酚酞啉在辣根过氧化酶发光检测中的应用

探讨酚酞啉对HRP(辣根过氧化酶)–Luminol–H2O2化学发光体系的增强作用。将酚酞啉加入辣根过氧化酶(HRP)–Luminol–H2O2化学发光体系,发光强度显著增强,发光持续时间达30 min以上。HRP–Luminol–H2O2化学发光体系的HRP质量浓度在5~800 pg/m L范围内与相对发光强度呈良好的线性关系,线性回归方程为lg I=1.07lgc+0.98,线性相关系数r=0.96,检出限为1.25 pg/m L。测定结果的相对标准偏差为2.8%~5.1%(n=10),加标回收率为93.5%~96.2%。酚酞啉可用于HRP及其相关标记物的定量分析。     

Thermally Activated Delayed Fluorescence in a Y3N@C80 Endohedral Fullerene: Time‐Resolved Luminescence and EPR Studies

The endohedral fullerene Y₃N@C₈₀ exhibits luminescence with reasonable quantum yield and extraordinary long lifetime. By variable‐temperature steady‐state and time‐resolved luminescence spectroscopy, it is demonstrated that above 60 K the Y₃N@C₈₀ exhibits thermally activated delayed fluorescence with maximum emission at 120 K and a negligible prompt fluorescence. Below 60 K, a phosphorescence with a lifetime of 192±1 ms is observed. Spin distribution and dynamics in the triplet excited state is investigated with X‐ and W‐band EPR and ENDOR spectroscopies and DFT computations. Finally, electroluminescence of the Y₃N@C₈₀/PFO film is demonstrated opening the possibility for red‐emitting fullerene‐based organic light‐emitting diodes (OLEDs).     

Biocatalytic System Made of 3D Chitin,Silica Nanopowder and Horseradish Peroxidase for the Removal of 17α-Ethinylestradiol: Determination of Process Efficiency and Degradation Mechanism

The occurrence of 17α-ethinylestradiol (EE2) in the environment and its removal have drawn special attention from the scientific community in recent years, due to its hazardous effects on human and wildlife around the world. Therefore, the aim of this study was to produce an efficient enzymatic system for the removal of EE2 from aqueous solutions. For the first time, commercial silica nanopowder and 3D fibrous chitinous scaffolds from Aplysina fistularis marine sponge were used as supports for horseradish peroxidase (HRP) immobilization. The effect of several process parameters onto the removal mechanism of EE2 by enzymatic conversion and adsorption of EE2 were investigated here, including system type, pH, temperature and concentrations of H₂O₂ and EE2. It was possible to fully remove EE2 from aqueous solutions using system SiO₂(HRP)–chitin(HRP) over a wide investigated pH range (5–9) and temperature ranges (4–45 °C). Moreover, the most suitable process conditions have been determined at pH 7, temperature 25 °C and H₂O₂ and EE2 concentrations equaling 2 mM and 1 mg/L, respectively. As determined, it was possible to reuse the nanoSiO₂(HRP)–chitin(HRP) system to obtain even 55% EE2 degradation efficiency after five consecutive catalytic cycles.     

TADF-Sensitized Fluorescent Enantiomers: A New Strategy for High-Efficiency Circularly Polarized Electroluminescence**

A promising strategy of thermally activated delayed fluorescence (TADF) sensitized circularly polarized luminescence (CPL) has been proposed for improving the electroluminescence efficiencies of circularly polarized fluorescent emitters. Compared with chiral TADF emitters which suffer from the dilemma of small ΔE_ST accompanied by small k_r, the TADF-sensitized CPL (TSCP) strategy using TADF molecules as sensitizers and CP-FL molecules as emitters might be the most promising method to construct high-performance circularly polarized organic light-emitting diodes (CP-OLEDs). Consequently, by taking advantage of the theoretically 100 % exciton utilization of TADF sensitizers, especially, by designing CP-FL emitters with high PLQY, narrow FWHM and large g_lum values, TSCP-type CP-OLEDs with excellent overall performances can be realized.     

Luminescent Chiral Exciplexes with Sky-Blue and Green Circularly Polarized-Thermally Activated Delayed Fluorescence

Luminescent exciplexes based on a chiral electron donor and achiral acceptors are reported as a new approach to design circularly polarized (CP) and thermally activated delayed fluorescence (TADF) emitters. This strategy results in rather high CP luminescence (CPL) values with g_lum up to 7×10⁻³, one order of magnitude higher in comparison to the CPL signal recorded for the chiral donor alone (g_lum ∼7×10⁻⁴). This increase occurs concomitantly with a CPL sign inversion, as a result of the strong charge-transfer emission character, as experimentally and theoretically rationalized by using a covalent chiral donor-acceptor model. Interestingly, blue, green-yellow and red chiral luminescent exciplexes can be obtained by modifying with the electron accepting character of the achiral unit while keeping the same chiral donor unit. These results bring new (inter)molecular guidelines to obtain simply and efficiently multi-color CP-TADF emitters.     

