ISOLATION OF V79 FIBROBLAST CELL LINES CONTAINING ELEVATED METALLOTHIONEIN LEVELS THAT HAVE INCREASED RESISTANCE TO THE CYTOTOXIC EFFECTS OF ULTRAVIOLET-A RADIATION

Abstract Isolated clones of V79 Chinese hamster lung fibroblasts, selected for resistance against cadmium toxicity, were exposed to monochromatic 365 nm ultraviolet-A (UVA, 320 nm to visible light) radiation and examined for cell survival. All three of the Cd-resistant V79 clones (V79Cd) tested exhibited significant increases in survival after irradiation compared with control cultures similar to the increased survival observed in Zn acetate-induced V79 cells. Dose-modifying factors calculated for these survival experiments were all approximately 1.5. When characterized for steady-state levels of metallothionein (MT) mRNA and associated Cd-binding activity, all of the Cd-resistant V79Cd clones demonstrated elevated constitutive levels of both, implicating MT as the mechanism responsible for the observed cellular resistance to Cd and also to 365 nm UVA radiation. However, whereas levels of intracellular MT protein correlated with differences in survival against Cd, MT intracellular levels did not correlate well with protection against 365 nm UVA. Increased cell survival after exposure to 365 nm UVA radiation mediated by MT appeared to reach a threshold level and MT only provided a limited degree of protection. Since UVA radiation is known to cause cell death mediated through the intracellular generation of reactive oxygen species (ROS), these results suggest that the role of MT in ameliorating cellular photooxidative damage produced by UVA is by reducing intracellular ROS.     

Combination of <Superscript>13</Superscript>C/<Superscript>113</Superscript>Cd NMR,potentiometry, and voltammetry in characterizing the interactions between Cd and two models of the main components of soil organic matter

This work allowed the characterization of the Cd-binding sites of two compounds taken as models for exudates, the main components of soil organic matter (SOM). The studied compounds were exopolysaccharides (EPS), specifically exudates of roots (polygalacturonic acid) and of soil bacteria (Phytagel). Potentiometric acid–base titrations were performed and fitting of the obtained results indicated the presence of two main classes of acidic sites, defined by their pK _a values, for both EPS but of a different nature when comparing the two compounds. The two studied exopolysaccharides presented different acidic/basic site ratios: 0.15 for Phytagel and 0.76 for polygalacturonic acid. Spectroscopic techniques (¹³C/¹¹³Cd NMR, FTIR) distinguished different Cd surroundings for each of the studied EPS, which is in agreement with the titration results. Furthermore, these analyses indicated the presence of –COOH and –OH groups in various proportions for each exopolysaccharide, which should be linked to their reactivity towards cadmium. Cadmium titrations (voltammetric measurements) also differentiated different binding sites for each compound and allowed the determination of the strength of the Cd-binding site of the EPS. Fitting of the results of such voltammetric measurements was performed using PROSECE (Programme d’Optimisation et de Speciation Chimique dans l’Environnement), a software coupling chemical speciation calculation and binding parameter optimization. The fitting, taking into account the Cd²⁺/H⁺ competition towards exopolysaccharides, confirmed the acid-base titrations and spectroscopic analyses by revealing two classes of binding sites: (i) one defined as a strong complexant regarding its Cd²⁺–EPS association (logK = 9–10.4) and with basic functionality regarding H⁺–EPS association (pK _a = 11.3–11.7), and (ii) one defined as a weak complexant (logK = 7.1–8.2) and with acidic functionality (pK _a = 3.7–4.0). Therefore the combination of spectroscopic analyses, voltammetry, and fitting allowed the precise characterization of the binding sites of the studied exopolysaccharides, mimicking the main SOM components. Furthermore, the binding parameters obtained by fitting can be used in biogeochemical models to better define the role of key SOM compounds like exudates of roots and of soil bacteria on trace metal transport or assimilation.     

