过氧化物模拟酶催化荧光反应性能的研究——FeTMPvP-HVA-H_2O_2体系

本文研究了moso-四(N-甲基-3-吡啶基)卟啉和铁的络合物作为辣根过氧化物酶的模拟酶催化过氧化氢氧化高香草酸的荧光反应的性能。拟订了用模拟酶催化测定H_2O_2和葡萄糖的荧光测定方法。其检出下限分别为1.1×10~(-7)mol/LH_2O_2和0.2μgml~(-1)葡萄糖。对血清中葡萄糖的含量进行了测定,结果令人满意。通过动力学研究,比较了该模拟酶与辣根过氧化物酶及其它模拟酶催化活性的相对大小。     

牛血红蛋白催化化学发光反应与FIA联用测定人血清中的血糖

基于葡萄糖氧化酶催化葡萄糖氧化并释放过氧化氢,再将其偶合到含鲁米诺的发光体系中;同时采用牛血红蛋白作为发光体系的催化剂,其发光强度在一定范围内与过氧化氢的浓度呈良好的线性关系,据此建立了一种间接测定人体血清中葡萄糖含量的新方法。本法与FIA技术联用,进样频率为112样/小时,线性范围为3.0×10-6~1.0×10-4mol.L-1,检出限可达1.4×10-7mol.L-1。     

催化显色反应的研究 (Ⅹ)铜(Ⅱ)-没食子酸-过氧化氢-活化剂体系

有关铜的催化显色反应已有不少研究,Cu~(2+)催化过氧化氢氧化多元酚的反应也有报导,但卤阴离子和Al~(3+)对该反应的活化作用是最近才发现的。本文研究了Cu~(2+)-没食子酸(GA)-H_2O_2催化显色体系,发现F~-、Cl~-、Br~-对该体系具有活化作用,适宜浓度的Cl~-可使该反应灵敏度提高十倍,若再加入Al~(3+),还可以大大加快反应速度。没食子酸与邻苯二酚相比性质较稳定,水中溶解度颇大,易于配制和保存。基于该反应拟订了测定铜的催化光度法,用于测定血清和纯锌样品中的痕量铜,结果尚好。     

基于聚多巴胺球负载Cu纳米粒子的催化作用分光光度法测定人血中葡萄糖含量北大核心CSCD

用水热法在聚多巴胺球(PDA)表面负载了Cu纳米粒子(Cu@PDA),制备了具有过氧化物酶活性的Cu@PDA复合物。此复合物能催化过氧化氢快速氧化无色还原态的四甲基联苯胺(TMB),使其转变为蓝色的氧化态TMB,其吸收峰波长为652 nm,其吸光度与过氧化氢的浓度有关。利用此催化反应结合葡萄糖氧化酶(Gox)催化经氧气氧化葡萄糖而产生过氧化氢的反应,提出了分光光度法测定血样中葡萄糖的方法。在总体积为10 mL的0.1 mol·L^(-1)磷酸盐缓冲溶液(pH 4.0)中含1.0 mg·L^(-1)Cu@PDA,4.0 g·L^(-1)Gox,0.5 mmol·L^(-1)TMB的条件下加入血样,于40℃反应10 min时,葡萄糖浓度在1.0~30.0 mmol·L^(-1)的范围内与相应的吸光度之间呈线性关系,检出限(3S/N)为0.3 mmol·L^(-1)。方法应用于人血样品的分析,所测得葡萄糖含量与医院实验室所给的值相符。     

基于聚多巴胺球负载Cu纳米粒子的催化作用分光光度法测定人血中葡萄糖含量

用水热法在聚多巴胺球(PDA)表面负载了Cu纳米粒子(Cu@PDA),制备了具有过氧化物酶活性的Cu@PDA复合物。此复合物能催化过氧化氢快速氧化无色还原态的四甲基联苯胺(TMB),使其转变为蓝色的氧化态TMB,其吸收峰波长为652 nm,其吸光度与过氧化氢的浓度有关。利用此催化反应结合葡萄糖氧化酶(Gox)催化经氧气氧化葡萄糖而产生过氧化氢的反应,提出了分光光度法测定血样中葡萄糖的方法。在总体积为10 mL的0.1 mol·L~(-1)磷酸盐缓冲溶液(pH 4.0)中含1.0 mg·L~(-1)Cu@PDA,4.0 g·L~(-1)Gox,0.5 mmol·L~(-1)TMB的条件下加入血样,于40℃反应10 min时,葡萄糖浓度在1.0~30.0 mmol·L~(-1)的范围内与相应的吸光度之间呈线性关系,检出限(3S/N)为0.3 mmol·L~(-1)。方法应用于人血样品的分析,所测得葡萄糖含量与医院实验室所给的值相符。     