酶联放大镧系螯合物发光法─辣根过氧化物酶(HRP)－铽－对羟基苯甲酸体系的研究及用于HRP和结核抗体的分析

酶联放大镧系螯合物发光法──辣根过氧化物酶（ＨＲＰ）－铽－对羟基苯甲酸体系的研究及用于ＨＲＰ和结核抗体的分析谢剑炜,鄢远,彭学军,许金钩,陈国珍（厦门大学化学系、现代分析化学研究所,厦门,３６１００５）关键词酶联放大镧系螯合物发光法,辣根过氧化物酶,...     

Electrochemical behavior of horseradish peroxidase on WS2 nanosheet‐modified electrode and electrocatalytic investigation

In this paper, a WS₂ nanosheet was modified on the surface of a carbon ionic liquid electrode (CILE), and horseradish peroxidase (HRP) was further fixed on the electrode with a Nafion film. Direct electrochemistry and bioelectrocatalysis of HRP incorporated on the modified electrode were investigated in detail. On Nafion/HRP/WS₂/CILE, a pair of well‐defined quasi‐reversible redox peaks appeared on the cyclic voltammogram, indicating that the presence of the WS₂ nanosheet on the electrode surface could provide a specific interface with large surface area for HRP and its direct electron transfer rate was greatly enhanced. The formal potential (E⁰) obtained was –0.179 V, which was the typical feature of heme Fe(III)/Fe(II) in HRP. The electron transfer coefficient (α) and the heterogeneous electron transfer rate constant (k_s) of HRP were calculated as 0.44 and 1.01 s^–1, respectively. This HRP‐modified electrode showed excellent electrocatalytic activity for the reduction of trichloroacetic acid and NaNO₂ with a wide linear range and low detection limit. Real samples were detected by this proposed method, indicating the successful fabrication of a new third‐generation electrochemical enzyme sensor utilizing the WS₂ nanosheet.     

Kinetics of phenol oxidation by peroxidase

Studies of the kinetic behavior of horseradish peroxidase (HRP) at pH 8 and at room temperature indicate that the reaction of phenol with H₂O₂ catalyzed by HRP exhibits normal Michaelis-Menten saturation kinetics. An irreversible reaction mechanism for the steady-state kinetics of HRP, which is consistent with the experimental data, is considered. The second-order rate constants for the reactions of HRP with H₂O₂ and compound II with phenol are 4.14 × 10⁵ M^-1s^-1 and 5.54 × 10⁴M^-1s^-1, respectively.     

Laser-induced luminescence of barite after thermal treatment

The photoluminescence (PL) of barite is a noncharacteristic property and cannot be used for the investigation of its structure. After thermal treatment of barite at 600°C several luminescent centers were observed, providing information about different impurities. UO ₂ ²⁺ was determined from the vibrational structure and the long decay time of the luminescence band. Two different types of uranyl were detected, thin films of uranyl mineral (most probably, reserfordin) and a solid solution of uranyl ion in barite crystal. Characteristic green luminescence of UO ₂ ²⁺ may be used as indicative feature for the prospecting of uranium deposits and for the sorting of barite ores with the aim of cleaning from harmful U impurities. Eu²⁺ was determined from the spectral position, the half-width and the characteristic decay time of the luminescence band. Mn²⁺ and Ag⁺ were determined by comparing luminescence bands spectral parameters to those of synthesized BaSO₄−Mn and BaSO₄−Ag. Fe³⁺ or Mn⁴⁺ were determined from the spectral-kinetic parameters of the luminescence bands. Dedicated to Professor Lisa Heller-Kallai on the occasion of her 65^th birthday     

Direct electrochemistry and electrocatalysis of horseradish peroxidase immobilized in sol–gel-derived tin oxide/gelatin composite films

