l'absorption de proie. La phosphatase alcaline a été détectée sur les membranes plasmiques apicales et latérales et dans les phagosomes anciens.

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Ultrastructural localization of acid and alkaline phosphatases in sterile septae of the anthozoa Pachycerianthus fimbriatus

Summary Acid and alkaline phosphatase activity has been localized in the cells of the sterile septae of starved and fed anthozoa. Acid phosphatase is present in lysosomes, in Golgi cisternae and old phagosomes of starved animals. In fed animals, the reaction is more intense and the number of lysosomes is increased. New phagosomes are loaded with lead phosphate. In starved animals, the alkaline phosphatase activity has been observed on the plasma membranes and in the old phagosomes.

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Ce travail a été rendu possible grâce à une bourse post-doctorale de recherche attribuée au titre des accords France-Québec, (Y.T.) et grâce à un octroi du Conseil des Recherches Médicales du Canada (J.H.)     

Microelements in seminal plasma of infertile men infected with Ureaplasma urealyticum

The purpose of this study was to assess the association among male infertility, infection of Ureaplasma urealyticum (Uu), and microelements in seminal fluid. Semen analysis and cultivation of Uu were carried out on 160 samples of seminal fluid. The concentrations of microelements, such as arsenic (As), molybdenum (Mo), magnesium (Mg), lead (Pb), copper (Cu), cadmium (Cd), and zinc (Zn) in the samples were measured by an inductively coupled plasma quantometer (ICP). The ratios Cu/Zn and Cd/Zn in the poor spermatic quality group were obviously higher than those in seminal plasma of the group with normal spermatic quality (p<0.01 and p<0.05, respectively), whereas the concentrations of As, Mg, Mo, and Pb showed no difference in the two groups. The ratios Cu/Zn and Cd/Zn and the concentrations of As and Mg in seminal plasma infected with Uu were markedly higher than those not infected with Uu (p<0.05, p<0.01, p<0.05, and p<0.05, respectively), whereas the concentrations of Mo and Pb showed no statistical difference. The ratios Cu/Zn and Cd/Zn and the concentrations of As and Mg in seminal plasma of the semen with poor spermatic quality and Uu infection were obviously higher than those not infected with Uu (p<0.05), whereas the concentrations of Mo and Pb showed no statistical difference. Abnormally high ratios Cu/Zn and Cd/Zn as well as an overdose of As were found to be predisposed to Uu infection. Uu infection resulted in an increase of the ratios Cu/Zn and Cd/Zn and the concentrations of As and Mg in seminal fluid, which therefore caused spermatic quality decline.     

Influence of aqueous extract of Hibiscus sabdariffa L. petal on cadmium toxicity in rats

The effects of chronic exposure to cadmium (Cd) on some selected biochemical parameters, as well as the possible protective role of aqueous extracts of Hibiscus sabdariffa L petal were studied in 12-wk-old male Wistar albino rats. Exposure to Cd caused a significant increase in plasma l-alanine aminotransferases (ALT) only but with a corresponding decrease in liver l-alanine and l-aspartate aminotransferases (L-ALT, L-AST) when compared to the Cd-free control. Total superoxide dismutase activity was decreased in the liver, testis, and prostate of Cd-exposed rats, whereas malondialdehyde (MDA) concentrations were increased relative to the Cd-free control. The metal significantly increased prostatic acid phosphatase activity in the prostate, but decreased the body weight gain of the rats and organ/body weight ratio for prostate and testis compared to the Cd-free control. Pretreatment of rats with aqueous extract of H. sabdariffa resulted in significantly less hepatotoxicity than with Cd alone as measured by plasma ALT and liver ALT and AST activities. The extract also protected the rats against Cd-induced liver, prostate, and testis lipoperoxidation as evidenced by significantly reduced MDA values in these organs, as well as reduced prostatic acid phosphatase activity in the prostate, when compared to the Cd-only exposed rats. Also, when compared to the organ/body weight ratios obtained from rats exposed to Cd alone the prostate and testis were protected by the extract as shown by enhanced prostate/body weight and testis/body weight ratios of Cd- and extract-treated rats. These data suggest that H. sabdarrifa L might be protective in Cd toxicity.     

