镉(Ⅱ)与牛血清白蛋白的相互作用及pH、钙(Ⅱ)、锌(Ⅱ)和表面活性剂的影响

本文用恢复电位法、荧光分光光度法及粘度法研究了镉(Ⅱ)与牛血清白蛋白(BSA)的配位平衡和pH的影响。结果表明BSA有两类镉结合部位(2个强结合和10个弱结合部位)。Cd—BSA配位反应受pH影响,在近中性pH范围Cd(Ⅱ)与BSA结合未接近饱和时,以CURFIT程序对结合曲线拟合发现,每摩尔BSA平均结合Cd(Ⅱ)摩尔数的对数值与游离镉浓度的对数间成线性关系,进一步用Marguardt法处理得到Cd(Ⅱ)与BSA结合平衡方程式:N =K[Cd²⁺]^m,K和m均为pH函数。构象研究表明H⁺和Cd(Ⅱ)的结合都能引起BSA构象改变。用凝胶色谱法测定了Zn(Ⅱ)、Ca(Ⅱ)、吐温—80和十二烷基硫酸钠(SDS)对Cd—BSA结合的影响。Z_n(Ⅱ)可以竞争Cd(Ⅱ)在BSA上结合部位使Cd(Ⅱ)活动化。SDS能够有效抑制Cd(Ⅱ)与BSA结合,这种作用可能与SDS引起BSA构象改变有关。     

螯合剂对结合在人血清白蛋白(HSA)上的铜离子活动化能力的研究

用凝胶色谱法研究了螯合剂使结合在人血清白蛋白上的铜(Ⅱ)活动化的能力,若用在螯合剂存在时和无螯合剂存在时蛋白部分结合金属量之比作为表征活动化能力的参数,有如下顺序:Trien>DTPA>EDTA>EGTA>NTA>IDA除Trien和DTPA外,其他几种螯合剂的活动化能力的对数值与其相应的条件稳定常数的对数值之间有一线性关系.Trien和DTPA的反常被认为与复杂的反应历程有关,这可能涉及三元配合物的形成。     

螯合剂对结合在人血清白蛋白(HSA)上的铜离子活动化能力的研究

用凝胶色谱法研究了螯合剂使结合在人血清白蛋白上的铜(Ⅱ)活动化的能力,若用在螯合剂存在时和无螯合剂存在时蛋白部分结合金属量之比作为表征活动化能力的参数,有如下顺序:Trien>DTPA>EDTA>EGTA>NTA>IDA除Trien和DTPA外,其他几种螯合剂的活动化能力的对数值与其相应的条件稳定常数的对数值之间有一线性关系.Trien和DTPA的反常被认为与复杂的反应历程有关,这可能涉及三元配合物的形成.     

火焰原子吸收光谱法研究五种螯合剂对染镉小鼠睾丸中镉钙铁锌浓度的影响

采用火焰原子吸收光谱法分别对染镉小鼠和N-对羟甲苯甲基-D-葡糖二硫代氨基甲酸钠(HBGD)、N-苯甲基-D-葡糖二硫代氨基甲酸钠(BGD)、二乙基二硫代氨基甲酸钠(DDTC)、二巯基丙醇(BAL)、乙二胺四乙酸(EDTA)等五种螯合剂治疗组小鼠睾丸中镉、钙、铁、锌等金属浓度进行了测定和研究.结果表明:镉染毒(Cd 2.5 mg·kg-1,腹腔注射)后,小鼠睾丸中镉、钙、铁、锌等金属浓度显著高于对照组(p＜0.05),染镉30 min后用各螯合剂治疗(0.400 mmol·kg-1,腹腔注射),24 h后除EDTA外,其余螯合剂能明显降低睾丸中镉的浓度并抑制钙、铁、锌浓度的增加(p＜0.05);染镉24 h后注射各螯合剂,治疗24 h后,HBGD、BGD和DDTC显著降低了小鼠睾丸中各金属浓度(p＜0.05);提示HBGD和BGD有望成为染镉中毒的理想解毒剂.     

