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The present study evaluates the role of graphene oxide's (GO's) peroxidase-like and inherent/carbocatalytic properties in oxidising silver nitrate (AgNO3) to create graphene nanocomposites with silver nanoparticles (GO/Ag nanocomposite). Activation of peroxidase-like catalytic function of GO required hydrogen peroxide (H2O2) and ammonia (NH3) in pH 4.0 disodium hydrogen phosphate (Na2HPO4). Carbocatalytic abilities of GO were triggered in pH 4.0 deionised distilled water (ddH2O). Transmission electron microscope (TEM), scanning electron microscope (SEM), cyclic voltammetry (CV) and UV-Vis spectroscopy aided in qualitatively and quantitatively assessing GO/Ag nanocomposites. TEM and SEM analysis demonstrated the successful use of GO's peroxidase-like and carbocatalytic properties to produce GO/Ag nanocomposite. UV-Vis analysis indicated a higher yield in optical density values for GO/Ag nanocomposites created using GO's carbocatalytic ability rather than its peroxidase-like counterpart. Additionally, CV demonstrated that GO/Ag nanocomposite fabricated here is a product of an irreversible electrochemical reaction. Our study outcomes show new opportunities for GO as a standalone catalyst in biosensing. We demonstrate a sustainable approach to obtain graphene nanocomposites exclusive of harmful chemicals or physical methods. [ABSTRACT FROM AUTHOR]

To circumvent the complicated natural peroxidases, palladium nanoparticles embedded in melamine cyanurate (MCA-Pd NPs) were synthesized. MCA-Pd NPs catalyzed the oxidation of ABTS2– by H2O2, and the solution turned green, which could be quantified via a typical absorption peak at 420 nm. MCA-Pd NPs had high peroxidase-like activity in a wider pH range than that of natural peroxidases. MCA-Pd NPs were used to develop a colorimetric sensor for H2O2 over the pH range of 7.0 to 11.0, which had same linear range, and their linear regression equations had similar slopes. Moreover, MCA-Pd NPs were applied to establish the biosensor for glucose by using glucose oxidase (GOx); it had a linear range of 5.0–120 μmol/L, with a linear regression equation of A = 0.04926 + 0.00536C (C: μmol/L, R = 0.9960) and a detection limit of 0.3 μmol/L (3σ/slope). When we applied it to detect glucose level in human blood, satisfactory results were obtained. [ABSTRACT FROM AUTHOR]

Foliar applications of osmoprotectants could be a valuable tool to counteract the deleterious effects of salinity and protect plant cells from oxidative damage. Some of the known osmoprotectants used in this field are salicylic acid, proline, and/or glycine betaine but there is no available empirical research exists to figure out which of these chemicals is the most effective in reducing the adverse effects of salt stress. To compare the ameliorating effects of foliar application of these osmoprotectants on growth, some physiological and biochemical parameters, and productivity of cucumber plants under NaCl stress (0, 50, or 100 mM), a factorial experiment was carried out during 2015 and 2016 seasons at the Experimental Farm of Horticulture Department, Faculty of Agriculture, Ain Shams University, Qalubia Governorate, Egypt. Increasing NaCl levels reduced plant growth parameters, leaf relative water content, leaf membrane stability, chlorophyll contents, some macro-nutrients content and yield and the lowest values were recorded with the use of 100 mM of sodium chloride treatment. Foliar application of salicylic acid at 1 mM, proline at 10 mM, or glycine betaine at 10 mM to cucumber plants ameliorated the harmful effects of NaCl stress on the vegetative growth and yield through enhancing both leaf relative water content and leaf membrane stability. In addition, these treatments improved both chlorophyll contents, and nutrient content, altogether resulted in a significant reduction in sodium (Na+) and chloride (Cl−) accumulation toxicity. Moreover, the expression of cucumber peroxidase isozymes was analyzed in cucumber leaves 1, 3 and 7 days after the second application. Native polyacrylamide gel electrophoresis analysis indicated that exogenous applications led to the differential regulation of peroxidase enzyme. In conclusion, salicylic acid was the most effective in attenuating the negative effects of the salt stress and in up-regulating peroxidase enzyme expression followed by proline and by glycine betaine. Peroxidase enzyme appeared to function as one of the main molecular mechanisms underlying the resistance response of cucumber to salinity. [ABSTRACT FROM AUTHOR]

Insect saliva is one of the first secretions to come in contact with plants during feeding. The composition and role of caterpillar saliva has not been as thoroughly studied as that of sucking insects. This study focuses on characterizing the proteome of the cabbage looper ( Trichoplusia ni ) saliva using iTRAQ labeling and LC-MS/MS. We also measured how the saliva proteome changed when larvae were reared on different diets – cabbage, tomato, and an artificial pinto bean diet. We identified 254 proteins in the saliva out of which 63 were differentially expressed. A large percentage (56%) of the proteins identified function in protein metabolism, followed by proteins involved in vesicle transport (6%) and oxidoreductase activity (5%), among other categories. Several proteins identified are antioxidants or reactive oxygen species (ROS) scavengers. Among these ROS scavengers, we identified a catalase and further analyzed its gene expression and enzymatic activity. We also applied commercial, purified catalase on tomato and measured the activity of defensive proteins – trypsin proteinase inhibitor, polyphenol oxidase and peroxidase. Catalase gene expression was significantly higher in the salivary glands of larvae fed on tomato. Also, catalase suppressed the induction of tomato trypsin proteinase inhibitor levels, but not the induction of polyphenol oxidase or peroxidase. These results add to our understanding of proteomic plasticity in saliva and its role in herbivore offense against plant defenses. [ABSTRACT FROM AUTHOR]

Novel and uniform urchin-like Co 9 S 8 nanomaterials assembled with needle-like nanorods were synthesized by a facile two-step hydrothermal process, and were found to exhibit intrinsic peroxidase-like activity. As a new peroxidase mimic, their catalytic properties, kinetics and mechanism were studied in depth by several techniques. The results showed that the catalytic activity of urchin-like Co 9 S 8 nanomaterials was highly efficient, which was arised from high electron-transfer ability of their unique morphology. Interestingly, using the peroxidase-like activity of urchin-like Co 9 S 8 nanomaterials, a novel off-on sensor for detection of Cu 2+ was constructed. The sensor showed a good linear range of 0.5–10 μM with a low detection limit of 0.09 μM. The proposed method was successfully applied for Cu 2+ determination in real samples with satisfactory results. [ABSTRACT FROM AUTHOR]

Cadmium (Cd) is a toxic element that poses a great threat to human health, while silicon (Si) is a beneficial element and has been shown to have a mitigation effect on plants under Cd toxicity. However, the mechanisms underlying the role of Si in alleviating Cd toxicity are still poorly understood in wheat. Therefore, growth status, photosynthesis parameters, root morphology, antioxidant system, and Cd2+ uptake and flux under Cd toxicity were studied through hydroponic experiment, aiming to explore the mitigation of Si on Cd toxicity in wheat seedlings. The results showed that Si supply improved plant biomass as well as photosynthetic but had little effects on root morphology of seedlings under Cd stress. Si addition decreased Cd contents both in shoots and roots. In situ measurements of Cd2+ flux showed that Si significantly inhibited the net Cd2+ influx in roots of wheat. Si also mitigated the oxidative stress in wheat leaves by decreasing malondialdialdehyde (MDA) and hydrogen peroxide (H2O2) contents as well as by increasing superoxide dismutase (SOD) and guaiacol peroxidase (POD) activity. Overall, the results revealed that Si could alleviate Cd toxicity in wheat seedlings by improving plant growth and antioxidant capacity and by decreasing Cd uptake and lipid peroxidation. [ABSTRACT FROM AUTHOR]

