OAP-H2O2-HRP伏安酶联免疫分析新体系测定人血清总甲状腺素

提出邻氨基酚（ＯＡＰ）－Ｈ２Ｏ２－辣根过氧化物酶（ＨＲＰ）发酶联免疫分析新体系并用于人血清中总甲状腺素的测定。本方法以线性扫描二阶导数伏安法检测ＨＲＰ催化Ｈ２Ｏ２氧化ＯＡＰ的产物,用于游离ＨＲＰ和ＨＲＰ标记物的测定,灵敏度的高于经典的ＥＬＩＳＡ显色光度法。测定游离ＨＲＰ的线性范围为１．０×１０＾１２￣４．０×１０＾－９ｇ／ｍＬ,检测限达６．０×１０＾１３ｇ／ｍＬ。本法对总甲状腺素测定的线性范围为１     

硝基磺酚S作为酶联免疫分析中辣根过氧化物酶测定底物的研究

提出一种新的ＨＲＰ底物－硝基磺酚Ｓ,初步探讨了在ＨＲＰ催化下被过氧化氢氧化的机制,以硝基磺酚Ｓ为底物测定了酶促反应动力学常数Ｋｍ和Ｋｓ分别为４．５１×１０＾－５ｍｏｌ／Ｌ和２８．１ｍｏｌ／Ｌ·ｓ＾－１。分别用于酶联免疫显色光度法和伏安酶联免疫分析法测定了ＩｇＧ－ＨＲＰ,其所能测定的最高稀释比分别为１：１．６×１０＾５和１：４．８×１０＾５。     

高锰酸钾—抗坏血酸化学发光体系测定抗坏血酸

本文研究了高锰酸钾－抗坏血酸化学发光体系，建立了测定抗坏血酸的化学发光分析新方法，线性响应的浓度范围为５．０×１０＾－７ｍｏｌ／Ｌ～４．０×１０＾－５ｍｏｌ／Ｌ，检出限为３．０×１０＾－７ｍｏｌ／Ｌ，对２．５×１０＾－６ｍｏｌ／Ｌ抗坏血酸进行１０次平行测定，相对标准偏差为１．６％，考察了２２种物质的干扰情况。用于维生素Ｃ片剂及注射液中抗坏血酸含量测定，结果令人满意。     

高锰酸钾－抗坏血酸化学发光体系测定抗坏血酸

本文研究了高锰酸钾－抗坏血酸化学发光体系，建立了测定抗坏血酸的化学发光分析新方法。线性响应的浓度范围为５．０×１０￣（－７）ｍｏｌ／Ｌ～４．０×１０￣（－５）ｍｏｌ／Ｌ检出限为３．０×１０￣（－７）ｍｏｌ／Ｌ对２．５×１０￣（－６）ｍｏｌ／Ｌ抗坏血酸进行１０次平行测定，相对标准偏差为１．６％。考察了２２种物质的干扰情况。用于维生素Ｃ片剂及注射液中抗坏血酸含量测定，结果令人满意。     

铕—敌鼠—DL—组氨酸—十六烷基三甲基溴化铵荧光体系及其应用

研究了铕－敌鼠－ＤＬ－组氨酸－ＣＴＭＡＢ荧光体系的荧光特性，测定了体系的影响因素和测定敌鼠的实验方法，体系的激发和发射波长分别为３３０ｎｍ和６１２ｎｍ，各组分的最佳浓度分别为１．０×１０＾－５ｍｏｌ／Ｌ，ＤＬ－组氨酸，１．０×１０＾－４ｍｏｌ／Ｌ，ＣＴＭＡＢ１．０×１０＾－５ｍｏｌ／Ｌ，ｐＨ为９．０，在此条件下，敌鼠浓度在６．０×１０＾－７～９．０×１０＾－５ｍｏｌ／Ｌ范围内同体系的荧光强度成线性     

四磺基铁酞菁做为过氧化物模拟酶在过氧化氢及葡萄糖测定中的应用

以四磺基铁酞菁（ＦｅＴＳＰｅ）作为过氧化物模拟酶催化Ｈ２Ｏ２与Ｌ－酪氨酸的荧光反应,建立了测定宽含量范围的Ｈ２Ｏ２及葡萄糖的方法。该法的线性范围为０．０－８．０×１０＾５ｍｏｌ／Ｌ,Ｈ２Ｏ２及０．０￣２．０×１０＾－４ｍｏｌ／Ｌ葡萄糖,检测限分别为５．０×１０＾－８ｍｏｌ／Ｌ及６．０×１０＾－６ｍｏｌ／Ｌ,将该经酶催化反应偶联,测定人血清中葡萄糖的含量,取得了满意的结果。     