牛血红蛋白催化分光光度法测定过氧化氢

在pH 4.8的0.1mol·L-1柠檬酸-0.2mol·L-1磷酸氢二钠缓冲介质中,于43℃水浴加热60min,牛血红蛋白对过氧化氢氧化邻苯二胺的反应有明显的催化作用。试验结果表明:过氧化氢浓度在8.24×10-6~2.31×10-4 mol·L-1范围内与相应吸光度的增加值ΔA呈线性关系,检出限(3s/k)为4.50×10-8 mol·L-1。据此提出了牛血红蛋白催化分光光度法测定过氧化氢的含量。采用此法测定消毒液和雨水中过氧化氢的含量,测定值的相对标准偏差(n=5)分别为0.081%和1.6%,消毒液中过氧化氢的测定值与滴定法测定结果相符。以上述2种试样作基体用标准加入法进行回收试验,测得回收率分别为99.3%和97.3%。     

维生素C氧化反应的研究: H2O2-Cu^2^+-磷酸体系中的反应动力学

研究了在铜离子存在时的弱酸性条件下, 用过氧化氢氧化维生素C, 并用直接分光光度法测定未变化的酸, 表明维生素C的催化氧化是以链式反应机制进行的.     

四磺基锰酞菁作为过氧化物模拟酶在过氧化氢测定中的应用

1　引　　言过氧化氢的测定在医学和环境科学中都具有很重要的意义。目前已有许多测定过氧化氢的方法 ,如光度法、荧光法、电分析法及化学发光法等。四磺基锰酞菁 (MnTSPc)是一种与金属卟啉有着类似母体结构的金属酞菁类化合物 ,它具有过氧化物模拟酶的性质。继前文用对羟基苯乙酸和L 酪氨酸两种经典荧光底物成功实现对过氧化氢进行荧光分析测定后 ,我们又发现 ,用邻苯二胺 (OPD)做底物 ,以MnTSPc催化其与过氧化氢反应 ,也可实现对过氧化氢的高灵敏测定。本文研究了此催化荧光反应的最佳条件 ,建立了测定过氧化氢的催化荧光分析新方法 …     

微波消解-磷钼蓝分光光度法测定城市污泥中总磷浓度

建立了微波消解-磷钼蓝分光光度法测定城市污泥中总磷的方法。在微波环境中硝酸-过氧化氢能够将城市污泥中无机磷盐和含磷有机物消解为正磷酸盐,在弱酸性条件下,正磷酸盐在铋盐的催化条件下与钼酸铵-抗坏血酸生成磷钼蓝,于分光光度计波长690nm处进行测定总磷浓度。方法的相对标准偏差0.63%～0.95%,加标回收率为101%～102%,能够满足对城市污泥中总磷浓度的测定要求。     

辣根过氧化物酶电极在H2O2分析中的应用

辣根过氧化物酶(HRP)能催化过氧化氢与氢供体间的氧化还原反应,是当今生物传感器研究的热点之一.HRP分子内含有α-D-葡萄糖和α-D-甘露糖,是一种糖蛋白,在pH 7.0下,能与具有识别α-D-葡萄糖和α-D-甘露糖功能的外源植物凝集素伴刀豆球蛋白(Con A)结合.通过Con A与HRP之间的识别作用在半胱氨酸修饰的金表面构造HRP多层自组装膜电极,以亚甲蓝(MB)溶液为介体,对电极进行了电化学表征,并用该酶电极测定了过氧化氢浓度.     