Horseradish peroxidase (HRP) was immobilized into a new type of sol–gel-derived nano-sized tin oxide/gelatin composite film (SnO₂ composite film) using a sol–gel film/enzyme/sol–gel film “sandwich” configuration. Direct electrochemistry and electrocatalysis of HRP incorporated into the composite films were investigated. HRP/SnO₂ composite film exhibited a pair of stable and quasi-reversible cyclic voltammetric peaks for the HRP Fe(III)/HRP Fe(II) redox couple with a formal potential of about −0.25 V (vs. SCE) in a pH 6.0 phosphate buffer solution. The electron transfer between the enzyme and the underlying electrode was greatly enhanced in the microenvironment with nano-SnO₂ particles and nanoporous structures. Morphologies and microstructures of the composite films and HRP/composite films were characterized with TEM, AFM. Electrochemical impedance spectroscopy (EIS) was also used to feature the HRP incorporated into composite films. FTIR and UV–Vis spectroscopy demonstrated that HRP in the composite film could retain its native secondary structure. With the advantages of organic–inorganic hybrid materials, the HRP/SnO₂ composite film modified electrode displayed good stability and electrocatalytic activity to the reduction of H₂O₂, The apparent Michaelis-Menten constant was estimated to be 0.345 mM, indicating a high affinity of HRP entrapped into the composite film toward H₂O₂.     

Amperometric Flow‐Biosensor for Cyanide Based on an Inhibitory Effect upon Bioelectrocatalytic Reduction of Oxygen by Peroxidase‐Modified Carbon‐Felt

A novel, simple and relative highly sensitive amperometric flow biosensor for cyanide was developed by using horseradish peroxidase (HRP)‐adsorbed carbon‐felt (CF), based on an inhibitory effect on the HRP‐catalyzed O₂ reduction. The HRP‐CF showed a sufficient bioelecrocatalytic activity for O₂ reduction in the potential region from 0 to ?0.5 V at pH 5.0, due to a direct electron transfer‐based O₂ reduction process via ferrous‐HRP and compound III. This HRP‐catalyzed O₂ reduction was reversibly inhibited by cyanide, which enabled to fabricate a novel and simple reagentless (i.e., no requirement of the ordinary substrate, H₂O₂, and the electron transfer mediators) flow‐biosensor for cyanide. When air‐saturated 0.1 M phosphate buffer (pH 5.0) was used as a carrier under the applied potential of ?0.2 V vs. Ag/AgCl, the steady‐state base‐current due to the HRP‐catalyzed O₂ reduction was reversibly inhibited by the cyanide injection (200 µL), resulting in peak‐shape current responses. The magnitude of the inhibition peak currents linearly increased with increasing concentrations of cyanide up to 1 µM, and the detection limit was found to be 0.04 µM (S/N=2). The apparent inhibition constant K_i′ was estimated to be 0.87 µM.     

无定形硒纳米粒子的交联酶模板法合成及其在生物传感器中的应用

本文以戊二醛交联的辣根过氧化物酶(HRP)为模板合成了均匀分散的无定形硒纳米粒子(粒径10~20 nm)，以所得合成产物为载体构建了HRP生物传感器。研究结果表明，无定形硒纳米粒子具有良好的生物相容性和吸附性，所得传感器灵敏度高，对测定底物的生物亲和性好。     

稀土铽离子对辣根过氧化物酶活力指数的影响(英)

0IntroductionHorseradishperoxidase(HRP)isaplanthemeenzymecatalyzingoxidationofawidevarietyofaro鄄maticmoleculesbyhydrogenperoxideanditisthemostwidelystudiedmemberoftheperoxidasefamily[1~3].ThecrystalstructureofHRPhasbeensolved[2].ThestructuralfeaturesofHR…     

Direct Electrochemistry of Horseradish Peroxidase Immobilized in Calcium Carbonate Microsphere Doped with Phospholipids

Protein electrochemistry affords a direct method to study the biological electron transfer processes. However, supplying a biocompatible environment to maintain the native state of protein is all‐important and challengeable. Here, we chose vaterite, one of the crystalline polymorphs of calcium carbonate, with highly porous nature and large specific surface area, which was doped with phospholipids, as the matrix to immobilize horseradish peroxidase (HRP). The integrity of HRP was kept during the simple immobilization procedure. By virtue of this organic/inorganic complex matrix, the direct electrochemistry of HRP was realized, and the activity of HRP for catalyzing reduction of O₂ and H₂O₂ was preserved.     