Evaluation of crop resistance to aqueous oil pollution

Root and shoot tolerance indices and photosynthesis characteristics (CO₂-dependent-O₂ evolution; H₂O→MV electron transport; whole-leaf photosynthesis by infrared gas analyzer) indicated that ADT-36 and CR-1009 rice varieties were more tolerant than other varieties (IR-50, IR-20, and Ponni). Vigna varieties (T-9 and Krishna) were most susceptible to aqueous oil pollution. Since the oil-polluted soil contained a potentially toxic concentration of cadmium, tolerant (ADT-36) and susceptible (IR-50) varieties of Oryza were further analyzed for cadmium-binding components. Higher accumulations of cadmium were found in the roots than in the leaves of both rice varieties. Cadmium is associated with proteins of about 50,000 and 33,000-dalton in molecular mass, of which the 33-kdalton protein was significantly induced only in ADT-36. It is suggested by the present study that the ADT-36 rice variety is most tolerant to aqueous oil pollution, as evidenced by higher STI (shoot tolerance index) and RTI (root tolerance index) values, induction of Cd-binding proteins, and comparatively normal photosynthesis rates.     

Physiological changes in barley plants under combined toxicity of aluminum, copper and cadmium

Crop production on acid soil is markedly reduced, further, a multiple heavy metal pollution except Al on acid soil is detected in many areas. The present study was undertaken to assess the toxicities of Al, Cd, and Cu separately and in combinations, three heavy metals very often coexisting on acid soil, and to identify their interactions in two kinds of barley seedlings differing in Al tolerance. The plant growth, metals accumulations, total soluble protein and sugar contents, MDA contents and the activities of SOD and POD were estimated in roots and leaves after 5-week supply of the heavy metals excess in the nutrient solution. The results indicated that the stress treatments including low pH (pH 4.5) alone all adversely affected plant growth and disturbed the cell metabolism seriously. The development of toxic symptoms corresponded to a high accumulation of Al, Cd, Cu and to a poor increase in soluble sugar contents but to a high increase in MDA contents, to the decrease in soluble protein contents and to the much elevated SOD and POD activities in both roots and leaves. In addition, binary metal combinations of Al + Cd and Al + Cu both produced the synergistic response for the growth of barley seedlings, in particular for Shang 70-119, while, ternary metal combination of Al + Cu + Cd produced different interactions in two kinds of genotypes, thus, the significant synergistic response was seen in Shang 70-119, but the antagonistic response was detected in Gebeina. The different responses to ternary metal combination of two genotypes may result from the different metal bioaccumulation patterns, hence, the existence of Cd and Cu promoted Al accumulation in Shang 70-119 but inhibited Al accumulation in Gebeina.     

Cadmium Distribution in a Timberline Forest in the Hengduan Mountains in the Eastern Tibetan Plateau

In order to study the regional distribution, characterization, and possible source of Cadmium in the eastern Tibetan Plateau, samples of leaves, xylem, twigs, bark, and roots of timberline forest trees (fir or spruce) and soils at depths from 0 to 40 cm were collected in eight sites in Hengduan Mountains. According to Cd contents, organs and tissues were divided into three groups: the highest-level organ (roots: 0.237 mg · kg^?1), the high-level organ/tissue (bark: 0.183 mg · kg^?1 and twigs: 0.159 mg · kg^?1), and the low-level organ/tissue (xylem: 0.054 mg · kg^?1and leaves: 0.048 mg · kg^?1). Cd contents in Transect A were a little higher than Transect B. Sites near pollutant sources and high-levels are more sensitive to Cd pollution and accumulated more Cd, while Cd contents in sites far away from pollutant sources and low-level organ/tissue were relatively low. It is suggested that the eastern Tibetan Plateau has already been polluted by the influence of general circulation to some extent. Nevertheless, low Cd contents in some sites prove that high mountains may barricade Cd pollutants from monsoon.     