Localisation ultrastructurale de l'activité phosphatasique alcaline dans les cellules interrénales d'un amphibien urodèle,Triturus cristatus

Résumé L'activité phosphatasique alcaline des cellules interrénales (corticosurrénaliennes) del'Amphibien Urodèle Triturus cristatus est presque exclusivement localisée sur les membranes plasmiques. Elle est plus ou moins intense selon les cellules; certaines cellules ne présentent aucune activité phosphatasique alcaline cytochimiquement décelable.Cette activité enzymatique est associée à toutes les villosités de la surface cellulaire, aux parois des vésicules de micropinocytose, ainsi qu'à de courts tubules, à signification peutêtre pinocytotique, situés dans le voisinage de la membrane plasmique. Le produit de la réaction enzymatique peut encore être observé à la périphérie de corps multivésiculaires; l'origine de la phosphatase alcaline dans ces structures est discutée.Les autres organites cytoplasmiques ne présentent aucune activité. D'autre part, la réaction enzymatique n'a pas lieu dans des cellules incubées dans un milieu dépourvu de substrat, ou préincubées dans une solution de L-cystéine.

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Ultrastructural localization of the alkaline phosphatase activity in the interrenal cells of the Amphibian Urodele, Triturus cristatus

Summary The alkaline phosphatase activity of interrenal (adrenocortical) cells of the Amphibian Urodele Triturus cristatus is almost exclusively located on the cytoplasmic membranes. This activity is variable from cell to cell, sometimes even missing.All the microvilli of the cellular surface, as well as the pinocytotic vesicles, show alkaline phosphatase activity. The product of the enzymatic reaction is also associated with several short tubules located in the vicinity of the cytoplasmic membrane; these tubules have probably a pinocytotic signification. The product of the enzymatic reaction can also be observed at the periphery of the multivesicular bodies; the origin of alkaline phosphatase activity in these structures is discussed.The other cytoplasmic organelles do not show any activity. The enzymatic reaction, on the other hand, does not take place in cells which have been incubated in a medium without substrate, or preincubated in a solution containing L-cysteine.

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Ce travail a été rendu possible grâce à une bourse de voyage des Accords France-Québec, et à l'octroi n MA 3994 du Conseil des Recherches Médicales du Canada.

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Nous remercions vivement M. Michel Couture et Melle Monique Walter pour leur aide précieuse.     

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.     

镉胁迫对小报春幼苗生长及生理特性的影响

该研究采用盆栽控制试验,测定不同浓度(0、5、50、100、150、200 mg/kg)土壤Cd胁迫下小报春幼苗生长及生理生化指标,以探究小报春(Primula forbesii Franch.)对重金属镉(Cd)污染的抗性和敏感性,为新型香花地被植物应用于Cd污染土壤提供理论依据。结果表明:(1)低浓度(5 mg/kg)Cd胁迫能促进小报春株高和根长的伸长,高浓度(≥150 mg/kg)Cd胁迫下株高、根长和生物量明显降低。(2)随Cd胁迫浓度增加,小报春幼苗叶片光合作用、叶绿素含量、SOD和CAT活性先上升后下降,而其超氧阴离子产生速率、过氧化氢、丙二醛含量、叶相对电导率和POD活性持续升高。(3)在Cd胁迫条件下,小报春体内K、Zn含量降低,叶和根中Ca、Mg含量显著增加,体内Cd含量明显升高,且根系Cd含量远远高于叶和叶柄。研究发现,小报春幼苗对Cd胁迫具有一定的耐性,低浓度(5 mg/kg)Cd胁迫对小报春生长影响较小,但高浓度(≥150 mg/kg)Cd胁迫对小报春产生明显的毒害作用,影响其正常生长;小报春可能通过增强体内抗氧化酶系统活性、增加叶和根中Ca、Mg含量和根系截留镉来减轻镉胁迫的伤害,提高自身耐受镉胁迫能力。     

Zn(II)-113Cd(II) and Zn(II)-Mg(II) hybrids of alkaline phosphatase. 31P and 113Cd NMR

