Peroxidase In Solution Research Articles

Effects of Zinc Compounds on the Enzymatic Activities of Lysozyme and Peroxidase and Their Antifungal Activities

This study aimed to investigate the effects of zinc compounds on the enzymatic activities of lysozyme, peroxidase, and the glucose oxidase–mediated peroxidase (GO-PO) system and their antifungal activities. Four different zinc compounds (zinc chloride, gluconate, lactate, and sulfate) were incubated with hen egg-white lysozyme (HEWL), bovine lactoperoxidase (bLPO), the GO-PO system, and human unstimulated whole saliva in solution and on a hydroxyapatite surface. Enzymatic activities of lysozyme, peroxidase, and the GO-PO system were measured through the hydrolysis of Micrococcus lysodeikticus, oxidation of fluorogenic 2′,7′-dichlorofluorescin, and glucose assay, respectively. Interactions between zinc and enzymes were analyzed by surface plasmon resonance (SPR). The minimum inhibitory concentration (MIC) and candidacidal activities of zinc compounds were examined against three Candida albicans strains. Zinc gluconate and sulfate significantly increased the enzymatic activities of salivary lysozyme in the solution assay and of HEWL and salivary lysozyme on the hydroxyapatite surface. However, all examined zinc compounds significantly decreased the enzymatic activities of bLPO and salivary peroxidase in solution and on the surface. SPR analyses revealed binding of zinc to lysozyme and peroxidase, with affinity differing according to the zinc compounds. The MIC of zinc compounds against C. albicans was 1.0–2.4 mM. Candidacidal activities were 17.7–38.8% and 23.7–47.0% at 1.0 and 10 mM concentrations, respectively. In conclusion, zinc compounds enhanced lysozyme activity but inhibited peroxidase activity. Zinc compounds exhibited concentration-dependent candidacidal activity against C. albicans. Zinc compounds are potential therapeutic agents for oral health, especially for geriatric patients.

Effects of molecular weight of hyaluronic acid on its viscosity and enzymatic activities of lysozyme and peroxidase

ObjectivesTo investigate the effects of the molecular weight of hyaluronic acid on its viscosity and enzymatic activities of lysozyme and peroxidase in solution and on the hydroxyapatite surface. DesignHyaluronic acids of four different molecular weights (10 kDa, 100 kDa, 1 MDa, and 2 MDa), hen egg-white lysozyme, bovine lactoperoxidase, and human whole saliva were used. Viscosity values of hyaluronic acids were measured using a cone-and-plate viscometer at six different concentrations (0.1–5.0 mg/mL). Enzymatic activities of lysozyme and peroxidase were examined by hydrolysis of fluorescein-labeled Micrococcus lysodeikticus and oxidation of fluorogenic 2′,7′-dichlorofluorescein to fluorescing 2′,7′-dichlorofluorescein, respectively. ResultsIn solution assays, only 2 MDa-hyaluronic acid significantly inhibited lysozyme activities in saliva. In surface assays, hyaluronic acids inhibited lysozyme and peroxidase activities; the inhibitory activities were more apparent with high-molecular-weight ones in saliva than in purified enzymes. The 100 kDa-hyaluronic acid at 5.0 mg/mL, 1 MDa-one at 0.5 mg/mL, and 2 MDa-one at 0.2 mg/mL showed viscosity values similar to those of human whole saliva at a shear rate range required for normal oral functions. The differences among the influences of the three conditions on the enzymatic activities were not statistically significant. ConclusionsHigh-molecular-weight hyaluronic acids at low concentration and low-molecular-weight ones at high concentration showed viscosity values similar to those of human whole saliva. Inhibitory effects of hyaluronic acids on lysozyme and peroxidase activities were more significant with high-molecular-weight ones on the surface and in saliva compared with in solution and on purified enzymes.

Multidisciplinary approach to study the effect of water status and mobility on the activity of peroxidase in solutions

The effect of water mobility on horseradish peroxidase (HRP) activity in solutions was investigated by measuring water activity (aw), freezable water content, 1H proton transverse relaxation time and water self-diffusivity determined by nuclear magnetic resonance. The effect of system mobility as described by viscosity and glass transition temperature (T′g) was also studied.The aw and viscosity of aqueous solutions were modulated using ligands (glucose, sorbitol and trehalose) and a thickener (maltodextrin).The effectiveness of a solute in the inhibition of HRP activity was better related to its ability to reduce the mobility of the system than to its water mobility depleting effect.The relationship among viscosity and peroxidase activity was influenced by the type of enzyme but not by the substrate. Bovine lactoperoxidase activity was hindered by viscosity changes more than HRP activity (tested in the same system) due to the higher molecular weight of the former enzyme.

Studies on the interaction of Nile red with horseradish peroxidase in solution

The interaction of an extrinsic probe (Nile red) with an enzyme (horseradish peroxidase) in solution was investigated using fluorescence techniques. Nile red fluorescence is very environmentally sensitive and the presence of domains of differing polarity within the enzyme was ascertained by the decomposition of the Nile red emission spectrum. Further evidence for the position of the probe inside the enzyme was obtained from a molecular modeling study. A decrease in the emission intensity of the dye during incubation with horseradish peroxidase was explained by the occurrence of resonance energy transfer between the Nile red and the heme group in the enzyme. This was supported by a calculation of the critical transfer distance and a comparison of the fluorescence intensity of the dye in both the holo- and apo-enzyme. These data were then applied to the study of the effect of temperature on the structure of the enzyme, where changes in conformation were elucidated.

