Publications

The deoxyribose assay: an assay both for ‘free’ hydroxyl radical and for site-specific hydroxyl radical production

Correspondence| August 01 1988 The deoxyribose assay: an assay both for 'free' hydroxyl radical and for site-specific hydroxyl radical production J M C Gutteridge; J M C Gutteridge Search for other works by this author on: This Site PubMed Google Scholar B Halliwell B Halliwell Search for other works by this author on: This Site PubMed Google Scholar Biochem J (1988) 253 (3): 932–933. https://doi.org/10.1042/bj2530932 Views Icon Views Article contents Figures & tables Video Audio Supplementary Data Peer Review Share Icon Share Facebook Twitter LinkedIn MailTo Cite Icon Cite Get Permissions Citation J M C Gutteridge, B Halliwell; The deoxyribose assay: an assay both for 'free' hydroxyl radical and for site-specific hydroxyl radical production. Biochem J 1 August 1988; 253 (3): 932–933. doi: https://doi.org/10.1042/bj2530932 Download citation file: Ris (Zotero) Reference Manager EasyBib Bookends Mendeley Papers EndNote RefWorks BibTex toolbar search Search Dropdown Menu toolbar search search input Search input auto suggest filter your search All ContentAll JournalsBiochemical Journal Search Advanced Search This content is only available as a PDF. © 1988 London: The Biochemical Society1988 Article PDF first page preview Close Modal You do not currently have access to this content.

Inhibition of Peroxynitrite-Mediated Tyrosine Nitration by Catechin Polyphenols

Peroxynitrite is a cytotoxic species generated by the reaction between superoxide and nitric oxide. The ability of catechins and their gallate esters to decrease peroxynitrite-induced nitration of tyrosine and to limit surface charge alteration of low density lipoprotein (LDL) was investigated. All compounds tested were found to be potent peroxynitrite scavengers preventing the nitration of tyrosine. The ability of the catechin polyphenols at 10 μM to minimise tyrosine nitration induced by peroxynitrite (500 μM) was ECG (38.1 ± 3.6%) ≈ EGCG (32.1 ± 7.5%) ≈ gallic acid (32.1 ± 1.9%) > catechin (23.9 ± 5.4%) ≈ epicatechin (22.9 ± 3.3%) ≈ EGC (19.9 ± 2.0%). Trolox (10 μM) was used as the standard for comparative purposes and was found to be less effective than the polyphenols in inhibiting tyrosine nitration (13.6 ± 2.9%). The catechin polyphenols were also found to offer protection from peroxynitrite-induced modification of critical amino acids of apolipoprotein B-100 of LDL which contribute towards its surface charge.

Allopurinol and oxypurinol are hydroxyl radical scavengers

Allopurinol is a scavenger of the highly reactive hydroxyl radical ( k 2 approx. 10 9 M −1 s −1 ). One product of attack of hydroxyl radical upon allopurinol is oxypurinol, which is a major metabolite of allopurinol. Oxypurinol is a better hydroxyl radical scavenger than is allopurinol ( k 2 approx. 4 × 10 9 M −1 s −1 ) and it also reacts with the myeloperoxidase‐derived oxidant hypochlorous acid. Hence the protective actions of allopurinol against reperfusion damage after hypoxia need not be entirely due to xanthine oxidase inhibition.

Artifacts in Cell Culture: Rapid Generation of Hydrogen Peroxide on Addition of (−)-Epigallocatechin, (−)-Epigallocatechin Gallate, (+)-Catechin, and Quercetin to Commonly Used Cell Culture Media

There is considerable current interest in the possible beneficial health effects of quercetin, catechins, epigallocatechins, epigallocatechin gallates, and related phenolic compounds found in teas, wines, and other plant products. As a result, many laboratories are studying the effects of these compounds on cells in culture. The present paper shows that addition of these compounds to commonly used cell culture media leads to generation of substantial amounts of hydrogen peroxide (H2O2). Dulbecco's modified Eagle medium gives the highest H2O2 level for all the compounds tested, with levels reaching >400 μM within 2 h for addition of 1 mM concentrations of gallic acid, epigallocatechin gallate, and epigallocatechin. Catechin and quercetin produced lower, but still significant, levels of H2O2. McCoy's 5A and RPMI 1640 media also promoted H2O2 production from the above phenolic compounds. This rapid generation of H2O2 could account for some or all of the reported effects of phenolic compounds on cells in culture.