Maize food allergy: lipid-transfer proteins,endochitinases, and alpha-zein precursor are relevant maize allergens in double-blind placebo-controlled maize-challenge-positive patients

Italian patients with maize anaphylaxis have been shown to have IgE toward two major maize allergens: an alpha-amylase inhibitor and a 9-kDa LTP. A complete study on maize food allergens in patients with positive maize double-blind, placebo-controlled food challenge (DBPCFC) is lacking. The objective was to utilize the three maize protein fractions to identify and characterize the most relevant IgE-binding proteins recognized by the sera of Italian and Swiss patients with either a positive maize-DBPCFC or a history of maize-induced anaphylaxis. Osborne’s protein fractions of maize were extracted to obtain water-soluble, total zein, and total protein fractions. Protein IgE-binding capacity was investigated by SDS-PAGE immunoblotting using the sera from DBPCFC-positive patients and from patients with maize-induced anaphylaxis. Purified maize LTP was used to inhibit the IgE immunoblotting of the three protein fractions. IgE immunoblotting demonstrated that the 9-kDa LTP was recognized by all the Italian patients and by none of the Swiss patients. Other allergens were: 14-kDa α-amylase inhibitor, 30-kDa endochitinases A and -B, 19 kDa zein-β precursor, and 26 kDa zein-α precursor; a newly described allergen, the globulin-2 precursor, identified in the total protein fraction. It is noteworthy that maize LTP and endochitinase were cross-reactive with grape LTP and one grape endochitinase. LTP was found to be the only major allergen in Italian patients with either positive maize challenge or a history of maize-induced anaphylaxis. We have identified other maize allergens in subjects with maize food allergy, as grape cross-reactive endochitinase, however, the clinical significance of these proteins needs to be investigated in larger groups of patients with allergy to these food items.     

耐高盐碱蓬对湿地土壤中金属元素的富集特征研究

基于秦皇岛滨海湿地人工种植耐高盐碱蓬修复工程试验,通过分析湿地土壤沉积物及耐高盐碱蓬中不同金属元素的含量与变化,研究了耐高盐碱蓬对金属元素的富集特征。结果显示:湿地沉积物中Fe、Mn、Cr、Pb和Zn的浓度较高,分别为8 210. 94、110. 04、8. 78、8. 25、10. 95 mg/kg,Cd的浓度最低,平均值为0. 022 mg/kg,湿地试验区土壤中同一金属元素变异程度较小,分布较均匀。碱蓬体内重金属含量根据采集地点的不同有差异,但与各站位中土壤的重金属分布特征存在正相关性;其中碱蓬内Fe、Mn、Zn、Cu的平均含量相对较高,且Fe、Mn、Ni、Cu、As、Cd、Pb元素在碱蓬根中的平均含量高于茎叶,而Cr在茎中的平均含量最高,Zn和Mo在叶中的含量最高。碱蓬的根、茎、叶对Cd的富集效果最好,其次为Mo、Cu,对Fe、Mn、Zn、As和Pb的富集效果相对较差,说明碱蓬对沉积物中不同金属元素的富集移出率存在差异。金属元素在碱蓬中的转移系数研究表明,Mo、Zn、Cd和Mn等元素可由根部转移到叶中,而Fe、Ni、Pb、As和Cu等金属元素固定在根部,该研究可为利用碱蓬修复湿地重金属污染提供理论基础。     

Radioindication method for determining the site of uptake of herbicide acetochlor in maize

Effect of acetochlor placement on the uptake and distribution of carbonyl-¹⁴C labelled compound in maize /Zea Mays L./ was followed in time-course experiments by using charcoal barrier to separate the roots from shoot in sand culture. The results confirm that the main entrance of acetochlor to maize is through the roots but the uptake by coleoptile also plays an important role in the selectivity of acetochlor. Phytotoxic symptoms caused by acetochlor show good correlation with the site of uptake.     

Cd同位素地质样品的预处理方法研究

考察了文献报道的地质样品中Cd同位素测试前的化学分离纯化和富集方法(预处理方法)中的离子交换树脂单柱法回收率低的原因,并在该方法的基础上做了改进,优化了离子交换柱的洗涤顺序和洗液用量.改进后的离子交换树脂单柱法Cd回收率达99.82%,主要干扰元素(Sn、In、Zn、Pb)和基质元素的去除效果均较理想,能更好地满足Cd同位素的测定要求.     