冠醚存在下稀土络合物荧光体系的荧光增强效应

本详细研究了冠醚对Ｅｕ－ＴＴＡ荧光体系的增敏作用和稀土离子Ｄｙ＾３＾＋或Ｈｏ＾３＾＋对Ｅｕ－ＴＴＡ－冠醚体系的荧光强增效应。对Ｅｕ－ＴＴＡ－冠醚和Ｅｕ－Ｄｙ（或Ｈｏ）－ＴＴＡ－冠醚体系，Ｅｕ的浓度分别在１．０×１０＾－＾８－４．０×１０＾－＾６ｍｏｌ／Ｌ，１．０×１０＾－＾９－１．０×１０＾－＾７ｍｏｌ／Ｌ和１．０×１０＾－＾８－９．０×１０＾－＾６ｍｏｌ／Ｌ范围内与体系的荧光强度呈线性关系，检     

用流动注射法研究巯基化合物-铈(Ⅳ)-氢化可的松的化学发光反应

采用流动注射法研究了巯基化合物－铈（Ⅳ）－氢化可的松（ＨＣＲｓ）体系的化学发光行 为，对影响化学发光强度的诸因素进行试验和探讨，建立了流动注射化学发光法检测谷胱甘 肽（ＧＳＨ），半胱氨酸（Ｃｙｓ）等含巯基化合物的新方法。检测ＧＳＨ和Ｃｙｓ和线性范围分别为 ２．０×１０－６～１０．０×１０－５ｍｏｌ／Ｌ和１．０×１０－６ｍｏｌ／Ｌ～１０．０×１０－５ｍｏｌ／Ｌ；检测限分别为２．０ ×１０－７ｍｏｌ／Ｌ和１．４×１０－６ｍｏｌ／Ｌ（Ｓ／Ｎ＝３）；ＧＳＨ、Ｃｙｓ加入血清中进行回收测定的回收率 为９０％～９５％，相对标准偏差小于４％。     

浸蜡石墨电极伏安法测定抗癌药物盐酸阿霉素

本用自制浸蜡光谱纯石墨电极伏安法测定盐酸阿霉素。在ＰＨ＝６．０的磷酸缓冲溶液体系中，盐酸阿霉素在５．０×１０＾－８ｍｏｌ／Ｌ－１．０×１０＾－６ｍｏｌ／Ｌ范围内浓度与电流呈线性检测限可达１．０×１０＾－８ｍｏｌ／Ｌ。方法选择性好，抗干扰能力强，不需样品分离可直接进行测定。结果令人满意。     

N－羟甲基中卟啉诱导产生抗体的酶学性质研究

以Ｎ－羟甲基中卟啉为半抗原，通过动物免疫、细胞融合等技术筛选得到的细胞株１１Ｄ~１，该细胞株产生的抗体具有结合铁中卟啉的能力，且表现出过氧化物酶活力。进一步研究表明，该抗体对半抗原及铁中卟啉的亲和力分别为：１．１×１０~（－６）ｍｏｌ／Ｌ和２．１×１０~（－６）ｍｏｌ／Ｌ；抗体－铁中卟啉复合物催化反应时，对底物过氧化氢的Ｋ_ｍ＝１３．０ｍｍｏｌ／Ｌ，Ｋ_（ｃａｔ）＝８６ｍｉｎ~（－１），与自然催化速度相比，活力提高１．８×１０~４（ｍｏｌ／Ｌ）~（－１）。     

Determination of L- and D-enantiomers of methotrexate using amperometric biosensors

In order to determine the enantiopurity of methotrexate (Mtx), seven biosensors were proposed for the assay of l-Mtx and three biosensors for the assay of d-Mtx. The biosensors were designed using physical and chemical immobilization of glutamate oxidase and/or l-amino acid oxidase (l-AAOD) and/or horseradish peroxidase (HRP) for the assay of l-methotherexate, and d-amino acid oxidase (d-AAOD) and HRP for the assay of d-Mtx. Electrode characteristics were obtained and compared for the different carbon paste based biosensors. The linear concentration ranges for the proposed biosensors were in the ranges of fmol l⁻¹ to pmol l⁻¹, magnitude order with limits of detection in the fmol l⁻¹ to nmol l⁻¹ concentration range. All biosensors were successful for the determination of the enantiopurity of Mtx as raw material, and in its pharmaceutical formulations (tablets and injections).     

