• Title/Summary/Keyword: oxygen free radical generating

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Establishment of New Method for the Assay of Glutamate-cysteine Ligase Activity in Crude Liver Extracts

  • Kwon Young-Hye;Stipanuk Martha H.
    • Toxicological Research
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    • v.22 no.1
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    • pp.39-45
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    • 2006
  • As the antioxidant and free radical scavenger, glutathione (GSH) participates in the preservation of cellular redox status and defense against reactive oxygen species and xenobiotics. Glutamate-cysteine ligase (GCL; also known as ${\gamma}$-glutamylcysteine synthetase, EC 6.3.2.2) is the rate limiting enzyme in GSH synthesis. In the present study, the accurate method for determination of GCL activity in crude liver extracts was developed by measuring both ${\gamma}$-glutamylcysteine and GSH from cysteine in the presence of glutamate, glycine and an ATP-generating system. We added glycine to promote the conversion of ${\gamma}$-glutamylcysteine to GSH, and to minimize the possibility of ${\gamma}$-glutamylcysteine metabolism to cysteine and oxoproline by ${\gamma}$-glutamylcyclotransferase. We established optimal conditions and substrate concentrations for the enzyme assay, and verified that inhibition of GCL by GSH did not interfere with this assay. Therefore, this assay of hepatic GCL under optimal conditions could provide a more accurate measurement of this enzyme activity in the crude liver extracts.

Effect of Lead Ion on The Hepatic Xanthine Oxidase Activity in Vitro (납이온이 잔틴 옥시다제 활성에 미치는 영향)

  • Huh, Keun;Shin, Uk-Seob;Lee, Sang-Hoon;Ann, Won-Hyo
    • YAKHAK HOEJI
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    • v.39 no.5
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    • pp.521-527
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    • 1995
  • This study was done to determine the effect of lead acetate on the activities of the hepatic cytosofic xanthine oxidase and aldehyde oxidase which were well known as oxygen free radical generating enzyme in vitro. Lead ion accelerated the formation of lipid peroxide and the increment of xanthine oxidase(type O) activity and the type conversion ratio from xanthine dehydrogenase to xanthine oxidase dose-dependently. But xanthine dehydrogenase(type D) activity was decreased. Aldehyde oxidase activity was not changed by lead ion. These data suggested that lead-induced cellular to)dcity may be concerned partially with xanthine oxidase mediated lipid peroxidation.

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Generation of Free Radicals by Interaction of Iron with Thiols in Human Plasma.

  • Lee, S. J.;K. Y. Chung;J. H. Chung.
    • Proceedings of the Korean Society of Food Hygiene and Safety Conference
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    • 2002.05a
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    • pp.138-138
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    • 2002
  • Oxidative stress has been associated with a number of diseases in human. Among the sources that can generate oxidative stress, it has been reported that iron can generate reactive oxygen species (ROS)with thiol. In iron overload state, increased thiol levels in plasma appeared to be associated with human mortality. In this study we examined whether iron could interact with thiols in plasma, generating ROS. In human plasma, unlike with Fe(III), Fe(II) increased lucigenin-enhanced chemiluminescence in concentration-dependent manner, and this was inhibited by SOD. Boiling of plasma did not affect chemiluminescence induced by Fe(II). Hovever, thiol depletion in plasma by pretreatment with N-ethylmaleimide (NEM)decreased Fe(II)-induced chemiluminescence significantly, suggesting that Fe(II) generated superoxide anion by the nonenzymatic reaction with plasma thiol. Consistent with this findings, albumin, the major thiol contributor in plasma, also generated ROS with Fe(II) and this generation was inhibited by pretreatment with NEM. Treatment with Fe(II) to plasma resulted un significant reduction of oxygen radical absorbance capacity (ORAC) value, suggest that total antioxidant capacity could diminished in iron overload state. In conclusion, In iron overload state, plasma may be affected by oxidative stress mediated by nonenzymatic reaction of Fe (II)with plasma thiol.