The effect of calcium chloride addition on the peroxidase (POD) thermal inactivation was evaluated in a fruit juice model solution and coconut water. The evaluation was conducted over the temperature range from 63 to 85 °C, considering the non-isothermal profile of processing. The accumulated lethality was calculated and the obtained data were well described by the first order kinetic model. The presence of calcium chloride resulted in an increase of POD thermal stabilities in both fruit juice model solution and green coconut water, being the inactivation kinetics also affected. Although all enzyme kinetics data of fruit juice solution can be described by the first order kinetic model, their response to thermal processing were different depending on the concentration of calcium chloride. The obtained results highlight the importance of evaluating engineering aspects of food formulation, as ingredients applied for nutritional or sensorial aspects can also impact the process performance. [ABSTRACT FROM AUTHOR]

In this study, a new spectrophotometric method was proposed for the measurement of hydrogen peroxide (H 2 O 2 ) in aqueous solutions. The method was based on the peroxidase (POD)-catalyzed reaction in which 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonate) (ABTS) was oxidized to form the stable green radical (ABTS + ). The generated ABTS + could be determined spectrophotometrically. The absorbance of the generated ABTS + at 415 nm, 650 nm, 732 nm and 820 nm were linear with H 2 O 2 concentrations in the range of 0–40 μM. The sensitivities of the proposed ABTS method for H 2 O 2 determination at 415 nm, 650 nm, 732 nm and 820 nm were 6.29 × 10 4 M −1 cm −1 , 2.00 × 10 4 M −1 cm −1 , 2.54 × 10 4 M −1 cm −1 and 1.89 × 10 4 M −1 cm −1 , respectively. The oxidation of ABTS to generate ABTS + in POD-catalyzed H 2 O 2 system at pH 6.0 was so fast that the determination time of the ABTS method was as short as 0.5 min. The stoichiometry of the reaction of H 2 O 2 and ABTS in the presence of POD was calculated as nearly 1:2 (1:1.92). The residual absorbance of the generated ABTS + was also found to be stable within 30 min in natural waters. Low H 2 O 2 concentration in rainwater could be both measured by the ABTS method and the DPD method with high accuracy, however, H 2 O 2 concentration in wastewater contained rhodamine B could only be accurately measured with the ABTS method at 732 nm. Moreover, waste solutions after H 2 O 2 analysis with the proposed ABTS method were non-hazardous towards E. coli . [ABSTRACT FROM AUTHOR]

Organic-inorganic hybrid nanoflowers, facilely made from bovine serum albumin and copper phosphate (BSA-Cu3(PO4)2·3H2O), have attracted considerable attention for the application of biocatalysts in recent years. The improved stability and activity of above-mentioned nanoflowers enhanced the efficiency of reversible addition-fragmentation chain transfer (RAFT) polymerization of functional methacrylate monomers with the assistance of acetylacetone (ACAC) and hydrogen peroxide (H2O2) in a mixed solvent of DMF and H2O. Such RAFT strategy can be employed for the polymerization of N,N-dimethylaminoethyl methacrylate (DMAEMA) and poly(ethylene glycol) methyl ether methacrylate (PEGMA500), in which both poly(DMAEMA) and poly(PEGMA500) could be synthesized in a controllable manner with typical RAFT features, e.g., precise control of molecular weight, specific molecular structure, and narrow polydispersity index (Mw/Mn). Significantly, the low-cost nanoflowers could be easily separated from reaction mixture after polymerization and will not adhere to resulting polymers as same as enzymes. Moreover, 1H NMR characterization of the retaining end groups of the resultant polymers and the chain extension experiments confirmed the mechanism of RAFT polymerization. The present biocatalytic system can serve as optimal alternatives of free enzymes in RAFT polymerization, which will hopefully enrich the methodology toward the construction of vinyl-based polymers with controlled radical polymerization (CRP). [ABSTRACT FROM AUTHOR]

Abstract: The effectiveness of gaseous ozone for inactivating peroxidase (POD) and polyphenoloxidase (PPO) in peach juice was investigated. The suitability of first‐order and Weibull models to describe inactivation kinetics was also analysed. Peach juice was exposed to ozone (0.11 and 0.20 mg O3 min−1 mL−1) in a bubble column up to 12 min at 20 ± 1 °C. Enzyme activities were reduced due to treatments. The magnitude of the inactivation increased with ozone level and exposure time. Reductions in activity after 12 min of treatment ranged between 99.5% and 99.8% for POD and between 93.9% and 97.3% for PPO, depending on ozone concentration. Inactivation curves were successfully fitted with the first‐order and Weibull models; although, based on the root‐mean‐square error, the corrected Akaike and the Bayesian Schwarz criterion, the Weibull model showed stronger capability in all cases. [ABSTRACT FROM AUTHOR]

The development of highly sensitive and low-cost biosensors for the detection of dopamine is of paramount importance for medical diagnostics. Herein, we report the preparation of a new peroxidase-like catalyst with a uniform heterostructure by using a technique involving electrospinning, annealing and solvothermal reaction. In this catalyst system, cobalt sulfide (CoS) nanoparticles were homogenously distributed and supported on the surface of cobalt ferrite (CoFe 2 O 4 ) nanotubes. The as-prepared CoFe 2 O 4 /CoS hybrid nanotubes showed remarkably high catalytic efficiency as peroxidase mimics toward the oxidation of 3,3,5,5-tetramethylbenzidine (TMB) in the presence of H 2 O 2 . Owing to the synergistic effect between the CoFe 2 O 4 and CoS component, the prepared CoFe 2 O 4 /CoS hybrid nanotubes exhibited enhanced peroxidase-like activity, exceeding that of either the CoS nanoparticles or CoFe 2 O 4 nanotubes alone. Dopamine has been widely investigated due to its unique function in the nervous system. Consequently, various approaches have been developed for the sensitive determination of dopamine. In this work, a simple and sensitive colorimetric route for the detection of dopamine was established based on the ability of dopamine to induce the reduction of oxidized TMB to TMB with consequent fading of the blue color. This method shows a wide linear range (0–50 μM) and a low detection limit of 0.58 μM. The unique heterostructure with spinel/sulfide interfaces represents a new concept for the construction of highly efficient and multifunctional biocatalysts. [ABSTRACT FROM AUTHOR]

The enzyme Mt PerII is a new peroxidase which has been isolated only recently from fungus Myceliophthora thermophila and has significant thermostability and stability at high H 2 O 2 concentrations. In the present work, an electrochemical kinetic study, based on cyclic voltammetry, is performed for the first time for the catalytic decomposition of H 2 O 2 by Mt PerII, at 18 °C. Leuco methylene blue (LMB) is used as a mediator and the catalytic and Michaelis constants are determined, assuming a Michaelis-Menten mechanism. Experimental evidence suggest the absence of inhibition by H 2 O 2 , for concentrations up to 16 mM, and increasing catalytic activity for temperatures up to 50 °C. Moreover, a modified electrode is constructed, by attempting the entrapment of Mt PerII on a dodecanothiol self-assembled monolayer on gold. The modified electrode is studied chronoamperometrically in solutions containing methylene blue mediator and different concentrations of H 2 O 2 . It is shown that adsorbed Mt PerII retains its activity and the modified electrode exhibits a considerably high linear region for the detection of H 2 O 2 . The experimental findings indicate that Mt PerII is a new candidate for analytical and industrial applications. [ABSTRACT FROM AUTHOR]

