In this study, antioxidant and free radical scavenging activities of the natural antioxidative compound, pyrogallol-phloroglucinol-6,6'-bieckol (PPB) isolated from brown algae, Ecklonia cava was assessed in vitro by measuring the radical scavenging activities (DPPH, alkyl, hydroxyl, and superoxide) using electron spin resonance (ESR) spectrometry, intracellular reactive oxygen species (ROS) scavenging activity, and DNA damage assay. According to the results of these experiments, the scavenging activity PPB against difference radicals was in the following order: DPPH, alkyl, hydroxyl, and superoxide radicals (IC50; 0.90, 2.54, 62.93 and 109.05 microM). The antioxidant activities of PPB were higher than that of the commercial antioxidant, ascorbic acid. Furthermore, PPB effectively inhibited DNA damage induced by H2O2. These results suggest that the natural antioxidative compound, PPB, can be used by the natural food industry.
Ascorbic Acid ; DNA Damage ; Electron Spin Resonance Spectroscopy ; Food Industry ; Phaeophyta ; Reactive Oxygen Species ; Spectrum Analysis ; Superoxides

Helicobacter pylori, a causative agent of gastroduodenal diseases, is a Gram-negative microaerophilic bacterium. Although H. pylori locates in the microaerophilic mucous layer, the bacteria would come into contact harmful reactive oxygen species generated by host immune system. It has been reported that H. pylori harbors various defense mechanisms which can protect bacterial cells from oxygen exposure. The change of the gene expression profile of sodB-negative isogenic mutant of H. pylori 26695 was analyzed by high resolution 2-DE followed by matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and tandem MS and microarray analysis. Eighteen genes and 41 genes were upregulated and downregulated respectively, either transcriptionally or translationally. Expression levels of three genes including trxB, yxjE and ribE that were changed both on a mRNA level and on a protein level were confirmed by RT-PCR analysis. However, change of expression levels of other major antioxidants such as KatA, AhpC and NapA were not detected, which means Sod is regulated by different way from that of KatA and AhpC. Mutant study of other antioxidant proteins may give us better understanding for the regulation of stress response in H. pylori.
Antioxidants ; Bacteria ; Defense Mechanisms ; Gene Expression* ; Helicobacter pylori ; Helicobacter* ; Immune System ; Mass Spectrometry ; Microarray Analysis ; Oxygen ; Reactive Oxygen Species ; Ribes ; RNA, Messenger ; Superoxide Dismutase* ; Superoxides* ; Transcriptome

capacitation of human sperm is essential for fertilization and is characterized visually by hyperactivated motility. There is a controversy whether reactive oxygen radicals cause infertility or stimulate sperm-zona interaction. We investigated the exact role of reactive oxygen radicals on hyperactivation (HA) of human sperm which could be a part of the capacitation process. Hyperactivation of human sperm was compared to the Ham's F-10 controls by the addition of superoxide anion and hydrogen peroxide generating enzymes on the percale treated sperms. The motility parameters of human sperms were estimated by computer assisted semen analysis system. The addition of xanthine + xanthine oxidase + catalase (generating system of superoxide anion and removal of hydrogen peroxide) on the sperms induced levels of HA (10.5% at 2 hour, 11.3% at 5 hour) which were about 2 times higher than those of controls (HA: 5.4% at 2 hour, 5.6% at 5 hour). The addition of glucose + glucose oxidase (generation of hydrogen peroxide) decreased the levels of HA (0.0% at 2 and 5 hour) significantly. Superoxide dismutase, the scavenger of superoxide anion inhibited HA significantly, whereas catalase, the scavenger of hydrogen peroxide promoted HA significantly These results suggest that the reactive oxygen radicals may be involved in hyperactivation of human sperms by the way that superoxide anion promotes and hydrogen peroxide inhibits hyperactivation of the fertile human sperms. It may be very important in the process of fertilization that promotion or inhibition of hyperactivation occurs at the proper time and location of female genital organ.
Catalase ; Female ; Fertilization ; Genitalia ; Glucose ; Glucose Oxidase ; Humans* ; Hydrogen ; Hydrogen Peroxide ; Infertility ; Reactive Oxygen Species* ; Semen Analysis ; Spermatozoa* ; Superoxide Dismutase ; Superoxides ; Xanthine ; Xanthine Oxidase