Electrochemical studies of 1,5-dihydroxynaphthalene as substrate for voltammetric enzyme immunoassay

Immunoassay is one of the biochemical analytical techniques using the specific antigen antibody com-plexation for analytical purposes. It has extensive ap-plication in clinical diagnostics, prevention and cure of diseases, and virus diagnostics. The presentation and progress of immunoassay methodology are one of the greatest achievements of bioanalytical chemistry. It is estimated that several-hundred millions of immuno-analytical determinations are carried out every year all over the world. E…     

Structure Determination of Alkynyl‐Protected Gold Nanocluster Au22(tBuC≡C)18 and Its Thermochromic Luminescence

An alkynyl‐protected gold nanocluster, Au₂₂(^tBuC≡C)₁₈ ( 1 ), has been synthesized and its structure has been determined by single‐crystal X‐ray diffraction. The molecular structure consists of a Au₁₃ cuboctahedron kernel and three [Au₃(^tBuC≡C)₄] trimeric staples. The cluster 1 has strong luminescence in the solid state with a 15 % quantum yield, and it displays interesting thermochromic luminescence as revealed by temperature‐dependent emission spectra. The enhanced room‐temperature emission is characterized as thermally activated delayed fluorescence.     

Enzyme-catalyzed reaction of voltammetric enzyme-linked immunoassay system based on OAP as substrate

The o-aminophenol (OAP)-H_2O_2-horseradish peroxidase (HRP) voltammetric enzyme-linked immunoassay new system has extremely high sensitivity. HRP can be measured with a detection limit of 6.0×10~-(10) g/L and a linear range of 1.0×10~(-9)—4.0×10~(-6) g/L. The pure product of H_2O_2 oxidizing OAP catalyzed by HRP was prepared with chemical method. The enzyme-catalyzed reaction has been investigated with electroanalytical chemistry, UV/Vis spectrum, IR spectrum, ~(13)C NMR, ~1H NMR, mass spectrum, elemental analysis, etc. Under the selected enzyme-catalyzed reaction conditions, the oxidation product of OAP with H_2_O2 catalyzed by HRP is 2-aminophe-noxazine-3-one. The processes of the enzyme-catalyzed reaction and the electroreduction of the product of the enzymecatalyzed reaction have been described.     

Apoferritin protein nanoparticles dually labeled with aptamer and horseradish peroxidase as a sensing probe for thrombin detection

A novel and ultrasensitive sandwich-type electrochemical aptasensor has been developed for the detection of thrombin, based on dual signal-amplification using HRP and apoferritin. Core/shell Fe₃O₄/Au magnetic nanoparticles (AuMNPs) loading aptamer1 (Apt1) was used as recognition elements, and apoferritin dually labeled with Aptamer2 (Apt2) and HRP was used as a detection probe. Sandwich-type complex, Apt1/thrombin/Apt2–apoferritin NPs–HRP was formed by the affinity reactions between AuMNPs–Apt1, thrombin, and Apt2–apoferritin–HRP. The complex was anchored on a screen-printed carbon electrode (SPCE). Differential pulse voltammetry (DPV) was used to monitor the electrode response. The proposed aptasensor yielded a linear current response to thrombin concentrations over a broad range of 0.5–100 pM with a detection limit of 0.07 pM (S/N = 3). The detection signal was amplified by using apoferritin and HRP. This nanoparticle-based aptasensor offers a new method for rapid, sensitive, selective, and inexpensive quantification of thrombin, and offers a promising potential in protein detection and disease diagnosis.     

Peroxidase catalyzed polymerization of phenol

effect of horseradish peroxidase (HRP) and H₂O₂ concentrations on the removal efficiency of phenol, defined as the percentage of phenol removed from solution as a function of time, has been investigated. When phenol and H₂O₂ react with an approximately one-to-one stoichiometry, the phenol is almost completely precipitated within 10 min. The reaction is inhibited at higher concentrations of H₂O₂. The removal efficiency increases with an increase in the concentration of HRP, but an increase in the time of treatment cannot be used to offset the reduction in removal efficiency at low concentrations of the enzyme, because of inactivation of the enzyme. One molecule of HRP is needed to remove approximately 1100 molecules of phenol when the reaction is conducted at pH 8.0 and at ambient temperature.     

Horseradish peroxidase immobilized through its carboxylic groups onto a polyacrylonitrile membrane: comparison of enzyme performances with inorganic beaded supports

A hydrophilic polyacrylonitrile (PAN) flat sheet membrane was aminated (8.5 μmol of NH₂/mg of dry support) for covalent binding of horseradish peroxidase (HRP), mediated by the soluble carbodiimide 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC). Silica microbeads derivatized by silanization, to yield an aminated support, and commercial aminated glass microbeads were also coupled to HRP with EDC or activated with glutaraldehyde. The immobilized enzyme activities were determined in a batch enzyme reactor with an external loop, the highest specific immobilized HRP activity being obtained on the glass support (55.8U/mg of protein). Continuous operational stability studies showed that hydrophilic PAN membrane led to the highest retention of HRP activity after an overall period of 35 h, with a normalized productivity of 59.5 μmol of H₂O₂ reduced/(h·U_{immob HRP}).