Supercritical carbon dioxide (SCCD) was applied for the inactivation of tissue enzymes in beetroot juice. The SCCD process parameters had a significant effect on the inactivation kinetics of polyphenol oxidase (PPO), peroxidase (POD), pectin esterase (PE) and polygalacturonase (PG). The use of traditional pasteurization (TP) resulted in a higher level of inactivation of the tested enzymes than SCCD. The lowest decimal reduction time was noted for PG and the highest for POD. The PG, PE, PPO and POD inactivation kinetic constants were calculated as 0.90, 0.85, 0.82 and 0.6 × 10 −2 min −1 for k-values and 256, 271, 281 and 384 min for D-values , respectively, based on the first order kinetic model. The highest degradation of bethacyanins, bethaxanthins and polyphenols occurred at 60 MPa at 55 °C, for 30 min: 58, 32 and 30%, respectively. Overall, SCCD treatment is a promising technique to obtain high quality non-acidified beetroot juice but further studies should concentrate on additional factors affecting the inactivation of tissue enzymes. [ABSTRACT FROM AUTHOR]

N6-methyladenosine (m6A) has been identified as the most frequent messenger RNA (mRNA) modification which is the reversible process of RNA methylation modification and related to biological rhythm regulation. In this work, an electrochemical immunosensor was developed for RNA methylation detection via using gold nanoparticles modified Au electrode (AuNPs/Au) as substrate electrode. Firstly, the hairpin DNA probe was assembled on the surface of AuNPs/Au via Au S bond. In the presence of the hairpin DNA, which is the template of assistant DNA and target RNA, the ligase can selectively ligate them as the template of the hairpin DNA to form a long DNA: RNA heteroduplex strand. Successively, anti-N6-methyladenosine antibody (anti-m6A antibody) could be specifically conjugated onto the RNA methylation site, and the horseradish peroxidase labeled goat anti mouse IgG (HRP-IgG) was further conjugated onto the anti-m6A antibody. In the detection buffer solution with H 2 O 2 and hydroquinone, HRP-IgG can catalyze hydroquinone oxidation to generate benzoquinone, resulting in a highly electrochemical reduction signal. This developed immunosensor shows a wide linear range for methylated RNA from 10 fM to 10 nM, low detection limit of 3.35 fM (S/N = 3), high specificity and satisfactory reproducibility. And this immunosensing method owns great potentials for providing the assay of the expression level of methylated RNA in micro-biological system. [ABSTRACT FROM AUTHOR]

Herein, by utilizing ionic liquid, we have shown for the first time that activated carbon can be used as a cheap and recyclable peroxidase mimic in the construction of colorimetric sensors for biomedical applications. The purposed nanozyme catalyzed the oxidation of 3,3′,5,5′-tetramethylbenzidine (TMB) in the presence of hydrogen peroxide (H 2 O 2 ) to produce a blue colored product which can be monitored at 652 nm. H 2 O 2 is the oxidative product of glucose in the presence of glucose oxidase. Therefore, the oxidation of glucose can be quantitatively related to the colorimetric response by combining these two reactions. Under the optimal experimental conditions, a linear response was observed for glucose concentrations ranging from 12 to 550 μmol/L, with a detection limit of 3.5 μmol/L. Furthermore, the specificity of the proposed method was demonstrated against common interfering compounds including uric acid, ascorbic acid, lactose, sucrose and maltose. The applicability of the proposed method was demonstrated for glucose detection in human serum samples. The proposed methodology can serve as a general way to design ionic liquid based nanozyme mimetics for various catalytic reactions and colorimetric sensing applications. [ABSTRACT FROM AUTHOR]

The rational design of metalloenzymes provides advantages not only for illustrating the structure and function relationship of native enzymes, but also for creating functional artificial enzymes comparable to native enzymes. Dye-decolorizing peroxidases (DyPs) are a new family of heme peroxidases and have received much attention recently. Inspired by the structural features of native DyPs with multiple Tyr and Trp residues, we herein aimed to design functional artificial DyPs using myoglobin (Mb), an O2 carrier as a protein scaffold, by further introduction of Tyr/Trp into the secondary sphere of the heme center in the F43Y Mb mutant. The latter has been shown to possess a novel Tyr–heme cross-link and exhibit enhanced peroxidase activity, which provides an ideal platform to design a series of derivatives, including F43Y/F46Y Mb, F43Y/I107Y Mb, F43Y/F138 W Mb and F43Y/I107Y/F138 W Mb. Our design revealed that the Tyr–heme cross-link was well-retained in the mutants except for F43Y/F46Y Mb, as confirmed by X-ray crystal structure analysis. More importantly, stopped-flow kinetic studies showed that these derivatives exhibit enhanced dye-decolorizing peroxidase activities compared to that of wild-type (WT) Mb. This is particularly the case for the double mutant F43Y/F138 W Mb, exhibiting an overall catalytic efficiency (kcat/Km) of 110 670 M−1 s−1, which is ∼144-fold and ∼20-fold that of WT Mb and F43Y Mb, respectively, and is ∼4.3-fold that of native DyP from Vibrio cholerae. Stopped-flow, electron paramagnetic resonance (EPR) and isothermal titration calorimetry (ITC) studies further provided insights into the activation of H2O2 and the binding of a substrate, reactive blue 19 (RB19), to the double mutant. This study provides valuable information for elucidating the structure and dye-decolorizing function relationship of peroxidases, and also clues for the design of other functional artificial heme enzymes. [ABSTRACT FROM AUTHOR]

Background: In the biorefnery utilizing lignocellulosic biomasses, lignin decomposition to value-added phenolic derivatives is a key issue, and recently biocatalytic delignifcation is emerging owing to its superior selectivity, low energy consumption, and unparalleled sustainability. However, besides heme-containing peroxidases and laccases, information about lignolytic biocatalysts is still limited till date. Results: Herein, we report a promiscuous activity of tyrosinase which is closely associated with delignifcation requiring high redox potentials (>1.4 V vs. normal hydrogen electrode [NHE]). The promiscuous activity of tyrosinase not only oxidizes veratryl alcohol, a commonly used nonphenolic substrate for assaying ligninolytic activity, to veratraldehyde but also cleaves the 4-O-5 and Cα--Cβ bonds in 4-phenoxyphenol and guaiacyl glycerol-β-guaiacyl ether (GGE) that are dimeric lignin model compounds. Cyclic voltammograms additionally verifed that the promiscuous activity oxidizes lignin-related high redox potential substrates. Conclusion: These results might be applicable for extending the versatility of tyrosinase toward biocatalytic delignifcation as well as suggesting a new perspective for sustainable lignin utilization. Furthermore, the results provide insight for exploring the previously unknown promiscuous activities of biocatalysts much more diverse than ever thought before, thereby innovatively expanding the applicable area of biocatalysis. [ABSTRACT FROM AUTHOR]

In the present work, an automatic methodology, allowing the evaluation of peroxidase activity, and the comparison of the effect of two substrates in this activity, was developed. Automation was carried out using a miniaturized micro-sequential injection analysis system, usually called μSIA-LOV (Lab-on-valve). This kind of system allows downscaling of current flow injection and sequential injection systems. For the automatic evaluation of peroxidase activity it was selected two different reactions using different substrates - 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonate (ABTS) and a mixture of 4-aminoantipyrine and phenol, that were implemented in a μSIA-LOV manifold. The experimental parameters such as reagent ratios and volumes, pH, temperature and reaction time, were optimized and kept constant for the two reactions. The use of these two reactions allowed the applicability of the system in two different ranges of concentrations (up to 0.20 U mL − 1 when it was used ABTS and up to 0.80 U mL − 1 for the reaction with 4-AAP and phenol) with different sensitivities and detection limits of 1.9 × 10 − 4 U mL − 1 and 9.2 × 10 − 3 U mL − 1 (using ABTS and 4-AAP with phenol, respectively). Horseradish, ginger and avocado samples were analyzed proving that the developed methodology constitutes a robust, versatile, rapid and simple alternative for the evaluation of peroxidase activity, adapted to the actual demands of reagents economy and reduced residue production, being in good agreement with the actual recommendations of green chemistry. [ABSTRACT FROM AUTHOR]

