基于原位电聚合硫堇的双酶型葡萄糖传感器的研究

提出了一种基于电聚合硫堇作为电子传递媒介的辣根过氧化物酶-葡萄糖氧化酶(HRP-GOD)的双酶型葡萄糖传感器的制备方法。该法采用碳糊包埋HRP和硫堇,通过原位电聚合形成聚硫堇修饰的碳糊电极,以壳聚糖膜修饰该电极表面,用戊二醛交联固定GOD制得葡萄糖传感器。葡萄糖在1.0×10-4~8.0×10-3mol/L浓度范围内与传感器的响应电流呈线性关系,检出限为5×10-5mol/L。该传感器有良好的选择性和稳定性,经一个月的连续使用后,仍能保持其初始活性的80%。     

Molecular dynamics simulation study on zwitterionic structure to maintain the normal conformations of Glutathione

Molecular dynamics simulations were applied to normal conformational Glutathione (GSH) and GSH over zwitterionic and hydrophobic surfaces respectively. Conformational analysis of GSH during the simulation time on RMSD, conformational flexibility and dihedral distribution were performed. The re- sults showed that zwitterionic structure maintains the normal conformations of GSH to a better extent, which should be a first good proof of the hypothesis of "maintain of normal structure".     

Peroxidase-catalyzed polymerization and depolymerization of coal in organic solvents

Peroxidases from horseradish roots (HRP) and soybean hulls (SBP) catalyze the efficient polymerization of a 4-kDa dimethylformamide (DMF)-soluble fraction of Mequininza (Spanish) lignite in 50% (v/v) DMF with an aqueous component consisting of acetate buffer, pH 5.0. Under these conditions, HRP and SBP catalyze the oxidation of free phenolic moieties in the coal matrix, thereby leading to oxidative polymerization of the low-molecular-weight coal polymers. The high fraction of nonphenolic aromatic moieties in coal inspired us to examine conditions whereby such coal components could also become oxidized. Oxidation of nonphenolic aromatic compounds was attempted using veratryl alcohol as a model substrate. SBP catalyzed the facile oxidation of veratryl alcohol at pH <3.HRP, however, was unable to elicit veratryl alcohol oxidation. The potential for SBP to catalyze interunit bond cleavage on complex polymeric substrates was examined using l-(3,4-dimethoxyphenyl)-2-(phenoxy)propan-1,3-diol (1) as a substrate. SBP catalyzed the Cα-Cβ and β-ether bond cleavage of this compound, suggesting that similar reactions on coal, itself, could lead to depolymerization. Depolymerization of a >50 Da coal fraction was achieved using SBP in 50% (v/v) DMF with an aqueous component adjusted to pH 2.2. Approximately 15% of the initial high-molecular-weight lignite fraction was depolymerized to polymers 4 Da in size. Hence, SBP is capable of catalyzing the depolymerization of coal in organic solvents, and this may have important ramifications in the generation of liquid fuels from coals.     

Biocatalytic Synthesis of Two-Photon Active Resveratrol Oligomer

Resveratrol (3,5,4′ trihydroxy trans-stilbene) is a plant based phenolic compound. Enzymatic oligomerization of trans-resveratrol using horseradish peroxidase followed by characterization of the oligomer is presented. The oligomerization reaction was monitored using UV-Visible absorption and fluorescence spectroscopies. The oligomer exhibits strong two-photon-induced fluorescence. Computational modeling using spin-density calculations was performed to investigate the most probable reaction sites and the nature of products formed in the oligomerization process.     

Detection of Laccase,Peroxidase, and Polyphenol Oxidase on a Single Polyacrylamide Gel Electrophoresis

Abstract

Laccase, polyphenol oxidase and peroxidase are widely distributed in both plants and micro-organisms. Among these enzymes, the identification of one particular activity in the presence of the others is often difficult as enzymes can often oxidise the same substrates. A method is described, that is suitable to differentiate the three activities on the same polyacrylamide gel electrophoresis. The method is based on the use of two substrates, 4-aminoN,N-diethylaniline and 4-tert-butyl-catechol, so that coloured spots appear corresponding to the different activities. Moreover, a comparison among different mushroom species belonging to the same genus is reported using the described method.     

Enzyme-Based Fiber Optic Sensor for Glucose Determination

Abstract

A new fiber-optic probe for the determination of glucose is developed. Glucose oxidase is immobilized on a preactivated Immunodyne membrane by direct application of the enzymatic solution to one side of the membrane. The membrane is then placed around the common end of a bifurcated glass fiber optic bundle, and immersed in the sample cell which contains the glucose sample, peroxidase and a colorless dye. Glucose is quantified by the color change when the reaction takes place.     

