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PubMed2002

Lignification related enzymes in Picea abies suspension cultures.

Activity of a number of enzymes related to lignin formation was measured in a Picea abies (L) Karsten suspension culture that is able to produce native-like lignin into the nutrient medium. This cell culture is an attractive model for studying lignin formation, as the process takes place independently of the complex macromolecular matrix of the native apoplast. Suspension culture proteins were fractionated into soluble cellular proteins, ionically and covalently bound cell wall proteins and nutrient medium proteins. The nutrient medium contained up to 5.3% of total coniferyl alcohol peroxidase (EC 1.11.1.7) activity and a significant NADH oxidase activity that is suggested to be responsible for hydrogen peroxide (H2O2) production. There also existed some malate dehydrogenase (EC 1.1.1.37) activity in the apoplast of suspension culture cells (in ionically and covalently bound cell wall protein fractions), possibly for the regeneration of NADH that is needed for peroxidase-catalysed H2O2 production. However, there is no proof of the existence of NADH in the apoplast. Nutrient medium peroxidases could be classified into acidic, slightly basic and highly basic isoenzyme groups by isoelectric focusing. Only acidic peroxidases were found in the covalently bound cell wall protein fraction. Several peroxidase isoenzymes across the whole pI range were detected in the protein fraction ionically bound to cell walls and in the soluble cellular protein fraction. One laccase-like isoenzyme with pI of approximately 8.5 was found in the nutrient medium that was able to form dehydrogenation polymer from coniferyl alcohol in the absence of H2O2. The total activity of this oxidase towards coniferyl alcohol was, however, several orders of magnitude smaller than that of peroxidases in vitro. According to 2D 1H-13C correlation NMR spectra, most of the abundant structural units of native lignin and released suspension culture lignin are present in the oxidase produced dehydrogenation polymer but in somewhat different amounts compared to peroxidase derived synthetic lignin preparations. A coniferin beta-glucosidase (EC 3.2.1.21) was observed to be secreted into the culture medium.

Journal Article↗
PubMed1999

Ceruloplasmin has a distinct active site for the catalyzing glutathione-dependent reduction of alkyl hydroperoxide.

Ceruloplasmin, a blue multi-copper alpha(2)-glycoprotein found in the plasma of all vertebrates, is capable of oxidizing aromatic amines and ferrous iron. Here, we report that human ceruloplasmin exhibits an alkyl hydroperoxide peroxidase activity, which is independent of the oxidase activity. The site-specific modification of the sulfhydryl of cysteine at position 699 in ceruloplasmin completely abolished the antioxidant activity, suggesting that ceruloplasmin is a peroxidase with a cysteinyl thiol as a functional nucleophile. The crystal structure of human ceruloplasmin reveals that the domain containing Cys-699 is apart from the multi-copper complex domains. Taken together, these data suggest that ceruloplasmin has a distinct active site for a glutathione-linked peroxidase activity apart from the copper complex site exerting ferroxidase activity.

Antioxidants↗
PubMed1981

A coupled NAD+-peroxidase spectrophotometric assay for cholesterol.

We describe a modified enzymic reagent for determination of cholesterol and cholesterol ester in serum and in high-density lipoprotein. In this new procedure, the hydrogen peroxide produced by the action of cholesterol oxidase is used as a substrate for NAD+ peroxidase. Spectrophotometric determination of the NADH consumed in this coupled reaction provides a direct and absolute measure of the cholesterol originally present in free form and that liberated by the action of cholesterol ester hydrolase.

Cholesterol↗
PubMed2002

Involvement of peroxidases in the formation of the brown coloration of heartwood in Juglans nigra.

Oxidase activities were investigated within the cross-section of walnut trunk in relation to the brown staining of heartwood, especially in the transition zone where the colour change occurs. The distribution of peroxidase activity was investigated using 3,3'-diaminobenzidine (DAB) or guaiacol as a substrate. Generally, the highest activity was found in the cambial zone and in the middle sapwood. This activity was mainly vacuolar. However, during autumn a peak of activity was observed in the transition zone with DAB, but not with guaiacol. Immunohistolocalization of the peroxidase revealed that the protein was present in the transition zone even if the enzymatic activity was not detectable. Flavan-3-ols were abundantly localized in the transition zone and it is hypothesized that they are physiological substrates of peroxidases. By contrast, polyphenoloxidases do not seem to be implicated in heartwood formation.