Localization and chemical forms of cadmium in plant samples by combining analytical electron microscopy and X-ray spectromicroscopy

Cadmium (Cd) is a metal of high toxicity for plants. Resolving its distribution and speciation in plants is essential for understanding the mechanisms involved in Cd tolerance, trafficking and accumulation. The model plant Arabidopsis thaliana was exposed to cadmium under controlled conditions. Elemental distributions in the roots and in the leaves were determined using scanning electron microscopy coupled with energy dispersive X-ray microanalysis (SEM-EDX), and synchrotron-based micro X-ray fluorescence (μ-XRF), which offers a better sensitivity. The chemical form(s) of cadmium was investigated using Cd L_III-edge (3538 eV) micro X-ray absorption near edge structure (μ-XANES) spectroscopy. Plant μ-XANES spectra were fitted by linear combination of Cd reference spectra. Biological sample preparation and conditioning is a critical point because of possible artifacts. In this work we compared freeze-dried samples analyzed at ambient temperature and frozen hydrated samples analyzed at −170 °C. Our results suggest that in the roots Cd is localized in vascular bundles, and coordinated to S ligands. In the leaves, trichomes (epidermal hairs) represent the main compartment of Cd accumulation. In these specialized cells, μ-XANES results show that the majority of Cd is bound to O/N ligands likely provided by the cell wall, and a minor fraction could be bound to S-containing ligands. No significant difference in Cd speciation was observed between freeze-dried and frozen hydrated samples. This work illustrates the interest and the sensitivity of Cd L_III-edge XANES spectroscopy, which is applied here for the first time to plant samples. Combining μ-XRF and Cd L_III-edge μ-XANES spectroscopy offers promising tools to study Cd storage and trafficking mechanisms in plants and other biological samples.     

Absorption,transportation and distribution of imidacloprid in maize

The neonicotinoid imidacloprid is a very important insecticide in maize cultivation in China. A laboratory experiment was conducted to investigate its absorption, transportation and distribution in maize plants. Plants were exposed to an aqueous solution of imidacloprid at five concentrations (10, 20, 50, 100 and 200 μg/mL). The residues of imidacloprid in different plant parts were determined by using a quick and effective method of high-performance liquid chromatography. Results showed that the average recoveries ranged from 85.16% to 102.23%, with relative standard deviations of 1.82–4.40% at three different spiking levels in each different matrix. The half-lives of imidacloprid in hydroponic maize water were from 5.33 to 11.55 days. The concentrations in roots, stems and leaves were from 5.61 to 7.48, 1.03 to 4.03 and 0 to 30.57 μg/mL after 6 h–7 days exposed in 10 μg/mL imidacloprid aqueous solutions, respectively. Our study showed that imidacloprid was strongly absorbed by roots and mainly accumulated in leaves. Quantity ratio and bioconcentration factors (BCFs) were also used to estimate the distribution and accumulation in maize. The values of quantity ratios were the highest in the leaves while lowest in the root after 7 days treatments, with the ranking of leaves > stem > root. The BCFs were 0.63–1.66, 0.52–0.92 and 3.20–6.78 in root, stem and leaves, respectively. These results demonstrated that the exposed time and imidacloprid application concentrations were also the main factors influencing the absorption. This study enhances our understandings of the uptake and distribution of imidacloprid in maize plants.     

土壤-温郁金体系中重金属的吸收富集研究

通过盆栽实验研究了铅（Pb）、镉（Cd）、铜（Cu）、汞（Hg）和砷（As）污染条件下,温郁金和种植土壤中重金属含量,分析了植物对重金属的吸收富集能力。结果表明,随着土壤中污染物的投加量增强,Cu、Pb、Cd、As元素主要集中在根部,而Hg元素由于其极易挥发的特性,其块根中含量反而随着Hg投加量的增加而减小。温郁金对不同元素的吸收富集能力不同,对Cd的富集能力最强,对Hg的富集能力最弱。     

Cadmium(II) specifically interacts with cellular signaling to induce proto-oncogenes c-fos and c-jun in rat PC12 cells