过氧化物模拟酶催化的苯基荧光酮氧化反应及其分析应用

在ＮＨ４Ｃｌ－ＮＨ４ＯＨ缓冲介质中，氯化血红素（Ｈｅｍｉｎ）有显著的过氧化物模拟酶活性，催化过程化氢氧化苯基荧光酮褪色。本文探讨了反应机理，比较Ｈｅｍｉｎ与天然酶催化性能，考察反应条件和共存物质影响，从而提出测定Ｈｅｍｉｎ和过氧化氢的高灵敏分光光度法，线性范围分别为０￣３．０×１０＾－８ｍｏｌ／Ｌ和０￣１．２×１０＾５ｍｏｌ／Ｌ；检测限（３σ）分别为１．８×１０＾－１０ｍｏｌ／Ｌ和１．４×１０＾７     

Construction of Supramolecular Nanostructures with High Catalytic Activity by Photoinduced Hierarchical Co-Assembly

Inspired by natural enzymes, hierarchical catalytic supramolecular nanostructures were developed by the co-assembly of hemin and glucose oxidase (or Au NPs) with the photosensitive ferrocene–tyrosine (Fc-Y) molecule. Illuminated by white light, the Fc-Y molecules are polymerized and co-assemble with hemin into truncated polyhedrons. The Au NPs grew in situ at the surface of the co-assembled polyhedrons, achieving ordered supramolecular nanostructures. Because the Au NPs can serve as an artificial glucose oxidase and the hemin could act as a peroxidase mimic, the supramolecular hybrid nanostructures were used to mimic natural enzymes and catalyze the glucose conversion cascade reaction. The hybrid Au NPs@Fc-Y&hemin polyhedrons showed superior catalytic activity, good reusability, and maintained the catalytic activity over a wide temperature and pH range. The study demonstrates a feasible strategy to construct hierarchical co-assembled supramolecular nanostructures as multi-enzyme mimics, with potential applications in biocatalysis and biosensing.     

轴向配基对HRP模拟酶Hemin催化荧光及化学发光反应影响的研究

以氯化血红素作为辣根过氧化物酶（ＨＲＰ）的模拟物,对二十种氨基酸和其他含氮有机配体的轴向配位效应进行了研究。发现在氯化血红素催化的荧光和化学发光体系中,组氨酸和咪唑均使ｈｅｍｉｎ的催化活性显著提高,而酪氨酸,色氨酸,Ｌ－半胱氨酸及卤代烷基吡啶等具有淬灭效应。     

A novel mimetic peroxidase catalyst by using magnetite-containing silica nanoparticles as carriers

A method for coating magnetite and mimetic enzyme (hemin) with amorphous silica to form a novel mimetic peroxidase (magnetite–hemin/SiO₂) has been developed by combining reverse microemulsion and the modified Stöber method. The magnetic silica nanoparticle supported hemin has a long-term stability toward temperature and good reusability. They can be easily separated from the reaction solution by using an external magnetic field and reused directly for next round of reaction. The peroxidase activity of the magnetite–hemin/SiO₂ was studied based on its catalytic effect on the reaction of p-hydroxyphenylacetic acid and H₂O₂. The results indicated that the catalytic activity of the new mimetic enzyme catalyst is higher than that of the free hemin. The possibility of its application was proven by the determination of H₂O₂, with the detection limits of 7.3 nmol L^?1 H₂O₂.     

基于酶催化反应的核酸定量新方法

近年来 ,将染料自缔合或诱导缔合用于核酸定量测定备受关注 [1～ 3 ] .但是将酶与染料的缔合用于核酸定量测定尚未见报道 .氯化血红素 (hemin)可作为辣根过氧化物酶 (HRP)的模拟酶 ,能催化 H2 O2氧化对 -羟基苯乙酸 (p- HPA)生成荧光产物——联二对 -羟基苯乙酸的反应 [4 ,5] .由于 hemin在碱性介质中是阴离子化合物 ,能与阳离子化合物如阿尔新蓝 (Alcian Blue 8GX)发生缔合作用 ,从而使自身的催化性质被抑制 .当加入带负电荷的脱氧核糖核酸 (DNA)时 ,由于阿尔新蓝与 DNA的强烈作用使hemin与阿尔新蓝的缔合物被破坏 ,hemin的催化活…     

Spectrophotometric Determination of Hydrogen Peroxide and Glucose Based on Hemin Peroxidase-Like Catalyzed Oxidation of Bromopyrogallol Red

In an ammonium buffer medium at pH 8.9–9.5, hemin exhibits mimetic peroxidase activity, and has a catalytic effect on the oxidative decoloration of bromopyrogallol red (BPR) with hydrogen peroxide. On this basis and in presence of ethanol as an effect-enhancing agent, a spectrophotometric determination of hydrogen peroxide is described with an apparent molar absorptivity of 4.00×10⁴?l?mol^?1?cm^?1 and a linear range from 3.2×10^?7 to 3.2×10^?5?mol?l^?1. BPR has advantages over some of widely used chromogenic substrates in aspects of sensitivity, simplicity and detection wavelength, while hemin has better stability than peroxidase. The system can be easily coupled with a glucose oxidase-catalyzed reaction, and glucose in the concentration range of 6.0×10^?7? 3.2×10^?5?mol?l^?1 is spectrophotometrically determined. The method has been applied to the analyses of synthetic water and human serum samples. The Michaelis parameters and the mechanism of the mimetic peroxidase reaction are also investigated.     