镉离子单克隆抗体的制备及其间接竞争酶联免疫分析方法的建立

制备镉离子单克隆抗体,建立快速检测水样中镉离子的间接竞争酶联免疫吸附分析方法(EnzymeLinked Immunosorbent Assay,ELISA)。选择双功能螯合剂1-(4-异硫氰苄基)乙烯基二胺-N,N,N’,N’-四乙酸(ITCBE),分别与载体蛋白(牛血清白蛋白,BSA;卵清蛋白,OVA)和镉离子连接,制备出免疫原(Cd-ITCBEBSA)和包被抗原(Cd-ITCBE-OVA),采用免疫原免疫BALB/C小鼠,利用细胞融合杂交瘤技术制备出针对镉离子的单克隆抗体;建立快速检测水样中镉离子的间接竞争ELISA方法。结果:方法的IC_(50)为13.67 ng·m L~(-1),检测限为0.48 ng·m L~(-1)。该抗体除与汞离子的螯合物的交叉反应率为79.29%之外,与Zn~(2+)、Cr~(3+)、Pb~(2+)、Cu~(2+)、Ni~(2+)、Al~(3+)、Co~(2+)、Fe~(3+)、Mg~(2+)、Ca~(2+)和Ba~(2+)均无交叉反应。超纯水、自来水和湖水样品中镉的加标回收率为87.78%~103.3%,相对标准差为1.60%~11.41%。建立的间接竞争酶联免疫吸附法灵敏、快速,可用于水样中镉离子的检测。     

螯合剂与牛血清白蛋白竞争镍作用的研究

用凝胶色谱法研究了13种螯合剂使牛血清白蛋白镍配合物BSANi(Ⅱ)中的镍活动化的能力,发现DTPA、NTA等氨羧类螯合剂有较好的竞争能力,而巯基类较差。用透析法研究了螯合剂与BSANi(Ⅱ)反应动力学机理,结果表明,在pH7.4和4℃时,L-组氨酸、EDTA为解离机理,而亚氨二乙酸,二乙三氨五乙酸为加合机理。测定了有关的动力学参数,依据竞争能力参数F讨论了可能作为治疗镍中毒的促排螯合剂。     

镉离子对三种黄酮小分子与蛋白质相互作用的生物无机化学效应

用荧光光谱法及紫外分光光度法研究了镉离子(Cd~(2+))对3种黄酮小分子槲皮素(Que)、金丝桃苷(Hyp)及芦丁(Rut)与牛血清白蛋白(BSA)相互作用的生物无机化学效应。对比了有无Cd~(2+)存在下,3种黄酮与血清蛋白的结合常数、结合位点数、作用力及蛋白质构象等参数,结果表明:Cd~(2+)与Hyp、Rut的结合部位位于其糖苷基团上,Cd~(2+)在3种黄酮与BSA分子之间起"离子架桥"作用并直接参与了黄酮与BSA之间的结合,使两者更有利于插入蛋白质的疏水腔内部;而Que母核C环3位的羟基却不利于此"离子架桥"作用。"离子架桥"是Cd~(2+)对黄酮小分子与BSA相互作用过程的主要参与机制。     

牛血清白蛋白与Cu(Ⅱ)、Cu(Ⅰ)相互作用的表面电化学研究

采用循环伏安法,研究了牛血清白蛋白(BSA)吸附到玻碳电极上构成的BSA修饰电极.考察了铜离子在BSA修饰电极上的电化学行为.结果表明,Cu(Ⅱ)、Cu(Ⅰ)通过非疏水(静电或者共价)作用与电极表面的BSA结合,Cu~(2+)在BSA修饰电极上与BSA的结合能力比Cu~+与BSA的结合能力强,使用该方法探究蛋白与金属离子、小分子的作用是可行的.     