Invited Review Free Radicals in Disease Processes: A Compilation of Cause and Consequence

(1993). Invited Review Free Radicals in Disease Processes: A Compilation of Cause and Consequence. Free Radical Research Communications: Vol. 19, No. 3, pp. 141-158.

Oxidative Damage to Human Plasma Proteins by Ozone

Exposure of human plasma to ozone produces oxidative protein damage, measured as protein carbonyl formation. Isolated human albumin or creatine phosphokinase are oxidized much faster than are total proteins. Consideration must be given to proteins as targets of oxidative injury by ozone in vivo.

Peroxynitrite mediates calcium‐dependent mitochondrial dysfunction and cell death via activation of calpains

Chondrocyte cell death is a hallmark of inflammatory and degenerative joint diseases such as rheumatoid arthritis (RA) and osteoarthritis (OA), but the molecular and cellular mechanisms involved have yet to be elucidated. Because 3-nitrotyrosine, a marker for reactive nitrogen species such as peroxynitrite, has been observed in OA and RA cartilage and has been associated with chondrocyte cell death, we investigated the mechanisms by which peroxynitrite induces cell death in human articular chondrocytes. The earliest biochemical event observed, subsequent to treatment with either peroxynitrite or the peroxynitrite generator SIN-1, was a rapid rise in intracellular calcium that lead to mitochondrial dysfunction and cell death. Although, chondrocyte death exhibited several classical hallmarks of apoptosis, including annexin V labeling, increased fraction of cells with subG1 DNA content and DNA condensation, we did not find evidence for caspase involvement either by Western blotting, fluorimetric assays, or caspase inhibition. Additionally, peroxynitrite did not inhibit cellular caspase activity. Furthermore, using other established assays of cell viability, including the MTT assay and release of lactate dehydrogenase, we found that the predominant mode of cell death involved calcium-dependent cysteine proteases, otherwise known as calpains. Our data show, for the first time, that peroxynitrite induces mitochondrial dysfunction in cells via a calcium-dependent process that leads to caspase-independent apoptosis mediated by calpains.

Protection against Doxorubicin-Induced Cardiotoxicity by Ergothioneine

Background: Anthracyclines such as doxorubicin remain a primary treatment for hematological malignancies and breast cancers. However, cardiotoxicity induced by anthracyclines, possibly leading to heart failure, severely limits their application. The pathological mechanisms of anthracycline-induced cardiac injury are believed to involve iron-overload-mediated formation of reactive oxygen species (ROS), mitochondrial dysfunction, and inflammation. The dietary thione, ergothioneine (ET), is avidly absorbed and accumulated in tissues, including the heart. Amongst other cytoprotective properties, ET was shown to scavenge ROS, decrease proinflammatory mediators, and chelate metal cations, including Fe2+, preventing them from partaking in redox activities, and may protect against mitochondrial damage and dysfunction. Plasma ET levels are also strongly correlated to a decreased risk of cardiovascular events in humans, suggesting a cardioprotective role. This evidence highlights ET's potential to counteract anthracycline cardiotoxicity. Methods and Findings: We investigated whether ET supplementation can protect against cardiac dysfunction in mice models of doxorubicin-induced cardiotoxicity and revealed that it had significant protective effects. Moreover, ET administration in a mouse breast cancer model did not exacerbate the growth of the tumor or interfere with the chemotherapeutic efficacy of doxorubicin. Conclusion: These results suggest that ET could be a viable co-therapy to alleviate the cardiotoxic effects of anthracyclines in the treatment of cancers.

1 Iron toxicity and oxygen radicals

No abstract is provided for this article.

