Effect of chitosan on peroxidase activity and isoenzyme profile in hairy root cultures of Armoracia lapathifolia

Hairy root cultures of Armoracia lapathifolia established by infection with Agrobacterium rhizogenes LBA 9402 present a level and isoenzyme pattern of peroxidases (POD) comparable to nontransformed roots. Elicitation with chitosan (10, 50, and 100 mg/L) was used in order to improve POD production. Total POD activity increased about 170% after 48h of treatment with chitosan 100 mg/L. Elicitation effect on soluble and ionically cell-wall-bound POD fractions of A. lapathifolia hairy roots was analyzed. POD activity of the ionically cell-wall-bound protein fraction increased in the presence of chitosan in a dose-response manner. No effect on soluble POD fractions was observed, but the isoenzyme pattern analyzed by isoelectrofocusing showed an increase of an acidic isoenzyme (pI=3.4) after the elicitation treatment. The ionically cell-wall-bound protein fraction showed only basic isoenzymes, with an increase of an isoenzyme of pI=8.7, after the elicitation treatment.     

甲醛对芦荟POD酶活性的影响

实验以玻璃箱模拟室内空间环境,应用乙酰丙酮分光光度法和邻苯三酚自氧化法测定在不同甲醛含量下芦荟对甲醛的吸收及其体内过氧化物酶(POD)活性的变化情况.结果表明,有芦荟存在时甲醛气体含量显著下降,芦荟体内的POD酶活性增强;随着通入甲醛气体量的增加,芦荟对甲醛的吸收量显著增加,POD活性的提高也越来越显著.说明芦荟体内POD酶对甲醛气体的胁迫存在着一定程度的生理应激反应.     

A biosensing platform based on horseradish peroxidase immobilized onto chitosan-wrapped single-walled carbon nanotubes

One-step, diameter-selective dispersion of single-walled carbon nanotubes (SWCNTs) has been accomplished through noncovalent complexation of the nanotubes with a water-soluble, biocompatible polymer chitosan at room temperature. Such chitosan-wrapped individual SWCNTs can be used for the immobilization of horseradish peroxidase (HRP) and be used to construct an electrode for direct bioelectrochemical sensing without an electron mediator. The direct electron transfer between HRP and the electrode surface was observed with a formal potential of approximately −0.35 V (vs. saturated calomel electrode) in phosphate buffer solution and the calculated heterogeneous electron transfer rate constant is approximately 23.5 s⁻¹. Experimental results indicate that the immobilized HRP retains its catalytic activity for the reduction of nitric oxide. Such an HRP–SWCNT–chitosan-based biosensor exhibited a rapid response time of less than 6 s and a good linear detection range for nitrite concentration, from 25 to 300 μM with a detection limit of 3 μM. The apparent Michaelis–Menten constant (K _m) and the maximum electrode sensitivity (i_max/K _m) are found to be 7.0 mM and 0.16 μA mM⁻¹, respectively. Both the unique electrical properties of SWCNTs and biocompatibility of chitosan enable the construction of an excellent biosensing platform for improved electrocatalysis of HRP, allowing, specifically, the detection of trace levels of nitric oxide.     

苹果果实大小与膜透性、过氧化物酶活性及丙二醛含量关系的研究

本文分析了苹果果实后熟衰老期间膜透性、过氧化物酶活性及丙二醛含量。结果表明,苹果果实大小不影响其过氧化物酶活性高低,但大果具有较大的过氧化物酶活性变化幅度,中等果具有较低小的过氧化物酶变化幅度。大果比中等果具有较高的膜透性与丙二醛含量。     

大球盖菇过氧化物酶及超氧化物歧化酶的研究

采用聚丙烯酰胺凝胶圆盘电泳法测定大球盖菇过氧化物酶和超氧化物歧化酶（SOD）同工酶活性,结果表明大球盖菇过氧化物酶有4种同工酶,比移分别是：0.13、0.18、0.25、0.32,其活性大小接近.SOD三种都有,比移分别为0.20、0.21、0.25,以Cu·Zn-SOD活性最大,Mn-SOD活性较小.Cu·Zn-SOD及Mn-SOD辅因子易于丢失,应用时应予以注意.     