高分子螯合剂稳定垃圾焚烧飞灰中的重金属

建立了用电感耦合等离子体质谱仪测定飞灰浸出液中重金属浓度的方法。该方法线性范围宽,0~500 ppb;检出限低,0.005~2.1 ppb;准确度高,加标回收率在86%~103%;可同时测定多种元素。以聚乙烯亚胺基黄原酸钠处理垃圾焚烧飞灰,考察高分子螯合剂添加量、反应时间和浸提剂的pH值对螯合效率的影响。螯合剂添加量为飞灰质量的2%时,高分子螯合剂对铜、镉的螯合效率大于99%,对铅、镍、铬的螯合效率大于96%,对锌、硒的螯合效率大于92%。经稳定化处理后,飞灰中所有重金属元素的浸出浓度均低于《生活垃圾填埋场污染控制标准》中的限值。     

三种抗氧化物质与牛血清白蛋白的相互作用

运用荧光光谱(FS)、紫外光谱(UV)和超滤(UF)法, 研究了阿魏酸、毛蕊异黄酮及芒柄花素三种抗氧化小分子物质与牛血清白蛋白(BSA)的结合反应. 以Lineweaver-Burk双倒数方程和能量传递原理分别计算了三者与BSA反应的结合常数(K_LB)和结合距离(r)以及由热力学参数的计算判断了三种小分子物质与BSA结合的作用力类型; 以Scatchard方程计算了三种小分子与BSA的结合常数(K_a)和结合位点数(n); 并对阿魏酸与毛蕊异黄酮及芒柄花素的竞争结合反应进行了研究. 实验结果表明: 这三种物质能与BSA结合形成非共价复合物, 导致BSA内源性荧光的静态猝灭; 它们与BSA的结合能力随着极性基团的增多、分子体积的增大而减弱; 静电作用和疏水作用可能是它们与BSA结合的主要作用力; 阿魏酸与芒柄花素或毛蕊异黄酮会产生与BSA的竞争结合效应.     

STUDIES ON THE COMPETITION FOR CADMIUM ION BETWEEN CHELATING AGENTS AND BOVINE SERUM ALBUMIN

Eleven chelating agents were studied for their capabilities to mobilize the cadmium bound tp bovine serum albumin(BSA).The parameter F,which is defined as the ratio between the percentages of cadmium bound to BSA in the presence and absence of chelating agents,can be used as the criterion to evaluate the mobilizing capability of chelating agent.The F values determined experimentally lead to a mobilizing capability order:DTPA>EDTA>EGTA>NTA>TR1EN>PEN>CYS>HIS>SAThe polyaminopolycarboxylate type chelators mobilize cadmium effectively.A linear relationship was found between 1gF and lg k'CdL (conditional stability constant of the cadmium chelate).     

Effects of three dithiocarbamates on tissue distribution and excretion of cadmium in mice

N-Benzyl-D-glucamine dithiocarbamate (BGD), N-p-hydroxymethylbenzyl-D-glucamine dithiocarbamate (HBGD), and N-p-carboxybenzyl-D-glucamine dithiocarbamate (CBGD) were compared for their relative efficacies in the distribution and excretion of cadmium in mice exposed to cadmium. Mice were injected intraperitoneally with 109CdCl2 (1 mg of Cd/kg and 2 microCi of 109Cd/one animal). Three days later, they were injected with chelating agents (400 mumol/kg) every other day for 2 weeks. After injections of BGD and HBGD, cadmium was excreted mainly in the feces through the bile, and the fecal excretion of cadmium by HBGD was significantly higher than that by BGD or CBGD. These chelating agents increased the urinary excretion of cadmium to a small extent. The hepatic cadmium content was decreased only after HBGD injection. Also, the injection of HBGD caused a much greater decrease in renal cadmium content than did BGD or CBGD. These chelating agents did not result in the redistribution of cadmium to the brain, testes, or heart. The growth of mice was only slightly retarded by injections of these chelating agents. The results of this study indicate that the injection of HBGD to mice pretreated with cadmium can remove cadmium from the body, mainly through fecal excretion, without redistribution of cadmium to other tissues such as the brain, testes, and heart, more effectively than BGD or CBGD.     