Catechol Oxidase↗
PubMed1978

Determination of uric acid on continuous-flow (AutoAnalyzer II and SMA) systems with a uricase/phenol/4-aminophenazone color test.

This report describes a new specific colorimetric procedure for uric acid assay with AutoAnalyzer II and SMA (Technicon) systems, made specific by the application of uricase. Hydrogen preroxide, formed in this reaction, effects the oxidative coupling of 4-aminophenazone and 2,4-dichlorophenol under the catalytic influence of peroxidase. The red dye formed is measured at 505 or 520 nm. A sample blank measurement is not necessary, and the reagents show very good stability. The test shows linearity up to 714 mumol of uric acid per liter. Results of thie method correlate very well with those by the uricase-ultraviolet and uricase--catalase methods. There is no interference by hemoglobin, bilirubin, lipemia, and various drugs, except a minor interference by alpha-methyldopa. Interference from ascorbate is eliminated by ascorbate oxidase. This method can be regarded as a considerably improved routine test for uric acid on continuous-flow systems in clinical laboratories as compared with the commonly used phosphotungstate method.

Aminopyrine↗
PubMed2000

Evaluation of free radical status in CSF in childhood meningitis.

The study was undertaken to investigate the possible role of free radicals and antioxidants in childhood meningitis. Sixty children suffering from acute bacterial meningitis (ABM) or tuberculous meningitis (TBM) according to their clinical and laboratory findings were enrolled in the study. The production of superoxide anions (O2.-), hydrogen peroxide (H2O2) and malondialdehyde (MDA) and the activities of xanthine oxidase (XO), superoxide dismutase (SOD) and glutathione peroxidase (GPx) were monitored in the study groups and findings compared with those in 20 age-matched controls. Children with ABM and TBM who died registered significant increases in the production of O2.- and MDA and in the activities of SOD and CPK compared with survivors. The rate of production of oxidants and MDA and the activities of XO, SOD and CPK were of a much higher magnitude in deceased ABM and in ABM survivors than in fatal TBM and survivors, respectively. The abnormalities in most of the biochemical parameters investigated were more marked in the children with ABM than in TBM and controls (p < 0.001). Increased MDA production and creatine phosphokinase (CPK) activity of different magnitudes in the two study groups suggest varying degrees of tissue damage. The alterations observed in 20 children who died (14 from ABM, 6 from TBM) revealed elevated levels of oxidants, antioxidants and toxicity markers, particularly in ABM patients, which suggests the possibility that natural or synthetic antioxidants might prevent disease progression and tissue damage in childhood meningitis.

Antioxidants↗
PubMedDate not supplied

[Behavior of some serum enzymes and proteins in experimental revascularization of acute myocardial infarct].

Effect of reoxygenation of the serum enzymatic activities and protein fractions in dog with acute myocardial infarction. The enzymatic activitites creatin phosphokinase (CPK; E.C. 2.7.3.2), glutamate oxalacetate transaminase (GOT; E.C. 2.6.1.1.), diamine oxidase (DAO; E.C. 1.4.3.6.), monoamine oxidase (MAO; E.C. 1.4.3.4.), catalase (E.C. 1.2.1.6.), peroxidase (E.C. 1.2.1.6.) and the protein fractions levels are measured in the dog serum during experimental myocardial infarction, followed by 2 hours of reoxygenation. The alpha, and alpha2 globuline serum proteins increase during anoxic period; after reperfusion alpha1 globuline content further increase again, while alpha2 globuline fraction decrease to control values. The enzymatic activities, particulary CPK, DAO and MAO, significantly enhance during coronary occlusion. After riperfusion the DAO, GOT and peroxidase activities decrease, while the CPK, atalase and MAO activities further increase, particulary at the early stage of the reperfusion. These results are discussed interms of cellular mechanism, induced by oxygen readmission.

Acute Disease↗
PubMedDate not supplied

[Effect of perftoran on experimental hepatitis].