Modulation of peroxynitrite‐ and hypochlorous acid‐induced inactivation of α₁‐antiproteinase by mercaptoethylguanidine

Peroxynitrite is a cytotoxic species that can be formed, among other mechanisms, by the rapid reaction of superoxide with nitric oxide. Peroxynitrite formation has been implicated in a wide range of neurodegenerative and chronic inflammatory diseases, as has the formation of hypochlorous acid by myeloperoxidase. There is considerable interest in the development of peroxynitrite scavengers as therapeutic agents. The thiol compound mercaptoethylguanidine has been suggested to fulfil this role since it has recently been shown to be not only a potent inhibitor of inducible nitric oxide synthase but also a scavenger of peroxynitrite. Indeed, it has been shown to be protective in some experimental models of circulatory shock and inflammation at plasma levels in the approximate range 100–300 μ M . One protein inactivated by peroxynitrite is the major inhibitor of serine proteinases in human body fluids, α 1 ‐antiproteinase. At high (250–1000 μ M ) concentrations, mercaptoethylguanidine was found to be effective in preventing peroxynitrite‐mediated tyrosine nitration and α 1 ‐AP inactivation. By contrast, lower concentrations of mercaptoethylguanidine (1–60 μ M ) enhanced the inactivation of α 1 ‐antiproteinase by peroxynitrite. At all concentrations tested (1–1000 μ M ), mercaptoethylguanidine decreased the inactivation of α 1 ‐antiproteinase by hypochlorous acid. We suggest that products of reaction of mercaptoethylguanidine with peroxynitrite or peroxynitrite‐derived products could cause damage to α 1 ‐antiproteinase, and possibly other proteins in vivo , whereas scavenging of hypochlorous acid by mercaptoethylguanidine could contribute to its anti‐inflammatory action in vivo . British Journal of Pharmacology (1999) 126 , 1646–1652; doi: 10.1038/sj.bjp.0702465

Human Fecal Water Inhibits COX-2 in Colonic HT-29 Cells: Role of Phenolic Compounds

The inducible enzyme cyclooxygenase-2 (COX-2) plays a major role in the regulation of inflammation and possibly in the development of colon cancer. The aim of the present study was to screen for COX-2 inhibitors in samples of fecal water (the aqueous phase of feces) and investigate whether phenolic compounds are responsible for any observed effects on COX-2. Volunteers (n = 20) were recruited and asked to supply a 24-h stool sample. Fecal water samples were prepared and analyzed by GC-MS for their content of phenolic compounds. These samples were also evaluated for their effects on COX-2 protein levels (Western blot) and prostaglandin (PG)E2 production in tumor necrosis-α–stimulated HT-29 cells and pure enzymatic activity in a COX-2–catalyzed prostaglandin biosynthesis in vitro assay. The major phenolic compounds identified were phenylpropionic acid, phenylacetic acid, cinnamic acid, and benzoic acid derivatives. Of 13 fecal water samples analyzed, 12 significantly decreased PGE2 production (range 5.4–39.7% inhibition, P-value < 0.05) compared with control cells and 13 of 14 samples analyzed decreased COX-2 protein levels in HT-29 cells (19–63% inhibition). Of the 20 fecal water samples, 2 also weakly inhibited enzymatic activity of purified COX-2 (22–24% inhibition). Three compounds identified in fecal water, 3-phenylpropionic acid, 3-hydroxyphenylacetic acid, and 3-(4-hydroxyphenyl)-propionic acid, decreased the protein level at 250 μmol/L (15–62% inhibition). This study shows for the first time that human fecal water contains components that can affect both the COX-2 protein level and enzymatic activity.

Cholesterol autoxidation

polyunsaturatedfatty acids which are found in the very fats that are recommended as being beneficial in reducing blood cholesterol levels.The chemistry of the lipids is explained in a simple fashion, their nomenclature and even the processes of extraction of edible oils and their purification and conversion into foods.fatty acids and British figures are similar -so it may be possible that the ingestion of polyunsaturated fatty acids intended to reduce the risk of coronary heart disease may, in fact, increase the risk of this and other disorders.