2，3－二羟墓－5－甲氧羰基苄基胺羧酰胺螯合剂促排铀酰的计算机模拟研究

用改进的计算机模型模拟研究了铀酰离子中毒血液体系在加入２，３－二羟基－５－甲氧羰基苄基胺羧酰胺螯合剂前后的各种配位平衡，讨论了铀酰离子的中毒浓度和螯合剂的剂量对螯合效率的关系，评估了这些螯合剂对铀酰离子的迁移能力和对体内必需金属离子的影响．     

2,3-二羟墓-5-甲氧羰基苄基胺羧酰胺螯合剂促排铀酰的计算机模拟研究

用改进的计算机模型模拟研究了铀酰离子中毒血液体系在加入２，３－二羟基－５－甲氧羰基苄基胺羧酰胺螯合剂前后的各种配位平衡，讨论了铀酰离子的中毒浓度和螯合剂的剂量对螯合效率的关系，评估了这些螯合剂对铀酰离子的迁移能力和对体内必需金属离子的影响。     

Analyzing several chelating agents and their effect on elemental composition of Lolium perenne and two growth media by capillary zone electrophoresis and inductively coupled plasma optical emission spectrometry

Capillary zone electrophoresis (CZE) along with ICP-OES was used for the determination of several chelating agents in clay and black soil, and in plants (Lolium perenne) grown in these matrices. Elemental levels in all the samples (Al, As, Ca, Cd, Cu, Fe, K, Mg, Mn, Pb, S and Zn) were investigated by inductively coupled plasma optical emission spectrometry (ICP-OES). Compared to previous methods, much shorter total analysis times were accomplished by the validation of an internal standard method for CZE analysis. The solubility of elements increased noticeably in the soils treated by the chelating agents before cultivation, and this phenomenon was strictly metal complex stability constant dependent. The most persistent chelating agents were triethylenetetraaminehexaacetic acid (TTHA) and 1,3-diaminopropanetetraacetic acid (PDTA). Both agents were bound to the growth medium and taken up by ryegrass, while only small proportions of them were degraded during cultivation. Diethylenetriaminepentaacetic acid (DTPA), ethylenediaminetetraacetic acid (EDTA) and nitrilotriamineacetic acid (NTA), mainly bound to the clay matrix during the 19 days cultivation, but after 40 days cultivation in black soil they were degraded to smaller acids. The results clearly show uptake of chelating agents by plants and are a clear indication of the degradation of chelating agents.     

Assembly of tetra, di and mononuclear molecular cadmium phosphonates using 2,4,6-triisopropylphenylphosponic acid and ancillary ligands