The effect of perftoran on the course of experimental acute hepatitis in albino rats was studied on the hepatitis models induced by allyl alcohol or P. acnes culture with typhoid fever endotoxin. Perftoran (10 ml/kg) favored more rapid cytolytic syndrome elimination by affecting the lipid peroxidation in rat liver. The drug inhibits the activity of prooxidant enzymes (xanthine oxidase and myeloperoxidase of Kupffer cells) and induces the synthesis of factors accounting for the antiperoxidation protection in hepatocytes such as catalase, glucose-6-phosphate dehydrogenase, and reduced glutathione.

Acute Disease↗
PubMed2000

Hydrogen peroxide generation by monoamine oxidases in rat white adipocytes: role on cAMP production.

In rat, white adipocytes monoamine oxidases (EC 1.4.3.4.) generate hydrogen peroxide (H(2)O(2)). Recent studies suggested that, in addition to its toxic features, H(2)O(2) may behave as a cell second messenger. In the present study, using fluorimetric and chemiluminescence (CL) assays, we showed that tyramine degradation by monoamine oxidases in intact adipocytes resulted in the concentration-dependent generation of H(2)O(2). In addition, we found that, in the presence of low tyramine concentrations, forskolin-dependent cAMP production was significantly increased as compared to that of the control and this increase was prevented by the monoamine oxidase inhibitor pargyline or by the H(2)O(2) trapping system homovanillic acid-peroxidase. Finally, we demonstrated that tyramine degradation by monoamine oxidases increased the ability of isoproterenol to induce cell lipolysis. Taken together, these data suggest that H(2)O(2) produced during substrate degradation by monoamine oxidases may participate in the regulation of adipocyte metabolism.

Adipocytes↗
PubMed1980

Role of hydroxyl radical in polymorphonuclear leukocyte-mediated bactericidal activity.

Among the antimicrobial systems of polymorphonuclear leukocytes (PMNs) are those that are oxygen-dependent. Recent interest has been directed at the hydroxyl radical(OH.), a highly reactive reduction product of oxygen, as a possible mediator of microbicidal activity. The role of OH. in PMN microbicidal activity is discussed in relation to the observations that (1) an enzymic OH. generating system consisting of xanthine oxidase plus acetaldehyde is bactericidal, (2) phagocytosing PMNs convert substrates like methional and 5,5-dimethyl pyrroline-1-oxide to products suggestive of OH.-mediated reactions and (3) PMN-mediated microbicidal activity is partially inhibited by scavengers of OH.

Acetaldehyde↗
PubMedDate not supplied

[A new oxidase method for analyzing L-lactate].

A new oxidase-coupled colorimetric method for analysis of L-lactate in biological fluids has been developed without use of peroxidase. The method is based on lactate oxidase-catalysed transformation of lactate to pyruvate which is determined photometrically in the next dye-producing reaction of 3-methyl-2-benzothiazolinone hydrazone (MBTH) in the presence of ferric ions. Sensitivity of the method is estimated as 0.1 micromole of analyte in 4-ml of reaction mixture. Linearity is observed in the range 0.1-1.0 micromole of L-lactate in sample (r = 0.99943; p < 0.0001). The developed method has been adapted for assay of L-lactic acid in kefirs and yogurts.

Catalysis↗
PubMed2003

Protection against hepatic ischemia-reperfusion injury in rats by oral pretreatment with quercetin.