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A Study on the Oxidative Damage Induced by UVB Irradiation to Mouse Skin (UVB 조사로 인한 마우스 피부조직의 산화적 손상)

  • Rhie Sung-Ja;Kim Young-Chul
    • Environmental Analysis Health and Toxicology
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    • v.21 no.2 s.53
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    • pp.165-172
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    • 2006
  • The backs with a hair cut of 6-week-old healthy ICR male mice were once exposed to a dose of $400mJ/cm^2$ UVB. An acute dermal inflammation was observed, and the inflamed skins were almost completely cured after 6 days of the exposure. At 24 hours after exposure, the epidermal keratinocytes showed a cell-membrane damage with the destruction of intercellular junctions, agglutination of tonofilaments within the cytoplasm and nucleus damage. The activity of XO showed a significant increase (p<0.05) in up to 144 hours. The activities of CAT and SOD showed a significant decrease (p<0.05) in up to 96 hours, but they were not significantly different from the normal value at 144 hours. The GST activity was significantly decreased (p<0.01) in up to 96 hours, not so at 24 hours. However, that was not significantly different from the normal value at 144 hours. There was a significant decrease (p<0.01) in the contents of TBARS at 48 and 96 hours, without any significant difference at 144 hours. While the content of GSH was significantly lower (p<0.05) at 24 hours, that was not significantly different thereafter up to 144 hours from the normal value. Therefore, it is assumed that skin damage with a dose of $400mJ/cm^2$ UVB irradiation might be caused by the oxidative stress which was resulted from the unbalance of oxygen fret radical generating and scavenging enzymes.

Effects of Cordyceps Sinensis on Antioxidation in the Livers of Hydrocortisone Acetate-Treated Rats (동충하초(冬蟲夏草)가 hydrocortisone으로 유발시킨 양허(陽虛) 동물모형(動物模型)에서 항산화(抗酸化) 작용(作用)에 미치는 영향(影響))

  • Lee, Gu-Hyong;Min, Gun-Woo;Yoon, Cheol-Ho;Seo, Un-Kyo;Jeong, Ji-Cheon;Han, Yeong-Hwan;Shin, Uk-Seob;Park, Jong-Hyuck
    • The Journal of Internal Korean Medicine
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    • v.22 no.1
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    • pp.63-71
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    • 2001
  • Objectives : Cordyceps Sinensis (CS) was tested for the effects of antioxidant enzymes and lipid peroxidation in the liver. Methods : We measured the changes in body weight, enzyme activity, lipid peroxide and the death rate in the hydrocortisone acetate-treated rats. Results : In vitro, CS didn't effect levels of lipid peroxide. the activities, and the ratio of type conversion of xanthine oxidase. In the hydrocortisone acetate-treated rats, lipid peroxide, the activities, the ratio of type conversion of xanthine oxidase, and the death rate all increased. But, glutathione peroxidase and superoxide dismutase decreased. In vitro, after CS was administered to hydrocortisone acetatetreated rats, the levels of lipid peroxide in the liver, and the death rate decreased. However, the activities, and the ratio of type conversion of xanthine oxidase decreased. The body weight, glutathione peroxidase, and superoxide dismutase in+creased. The effects of Sinensis Cordyceps Broth did better than the effects of Sinensis Cordyceps Mycelia. Conclusions : These results suggest that CS decrease the activities of free radical generating enzymes which form lipid peroxide and increase the activities of oxygen free radical scavenging enzymes.

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Hepatotoxic Effects of 1-Furan-2-yl-3-pyridin-2-yl-propenone, a New Anti-Inflammatory Agent, in Mice