The reaction of ArPO(3)H(2) (Ar = 2,4,6-iPr(3)-C(6)H(2)) with Cd(CH(3)COO)(2).2H(2)O using various co-ligands such as methanol, dimethylformamide (DMF) and 3,5-dimethylpyrazole (DMPZH) resulted in the formation of tetranuclear assemblies [Cd(4)(ArPO(3))(2)(ArPO(3)H)(4)(CH(3)OH)(4)].3(CH(3)OH) (1), [Cd(4)(ArPO(3))(2)(ArPO(3)H)(4)(DMF)(4)].3(DMF) (2) and [Cd(4)(ArPO(3))(2)(ArPO(3)H)(4)(DMF)(2)(DMPZH)(2)].2(DMF).2(H(2)O) (3). In all of these compounds the tetranuclear cadmium array, containing two five-coordinate and two six-coordinate cadmium atoms, is held together by two mu(4) capping [ArPO(3)](2-) and four anisobidentate mu(2) [ArPO(2)(OH)](-) ligands. Each cadmium atom is bound to an additional ancillary ligand. The reaction of ArPO(3)H(2) with Cd(CH(3)COO)(2).2H(2)O in the presence of the chelating ligand 2,2'-bipyridine (bipy) leads to the exclusive formation of the dinuclear assembly [Cd(2)(ArPO(3)H)(4)(bipy)(2)].(CH(3)OH)(H(2)O) (4). The latter contains an eight-membered Cd(2)P(2)O(4) inorganic ring formed as a result of the bridging coordination action of two anisobidentate mu(2) [ArPO(2)(OH)](-) ligands. Each cadmium atom is bound by one chelating bipy and one monodentate [ArPO(2)(OH)](-) ligands. Use of four equivalents of 3,5-dimethylpyrazole leads to the formation of the mononuclear derivative [Cd(ArPO(3)H)(2)(DMPZH)(4)] (5). The molecular structure of the latter comprises of a central cadmium atom surrounded by six monodentate ligands. Four of these are neutral pyrazole ligands that occupy the equatorial plane; the remaining two are anionic phosphinate ligands which are present trans to each other. The thermal analysis of 1 and 4 reveals that the char residue obtained at 600 degrees C consists predominantly of Cd(2)P(2)O(7).     

Synthesis and metal binding properties of salicylate-, catecholate-, and hydroxypyridinonate-functionalized dendrimers

The synthesis, characterization, and metal-binding studies of chelate-functionalized dendrimers is reported. Salicylate, catecholate, and hydroxypyridinonate bidentate chelators have been coupled to the surface of both poly(propyleneimine) (Astramol) and poly(amidoamine) (Starburst, PAMAM) dendrimers up to the fifth generation (64 endgroups). A general method has been developed for the facile and high quality chromatographic purification of poly(propyleneimine) and poly(amidoamine) dendrimer derivatives. One- and two-dimensional (TOCSY) 1H NMR experiments and electrospray ionization mass spectrometry (ESI-MS) have confirmed the exhaustive coupling of these chelators to the primary amine functionalities of the dendrimers. Spectrophotometric titrations were used to investigate the metal binding ability of these macrochelates. Spectral analysis shows that ferric iron binding to these ligands is localized to the chelating endgroups. The ability of these dendritic polymers to bind large numbers of metal ions may lead to applications as metal sequestering agents for waste remediation technologies.     

工程化蛋白质纳米孔实时监测镉离子与谷胱甘肽的单分子反应

本研究利用合成的全-6-季铵基-β-环糊精(Per-6-quaternary ammonium-β-cyclodextrin,p-QABCD)装备基因工程化的α-溶血素(α-Hemolysin,αHL)蛋白质纳米孔(M113R)7,构建全新的单分子纳米孔反应器,在单分子水平实现对谷胱甘肽(GSH)和镉离子(Cd2+)的络合反应的实时原位监测,并辨认络合反应的不同路径、反应中间产物和最终产物.结果表明,溶液的pH值显著影响GSH与Cd2+络合产物的络合比例.pH=7.4时,GSH与Cd2+络合反应的最终产物为Cd(GSH)2;pH=9.0时,最终产物为Cd(GSH)2和Cd2(GSH)2.其中,Cd2(GSH)2的形成遵循两种路径:(1)一个Cd2+首先结合两个GSH分子的巯基形成Cd(GSH)2,然后,第二个Cd2+结合去质子化的氨基形成Cd2(GSH)2;(2)两个Cd2+分别结合同一个GSH分子的巯基和去质子化的氨基形成Cd2(GSH)1,然后,第二个GSH分子的巯基和去质子化的氨基结合Cd2(GSH)1的Cd2+形成Cd2(GSH)2.本研究实现了在单分子水平无标记和无化学修饰研究金属离子和生物小分子的反应,对理解细胞内重金属的解毒机理和拓展纳米孔单分子技术的研究领域具有重要意义.