Cadmium(II) compounds are carcinogens but only weakly mutagens. Because Cd(II) at low micromolar concentrations stimulates cell growth and induces some proto-oncogenes with several mammalian cell lines, the stimulation of cell proliferation is discussed as a major mechanism of the carcinogenicity of this metal. In the present work, the induction of the cellular proto-oncogenes c-fos and c-jun by Cd(II) was studied in rat PC12 cells. Several cellular signal transduction elements were tested as mediators of the proto-oncogene induction by cadmium. Cd(II) does not evoke mobilization of free intracellular Ca²⁺ in PC12 cells, and there was no impairment of the effect of Cd(II) by inhibitors of protein kinase A or of MAPK kinase. However, bisindolylmaleimide I, a specific inhibitor of protein kinase C abolished the proto-oncogene induction by Cd(II). Hence, a critical role for protein kinase C in the mitogenic effect of Cd(II) is inferred, and a substitution of structural Zn²⁺ ions by Cd²⁺ ions in this enzyme is discussed as the putative mechanism. Received: 30 July 1997 / Revised: 9 October 1997 / Accepted: 11 October 1997     

Separation of injectable salidroside by column chromatography of macroporous resins for treating myocardial ischemia

The objective of the present study is to develop a method for large-scale separating and purifying salidroside from rhodiola kirilowii roots and for preparing injectable medicinal ingredient.Crude extract of salidroside was prepared by water-ethanol system,and purified by column chromatography of macroporous resins.Static adsorption and desorption studies were performed on six kinds of macroporous resins,and SP825 resin was chosen,followed by optimizing process parameters.The optimum sample volume,feed concentration,ratio of diameter to height,and feeding flow rate were 1.5 bed volumes(BV),15 mg/mL,1:10 and 1 BV/h,respectively.Dynamic desorption was performed consecutively with 8 BV of distilled water,3 BV of 5% ethanol and 8 BV of 10% ethanol at a flow rate of 2 BV/h.After three cycles in separating 3.5 tons of rhodiola kirilowii roots,salidroside purity was increased from 3.4% in the crude extract to 93.6% in purified salidroside product.This study provides a novel method to separate salidroside for injectable use.     

Cadmium(II) specifically interacts with cellular signaling to induce proto-oncogenes c-fos and c-jun in rat PC12 cells

Cadmium(II) compounds are carcinogens but only weakly mutagens. Because Cd(II) at low micromolar concentrations stimulates cell growth and induces some proto-oncogenes with several mammalian cell lines, the stimulation of cell proliferation is discussed as a major mechanism of the carcinogenicity of this metal. In the present work, the induction of the cellular proto-oncogenes c-fos and c-jun by Cd(II) was studied in rat PC12 cells. Several cellular signal transduction elements were tested as mediators of the proto-oncogene induction by cadmium. Cd(II) does not evoke mobilization of free intracellular Ca²⁺ in PC12 cells, and there was no impairment of the effect of Cd(II) by inhibitors of protein kinase A or of MAPK kinase. However, bisindolylmaleimide I, a specific inhibitor of protein kinase C abolished the proto-oncogene induction by Cd(II). Hence, a critical role for protein kinase C in the mitogenic effect of Cd(II) is inferred, and a substitution of structural Zn²⁺ ions by Cd²⁺ ions in this enzyme is discussed as the putative mechanism. Received: 30 July 1997 / Revised: 9 October 1997 / Accepted: 11 October 1997     

Kinetic Studies on Ligand Substitution Reactions of Metallothioneins with DTPA and EDTA

Cd, Zn-thionelns were isolated from the rat liver by improved Winge method and purified further. The composition was determined. The mobilization of Cd ions from the Cd, Zn-metallothioneins was studied with spectrophotometric method. The results show that the reaction of DTP A or EDTA with Cd, Zn-thioneins occurs through three steps. The first step follows pseudo-first order reaction with association mechanism. The formation constants and the dissociation rate constants of the relevant intermediate ternary complexes and apparent rate constants, Kapp were determined. The properties and structure of Cd, Zn-thioneins and ligands with low molecular weights affect the values of Kapp.     