  • Jeon, Tae-Won;Kim, Chun-Hwa;Lee, Sang-Kyu;Shin, Sil;Choi, Jae-Ho;Kang, Won-Ku;Kim, Sang-Hyun;Kang, Mi-Jeong;Lee, Eung-Seok;Jeong, Tae-Cheon
    • Biomolecules & Therapeutics
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    • v.17 no.3
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    • pp.318-324
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    • 2009
  • 1-Furan-2-yl-3-pyridin-2-yl-propenone (FPP-3) has recently been synthesized and characterized to have an anti-inflammatory activity through the inhibition of the production of nitric oxide. In the present study, adverse effects of FPP-3 on hepatic functions were determined in female BALB/c mice. When mice were administered with FPP-3 at 125, 250 or 500 mg/kg for 7 consecutive days orally, FPP-3 significantly increased absolute and relative weights of liver with a dose-dependent manner. In addition, FPP-3 administration dramatically increased the hepatotoxicity parameters in serum at 500 mg/kg, in association of hepatic necrosis. FPP-3 significantly induced several phase I enzyme activities. To elucidate the possible mechanism(s) involved in FPP-3 induced hepatotoxicity, we investigated the hepatic activities of free radical generating and scavenging enzymes and the level of hepatic lipid peroxidation. FPP-3 treatment significantly elevated the hepatic lipid peroxidation, measured as the thiobarbituric acid-reactive substance, and the activity of superoxide dismutase. Taken together, the present data indicated that reactive oxygen species might be involved in FPP-3-induced hepatotoxicity.

Oxygen Toxicity of Superoxide Dismutase-Deficient Saccharomyces cerevisiae by Paraquat (Paraquat에 의해 유도된 Superoxide Dismutase 결핍 효모의 산소 독성)

  • 김지면;남두현용철순허근
    • KSBB Journal
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    • v.10 no.5
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    • pp.561-567
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    • 1995
  • Using superoxide dismutase (SOD)-deficient mutants of Saccharomyces cerevisiae, the oxygen toxicity induced by paraquat was studied. In aerobic culture condition, yeasts lacking MnSOD (milochondrial SOD) showed more significant growth retardation than CuZnSOD (cytoplasmic SOD)-deficient yeasts. However, not so big differences in growth pattern of those mutants compared with wild type were observed under anaerobic condition. When exposed to paraquat, the growth of yeasts lacking CuZnSOD was severely affected by higher than 0.01mM of paraquat in culture medium. By the analysis of several cellular components ivolved in free radical generating and scavenging system, it was found that, under aerobic condition, the content of lipid peroxides in cell membrane as well as cellular activity of glutathion peroxidase of CuZnSOD-deficient mutants was increased in the presence of paraquat, although significant decrease of catalase activity was observed in those stratns. In MnSOD-deficient yeast, however, increment in cellular activity of glutathion peroxldase and catalase by paraquat was observed without any deterioration of membrane lipid. It implies that the lack of mitochondrial SOD could be compensated by both of glutathion peroxldase and catalase, but that only glutathion peroxidase might act for CuZnSOD in cytoplasm. In contrast, all of SOD-deficient mutants showed a significant decrease in catalase activity, but slight increase in the activities of glutathion peroxidase, when cultivated anaerobically in the medium containing paraquat. Nevertheless, any significant changes of lipid peroxides in cell membranes were not observed during anaerobic cultivation of SOD-deficient mutants. It suggests that a little amount of free radicals generated by paraquat under anaerobic condition could be sufficiently overcome by glutathion peroxidase but not by catalase.

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Increased Antioxidative Activities against Oxidative Stress in Saccharomyces cerevisiae KNU5377 (산화 스트레스 대한 Saccharomyces cerevisiae KNU5377의 항산화 활성의 증가)

  • Kim, Il-Sup;Yun, Hae-Sun;Yang, Ji-Young;Lee, Oh-Seok;Park, Heui-Dong;Jin, Ing-Nyol;Yoon, Ho-Sung
    • Journal of Life Science
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    • v.19 no.4
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    • pp.429-435
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    • 2009
  • Oxidative stress is a consequence of an imbalance of the defense system against cellular damage generated by reactive oxygen species (ROSs) such as superoxide anions (menadione; MD). Most organisms have evolved a variety of defense systems to protect cells from adverse conditions. In order to evaluate stress tolerance against oxidative stress generating MD, comparative analyses of antioxidant capacity, or free radical scavenger ability, were performed between S. cerevisiae KNU5377 (KNU5377) and three wild-type S. cerevisiae strains. In a medium containing 0.4 mM MD, the KNU5377 strain showed higher cell viability and antioxidant ability, and contained higher levels of trehalose, superoxide dismutase, thioredoxin system, glucose-6-phosphate dehydrogenase, and some heat shock proteins. The KNU5377 strain also produced a lower level of oxidative stress biomarker than the other three yeast strains. These results indicate that S. cerevisiae KNU5377 has a higher level of tolerance to oxidative stress due to the increased expression of cell rescue proteins and molecules, thus alleviating cellular damage more efficiently than other S. cerevisiae strains.