ANTIOXIDANT AND PRO-OXIDANT ACTIONS OF DIETARY COMPONENTS

Antioxidants: The Basics-what they are and how to Evaluate them

Words such as “antioxidant” and “oxidative stress” are difficult to define. The term “antioxidant” used in the literature is restricted to chain-breaking antioxidant inhibitors of lipid peroxidation. Food scientists frequently equate antioxidants to inhibitors of lipid peroxidation because they use antioxidants to prevent rancidity. Free radicals generated in vivo damage proteins, DNA, and other molecules in addition to lipids. A broader definition is: an antioxidant is any substance that when present at low concentrations compared to those of an oxidizable substrate delays or prevents oxidation of that substrate. Mechanisms of antioxidant action can include: removal of O2, scavenging reactive oxygen/nitrogen species or their inhibiting ROS/RNS formation, binding metal ions needed for catalysis of ROS generation, and up regulation of endogenous antioxidant defenses. This chapter emphasizes the importance of the source of stress and the target (“oxidizable substrate”) measured; but also mentions the cases that the definition does not include. When ROS/RNS are generated in vivo, many antioxidants come into play. The chapter discusses the antioxidants known, or proposed, to be important in aerobic organisms. There is outline on some approaches to the characterization of direct antioxidant activity and discussion on biologically relevant ROS/RNS—superoxide and hydrogen peroxide, hydroxyl radical, peroxyl radicals, lipid peroxidation, hypochlorous acid, heme proteins/peroxides, peroxynitrite, and singlet oxygen—measuring their scavenging, their reactions, some inhibitions on radical formation and other properties. Some approaches have been listed to help prove that a given compound act as an antioxidant in vivo, including two approaches for putative antioxidants. Specific assays are being developed to measure rates of oxidative damage to protein, DNA, and lipid. Steady-state and total body oxidative damage to these targets can now be approximated, providing a tool to examine the effects of “antioxidants” in vivo.

The Lively Membranes

Formation of Hydroxyl Radicals in Biological Systems. Does Myoglobin Stimulate Hydroxyl Radical Formation from Hydrogen Peroxide?

Incubation of horse-heart oxymyoglobin or metmyoglobin with excess H2O2 causes formation of myoglobin(IV), followed by haem degradation. At the time when haem degradation is observed, hydroxyl radicals (.OH) can be detected in the reaction mixture by their ability to degrade the sugar deoxyribose. Detection of hydroxyl radicals can be decreased by transferrin or by OH scavengers (mannitol, arginine, phenylalanine) but not by urea. Neither transferrin nor any of these scavengers inhibit the haem degradation. It is concluded that intact oxymyoglobin or metmyoglobin molecules do not react with H2O2 to form OH detectable by deoxyribose, but that H2O2 eventually leads to release of iron ions from the proteins. These released iron ions can react to form OH outside the protein or close to its surface. Salicylate and the iron chelator desferrioxamine stabilize myoglobin and prevent haem degradation. The biological importance of OH generated using iron ions released from myoglobin by H2O2 is discussed in relation to myocardial reoxygenation injury.

Ferroxidase and Ascorbate Oxidase Activities of Caeruloplasmin in Synovial Fluid from Rheumatoid Patients

1. The protein caeruloplasmin inhibits certain free radical reactions, in part probably due to its ability to oxidize Fe2+ into Fe3+ (ferroxidase activity). Since caeruloplasmin is present in synovial fluid from rheumatoid patients, we investigated its properties in relation to protection of the joint from damage by oxygen radicals produced by activated phagocytes. 2. The ferroxidase and ascorbate oxidase activities of serum from both normal and rheumatoid patients could be accounted for by the caeruloplasmin present, as determined immunologically. More caeruloplasmin is present in the serum of rheumatoid patients than in normal serum. 3. Synovial fluid from rheumatoid patients contains caeruloplasmin protein but its ferroxidase and ascorbate oxidase activities are abnormally low. It is suggested that ferroxidase deficiency contributes to radical damage in the rheumatoid joint.

Bleomycin‐detectable iron in serum from leukaemic patients before and after chemotherapy Therapeutic implications for treatment with oxidant‐generating drugs

Patients with acute myeloid leukaemia show elevated plasma iron and, in 2/6 cases studied, low-molecular-mass iron complexes capable of stimulating radical reactions were present in the plasma. Shortly after the onset of chemotherapy, there is a sharp rise in transferrin saturation and all patients studied showed low-molecular-mass iron in their plasma. It is proposed that such iron could interact with oxidants generated by certain drugs (e.g. adriamycin or daunorubicin) to facilitate tissue damage, and that some of the side-effects of chemotherapy might be ameliorated by careful co-administration of small doses of desferrioxamine.