OBJECTIVE: To investigate the possible protection provided by oral quercetin pretreatment against hepatic ischemia-reperfusion injury in rats. METHODS: The quercetin (0.13 mmol/kg) was orally administrated in 50 min prior to hepatic ischemia-reperfusion injury. Ascorbic acid was also similarly administered. The hepatic content of quercetin was assayed by high performance liquid chromatography (HPLC). Plasma glutamic pyruvic transaminase (GPT), glutamic oxaloacetic transaminase (GOT) activities and malondialdehyde (MDA) concentration were measured as markers of hepatic ischemia-reperfusion injury. Meanwhile, hepatic content of glutathione (GSH), activities of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and xanthine oxidase (XO), total antioxidant capacity (TAOC), contents of reactive oxygen species (ROS) and MDA, DNA fragmentation were also determined. RESULTS: Hepatic content of quercetin after intragastric administration of quercetin was increased significantly. The increases in plasma GPT, GOT activities and MDA concentration after hepatic ischemia-reperfusion injury were reduced significantly by pretreatment with quercetin. Hepatic content of GSH and activities of SOD, GSH-Px and TAOC were restored remarkably while the ROS and MDA contents were significantly diminished by quercetin pretreatment after ischemia-reperfusion injury. However, quercetin pretreatment did not reduce significantly hepatic XO activity and DNA fragmentation. Ascorbic acid pretreatment had also protective effects against hepatic ischemia-reperfusion injury by restoring hepatic content of GSH, TAOC and diminishing ROS and MDA formation and DNA fragmentation. CONCLUSION: It is indicated that quercetin can protect the liver against ischemia-reperfusion injury after oral pretreatment and the underlying mechanism is associated with improved hepatic antioxidant capacity.

Administration, Oral↗
PubMed1990

Influence of the reaction medium on the product distribution of peroxidase-catalysed oxidation of p-cresol.

p-Cresol was oxidized by hydrogen peroxide in a reaction catalysed by horseradish peroxidase and the low molecular weight products were investigated. In aqueous media Pummerer's ketone (I) was the dominating product but in organic media the product distribution was quite different; 2,2'-dihydroxy-5,5'-dimethyldiphenyl (II) was the main low molecular weight product. Similar product distributions were obtained with peroxidase adsorbed on a solid support and suspended in toluene and with peroxidase solubilized in a microemulsion containing the same solvent. The best selectivity for the formation of (II) was obtained when the enzyme was adsorbed on Celite and suspended in water-saturated chloroform with 0.5% (v/v) extra water added. The yield of low molecular weight products in this case was 28%; of this fraction, 95% was (II).

Cholesterol Oxidase↗
PubMed2006

Effects of Ginkgo biloba on plasma oxidant injury induced by bleomycin in rats.

Bleomycin is an anti-neoplastic agent and its clinical usage is limited by its toxicity, which is mostly induced by oxygen radicals. The aim of this study was to investigate the effect of Ginkgo biloba on plasma indices of oxidants induced by bleomycin in rats. Male Sprague-Dawley rats were divided into five groups: none medicated or 0.9% NaCl injected or only Ginkgo biloba (orally, 100 mg/kg per day for 14 days) or only a single dose of bleomycin (intratracheal, 2.5 U/kg) or Gingko biloba and bleomycin-treated groups. After 14 days, blood was taken before the rats were sacrificed. The plasma was removed and stored at -85 degrees C until the study day. Plasma superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and xanthine oxidase (XO) enzyme activities with malondialdehyde and nitric oxide (NO) levels were studied. The levels of malondialdehyde and NO with activity of XO were higher in plasma of bleomycin group than the other groups (P <0.05). The activities of SOD and GSH-Px were increased in the bleomycin plus Gingko biloba group in comparison with the bleomycin group (P <0.05). There was a positive correlation between malondialdehyde and NO levels in the bleomycin group (r =0.859, P <0.05). There were positive correlations between SOD and GSH-Px activities (r =0.760, P <0.05) and between XO activity and malondialdehyde level (r =0.822, P <0.05) in the bleomycin plus Gingko biloba group. In conclusion, it was thought that bleomycin induced oxidative stress can be prevented by Gingko biloba treatment via high anti-oxidant enzyme activity together with decreased radical production from XO.

Animals↗
PubMed2001

Tyrosine cross-linking of extracellular matrix is catalyzed by Duox, a multidomain oxidase/peroxidase with homology to the phagocyte oxidase subunit gp91phox.