Radiation synthesis of a water-soluble temperature sensitive polymer, activated copolymer and applications in immobilization of proteins

In this work the radiation polymerization of N-isopropylacrylamide (NIPAAM) in aqueous solutions has been carried out and a water-soluble, temperature sensitive polymer and copolymer were obtained by using γ-rays from Co-60 source at room temperature. We have gained the optimum dose and dose—rate of radiation synthesis of linear polyNIPAAM through determining conversion yield and viscosity. In order to immobilize protein (BSA) and enzyme (HRP) into this water-soluble polymer, we prepared an activated copolymer, poly ( N-isopropylacrylamide-co-N-acryloxysuccinimide). The BSA and HRP has been immobilized onto the activated copolymer. The BSA (HRP) / copolymer conjugates still kept the original thermally sensitive properties of the linear polyNIPAAM. The conjugation yield of BSA to the activated copolymer decreased with increasing of dose. The thermal stability of the immobilized HRP was stable at 0 °C for a long time and has, at least, 4 days stability at room temperature. Immobilized HRP activity was lowered when the temperature was raised above its LCST. This phenomenon was reversible and the immobilized HRP regained activity below its LCST. The optimum pH of the immobilized HRP shifted from ca.5 upward to ca.7.     

Potential control of horseradish peroxidase immobilization on gold electrode

Based on the self-assembled monolayer (SAM) technique, a number of methods for immobilizing protein onto electrodes have been recently reported, such as entrapment method in which the protein was wrapped with regenerated silk fibroin[1], self-assem- bled monolayer[2] and silica sol-gel[3], layer-by-layer self-assembly method in which the protein was ad-sorbed to opposite charged macromolecules due to electrostatic attraction[4], reversed micelle[5], cross- linked method[6] and surface spin-coa…     

Novel hexacentered phosphazene compound as selective Fe3+ ions sensor with high quantum yield: Synthesis and application

A novel compound hexa-rhodamine substituted phosphazene (HRP) with six active centers on a cyclotriphosphazene ring was synthesized using the alkyne-azide “click” reaction. The structure of HRP was characterized using spectroscopic techniques. The optical sensor properties of HRP for metal ions were investigated using UV-Vis and Fluorescence spectroscopy. It was determined that HRP is a selective sensor with colorimetric and fluorescent properties for Fe³⁺ ions. Limit of detection (LOD) of HRP was determined as 6.94?×?10^?9 M using fluorescence intensities in the presence of different concentrations of Fe³⁺ ions. It was determined that HRP-Fe³⁺ complex has high quantum yield and excellent photostability.     

超顺磁性氧化石墨烯复合材料固定辣根过氧化物酶催化去除氯酚

采用超声辅助共沉淀法成功地将磁性Fe3O4纳米颗粒沉积在氧化石墨烯表面,利用透射电镜、磁滞回归曲线和X射线光电子能谱对材料进行了表征。将该材料作为载体固定辣根过氧化物酶,考察了固定化酶催化2-氯酚、4-氯酚和2,4-二氯酚降解反应,研究了溶液pH值、反应温度、反应时间、H2O2和氯酚浓度以及固定化酶用量对酚类物质去除率的影响。基于取代基数量和位置不同,去除率排序为2-氯酚<4-氯酚<2,4-二氯酚。另外,采用GC-MS研究了降解过程中的氧化产物。固定化酶的生化性质研究表明,固定化酶比游离酶具有更好的储存稳定性、pH稳定性和热稳定性。经过4次循环利用,固定化酶仍保留66%的活性,说明磁性纳米材料可以分离回收并重复利用,在污水处理领域具有应用前景。     

Disposable biosensors for determination of biogenic amines

This work reports monoamine oxidase (MAO)/horseradish peroxidase (HRP) and diamine oxidase (DAO)/horseradish peroxidase (HRP) based biosensors using screen-printed carbon electrodes for the determination of biogenic amines (BA). The enzymes have been covalently immobilized onto the carbon working electrode, previously modified by an aryl diazonium salt, using hydroxysuccinimide and carbodiimide. The detection has been performed by measuring the cathodic current due to the reduction of the mediator hydroxymethylferrocene at a low potential, 250 mV vs screen-printed Ag/AgCl reference electrode. The experimental conditions for the enzymes immobilization, as well as for the main variables that can influence the chronoamperometric current have been optimized by the experimental design methodology. Under these optimum conditions, the disposable biosensors have been characterized. A linear response range from 0.2 up to 1.6 μM and from 0.4 to 2.4 μM of histamine was obtained for DAO/HRP and MAO/HRP based biosensors, respectively. The biosensor construction was highly reproducible, yielding relative standard deviations of 10% and 11% in terms of sensitivity for DAO/HRP and MAO/HRP based biosensors, respectively. The capability of detection, 0.18 ± 0.01 μM in the case of DAO/HRP and 0.40 ± 0.04 μM (α = 0.05 and β = 0.005) for MAO/HRP based biosensors, and the biosensor sensitivity towards different BA has also been analyzed. Finally, the developed biosensors have been applied to the determination of the total amine content in fish samples.     