Allopurinol Decreases Liver Damage Induced by Dermal Scald Burn Injury (피부 화상으로 유도된 간 손상에서 Allopurinol의 효과)

  • Cho, Hyun-Gug;Yoon, Chong-Guk;Park, Won-Hark
    • Applied Microscopy
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    • v.31 no.1
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    • pp.37-47
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    • 2001
  • In order to investigate a pathogenesis of liver damage induced by skin burn, thermal injury was induced by scald burn on entirely dorsal surface in rats (total burn surface area $20\sim25\%$) except for inhalated injury. At 5 and 24 h after scald burn, biochemical assay and morphological changes in serum and liver tissue were examined. Skin burn increased liver weight (% of body weight, p<0.05) and the activity of serum aniline amino-transferase (ALT, p<0.05), in addition, the activity of xanthine oxidase (XO), an enzyme of oxygen free radical generating system, was elevated (p<0.01) in serum, but not in skin and in liver. Postburn treatment of allopurinol intraperitoneally decreased liver weight, serum ALT activity and serum XO activity. Scald burn induced ultrastructurally swelling of endoplasmic reticulum, ribosome detachment, accumulation of lipid, dilatation of bile canaliculi and intercellular space, neutrophil infiltration, activation of Kupffer's cells and degeneration of hepatocytic microvilli. Futhermore , thermal injury decreased not only the protein concentration in plasma but also the number of intravascular leukocytes, that indicates induction of edema formation with protein exudation and inflammation by neutrophil infiltration into the internal organs. However allopurinol injection after burn inhibited post burn ultrastructural changes. These data suggest that acute dermal scald burn injury leads to liver damage, that is related to elevation of xanthine oxidase activity in serum. Xanthine oxidase may be a key role in the pathogenesis of liver damage induced by skin burn.

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Effect of Acanthopanax senticosus Extracts on Blood Sugar and Serum Lipid Profiles of Streptozotocin-Induced Diabetic Rats (Streptozotocin으로 유발한 당뇨 흰쥐의 혈당 및 혈청 지질함량에 미치는 가시오가피 추출물의 영향)

  • Kim Soon-Dong;Lee Sang-Il;Shin Kyung-Ok
    • Journal of the East Asian Society of Dietary Life
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    • v.15 no.5
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    • pp.549-557
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    • 2005
  • Effects of Acanthopanax senticosus extract (AS) on blood sugar content and serum lipid profiles of streptozotocin-induced diabetic rats were investigated. Experimental groups were classified into four groups, that is, normal control (NC) group, diabetic mellitus (DM) group, AS-fed group and DMAS-fed group. The AS group showed lower feed efficiency than the NC group, but the efficiency of DMAS group was higher than DM group. DMAS group showed the decreased water intake and urine by $45.5\%$ and $23.7\%$ respectively, compared with DM group. Compared with DM group, DMAS group decreased blood sugar by $46.9\%$ and triglyceride by $17.8\%$, total cholesterol by $10.0\%$ and LDL cholesterol by $22.0\%$ in serum, but increased serum HDL cholesterol by $14.4\%$ The relative percentage of liver or kidney per body weight, and the serum ALT activity in DMAS group were lower than those of DM group. There were no significant differences in hepatic glutathione(GSH) contents and total xanthine oxidase(XOD) activities among experimental groups. The hepatic lipid peroxide(LPO) content in DMAS group decreased by $54.6\%$ compared with that in DM group. The XOD (O type) and the ratio of O type to total type of both STZ-treated groups (DM and DMAS) were higher than those of NC group, but less conversion of D to O type was observed in DMAS group than in DM group. There was no significant difference in GST activity between NC and AS, but STZ-treated groups showed lower glutathione S-transferase(GST) activity than NC. In conclusion, it seems that AS reduces blood sugar by inhibiting the activity of xanthine oxidase type O as an oxygen-free radical generating system which induces the tissue damage. Antidiabetic effect of AS may regulate diabetes-induced high lipid profiles in blood.

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