Euphorbia tirucalli peroxidase (ETP) was immobilized on chitosan beads having magnetic properties for the ease of separation and increasing the reusability of ETP for cost effective assay conditions. The present work reports immobilization of ETP on polymeric support chitosan-cobalt oxide beads subsequently activated with 0.05% cynuric chloride. The magnetic immobilized enzyme was characterized by Fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD) analysis and scanning electron microscopy (SEM). The immobilized ETP can be reused up to 10 cycles with retention of more than 60% activity. The optimum pH was shifted from 6.0 to 5.5 for soluble ETP to immobilized ETP and optimum temperature from 50 °C and 55 °C for the immobilized ETP. Based on response surface methodology, the optimal immobilization conditions obtained were: enzyme concentration, 2 mg/286 mg beads; optimal pH, 4.93; temperature, 28.88; cynuric chloride concentration, 0.17%; reaction time, 14.4 h, which resulted 74.51% maximum immobilization. The enzyme magnetic nanoparticles could be separated magnetically for easy reuse. Immobilization of ETP onto the magnetic nanoparticles could be useful for biotechnological applications and bioassay due to its reusability and improved stability. [ABSTRACT FROM AUTHOR]

Simultaneous, sensitive and quantitative detection of biomarkers in infectious disease is crucial for guiding antimicrobial treatment and predicting prognosis. This work reported an ultrasensitive and quantitative microfluidic immunoassay combined with the streptavidin-biotin-peroxidase (SA-B-HRP) nanocomplex-signal amplification system (MIS) to detect two inflammatory biomarkers, procalcitonin (PCT, for discriminating bacterial infections from nonbacterial infections) and interleukin-6 (IL-6, for monitoring the kinetics of infectious disease) simultaneously. The amplification system was based on the one step self-assembly of SA and B-HRP to form the SA-B-HRP nanocomplex, which effectively amplified the chemiluminescent signals. The linear ranges for PCT and IL-6 detections by MIS were 250–1.28 × 10 5 pg mL −1 and 5–1280 pg mL −1 , and the limit of detection (LOD) were 48.9 pg mL −1 and 1.0 pg mL −1 , respectively, both of which were significantly improved compared with microfluidic immunoassays without amplification system (MI). More importantly, PCT and IL-6 in human serum could be simultaneously detected in the same run by MIS, which could greatly improve the detection efficiency and reduce the cost. Given the advantages of high sensitivity, multiplex and quantitative detection, MIS could be potentially applied for detection of biomarkers at low concentration in clinical diagnosis. [ABSTRACT FROM AUTHOR]

Root hair polar growth is endogenously controlled by auxin and sustained by oscillating levels of reactive oxygen species (ROS). These cells extend several hundred-fold their original size toward signals important for plant survival. Although their final cell size is of fundamental importance, the molecular mechanisms that control it remain largely unknown. Here we show that ROS production is controlled by the transcription factor RSL4, which in turn is transcriptionally regulated by auxin through several auxin response factors (ARFs). In this manner, auxin controls ROS-mediated polar growth by activating RSL4, which then up-regulates the expression of genes encoding NADPH oxidases (also known as RESPIRATORY BURST OXIDASE HOMOLOG proteins) and class III peroxidases, which catalyze ROS production. Chemical or genetic interference with ROS balance or peroxidase activity affects root hair final cell size. Overall, our findings establish amolecular link between auxin and ROS-mediated polar root hair growth. [ABSTRACT FROM AUTHOR]

The heme peroxidase HPX15 is an evolutionary conserved anopheline lineage-specific gene. Previously, we found that this gene is present in the genome of 19 worldwide distributed different species of Anopheles mosquito and its orthologs are absent in other mosquitoes, insects, or human. In addition, 65-99% amino acid identity among these 19 orthologs permitted us to hypothesize that the functional aspects of this gene might be also conserved in different anophelines. In this study, we found that Anopheles stephensi AsHPX15 gene is mainly expressed in the midgut and highly induced after uninfected or Plasmodium berghei-infected blood feeding. RNA interference-mediated silencing of midgut AsHPX15 gene drastically reduced the number of developing P. berghei oocysts. An antiplasmodial gene nitric oxide synthase was induced 13-fold in silenced midguts when compared to the unsilenced controls. Interestingly, the induction of antiplasmodial immunity in AsHPX15-silenced midguts is in absolute agreement with Anopheles gambiae. In A. gambiae, AgHPX15 catalyzes the formation of a dityrosine network at luminal side of the midgut that suppresses the activation of mosquito immunity against the bolus bacteria. Thus, a low-immunity zone created by this mechanism indirectly supports Plasmodium development inside the midgut lumen. These indistinguishable functional behaviors and conserved homology indicates that HPX15 might be a potent target to manipulate the antiplasmodial immunity of the anopheline midgut, and it will open new frontiers in the field of malaria control. [ABSTRACT FROM AUTHOR]

In this paper, a novel strategy for the fabrication of reduced graphene oxide (rGO)/Cu8S5/polypyrrole (PPy) composite nanosheets with Cu8S5 nanoparticles and PPy layer anchored on the surface of rGO as peroxidase-like nanocatalyst is reported. During the synthesis, graphene oxide (GO)/CuO composite nanosheets are prepared first and used as templates, then the sulfuration of CuO and polymerization of pyrrole are accompanied with the reduction of GO, resulting in ternary rGO/Cu8S5/PPy composite nanosheets. The synthesized Cu8S5 nanoparticles with a diameter in the range from tens to hundreds of nanometers are dispersed within PPy decorated rGO nanosheets. The resultant ternary rGO/Cu8S5/PPy composite nanosheets exhibit a higher peroxidase-like catalytic activity toward the oxidation of 3,3′,5,5′-tetramethylbenzidine in the presence of H2O2 than GO/CuO and rGO/CuS composite nanosheets, revealing a synergistic effect on their activity. The as-prepared rGO/Cu8S5/PPy platform provides a simple colorimetric approach for the detection of H2O2 and phenol with a high sensitivity. This work offers a new way for the fabrication of rGO-based nanocomposite with superior enzyme-like activity, which displays great potential applications in biocatalysis and environmental monitoring. [ABSTRACT FROM AUTHOR]

The Trypanosoma cruzi ascorbate peroxidase is, by sequence analysis, a hybrid type A member of class I heme peroxidases [TcAPx-cytochrome c peroxidase (CcP)], suggesting both ascorbate (Asc) and cytochrome c (Cc) peroxidase activity. Here, we show that the enzyme reacts fast with H2O2 (k = 2.9 × 107 M-1.s-1) and catalytically decomposes H2O2 using Cc as the reducing substrate with higher efficiency than Asc (kcat/Km = 2.1 × 105 versus 3.5 × 104 M-1.s-1, respectively). Visible-absorption spectra of purified recombinant TcAPx-CcP after H2O2 reaction denote the formation of a compound I-like product, characteristic of the generation of a tryptophanyl radical-cation (Trp233•+). Mutation of Trp233 to phenylalanine (W233F) completely abolishes the Cc-dependent peroxidase activity. In addition to Trp233•+, a Cys222-derived radical was identified by electron paramagnetic resonance spin trapping, immunospin trapping, andMS analysis after equimolar H2O2 addition, supporting an alternative electron transfer (ET) pathway from the heme. Molecular dynamics studies revealed that ET between Trp233 and Cys222 is possible and likely to participate in the catalytic cycle. Recognizing the ability of TcAPx-CcP to use alternative reducing substrates, we searched for its subcellular localization in the infective parasite stages (intracellular amastigotes and extracellular trypomastigotes). TcAPx-CcP was found closely associated with mitochondrial membranes and, most interestingly, with the outer leaflet of the plasma membrane, suggesting a role at the host-parasite interface. TcAPx-CcP overexpressers were significantly more infective to macrophages and cardiomyocytes, as well as in the mouse model of Chagas disease, supporting the involvement of TcAPx-CcP in pathogen virulence as part of the parasite antioxidant armamentarium. [ABSTRACT FROM AUTHOR]