High molecular weight homologues of gp91phox, the superoxide-generating subunit of phagocyte nicotinamide adenine dinucleotide phosphate (NADPH)-oxidase, have been identified in human (h) and Caenorhabditis elegans (Ce), and are termed Duox for "dual oxidase" because they have both a peroxidase homology domain and a gp91phox domain. A topology model predicts that the enzyme will utilize cytosolic NADPH to generate reactive oxygen, but the function of the ecto peroxidase domain was unknown. Ce-Duox1 is expressed in hypodermal cells underlying the cuticle of larval animals. To investigate function, RNA interference (RNAi) was carried out in C. elegans. RNAi animals showed complex phenotypes similar to those described previously in mutations in collagen biosynthesis that are known to affect the cuticle, an extracellular matrix. Electron micrographs showed gross abnormalities in the cuticle of RNAi animals. In cuticle, collagen and other proteins are cross-linked via di- and trityrosine linkages, and these linkages were absent in RNAi animals. The expressed peroxidase domains of both Ce-Duox1 and h-Duox showed peroxidase activity and catalyzed cross-linking of free tyrosine ethyl ester. Thus, Ce-Duox catalyzes the cross-linking of tyrosine residues involved in the stabilization of cuticular extracellular matrix.

Amino Acid Sequence↗
PubMed2002

The indices of endogenous oxidative and antioxidative processes in plasma from schizophrenic patients. The possible role of oxidant/antioxidant imbalance.

There is great evidence in recent years that oxygen free radicals play an important role in the pathophysiology of schizophrenia. The present study was performed to assess the changes in plasma nitric oxide (NO) and thiobarbituric acid-reactive substances (TBARS) levels, and superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and xanthine oxidase (XO) activities in schizophrenic patients compared to age- and sex-matched normal controls. A hundred patients with schizophrenia and 51 healthy volunteers were included in the study. XO, SOD, and GSH-Px activities as well as NO and TBARS levels were estimated by standard biochemical techniques in the plasma of normal healthy controls and schizophrenia patients. In schizophrenia, increased plasma XO activity (P < .0001) and NO levels (P < .0001), decreased SOD activity (P < .0001), and unchanged GSH-Px activity were detected compared to control group. Plasma TBARS levels were increased in schizophrenic patients (P < .01), especially in the residual subtype. TBARS levels in nonsmoker schizophrenic patients were found to be higher than nonsmoker controls. Although TBARS levels in both patients and controls were found to be higher in smokers as compared to nonsmokers, it was not statistically significant. No effects of duration of the illness, gender, and low and high dose of daily neuroleptic treatment equivalent to chlorpromazine on oxidant and antioxidant parameters were observed. Because the dose and the duration of treatment with drugs have no influence on the results, it can be interpreted that the findings are more likely to be related mainly to the underlying disease. These findings indicated a possible role of increased oxidative stress and diminished enzymatic antioxidants, both of which may be relevant to the pathophysiology of schizophrenia. On the other hand, increased NO production by nitric oxide synthetases (NOSs) suggests a possible role of NO in the pathophysiological process of schizophrenia. These findings may also suggest some clues for the new treatment strategies with antioxidants and NO synthase (NOS) inhibitors in schizophrenia.

Adolescent↗
PubMed2004

Effect of fish oil supplementation on plasma oxidant/antioxidant status in rats.

The aim of this study was to investigate effect of dietary omega-3 fatty acid supplementation on the indices of in vivo lipid peroxidation and oxidant/antioxidant status of plasma in rats. The plasma thiobarbituric acid reactive substances (TBARS) and nitric oxide (NO) levels, and activities of xanthine oxidase (XO), superoxide dismutase (SOD) and glutathione peroxidase (GSH-PX) were studied in male Wistar Albino rats after ingestion of 0.4 g/kg fish oil (rich in omega-3 fatty acids, eicosapentaenoic acid and docosahexaenoic acid) for 30 days and compared to untreated control rats. The rats in the treated group had significantly higher SOD activity (P < 0.001), NO levels (P < 0.01) and decreased TBARS levels (P < 0.05) with respect to controls whereas GSH-Px and XO activities were not significantly different between the groups. None of the measured parameters had significant correlation with each other in both groups. We conclude that dietary supplementation of omega-3 fatty acids may enhance resistance to free radical attack and reduce lipid peroxidation. These results support the notion that omega-3 fatty acids may be effective dietary supplements in the management of various diseases in which oxidant/antioxidant defence mechanisms are decelerated.

Animals↗