辣根过氧化物酶修饰电极在离子液体[EMIM]BF4中的直接电化学

将壳聚糖(Chi)-辣根过氧化物酶(HRP)-多壁碳纳米管(MWCNTs)的复合物修饰在玻碳电极(GCE)表面, 制备了HRP修饰电极(Chi-HRP-MWCNTs/GCE), 并将其用于在亲水性离子液体1-乙基-3-甲基咪唑四氟硼酸([EMIM]BF4)中HRP的直接电化学研究. 紫外可见光谱和红外光谱表明, HRP在复合物内保持了其原始构象. 电化学研究表明, 该修饰电极在[EMIM]BF4中的循环伏安图上出现了一对峰形良好、几乎对称的氧化还原峰, 式量电位为－0.247 V (vs. Ag/AgCl), 说明包埋在Chi-MWCNTs中的HRP与电极之间发生了直接电子传递; HRP在电极表面直接电子转移的速率常数ks为3.12 s－1; 在65 ℃的[EMIM]BF4中HRP仍然保持其活性; HRP修饰电极对过氧化氢的还原具有电催化作用, 其表观米氏常数Km为5.6×10－5 mol&#8226;L－1, 催化电流与过氧化氢浓度在5.0×10－7～5.0×10－5 mol&#8226;L－1范围内呈线性关系, 检出限为2.0×10－7 mol&#8226;L－1. 该研究为非水相生物传感器的构制提供了一种新途径.     

Facile preparation of hydrophilic glutathione modified magnetic nanomaterials for specific enrichment of glycopeptides

Glutathione modified magnetic nanoparticles (Fe₃O₄@Au-GSH) were synthesized through a simple process and exploited to enrich glycopeptides from complex samples.     

Facile preparation of hydrophilic glutathione modified magnetic nanomaterials for specific enrichment of glycopeptides

Investigations of glycosylated proteins or peptides and their related biological pathways provide new possibilities for illuminating the physiological and pathological mechanisms of glycosylation modification. However, open-ended and in-depth analysis of glycoproteomics is usually subjected to the low-abundance of glycopeptides, heterogeneous glycans, and a variety of interference molecules. In order to alleviate the influence of these obstacles, effective preconcentration of glycopeptides are indispensable. Here, we employed a hydrophilic interaction liquid chromatography (HILIC)-based method to universally capture glycopeptides. Glutathione modified magnetic nanoparticles (Fe₃O₄@Au-GSH) were synthesized through a simple process and exploited to enrich glycopeptides from complex samples. The prepared materials showed excellent ability to trap glycopeptides from standard glycoproteins digests, low detection limit (10 fmol/μL), and good selectivity (HRP:BSA = 1:100). These results indicated that glutathione-based magnetic nanoparticles synthesized in this work had great potential for glycopeptides enrichment.     

Isoform-specific quantification of endothelins in HUVEC culture supernatants by on-line high-performance liquid chromatography/electrospray mass spectrometry

A method for the quantitative analysis of endothelin peptides in human umbilical vein endothelial cell (HUVEC) culture supernatants is reported. The analysis is isoform-specific and employs solid-phase extraction and subsequent HPLC fractionation followed by HPLC-ESIMS analysis. The peptide vasoactive-intestinal-contractor (VIC) was used as internal standard for the HPLC-ESIMS analysis. Linearity of calibration curves was from 50 fmol to 25 pmol. The limit of detection of the HPLC-ESIMS step using a buffer matrix was estimated at 50 fmol (S/N > 3). The overall limit of detection for supernatants of HUVEC was 500 fmol/mL. In HUVEC culture supernatants only ions of endothelin-1 (ET1) were observed. Basal levels were determined to be 1.8 +/- 0.3 pmol/mL. Quantitative results obtained for ET1 were in agreement with those obtained by using a standard addition method and by an ELISA method.