HBP23, a 23-kDa heme-binding protein identified in rats, is a member of the peroxiredoxin (Prx) family, the primary peroxidases involved in hydrogen peroxide catabolism. Although HBP23 has a characteristic Cys-Pro heme-binding motif, the significance of heme binding to Prx family proteins remains to be elucidated. Here, we examined the effect of heme binding to human peroxiredoxin-1 (PRX1), which has 97% amino acid identity to HBP23. PRX1 was expressed in Escherichia coli and purified to homogeneity. Spectroscopic titration demonstrated that PRX1 binds heme with a 1:1 stoichiometry and a dissociation constant of 0.17 μM. UV–vis spectra of heme-PRX1 suggested that Cys52 is the axial ligand of ferric heme. PRX1 peroxidase activity was lost upon heme binding, reflecting the fact that Cys52 is not only the heme-binding site but also the active center of peroxidase activity. Interestingly, heme binding to PRX1 caused a decrease in the toxicity and degradation of heme, significantly suppressing H 2 O 2 -dependent heme peroxidase activity and degradation of PRX1-bound heme compared with that of free hemin. By virtue of its cytosolic abundance (∼20 μM), PRX1 thus functions as a scavenger of cytosolic hemin (<1 μM). Collectively, our results indicate that PRX1 has a dual role; Cys-dependent peroxidase activity and cytosolic heme scavenger. [ABSTRACT FROM AUTHOR]

This study investigated the non-thermal effects of electricity on enzymatic inactivation kinetics of peroxidase and polyphenol oxidase and on degradation of total phenolic and total flavonoid compounds during ohmic heating of sugarcane juice. A kinetic modeling was carried out from the adjustment of various models applicable to enzyme inactivation, and the Weibull distribution model was the most suitable to describe the inactivation of peroxidase. The presence of an electric field with low intensity significantly influenced the biochemical reactions occurred during peroxidase activation at 60 °C and inactivation at 80 °C. Polyphenol oxidase, a more thermolabile enzyme, was almost totally inactivated before reaching the target temperature (holding phase) of the experiments. The maximum total phenolic and total flavonoid degradation were approximately 23 and 39%, respectively. Statistical analyses showed no differences between conventional and ohmic heating for total phenolic and total flavonoid compounds. [ABSTRACT FROM AUTHOR]

The present study explored the dietary administration effects of guanosine monophosphate (GMP) on growth, digestibility, innate immune responses and stress resistance of juvenile red sea bream, Pagrus major . A semi-purified basal diet supplemented with 0% (Control), 0.1% (GMP-0.1), 0.2% (GMP-0.2), 0.4% (GMP-0.4) and 0.8% (GMP-0.8) purified GMP to formulate five experimental diets. Each diet was randomly allocated to triplicate groups of fish (mean initial weight 3.4 g) for 56 days. The obtained results clearly indicated that, growth performance of red sea bream enhanced by dietary GMP supplementation compared to control and significantly higher final weight was found in fish fed diet group GMP-0.4. Specific growth rate (SGR) and percent weight gain (%WG) also significantly higher in diet group GMP-0.4 in compared to control and it was not differed (P > 0.05) with diet group GMP-0.8. Feed intake significantly increased with the supplementation of GMP. Feed conversion efficiency (FCE) and protein efficiency ratio (PER) also improved (P < 0.05) when fish fed the diets containing GMP and diet group GMP-0.4 showed the significantly higher value in compared to control. The Apparent digestibility coefficients (dry matter, protein and lipid) also improved by GMP supplementation and the significantly higher protein digestibility was observed in fish fed diet groups GMP-0.2, GMP-0.4 and GMP-0.8. Among the measured non specific immune parameters peroxidase activity (PA), respiratory burst activity (NBT), Bactericidal activity (BA) were significantly affected by dietary supplementation and highest value obtained in diet group GMP-0.4. Total serum protein, lysozyme activity (LA), and agglutination antibody titer also increased (P > 0.05) by GMP supplementation. In contrast, catalase activity decreased with GMP supplementation. In terms of oxidative stress GMP-0.2 showed best condition with low oxidative stress and high antioxidant level. Moreover, the fish fed GMP supplemented diets had better improvement (P < 0.05) in body protein contents, hepatosomatic index, hematocrit content and glutamyl oxaloacetic transaminase (GOT) and glutamic-pyruvate transaminase (GPT) level than the control group. Supplementation also improved (P < 0.05) freshwater stress resistances. Quadratic regression analysis of WG and LA revealed that, the optimal levels of dietary GMP were 0.45 and 0.48%, respectively, for juvenile red sea bream, which is also in line with the most of the growth performance and health parameters of the fish. [ABSTRACT FROM AUTHOR]

Improving the performance of conventional enzyme-linked immunosorbent assay (ELISA) is of great importance to meet the demand of early clinical diagnosis of various diseases. Herein, we report a feasible enzyme-free immunosorbent assay (EFISA) system using antibody conjugated Au-Pt/SiO 2 nanocomposites (APS NCs) as labels. In this system, Au-Pt/SiO 2 nanospheres (APS NPs) were first synthesized by wet chemical method and exhibited intrinsic peroxidase and catalase-like activity with excellent water-solubility. Then APS NCs were utilized as labels to replace HRP conjugated antibody, and Fe 3 O 4 magnetic beads (MBs) to entrap the analyte. To discuss the performance of EFISA system, Human IgG was served as a model analyte, and porcine circovirus type 2 (PCV2) serums as real samples. The system boosted the detection limit of HIgG to 75 pg mL −1 with a RSD below 5%, a 264-fold improvement as compared with conventional ELISA. This is the first time that APS NCs have been used and successfully optimized for the sensitive dilution detection of PCV2 antibody (5:10 7 ) in ELISA. Besides, APS NCs have advantages related to low cost, easy preparation, good stability and tunable catalytic activity, which make them a potent enzyme mimetic candidate and may find potential applications in bioassays and clinical diagnostics. [ABSTRACT FROM AUTHOR]