Effect of surfactants on peroxidase activity: IV. Effect of milk lipids and fatty acid salts

The effect of surfactants, lipids and fatty acid salts isolated from cow milk on the activity of hemecontaining horseradish peroxidase in solution was studied. As the surfactant concentration increases, the rate of the enzymic reaction successively decreases, increases, and again decreases, down to zero in the case of the fatty acid salts. The initial deceleration of the reaction rate results from the enzyme inhibition. The subsequent increase is caused by an improved accessibility for the substrate and the enhanced activity of the catalytic site of the enzyme due to its immobilization in the surfactant aggregates. A shielding of the protein by these aggregates can explain the secondary deceleration of the enzymic reaction rate. The general character of the dependence is similar and does not depend on the surfactant structure for a series of fatty acid salts and phospholipids; however, it is quite different in the case of cholesterol and sphingomyelin.

Direct electrocatalytic determination of dissolved peroxidases

A novel voltammetric approach is described for the quantification of dissolved redox enzymes. The measurement of horseradish peroxidase and microperoxidase is based on the electrocatalytic current generated when the enzyme reduces hydrogen peroxide to water and oxidises at low potential a mediator which is regenerated in a cathodic electrode reaction. Since the mediator regeneration is sufficiently fast, the overall reaction rate is determined by the enzyme activity. As a consequence stationary current-time curves are obtained in dependence on the activity of peroxidase in solution. For the chemical modification the electrode was modified with 1-(N,N-dimethylamine)-4-(4-morpholine)benzene, a mediator of low solubility in reduced form and with favourably low formal potential. At a working potential of − 50 mV vs. 0.1 M Ag AgCl cathodic steady-state currents were obtained after injection of peroxidase in the presence of 1 mmol/1 hydrogen peroxide. The detection limit for horseradish peroxidase and microperoxidase-11 were 10ng/ml (250 pmol/1) and 20 ng/ml (10 nmol/1), respectively.

4757016 Process for stabilizing the activity of peroxidase in solution: Dagmar Klenner, Gerd Kleinhammer, Rolf Deeg, Tutzing, Federal Republic of Germany assigned to Boehringer Mannheim GmbH

4757016 Process for stabilizing the activity of peroxidase in solution: Dagmar Klenner, Gerd Kleinhammer, Rolf Deeg, Tutzing, Federal Republic of Germany assigned to Boehringer Mannheim GmbH

4764468 Process for stabilizing the activity of peroxidase in solution: Rainer Wehner, Johann Mattersberger, Dagmar Klenner, Gerlinde Deutsch-Weyer, Gauting, Federal Republic of Germany assigned to Boehringer Mannheim GmbH

4764468 Process for stabilizing the activity of peroxidase in solution: Rainer Wehner, Johann Mattersberger, Dagmar Klenner, Gerlinde Deutsch-Weyer, Gauting, Federal Republic of Germany assigned to Boehringer Mannheim GmbH

Improvement of survival rates of bovine blastocysts with sucrose for glycerol dilution after a fast freezing and thawing method

To investigate the effects of the molecular weight of hyaluronic acid on its viscosity and enzymatic activities of lysozyme and peroxidase in solution and on the hydroxyapatite surface.Hyaluronic acids of four different molecular weights (10 kDa, 100 kDa, 1 MDa, and 2 MDa), hen egg-white lysozyme, bovine lactoperoxidase, and human whole saliva were used. Viscosity values of hyaluronic acids were measured using a cone-and-plate viscometer at six different concentrations (0.1–5.0 mg/mL). Enzymatic activities of lysozyme and peroxidase were examined by hydrolysis of fluorescein-labeled Micrococcus lysodeikticus and oxidation of fluorogenic 2′,7′-dichlorofluorescein to fluorescing 2′,7′-dichlorofluorescein, respectively.In solution assays, only 2 MDa-hyaluronic acid significantly inhibited lysozyme activities in saliva. In surface assays, hyaluronic acids inhibited lysozyme and peroxidase activities; the inhibitory activities were more apparent with high-molecular-weight ones in saliva than in purified enzymes. The 100 kDa-hyaluronic acid at 5.0 mg/mL, 1 MDa-one at 0.5 mg/mL, and 2 MDa-one at 0.2 mg/mL showed viscosity values similar to those of human whole saliva at a shear rate range required for normal oral functions. The differences among the influences of the three conditions on the enzymatic activities were not statistically significant.High-molecular-weight hyaluronic acids at low concentration and low-molecular-weight ones at high concentration showed viscosity values similar to those of human whole saliva. Inhibitory effects of hyaluronic acids on lysozyme and peroxidase activities were more significant with high-molecular-weight ones on the surface and in saliva compared with in solution and on purified enzymes.

UPTAKE AND INTRACELLULAR DIGESTION OF PROTEIN (PEROXIDASE) IN PLANARIANS

1. Stages in the ingestion of protein, and the formation and ultimate dis-appearance of spherules in the phagocytic cells of Dugesia tigrina were visualized histochemically by the peroxidase technique. 2. The formation of spherules involved the coalescence of numerous small peroxidase-positive granules. Typical spherules were present three hours after feeding kidney from a mouse previously injected with peroxidase, and maximal size of the spherules was achieved by the end of 24 hours. 3. Peroxidase-positive material was confined to the phagocytic cells and disappeared gradually until none remained 8 days following feeding. 4. Pharyngectomized planarians exposed to a medium containing peroxidase in solution absorbed the protein through the epidermis, and formed typical spherules in the phagocytic cells. It is suggested that this may indicate a role of transepidermal absorption of nutrients in regenerating planarians.