One-step synthesis of N-acetylcysteine and glutathione derivatives using the Ugi reaction

Fully protected natural and unnatural N-acetylcysteine, dipeptide Cys-Gly, glutathione, and homoglutathione derivatives were synthesized by the Ugi four-component reaction using various benzylthio aldehydes and ketones as carbonyl building blocks. The scope and limitations of the method were investigated. Formation of imidazoline by-products in the Ugi reaction was discussed. 2,2,2-Trifluoroethanol was shown to be a superior reaction media than methanol in some reactions. Also, the 4-methyl-2,6,7-trioxabicyclo[2.2.2]octyl derivative (OBO-ester) of isocyanoacetic acid was shown to be superior to use than ethyl isocyanoacetate as a peptide synthesis precursor in cases when higher reactivity of an isocyanide is required.     

Palladium Nanoparticles/Graphdiyne Oxide Nanocomposite with Excellent Peroxidase-like Activity and Its Application for Glutathione Detection

In this study, palladium nanoparticles loaded graphdiyne oxide (Pd/GDYO) nanocomposite were fabricated by in-situ reduction of palladium chloride in the dispersion of GDYO, and characte-rized by Raman spectra, transmission electron microscopy, X-ray diffraction and X-ray photoelectron spectroscopy. The synthesized Pd/GDYO was first found to have catalytic activities similar to those of the peroxidase enzyme, which can catalyze the oxidation of peroxidase substrate 3,3',5,5'-tetramethylbenzidine(TMB) in the presence of hydrogen peroxide(H₂O₂). Steady-state kinetic studies showed that the catalytic reaction of Pd/GDYO follows a ping-pong mechanism, and Pd/GDYO has a stronger activity to TMB with a Michaelis constant(K_m) value of 5.32×10^-4 mmol/L. Based on the shielding effect of glutathione(GSH) on the Pd/GDYO-H₂O₂-TMB reaction system, a colorimetric detection method for GSH was deve-loped with a wide linear range from 0.1 μmol/L to 40 μmol/L and a limit of detection of 0.1 μmol/L. The method was successfully applied for fast and accurate detection of GSH in injection powder drugs. It was expected that this peroxidase-like Pd/GDYO nano- composite would have wide applications in the fields of biomedicine and environmental chemistry.     

A surface-enhanced Raman scattering method for detection of trace glutathione on the basis of immobilized silver nanoparticles and crystal violet probe

Unsatisfactory sensitivity and stability for molecules with low polarizability is still a problem limiting the practical applications of surface-enhanced Raman scattering (SERS) technique. By preparing immobilized silver nanoparticles (Fe₃O₄/Ag) through depositing silver on the surface of magnetite particles, a highly sensitive and selective SERS method for the detection of trace glutathione (GSH) was proposed on the basis of a system of Fe₃O₄/Ag nanoparticles and crystal violet (CV), in which the target GSH competed with the CV probe for the adsorption on the Fe₃O₄/Ag nanoparticles. Raman insensitive GSH replaced the highly Raman sensitive CV adsorbed on the surface of Fe₃O₄/Ag particles. This replacement led to a strong decrease of the CV SERS signal, which was used to determine the concentration of GSH. Under optimal conditions, a linear response was established between the intensity decrease of the CV SERS signal and the GSH concentration in the range of 50–700 nmol L⁻¹ with a detection limit of 40 nmol L⁻¹. The use of a Fe₃O₄/Ag substrate provided not only a great SERS enhancement but also a good stability, which guarantees the reproducibility of the proposed method. Its use for the determination of GSH in practical blood samples and cell extract yielded satisfactory results.     

Mechanism of Chicoric Acid Electrochemical Oxidation and Identification of Oxidation Products by Liquid Chromatography and Mass Spectrometry

Electrochemical oxidation of chicoric acid (ChA) was investigated using cyclic voltammetry and chronoamperometry at a glassy carbon electrode. Chicoric acid generates single quasi‐reversible redox wave in cyclic voltammetry over a wide pH range, and an ECEC‐dimerization mechanism is proposed. Effect of glutathione (GSH) on the electrochemical oxidation of chicoric acid (ChA) was investigated in Britton−Robinson buffer solution. Ultra‐high performance liquid chromatography (UPLC) coupled with mass spectrometry (MS) was used to show that the naturally occurring chicoric acid (ChA) underwent an electrochemical oxidation in the presence of glutathione (GSH) to form mono‐, bi‐, tri‐, and four‐glutathione conjugates of chicoric acid and a mono‐glutathione conjugate of a chicoric acid dimer. The obtained results are useful for understanding and predicting the oxidative degradation pathway of chicoric acid.