The spectroscopic and kinetic characterization of two intermediates from the H2O2 oxidation of three dimethyl ester [(proto), (meso), (deuteroporphyrinato) (picdien)]Fe(III) complexes ([FePPPic], [FeMPPic] and [FeDPPic], respectively) pinch-porphyrin peroxidase enzyme models, with s = 5/2 and 3/2 Fe(III) quantum mixed spin (qms) ground states is described herein. The kinetic study by UV/Vis at λmax = 465 nm showed two different types of kinetics during the oxidation process in the guaiacol test for peroxidases (1-3 + guaiacol + H2O2 Ñ oxidation guaiacol products). The first intermediate was observed during the first 24 s of the reaction. When the reaction conditions were changed to higher concentration of pinch-porphyrins and hydrogen peroxide only one type of kinetics was observed. Next, the reaction was performed only between pinch-porphyrins-Fe(III) and H2O2, resulting in only two types of kinetics that were developed during the first 0-4 s. After this time a self-oxidation process was observed. Our hypotheses state that the formation of the π-cation radicals, reaction intermediates of the pinch-porphyrin-Fe(III) family with the ligand picdien [N,N'-bis-pyridin-2-ylmethyl-propane-1,3-diamine], occurred with unique kinetics that are different from the overall process and was involved in the oxidation pathway. UV-Vis, 1H-NMR and ESR spectra confirmed the formation of such intermediates. The results in this paper highlight the link between different spectroscopic techniques that positively depict the kinetic traits of artificial compounds with enzyme-like activity. [ABSTRACT FROM AUTHOR]

We exposed the microalgal strain, Chlorella sp. MM3, to unused or used engine oil, or their water accommodated fractions (WAFs) to determine growth inhibition and response of antioxidant enzymes. Oil type and oil concentration greatly affected the microalgal growth. Used oil at 0.04 % (0.4 g L) resulted in 50 % inhibition in algal growth, measured in terms of chlorophyll-a, while the corresponding concentration of unused oil was nontoxic. Similarly, used oil WAF showed significant toxicity to the algal growth at 10 % level, whereas WAF from unused oil was nontoxic even at 100 % concentration. Peroxidase enzyme in the microalga significantly increased with used oil at concentrations above 0.04 g L whereas the induction of superoxide dismutase and catalase was apparent only at 0.06 g L. Activities of the antioxidant enzymes increased significantly when the microalga was exposed to 75 and 100 % WAF obtained from used oil. The used oil toxicity on microalga could be due to the presence of toxic soluble mono- and polyaromatic compounds, heavy metals, and other compounds attained by the oil during its use in the motor engines. [ABSTRACT FROM AUTHOR]

The effects of molecular confinement and crowding on enzyme kinetics were studied at length scales and under conditions similar to those found in biological cells. These experiments were carried out using a nanofluidic network of channels constituting a nanofluidic gradient mixer, providing the basis for measuring multiple experimental conditions simultaneously. The 100 nm × 40 μm nanochannels were wet etched directly into borosilicate glass, then annealed and characterized with fluorescein emission prior to kinetic measurements. The nanofluidic gradient mixer was then used to measure the kinetics of the conversion of the horseradish peroxidase (HRP)-catalyzed conversion of non-fluorescent Amplex Red (AR) to the fluorescent product resorufin in the presence of hydrogen peroxide (H2O2). The design of the gradient mixer allows reaction kinetics to be studied under multiple (five) unique solution compositions in a single experiment. To characterize the efficiency of the device the effects of confinement on HRP-catalyzed AR conversion kinetics were studied by varying the starting ratio of AR : H2O2. Equimolar concentrations of Amplex Red and H2O2 yielded the highest reaction rates followed by 2 : 1, 1 : 2, 5 : 1, and finally 1 : 5 [AR] : [H2O2]. Under all conditions, initial reaction velocities were decreased by excess H2O2. Crowding effects on kinetics were studied by increasing solution viscosity in the nanochannels in the range 1.0–1.6 cP with sucrose. Increasing the solution viscosities in these confined geometries decreases the initial reaction velocity at the highest concentration from 3.79 μM min−1 at 1.00 cP to 0.192 μM min−1 at 1.59 cP. Variations in reaction velocity are interpreted in the context of models for HRP catalysis and for molecular crowding. [ABSTRACT FROM AUTHOR]

Lead (Pb) is one of the most important toxic heavy metals that reduce plant growth and development. Therefore, finding compounds that can alleviate the toxic effects of Pb is necessary. Nitric oxide (NO) is a signaling molecule that plays important roles in different physiological processes in plants, such as the response to biotic and abiotic stresses. A pot experiment was conducted to study the effects of exogenously applied NO on Pb stress tolerance in cowpea. Seeds were soaked at various concentrations of sodium nitroprusside (SNP; NO donor) (0, 0.5 and 1 mM) for 20 h, and then sown in pots containing 0 or 200 mg kg-1 Pb (NO3)2. The obtained results showed that Pb stress significantly reduced the chlorophyll value, relative water content (RWC) and net photosynthetic rate, but increased lipid peroxidation, proline content and superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX) and glutathione reductase (GR) activities. Nonetheless, different levels of NO significantly lowered Pb toxicity by further increasing SOD, CAT, APX and GR activities as well as proline accumulation. NO pretreatment also raised the chlorophyll value, RWC and net photosynthetic rate but reduced lipid peroxidation. Both NO levels (0.5 and 1 mM) were effective in Pb stress tolerance; however, 0.5 mM was more effective. These results indicate that NO pretreatment plays an important role in protecting cowpea plants against Pb toxicity by enhancing antioxidant enzyme activities and proline accumulation. [ABSTRACT FROM AUTHOR]

Variability in the concentration of total proteins and activities of peroxides enzyme in two-year-old needles of Austrian pine (Pinus nigra Arnold), were studied in the needles of eight different half-sib lines of Austrian pine. The samples were taken from a twenty-two-year-old seedling seed orchard on Jelova Gora (Western Serbia). The results of this study show the pronounced variability of the concentration of total proteins, and enzyme activity of peroxides in the study half-sib lines. This infers that proteins have different levels of expression in individual Austrian pine lines. Also the enzyme of peroxides shows different activity depending on the genetic constitution of the trees of the analyzed half-sib lines. Based on the seed orchard size and the planting pattern, it can be assumed that the effect of external factors is reduced to a minimum. The peroxides present as soluble in the apoplast or with covalent bond to cell wall, have an important role in lignification, suberisation, healing of injuries, defense against pathogens and atmospheric pollutants. [ABSTRACT FROM AUTHOR]

The asymmetric ligand (E)-4-bromo-2-(((2-((5-bromo-2-hydroxybenzyl)(methyl)amino)ethyl)imino)methyl)phenol has been prepared by a novel seven-step route. All organic compounds isolated in each step have been characterised by elemental analysis, infrared and 1H NMR spectroscopy, and mass spectrometry. Interaction of this ligand with manganese has been investigated employing an electrochemical method. This method leads to the formation of a neutral manganese(II) complex 7 in high yield and purity. The complex has been thoroughly characterised by elemental analysis, infrared spectroscopy, mass spectrometry, magnetic susceptibility measurements, and cyclic voltammetry. Complex 7 behaves as peroxidase mimic in the presence of the water-soluble trap ABTS, probably due to its ease to coordinate the substrate molecule. [ABSTRACT FROM AUTHOR]

The present work describes the work on three phase partitioning (TPP) of a peroxidase enzyme from orange peels (Citrus sinenses). TPP is a bioseparation technique which has been used for the extraction, concentration and purification of enzymes as well as natural products. Three phase system is formed with simultaneous addition of ammonium sulfate directly to crude broth/feed followed by addition of t-butanol. Effect of different process parameters such as time, ammonium sulfate concentration, pH, broth to t-butanol ratio, temperature and speed of agitation have been studied on purification fold and activity recovery. Maximum purity of 18.20-fold with 93.96% recovery has been obtained with optimized parameters. Optimized system comprised of solution saturated with 50% ammonium sulfate at pH 6 and temperature 30 °C with broth to t-butanol ratio 1:1.5 (v/v) with 200rpm with for 80min. SDS PAGE analysis of partially purified enzyme shows molecular weight of 26 kDa. TPP was found to be an attractive technique for the extraction and partial purification of peroxidase from orange peels. [ABSTRACT FROM AUTHOR]