PURPOSE: Rebamipide is a propionic acid derivative that inhibits superoxide production and removes hydroxyl radicals. This study was performed to investigate the effects of adding rebamipide to semen, in an effort to determine if reactive oxygen species (ROS) production and lipid peroxidation of the sperm cell membrane as well as an improvement in seminal parameter and fertilizing capacity under oxidative stress was inhibited. MATERIALS AND MTHODS: Semen was collected from 30 normal healthy volunteers by masturbation after at least 48 hours abstinence. After liquefaction of the semen at room temperature, the prepared sperm was diluted with a sperm wash media to a uniform density of 20x106/ml. The semen was treated with 0.25ml of 0.2mM FeSO4 and 1mM sodium ascorbate for 60 min in the presence of various rebamipide concentrations (0, 10, 30, 100, and 300microM). ROS production, sperm motility, vitality, fertilizing capacity and the level of lipid peroxidation were analyzed by chemiluminescence, computer assisted semen analysis, eosin-nigrosin staining, a hypo-osmotic swelling test and the thiobarbituric acid method, respectively. RESULTS: Rebamipide at 100 and 300microM increased the sperm motility (p<0.05) but did not affect the sperm vitality. The ROS production and lipid peroxidation in the sperms treated with FeSO4/sodium ascorbate were inhibited by rebamipide in a dose-dependent fashion (p<0.05 in each). The total swelling rate of the hypo-osmotic swelling test was also increased by high rebamipide concentrations (100 and 300microM), respectively 49.2 17.9 and 50.8 21.7% (p<0.05). CONCLUSIONS: These results suggest rebamipide is an effective free radical scavenger and may be useful as an oral antioxidant in patients with male infertility due to increased ROS generation. However, further study to be possible the clinical use of rebamipide for improve the fertilizing capacity in male infertility is required.
Antioxidants* ; Ascorbic Acid ; Cell Membrane ; Diethylpropion ; Healthy Volunteers ; Humans* ; Infertility, Male ; Lipid Peroxidation ; Luminescence ; Male ; Masturbation ; Oxidative Stress ; Reactive Oxygen Species ; Semen Analysis ; Semen* ; Sperm Motility ; Spermatozoa ; Superoxides

PURPOSE: Rebamipide is a propionic acid derivative that inhibits superoxide production and removes hydroxyl radicals. This study was performed to investigate the effects of adding rebamipide to semen, in an effort to determine if reactive oxygen species (ROS) production and lipid peroxidation of the sperm cell membrane as well as an improvement in seminal parameter and fertilizing capacity under oxidative stress was inhibited. MATERIALS AND MTHODS: Semen was collected from 30 normal healthy volunteers by masturbation after at least 48 hours abstinence. After liquefaction of the semen at room temperature, the prepared sperm was diluted with a sperm wash media to a uniform density of 20x106/ml. The semen was treated with 0.25ml of 0.2mM FeSO4 and 1mM sodium ascorbate for 60 min in the presence of various rebamipide concentrations (0, 10, 30, 100, and 300microM). ROS production, sperm motility, vitality, fertilizing capacity and the level of lipid peroxidation were analyzed by chemiluminescence, computer assisted semen analysis, eosin-nigrosin staining, a hypo-osmotic swelling test and the thiobarbituric acid method, respectively. RESULTS: Rebamipide at 100 and 300microM increased the sperm motility (p<0.05) but did not affect the sperm vitality. The ROS production and lipid peroxidation in the sperms treated with FeSO4/sodium ascorbate were inhibited by rebamipide in a dose-dependent fashion (p<0.05 in each). The total swelling rate of the hypo-osmotic swelling test was also increased by high rebamipide concentrations (100 and 300microM), respectively 49.2 17.9 and 50.8 21.7% (p<0.05). CONCLUSIONS: These results suggest rebamipide is an effective free radical scavenger and may be useful as an oral antioxidant in patients with male infertility due to increased ROS generation. However, further study to be possible the clinical use of rebamipide for improve the fertilizing capacity in male infertility is required.
Antioxidants* ; Ascorbic Acid ; Cell Membrane ; Diethylpropion ; Healthy Volunteers ; Humans* ; Infertility, Male ; Lipid Peroxidation ; Luminescence ; Male ; Masturbation ; Oxidative Stress ; Reactive Oxygen Species ; Semen Analysis ; Semen* ; Sperm Motility ; Spermatozoa ; Superoxides