Methods have been developed for the addition of different metal ion species to the three distinct pairs of metal sites (A, B, and C) found in the dimer of apoalkaline phosphatase. This allows the preparation of hybrid alkaline phosphatases in which A and B sites of each monomer contain two different species of metal ion or the A and B sites of one monomer contain the same species of metal ion, while the adjacent monomer contains a second species. The following hybrids have been characterized in detail: (Zn(II)ACd(II)B)2 alkaline phosphatase, (Zn(II)AMg(II)B)2 alkaline phosphatase, (Cd(II)AZn(II)B)2 alkaline phosphatase, and (Zn(II)AZn(II]B)(Cd(II)ACd(II)B) alkaline phosphatase. 31P and, where appropriate, 113Cd NMR have been used to monitor the behavior of the covalent (E-P) and noncovalent (E X P) phosphointermediates and of the A and B metal ions. From the pH dependencies of the E-P in equilibrium E X P in equilibrium E + Pi equilibria, it is clear that A site metal is the dominant influence in dephosphorylation of E-P and may have a coordinated water molecule, which ionizes to ZnOH- at a low pH providing the nucleophile for dephosphorylation. A site metal also serves to coordinate phosphate in the E X P complex. B site metal has a much smaller effect on dephosphorylation rates, although it does dramatically alter the Pi dissociation rate, which is the rate-limiting step for the native enzyme at alkaline pH, and is probably important in neutralizing the charge on the phosphoseryl residue, thus potentiating the nucleophilic attack of the OH- bound at A site. Phosphate dissociation is slowed markedly by replacement of B site zinc by cadmium. There is clear evidence for long range effects of subunit-subunit interactions, since metal ion and phosphate binding at one active center alters the environments of A and B site metal ions and phosphoserine at the other active site.     

Genetic basis of Cd tolerance and hyperaccumulation in Arabidopsis halleri

The genetic basis of Cd tolerance and hyperaccumulation was investigated in Arabidopsis halleri. The study was conducted in hydroponic culture with a backcross progeny, derived from a cross between A. halleri and a non-tolerant and non-accumulating related species Arabidopsis lyrata ssp. petraea, as well as with the parents of the backcross. The backcross progeny segregates for both cadmium (Cd) tolerance and accumulation. The results support that (i) Cd tolerance may be governed by more than one major gene, (ii) Cd tolerance and Cd accumulation are independent characters, (iii) Cd and Zn tolerances co-segregate suggesting that they are under pleiotropic genetic control, at least to a certain degree, (iv) the same result was obtained for Cd and Zn accumulation.     

Influence de la concentration de la solution nutritive dans la phase de réinsertionin vivo des vitroplants d'Erica × darleyensis

Résumé La réinsertion de vitroplants d'Erica × darleyensis est suivie en chambre climatisée et sur perlite arrosée de solutions nutritives, en analysant pendant deux mois le développement des plantes. Quatre solutions nutritives ont été testées. Seule en varie les concentrations (A, A/2, A/4 et A/10); le rapport entre les différents éléments reste constant. Les vitroplants issus d'un milieu de multiplication gélosé A/2, transplantés sur les milieux de réinsertion subisent un choc nutritionnel: même les vitroplants transférésin vivo sur solution dont la concentration en macroéléments est le double de celle du milieu de multiplicationin vitro, enregistrent une diminution des teneurs minérales. L'influence de la nature des substrats de ces différents milieux (gélose ou perlite) semble prédominante à cet égard. Des milieux de réinsertion moyennement concentrés (A/2 ou A/4) constituent les meilleures références pour la précocité et l'optimisation de la production des rameaux. La production des rameaux est reliée à certains paramètres internes nutritionnels tels que: rapports élevés K⁺/(Ca²⁺+Mg²⁺) et Mn²⁺/Fe³⁺, et de hautes teneurs en azote et phosphore.

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Influence ofin vivo nutrient concentration inErica × darleyensis vitroplant transfer on perlite mediaI. Development and nutrient contents of transferred vitroplants

Development ofErica × darleyensis vitroplants was studied on perlite nutritive media in growth chamber conditions, during two months. Four nutritive solutions were examined. Only solution concentrations vary (solutions A, A/2, A/4 and A/10); the ratio between nutrients remains constant. Vitroplants from a A/2 agar-agar propagation medium, transferred on perlite media underwent a nutritional stress: even vitroplants transferred on a perlite medium whose concentration is twice that of the propagation medium, show a decrease in mineral content. The influence of the kinds of substrates (agar-agar or perlite) seems to be prevailing in this respect. Perlite media irrigated with mildly concentrated solutions (A/2 or A/4) are the best references for earliness and optimization of ramification. Branch production is correlated to internal nutritional parameters such as: high K⁺/(Ca²⁺+Mg²⁺) and Mn²⁺/Fe³⁺), a high nitrogen and phosphorus contents.