Thermobifida fusca is a mesothermophilic organism known for its ability to degrade plant biomass and other organics, and it was demonstrated that it represents a rich resource of genes encoding for potent enzymes for biocatalysis. The thermostable catalase from T. fusca has been cloned and overexpressed in Escherichia coli with a yield of 400 mg/L. Heat treatment of disrupted cells at 60 °C for 1 h resulted in enzyme preparation of high purity; hence, no chromatography steps are needed for large-scale production. Except for catalyzing the dismutation of hydrogen peroxide, TfuCat was also found to catalyze oxidations of phenolic compounds. The catalase activity was comparable to other described catalases while peroxidase activity was quite remarkable with a k of nearly 1000 s for catechol. Site directed mutagenesis was used to alter the ratio of peroxidase/catalase activity. Resistance to inhibition by classic catalase inhibitors and an apparent melting temperature of 74 °C classifies this enzyme as a robust biocatalyst. As such, it could compete with other commercially available catalases while the relatively high peroxidase activity also offers new biocatalytic possibilities. [ABSTRACT FROM AUTHOR]

Dye-decolorizing peroxidase (DyP) of Auricularia auricula-judae has been expressed in Escherichia coli as a representative of a new DyP family, and subjected to mutagenic, spectroscopic, crystallographic and computational studies. The crystal structure of DyP shows a buried haem cofactor, and surface tryptophan and tyrosine residues potentially involved in long-range electron transfer from bulky dyes. Simulations using PELE (Protein Energy Landscape Exploration) software provided several binding-energy optima for the anthraquinone-type RB19 (Reactive Blue 19) near the above aromatic residues and the haem access-channel. Subsequent QM/MM (quantum mechanics/molecular mechanics) calculations showed a higher tendency of Trp-377 than other exposed haem-neighbouring residues to harbour a catalytic protein radical, and identified the electron-transfer pathway. The existence of such a radical in H2O2-activated DyP was shown by low-temperature EPR, being identified as a mixed tryptophanyl/tyrosyl radical in multifrequency experiments. The signal was dominated by the Trp-377 neutral radical contribution, which disappeared in the W377S variant, and included a tyrosyl contribution assigned to Tyr-337 after analysing the W377S spectra. Kinetics of substrate oxidation by DyP suggests the existence of high- and low-turnover sites. The high-turnover site for oxidation of RB19 (kcat> 200 s-1) and other DyP substrates was assigned to Trp-377 since it was absent from the W377S variant. The low-turnover site/s (RB19 kcat ~20 s-1) could correspond to the haem access-channel, since activity was decreased when the haem channel was occluded by the G169L mutation. If a tyrosine residue is also involved, it will be different from Tyr-337 since all activities are largely unaffected in the Y337S variant. [ABSTRACT FROM AUTHOR]

Sperm whale myoglobin (Mb) functions as an oxygen-storage protein, but in the ferric state it possesses a weak peroxidase activity which enables it to carry out H2O2-dependent dehalogenation reactions. Hemoglobin/dehaloperoxidase from Amphitrite ornata (DHP) is a dual-function protein represented by two isoproteins DHP A and DHP B; its peroxidase activity is at least ten times stronger than that of Mb and plays a physiological role. The `DHP A-like' K42Y Mb mutant (K42Y) and the `DHP B-like' K42N mutant (K42N) were engineered in sperm whale Mb to mimic the extended heme environments of DHP A and DHP B, respectively. The peroxidase reaction rates increased ∼3.5-fold and ∼5.5-fold in K42Y and K42N versus Mb, respectively. The crystal structures of the K42Y and K42N mutants revealed that the substitutions at position 42 slightly elongate not only the distances between the distal His55 and the heme iron but also the hydrogen-bonding distances between His55 and the Fe-coordinated water. The enhanced peroxidase activity of K42Y and K42N thus might be attributed in part to the weaker binding of the axial water molecule that competes with hydrogen peroxide for the binding site at the heme in the ferric state. This is likely to be the mechanism by which the relationship `longer distal histidine to Fe distance - better peroxidase activity', which was previously proposed for heme proteins by Matsui et al. (1999) ( J. Biol. Chem. 274, 2838-2844), works. Furthermore, positive cooperativity in K42N was observed when its dehaloperoxidase activity was measured as a function of the concentration of the substrate trichlorophenol. This serendipitously engineered cooperativity was rationalized by K42N dimerization through the formation of a dityrosine bond induced by excess H2O2. [ABSTRACT FROM AUTHOR]

Here, we describe the design of a novel mimic peroxidase, nanocomposites composed by 5,10,15,20-tetrakis(4-carboxyl phenyl)-porphyrin (H2TCPP) and cadmium sulfide (CdS). The H2TCPP-CdS nanocomposites can catalyze oxidation of substrate 3,3,5,5-tetramethylbenzidine (TMB) in the presence of H2O2 and form a blue product which can be seen by the naked eye in 5 min. The mechanism of the catalytic reaction originated from the generation of hydroxyl radical (⋅OH), which is a powerful oxidizing agent to oxidize TMB to produce a blue product. Then, we developed a colorimetric method that is highly sensitive and selective to detect glucose, combined with glucose oxidase (GOx). The proposed method allowed the detection of H2O2 concentration in the range of 4 x 10-6-1.4 x 10-5 M and glucose in the range of 1.875 x 10-5-1 x 10-4 M with detectable H2O2 concentration as low as 4.6 x 10-7 M and glucose as low as 7.02 x 10-6 M, respectively. The results provided the theoretical basis of practical application in glucose detecting and peroxidase mimetic enzymes. [ABSTRACT FROM AUTHOR]

Abstract: Stable and easily-handled synthetic materials mimicking natural enzymes activity would find important biotechnological applications. This article describes the synthesis and characterization of magnetic molecularly imprinted catalytic polymers that exhibit peroxidase-like activity towards 5-hydroxyindole-3-acetic acid (5-HIAA) oxidation. This multifunctional material is obtained from highly crystalline magnetite nuclei coated with a silica layer to protect the iron nucleus from oxidation and to provide anchoring for hydroxyl surface groups. After acrylic functionalization via sol–gel process, a molecularly imprinted polymer with hemin as catalytic center and 5-HIAA as template has been successfully attached to the structure. The resulting hybrid composite is magnetically separable and possesses excellent catalytic ability for the selective oxidation of the indoleamine metabolite tumor marker, showing Michaelis–Menten kinetics with this molecule but not towards other structural analogs. Therefore, it can be considered an artificial peroxidase enzyme. [Copyright &y& Elsevier]

A thermo-sensitive hydrogel composed of chitosan and α, β-glycerophosphate was successfully synthesized and used as basic substrate for primary culture and subculture of lymphoid cells from shrimp, Penaeus chinensis. When the ambient temperature increases from 4°C to 25°C, the chitosan/α, β-glycerophosphate solution transforms into a semi-solid three-dimensional hydrogel with a rough surface. Primary lymphoid cells formed were round and grew homogenously over the surface of the hydrogel. The peroxidase activity indicated that cells seeded on the hydrogel suffered less stress than cells suspended in control culture flasks. The higher relative viability and alkaline phosphatase activity of primary cultured lymphoid cells indicated that the chitosan/α, β-glycerophosphate hydrogel enhanced the growth and proliferation of lymphoid cells. Subsequent subculture of lymphoid cells starting from crumbled primary hydrogel culture was also successful. [ABSTRACT FROM AUTHOR]