To determine whether oxygen free radicals are responsible for the pathogenesis of the cholestasis induced by ligation of common bile duct (CBD) variables which reflect the hepatic function in the serum, the amount of superoxide radical production, and xanthine oxidase(XO) activity were studied. The activity of serum alanine aminotransferase, bilirubin level in the serum and the amount of superoxide radical production were lower in a CBD ligation with allopurinol treated group than in a CBD ligation without allopurinol treated group. Abnormalities of the microscopic structures were reduced in a CBD ligation with allopurinol treated group than in a CBD ligation without allopurinol treated group. Allopurinol, an inhibitor of XO, prevented the hepatic damage induced by CBD ligation through the inhibition of XO. These experiments demonstrate that oxygen free radicals are responsible for the pathogenesis of the cholestatic liver.
Allopurinol/*pharmacology ; Animal ; Bile Ducts ; Cholestasis/*pathology ; Enzyme Inhibitors/*pharmacology ; Free Radicals ; Ligation ; Liver/*pathology ; Male ; Rats ; Rats, Sprague-Dawley ; Superoxides/metabolism ; Xanthine Oxidase/analysis/*antagonists & inhibitors

OBJECTIVETo investigate the influence of pinacidil preconditioning on the protection of the structure and respiratory function of injured myocardial mitochondria in scalded rats.

METHODSSeventy-five healthy Wistar rats, weighed 250 approximately 300 g, were randomly divided into three groups: i.e. control (n = 9, with intraperitoneal injection of 50 microg/kg isotonic saline), scald (n = 33, with 30% TBSA full thickness scald) and pre-conditioning (n = 33, with same extent of scald injury after intraperitoneal injection of 50 microg/kg pinacidil) groups. Mitochondrial ultrastructure was observed by transmission electron microscope. The mitochondrial respiratory function, the MDA content and the superoxide anion level were determined with corresponding methods.

RESULTSThe degree of injury to rat myocardial mitochondria in pre-conditioning group was less intensive than that in scald group (P < 0.05 or 0.01). The respiratory control rate in pre-conditioning group was obviously higher than that in scald group (P < 0.05), and the contents of MDA and superoxide anion in pre-conditioning group were markedly lower than those in scald group (P < 0.05 or 0.01), as evidenced by their contents at 3 post scalding hours (0.60 +/- 0.09 micromol/g and 0.127 +/- 0.020) were obviously lower than those in scald group (0.83 +/- 0.07 micromol/g and 0.169 +/- 0.015) (P < 0.01).

CONCLUSIONPinacidil preconditioning was beneficial in the protection of myocardial mitochondria in scalded rats, and it might be related to the pre-opening of potassium channel which was sensitive to mitochondrial ATP.

Animals ; Burns ; drug therapy ; metabolism ; pathology ; Cell Respiration ; drug effects ; Disease Models, Animal ; Mitochondria, Heart ; metabolism ; pathology ; Pinacidil ; therapeutic use ; Rats ; Rats, Wistar ; Superoxides ; analysis

OBJECTIVETo study the effect of terephthalic acid (TPA) on lipid metabolism in Sprague-Dawley (SD) rats.

METHODSFive groups of SD rats that ingested 0%, 0.04%, 0.2%, 1%, and 5% TPA, respectively, were included in a 90-day subchronic feeding study. Effects of TPA on levels of serum protein, total cholesterol (TC), triglycerides (TG), high-density lipoprotein cholesterol (HDL), total antioxidative capability (T-AOC), superoxide dismutase (SOD) and malondialdehyde (MDA) were observed. Urine samples were collected and analyzed for concentration of ion.

RESULTSTPA decreased the level of serum T-AOC in a dose dependent manner. The contents of serum and bladder MDA significantly decreased in 1% and 5% TPA ingestion groups. Serum CuZn superoxide dismutase (CuZnSOD) lowered in groups of 0.2%, 1%, and 5% TPA. TPA subchronic feeding had no significant influences on serum TC, LDL or HDL, but increased serum TG, TP and ALB after administration of 0.04% and/or 0.2% TPA. Concentrations of urinary Ca2+, Mg2+, Na+, and K+ were elevated in 1% and 5% TPA groups.

CONCLUSIONAntioxidative potential decreased after TPA exposure. MDA increase in serum and bladder tissues was one of the most important reactions in rats which could protect themselves against TPA impairment. The decrease of serum CuZnSOD was related to the excretion of Zn2+.