Determination of113mCd in natural water

A method has been developed for the determination of^113mCd from fallout in large-volume environmental water samples. In the analytical procedure a known amount of cadmium carrier is added to the sample and the cadmium is preconcentrated as insoluble hydroxide by coprecipitation with ferric hydroxide. The separated cadmium is purified from other interfering elements by an anion—exchange procedure. Finally, cadmium is coprecipitated with copper sulfide, and the radioactivity of^113mCd is measured in a lowbackground, gas-flow proportional beta counter. The cadmium recovery is established by determining the amount of Cd in the precipitate using an energy dispersive x-ray fluorescence spectrometer. For a 24hr counting period a lower limit of detection around 50μBq^113mCd/1 of sample has been attained. Samples of Lake Michigan water were analyzed: the concentration of^113mCd in 1980 was 177±5 μBq/l.     

Effect of selenium oxidation state on cadmium translocation in chamomile plants

Syntheses and spectral characteristics of cadmium(II) compounds (CdSeO₄, CdSeO₃, and Cd(NCSe)₂(nia)₂) containing selenium in oxidation states (VI), (IV), and (-II) are described. In Cd(NCSe)₂(nia)₂, nicotinamide (nia) and selenocyanate anions are bonded to Cd atom as N-donor monodentate ligands. Nicotinamide is coordinated through the ring nitrogen atom. The effects of these selenium compounds as well as Cd(NCS)₂(nia)₂ on the growth and Cd accumulation in roots and shoots of hydroponically cultivated chamomile plants (cultivar Lutea) were studied. In the applied concentration range (12–60 μmol dm⁻³) Cd(NCS)₂(nia)₂ affected neither the length nor the dry mass of roots and shoots. Other compounds applied at 24 μmol dm⁻³ and 60 μmol dm⁻³ significantly reduced dry mass of roots and shoots. Selenium oxidation state in the cadmium compounds affected Cd accumulation in plant organs as well as Cd translocation within the plants, which was reflected in the values of bioaccumulation (BAF) and translocation factors (S/R). Cd amount accumulated by shoots was lower than that in the roots. The highest BAF values determined for Cd accumulation in shoots were obtained with CdSeO₄. Substitution of S with Se in the Cd(NCX)₂(nia)₂ (X = Se or S) caused an increase of Cd translocation into the shoots. Presented at the XVIIIth Slovak Spectroscopic Conference, Spišská Nová Ves, 15–18 October 2006.     

Electrochemical preconcentration of ultra-trace Cd2+ from environmental and biological samples prior to its determination using carbon paste electrode impregnated with ion imprinted polymer nanoparticles

Cadmium (Cd) is known as one of the most toxic elements among the heavy metals. This study proposes an electrochemical preconcentration method using the ion imprinted polymer nanoparticles modified carbon paste electrode (IIP-CPE) for the stripping voltammetry determination of Cd²⁺ in the environmental and biological samples. These IIP NPs were synthesised using ethylene glycol dimethacrylate as the cross-linker, 2,2?-azobisisobutyronitrile as the free radical initiator and 4?-(4-vinylphenyl)-2,2?:6?,2?-terpyridine as the Cd-binding ligand. The calibration plot was linear in the concentration range of 4–500 nM (R² = 0.9936) with a sensitivity of 918 µA µM^?1 cm^?2. The LOD based on 3S_b/m (where m is the slope of the calibration curve and S_b is the standard deviation for five blank measurements) was found to be 1.94 nM. The relative standard deviations (RSDs) for single-electrode repeatability and electrode-to-electrode reproducibility were 3.0% and 5.6% (n = 5), respectively. The accuracy of the current method was confirmed by the analysis of urine quality control material (QCM, Seronorm^TM Trace Elements Urine, REF NO 201205, Norway) and spiked blood, rice and water samples. Recoveries were found to be above 95.0% for all samples, which confirms the good performance of the proposed voltammetry method.