Peroxidases are heme containing enzymes that are produced by a number of organisms. A plant peroxidase was purified from leaves of chard(Beta vulgarissubspeciescicla)by ammonium sulphate precipitation, dialyses, and CM-sephadex cation exchange chromatography. Purification fold was approximately 36 with 14.4% yield. Optimum pH, optimum temperature, optimum ionic strength, and stable pH values of purified enzyme were determined for the guaiacol substrate. Michaelis constant (KM) and maximum rate (Vmax) values were determined from the Lineweaver–Burk graph for pyrogallol and guaiacol substrates. Phenolic and flavonoid contents in chard that connected with antioxidant properties were determined as 17.5 μg (GAE/mg extract) and 11.7 μg (QE/mg extract), respectively. In conclusion, chard is shown to be a novel rich source of phenolic antioxidants and has a high peroxidase activity. [ABSTRACT FROM AUTHOR]

Some guanine-rich DNA sequences, which are called DNAzymes, can adopt G-quadruplex structures and exhibit peroxidase activity by binding with hemin. Although known DNAzymes show less activity than horseradish peroxidase, they have the potential to be widely used for the detection of target molecules in enzyme-linked immunosorbent assays if sequences that exhibit higher activity can be identified. However, techniques for achieving this have not yet been described. Therefore, we compared the DNAzyme activities of more than 1000 novelistically designed sequences with that of the original DNAzyme by using an electrochemical detection system on a 12K DNA microarray platform. To the best of our knowledge, this is the first description of an array-based assessment of peroxidase activity of G-quadruplex-hemin complexes. By using this novel assay system, more than 200 different mutants were found that had significantly higher activities than the original DNAzyme sequence. This microarray-based DNAzyme evaluation system is useful for identifying highly active new DNAzymes that might have potential as tools for developing DNA-based biosensors with aptamers. [ABSTRACT FROM AUTHOR]

Deuterohaemin-alanine-histidine-threonine-valine-glutamic acid-lysine (DhHP-6) is a synthetic heme-containing peroxidase mimic that exhibits a high peroxidase enzyme activity. Compared to other microperoxidases, DhHP-6 has a poor stability and tends to aggregate in aqueous solutions. In this study, poly(ethylene glycol) (PEG) was used to improve the properties of DhHP-6. Factors that affected the PEGylation product yield were investigated. PEGylated DhHP-6 (mPEG-DhHP-6) was characterized by reversed-phase high-pressure liquid chromatography (RP-HPLC), matrix-assisted laser desorption/ionization time of flight mass spectra (MALDI-TOF-MS), and ultraviolet-visible (UV-vis) spectroscopy. The results show that the optimal PEGylation reaction conditions were achieved when the PEGylation was conducted in a borate buffer solution at pH 8.0 and 25°C for 4 h with a feeding ratio of 2 equiv of active PEG. After PEGylation, mPEG-DhHP-6 showed a great improvement in its stability with little activity loss. The UV-vis spectra of DhHP-6 and mPEG-DhHP-6 in different pH solutions showed that the aggregation of DhHP-6 was partly suppressed after PEGylation. © 2012 Wiley Periodicals, Inc. J. Appl. Polym. Sci., 2013 [ABSTRACT FROM AUTHOR]

Abstract: Button mushrooms (Agaricus bisporus) were dipped for 2min in different concentrations (1%, 2% and 3%) of alginate, then placed in jars and ventilated continuously with 100% O2 for up to 16d at 4°C. Mushroom respiration rate, weight loss, texture, color, percent open caps, chemical properties and activities of polyphenol oxidase (PPO), and peroxidase (POD) were measured. The results indicate that treatment with alginate coating (2%)+100% O2 maintained a high level of firmness, and delayed browning and cap opening. In addition, alginate coating (2%)+100% O2 also delayed changes in the soluble solids concentration, total sugars and ascorbic acid, and inhibited the activity of PPO and POD throughout storage. Our study suggests that the use of alginate coating under a high oxygen modified atmosphere has the potential to maintain button mushroom quality and extend its postharvest life to 16d. [Copyright &y& Elsevier]

Abstract: Cytochrome c has been extensively used as model of peroxidase reaction. The peroxidase activity and stability of a triple mutant CYC-3 (N52I, Y67F and M80A) were studied and compared to those of wild type protein (Wt-16). The CYC-3variant showed ten-fold increased activity in styrene oxidation. An intermediary specie that resembles to Cpd 0 (FeIII eroxidases was detected through EPR measurements during the reaction of CYC-3 with H2O2. Using molecular dynamics (MD) it was found that mutations in CYC-3 induce conformational changes in the M80 loop promoting the rotation of D ring propionate toward heme iron and the inclusion of transient water molecules that could explain the formation of Cpd 0 intermediate. The effects of these conformational changes on the activity increase are discussed. [Copyright &y& Elsevier]

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The exact mechanism of cell death in neurodegenerative diseases remains obscure, but the aberrant assembly of proteins into fibrillar aggregates is accused to be the primary cause of pathogenesis. Furthermore, the structural determinants of protein fibrils that are responsible for cell dysfunction are not yet clear. In the present study, using α-crystallin-/α-chymotrypsin-based experimental systems, we reported non-specific peroxidase behavior of 'heme-amyloid fibril' complex using DNA, serotonin and DOPA as potential substrates. Also, amyloid-mediated inhibition of catalase activity was documented. More importantly, we suggested that uncontrollable peroxidase activity may be involved in neurodegenerative cell toxicity via several independent (mechanistic) routes. According to the results and literature, it is reasonable to assume that oxidative stress plays at least partially a causative rather than merely bystander role in the neurofibrillary pathology in AD/PD in vivo. Since the consequence of heme-amyloid interaction has yet to be identified, additional in vitro /in vivo data on it may help us to manage amyloid aggregation processes and postpone/attenuate the onset/extent of irreversible part of neurodegenerative pathogenesis. Graphical Abstract: [Figure not available: see fulltext.] [ABSTRACT FROM AUTHOR]

Abstract: PP1084 protein was exposed to gamma irradiation ranging from 5 to 500kGy. Native PAGE showed minor structural changes in PP1084 at 5kGy, and major structural changes at >15kGy. Size-exclusion chromatography (SEC) showed the formation of a new shoulder peak when the protein was irradiated with 15 and 30kGy, and a double peak appeared at 100kGy. The results of PAGE and SEC imply that PP1084 protein is degraded by gamma irradiation, with simultaneous oligomerization. PP1084 chaperone activity reached the highest level at 30kGy of gamma irradiation, and then, decreased in a dose-dependent manner with increasing gamma irradiation. However, the peroxidase activity significantly decreased following exposure to all intensities of gamma irradiation. The improvement of chaperone activity using gamma irradiation might be promoted by the oligomeric structures containing covalently cross-linked amino acids. Consequently, PP1084 modification using gamma irradiation could elevate chaperone activity by about 3–4 folds compared to the non-irradiated protein. [Copyright &y& Elsevier]

The objective of this work was to determine the inactivation kinetics of peroxidase enzyme and the change in color and texture in golden potato tubers (Solanum tuberosum phureja group) exposed to blanching with water at 80°C, 90°C and to saturated steam at 93°C. The heat transfer coefficients for the blanch water ranged between 214 and 230 W °C-1 m-2 while for saturated steam they showed an average of 84.5 W °C-1 m-2. At different pre-cooking times the golden potato tuber showed an area that developed enzymatic browning, which was fitted to the Michaelis-Menten kinetic model and another area where the color change decreased due to enzyme inactivation, which corresponded to first order kinetics, in a similar way as the texture behavior. The inactivation of peroxidase enzyme followed the Lumry-Eyring mechanism. [ABSTRACT FROM AUTHOR]