Animals ; Antioxidants ; analysis ; Blood Proteins ; analysis ; Cholesterol ; blood ; Female ; Ions ; urine ; Lipid Metabolism ; drug effects ; Lipoproteins ; blood ; Male ; Malondialdehyde ; blood ; Phthalic Acids ; toxicity ; Rats ; Rats, Sprague-Dawley ; Superoxides ; blood ; Triglycerides ; blood ; Weight Gain

The use of lower extremity tourniquets for procedures of the lower leg is considered routine in orthopedic surgery, yet, lower extremity tourniquets are not benign. While the tourniquet is inflated, metabolic changes such as increased PaCO2, lactic acid, and serum potassium and decreased level of PaO2. and pH occur in the ischemic limb. Deflation of tourniquet results in release of anaerobic metabolic products during ischemia into systemic circulation. In this ischemia/reperfusion situation, oxygen free radicals could potentially be produced during the reperfusion period by several mechanisms. One of these mechanisms is release of intracellular superoxide or hydrogen peroxide by activated neutrophils in the area. These reactive oxygen species(ROS) could be a causative factor for the postreperfusion no-flow, lung injury, induction of tourniquet shock, etc, The purpose of this clinical study was to investigate the effect of tourniquet deflation on the hemodynamic changes, changes of blood gas analysis, and hydrogen peroxide production using flow cytometric analysis of fluorescent DCF(Dichlorofluorescein). Quantitative Analysis of fluorescent DCF was performed in resting and fMLP(N-formyl-methyonyl-leucyl-phenylalanine) or PMA(phorbol myristate acetate) stimuliated neutrophils. The results were as follows: 1)The hemodynamic changes (systolic and diastolic BP, pulse rate) did not show any significant difference before and after tourniquet release(P>0.05). 2)Arterial pH deceased significantly until 10min and PaC was increased significantly until Smin after toumiquet release(P<0.05). 3) Arterial PO2, bicarbonate, base excess showed no significant change before and after tour- niquet release(P>0.05). 4) Hydrogen peroxide production which was estimated by fluorescent DCF in neutrophils did not show any significant change before and after tourniquet release(P>0.05). These results indicate that tourniquet application(400mmHg, less than 2hours) could not release significant hydrogen peroxide during reperfusion period after tourniquet release.
Blood Gas Analysis ; Extremities ; Flow Cytometry ; Free Radicals ; Hemodynamics ; Hydrogen Peroxide* ; Hydrogen* ; Hydrogen-Ion Concentration ; Ischemia ; Lactic Acid ; Leg ; Lower Extremity ; Lung Injury ; Myristic Acid ; Neutrophils* ; Orthopedics ; Oxygen ; Potassium ; Reperfusion ; Shock ; Superoxides ; Tourniquets*

This study was performed to evaluate the effect of granulocyte-colony stimulating factor on neutrophil functions in diabetic patients with active foot infections in vitro. Twelve diabetic patients with foot infections and 12 normal volunteers were enrolled. Neutrophils from peripheral blood were incubated with granulocyte colony-stimulating factor (G-CSF, 50 ng/mL) for 20 min. Superoxide production of neutrophils was measured by the reduction of ferricytochrome C. Neutrophil phagocytosis was assayed using Staphylococcus aureus and the weighted phagocytic index was calculated. Superoxide production of neutrophils in diabetic patients with foot infections was 7.7 (unit: nmol/2 x 10(5) cells/60 min), which was significantly lower than that in controls (12.0) (p<0.05). G-CSF increased neutrophil superoxide production to 12.1 in diabetic patients with foot infections and to 19.8 in controls (p<0.05 for each). Weighted phagocytic index in diabetic patients with foot infections was 0.77, which was not significantly different from that of the controls (0.69). Weighted phagocytic index was increased significantly by G-CSF to 0.88 in diabetic patients with foot infections and to 0.79 in controls (p<0.05 for each). In conclusion, G-CSF significantly enhanced neutrophil functions in diabetic patients with foot infections in vitro.
Adult ; Aged ; Bacterial Infections/immunology* ; Diabetes Mellitus/immunology* ; Female ; Foot Diseases/immunology* ; Granulocyte Colony Stimulating Factor, Recombinant/pharmacology* ; Human ; Male ; Middle Age ; Neutrophils/immunology ; Neutrophils/drug effects* ; Receptors, IgG/analysis ; Superoxides/metabolism