Characterization of G-quadruplex/hemin peroxidase: substrate specificity and inactivation kinetics

Recently, G-quadruplex/hemin (G4/hemin) complexes have been found to exhibit peroxidase activity, and this feature has been extensively exploited for colorimetric detection of various targets. To further understand and characterize this important DNAzyme, its substrate specificity, inactivation mechanism, and kinetics have been examined by comparison with horseradish peroxidase (HRP). G4/hemin DNAzyme exhibits broader substrate specificity and much higher inactivation rate than HRP because of the exposure of the catalytic hemin center. The inactivation of G4/hemin DNAzyme is mainly attributed to the degradation of hemin by H(2)O(2) rather than the destruction of G4. Both the inactivation rate and catalytic oxidation rate of G4/hemin DNAzyme depend on the concentration of H(2)O(2), which suggests that active intermediates formed by G4/hemin and H(2)O(2) are the branch point of catalysis and inactivation. Reducing substrates greatly inhibit the inactivation of G4/hemin DNAzyme by rapidly reacting with the active intermediates. A possible catalytic and inactivation process of G4/hemin has been proposed. These results imply a potential cause for the hemin-mediated cellular injury and provide insightful information for the future application of G4/hemin DNAzyme.     

Application ofo-Hydroxyphenylfluorone as a Fluorogenic Indicator to the Determination of Hydrogen Peroxide and Oxidase Substrates Based on the Catalytic Effect of Hemin

A highly sensitive fluorescence quenching method has been developed for selective determination of hydrogen peroxide based on the catalytic effect of hemin on theo-hydroxyphenylfluorone (a new fluorogenic substrate) and hydrogen peroxide system. Under optimum conditions, the calibration graph was linear over the range 0–1.0 × 10⁻⁶mol/liter hydrogen peroxide, with a limit of detection of 8.0 × 10⁻⁹mol/liter in a 10-min reaction period. It can easily be incorporated into the determination of biochemical substances that produce hydrogen peroxide under catalytic oxidation by their oxidase. This possibility has been tested for the determination of glucose in human sera as an example.     

氯化血红素催化氧化巯基形成二硫键

对氯化血红素催化氧化巯基形成二硫键的反应进行了研究,发现N,N-二异丙基乙胺(DIEA)的加入可以提高氯化血红素的催化活性,并降低其在氧化过程中的自聚现象.在室温及少量DIEA存在下,将氯化血红素和巯基乙酸甲酯按摩尔比1∶4混合于p H=8.0的水溶液中,敞口搅拌反应20 min,可以催化空气氧化90%的巯基乙酸甲酯形成相应的分子间二硫键产物.该催化氧化体系还可应用于多肽合成中,在相同条件下,只需2 h即可完成还原型催产素和利那洛肽的氧化环合,生成高产率的催产素和利那洛肽环肽.与传统的氧化方法相比,氯化血红素催化氧化的方法具有高效、环保的优点,为多肽合成中二硫键的形成提供了一种新方法.     

The electrogenerated chemiluminescent behavior of hemin and its catalytic activity for the electrogenerated chemiluminescence of lucigenin

The electrogenerated chemiluminescent (ECL) behavior of hemin at a platinum electrode in the alkaline solution has been investigated in detail. Under the optimum conditions the linear response range of hemin is 1.0×10⁻⁵–1.0×10⁻⁸ g ml⁻¹, the detection limit was 1.0×10⁻⁸ g ml⁻¹, and the relative standard derivation for 1×10⁻⁷ g ml⁻¹ hemin was 2.8%. It has been also found that hemin would catalyze the ECL of lucigenin at a platinum electrode in a neutral solution in the presence of hydrogen peroxide, the catalytic ECL intensity was linear with the concentration of hemin in the range of 1.0×10⁻¹⁴–1.0×10⁻¹⁰ g ml⁻¹. IgG labeled with hemin was used to examine the ECL catalytic activity of hemin after conjugating to protein, and the results showed that hemin retained ECL catalytic activity when conjugated to protein.