An Ultrasensitive Virus ELISA Based on a magnetic Mesoporous Silica Nanoprobe

Detection of infectious viruses relies on quantitative polymerase chain reaction (qPCR). However, qPCR requires costly equipment, a clean operating environment and experienced technicians, limiting its wide applicability. On the other hand, enzyme-linked immunosorbent assay (ELISA) is widely used in biological laboratories due to its relatively high sensitivity and ease of operation. However, ELISA-based detection of the virus is hampered because it is lower sensitive than qPCR. Herein, a nanoprobe ELISA (NP-ELISA) based on a mesoporous silica nanoprobe, which is constructed by first being loaded with peroxidase and further coated with positively charged polymer polyethyleneimine, and finally functionalized with antivirus antibodies, is designed. Results show that each NP probe is encapsulating 170 peroxidase molecules and presents 200 antibody molecules on the surface. The limit of detection (LOD) of NP-ELISA (LOD = 1450 PFU mL⁻¹) for the detection of real virus samples is tenfold sensitive than that of standard ELISA (LOD = 14, 414 PFU mL⁻¹) and the assay time for NP-ELISA is reduced by 1 h as compared with standard one. Therefore, the NP-ELISA provides a rapid and sensitive immunoassay platform that can readily be implemented for biological laboratory research as well as for on-site clinical diagnostics.     

Fabrication of Bienzymatic Glucose Biosensor Based on Novel Gold Nanoparticles‐Bacteria Cellulose Nanofibers Nanocomposite

An exploration of gold nanoparticles–bacterial cellulose nanofibers (Au‐BC) nanocomposite as a platform for amperometric determination of glucose is presented. Two enzymes, glucose oxidase (GOx) and horseradish peroxidase (HRP) were immobilized in Au‐BC nanocomposite modified glassy carbon electrode at the same time. A sensitive and fast amperometric response to glucose was observed in the presence of electron mediator (HQ). Both of GOx and HRP kept their biocatalytic activities very well in Au‐BC nanocomposite. The detection limit for glucose in optimized conditions was as low as 2.3 µM with a linear range from 10 µM to 400 µM. The biosensor was successfully applied to the determination of glucose in human blood samples.     

Analytical Biosensing of Hydrogen Peroxide on Brilliant Cresyl Blue/Multiwalled Carbon Nanotubes Modified Glassy Carbon Electrode

A cationic quinine‐imide dye brilliant cresyl blue (BCB) and horseradish peroxidase (HRP) were co‐immobilized within ormosil on multiwalled carbon nanotubes modified glassy carbon electrode for the fabrication of highly sensitive and selective hydrogen peroxide biosensor. The presence of epoxy group in ormosil as organic moiety improves the mechanical strength and transparency of the film and amino group provides biocompatible microenvironment for the immobilization of enzyme. The presence of MWCNTs improved the conductivity of the nanocomposite film. The surface characterization of MWCNT modified ormosil nanocomposite film was performed with scanning electron microscopy (SEM) and atomic force microscopy (AFM). Cyclic voltammetry and amperometry measurements were used to study and optimize the performance of the resulting peroxide biosensor. The apparent Michaelis–Menten constant was determined to be 1.5 mM. The proposed H₂O₂ biosensor exhibited wide linear range from 3×10^?7 to 1×10^?4 M, and low detection limit 1×10^?7 M (S/N=3) with fast response time <5 s. The probable interferences in bio‐matrix were selected to test the selectivity and no significant response was observed in the biosensor. This biosensor possessed good analytical performance and long term storage stability.     

Electrografted Poly(N‐mercaptoethyl acrylamide) and Au Nanoparticles‐Based Organic/Inorganic Film: A Platform for the High‐Performance Electrochemical Biosensors

In this study, we describe the use of the combination of eletrografting poly(N‐mercaptoethyl acrylamide) and Au nanoparticles in the construction of high‐performance biosensors. The poly(N‐mercaptoethyl acrylamide) was electrografted onto the glassy carbon electrode surface, which provided a strongly adhering primer film for the stable attachment of Au nanoparticles and horseradish peroxidase (HRP) enzymes. The performances of the biosensors based on the HRP immobilized in the Au/poly(N‐mercaptoethyl acrylamide) composite film were investigated. A couple of redox peaks were obtained, indicating that the Au nanoparticles could facilitate the direct‐electron transfer between HRP and the underlying electrode. The biosensor showed an excellent electrocatalytic activity toward the reduction of hydrogen oxide and long‐term stability, owing to the stable electrografted film and biocompatible Au nanoparticles. Our results demonstrate that the combination of electrografting and Au nanoparticles provides a promising platform for the immobilization of biomolecules and analysis of redox enzymes for their sensing applications.