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A mixture of xanthine or hypoxanthine and xanthine oxidase generates the superoxide radical, O2 • , and H2O2. In the presence of iron salts, O2 • and H2O2 can interact to produce the hydroxyl radical, OH·. Superoxide-dependent formation of OH· can be measured by its ability to hydroxylate salicylate as followed by an improved colorimetric assay described in this paper. A more accurate analysis of OH· can be obtained using its ability to hydroxylate phenol, the hydroxylated products being separated and measured after derivatization using gas-liquid chromatography and electron-capture detection. The derivatization and separation techniques are described.
F2-isoprostanes (F2-IPs) are formed by the free radicalcatalysed oxidation of arachidonic acid. The measurement of F2-IPs, especially 8-epi-PGF2α, is recognised as a reliable marker of lipid peroxidation and is currently used as a sensitive index of oxidative stress in vivo. The majority of 8-epi-PGF2α present in the circulation occurs in association with lipoproteins which are synthesised in the liver. Since lipoproteins are derived from dietary fatty acids and triglycerides, it is possible that 8-epi-PGF2α generated in polyunsaturated fatty acid-rich food (during initial processing/packaging or during meal preparation) may become incorporated within these lipoproteins during synthesis. In view of the growing use of 8-epi-PGF2α as a marker of lipid peroxidation in vivo in nutritional or clinical studies, it is therefore important to investigate the possibility that the circulating levels measured could be confounded by the presence of 8-epi-PGF2α in food. In this study we evaluated the levels of 8-epi-PGF2α present in several popular fastfoods, using a combination of solid phase extraction and gas chromatography-mass spectrometry. Fastfoods were selected to represent meals prepared from vegetable-, chicken-, fish- and meat-derived ingredients. Total (free + esterified) 8-epi-PGF2α levels ranged from 0.09 to 0.73 pmol/g (122–644 pmol/mmol arachidonic acid), with the highest levels present in beef-derived meals. Further investigation of hamburgers and cheeseburgers revealed 8-epi-PGF2α levels of 1.83 ± 0.24 and 0.84 ± 0.03 nmol/mmol arachidonic acid, respectively. Lower concentrations of vitamin E were found in the hamburgers. The postprandial contribution to plasma 8-epi-PGF2α levels following ingestion of 100 g portions of these fast-foods would therefore be expected to be no greater than the low picomole range, and would be unlikely to influence the normal endogenous levels of 8-epi-PGF2α, and those produced during oxidative stress.
Both ferrioxamine and desferrioxamine react with the hydroxyl radical with a second order rate constant equal to 1.3 × 1010 M−1 s−1. Conditions for the use of desferrioxamine as a probe for the role of iron salts in the formation of hydroxyl radicals in biochemical systems are discussed.
Chloroplasts isolated from leaves of spinach‐beet ( Beta vulgaris L. ssp. vulgaris ) do not catalyse the hydroxylation of p ‐coumaric acid in the dark unless a reductant (such as ascorbate, NADH or NADPH) is added. Superoxide dismutase has no effect on this reaction. Illuminated chloroplasts catalyse the hydroxylation in the absence of added reductant. This reaction is completely inhibited by superoxide dismutase, but catalase has little effect. Both hydroxylation in the light and hydroxylation in the dark in the presence of reductants are inhibited by diethyldithiocarbamate, EDTA, cyanide and 2‐mercaptoethanol. It is proposed that O 2 .− generated by illuminated chloroplasts is involved in the provision of a reductant to the enzyme phenolase.
THE mechanism of nigral cell death in Parkinson's disease (PD) remains unknown, but it is increasingly proposed that free radical reactions are important in the disease pathology. One of the most striking features of PD is an approximate 40% decrease in the levels of reduced glutathione (GSH) which occurs early in the development of the disease. We describe a possible mechanism of GSH depletion which results from the reaction of L-DOPA and dopamine with the superoxide free radical (O2-) and leads to a very rapid loss of GSH.
Conference Article| February 01 1976 Production of the Superoxide Radical by Horseradish Peroxidase BARRY HALLIWELL; BARRY HALLIWELL 1Department of Biochemistry, King's College London, Strand, London WC2R2LS, U.K. Search for other works by this author on: This Site PubMed Google Scholar SUNIL AHLUWALIA SUNIL AHLUWALIA 2Department of Biochemistry, King's College London, Strand, London WC2R2LS, U.K. Search for other works by this author on: This Site PubMed Google Scholar Biochem Soc Trans (1976) 4 (1): 73–74. https://doi.org/10.1042/bst0040073 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 BARRY HALLIWELL, SUNIL AHLUWALIA; Production of the Superoxide Radical by Horseradish Peroxidase. Biochem Soc Trans 1 February 1976; 4 (1): 73–74. doi: https://doi.org/10.1042/bst0040073 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 Society Transactions Search Advanced Search This content is only available as a PDF. © 1976 Biochemical Society1976 Article PDF first page preview Close Modal You do not currently have access to this content.
A bstract : In the late 1950's free radicals and antioxidants were almost unheard of in the clinical and biological sciences but chemists had known about them for years in the context of radiation, polymer and combustion technology. Daniel Gilbert, Rebeca Gerschman and their colleagues related the toxic effects of elevated oxygen levels on aerobes to those of ionizing radiation, and proposed that oxygen toxicity is due to free radical formation, in a pioneering paper in 1956. Biochemistry owes much of its early expansion to the development and application of chromatographic and electrophoretic techniques, especially as applied to the study of proteins. Thus, superoxide dismutase (SOD) enzymes (MnSOD, CuZnSOD, FeSOD) were quickly identified. By the 1980's Molecular Biology had evolved from within biochemistry and microbiology to become a dominant new discipline, with DNA sequencing, recombinant DNA technology, cloning, and the development of PCR representing milestones in its advance. As a biological tool to explore reaction mechanisms, SOD was a unique and valuable asset. Its ability to inhibit radical reactions leading to oxidative damage in vitro often turned out to be due to its ability to prevent reduction of iron ions by superoxide. Nitric oxide (NO·) provided the next clue as to how SOD might be playing a critical biological role. Although NO· is sluggish in its reactions with most biomolecules it is astoundingly reactive with free radicals, including superoxide. Overall, this high reactivity of NO· with radicals may be beneficial in vivo , e.g. by scavenging peroxyl radicals and inhibiting lipid peroxidation. If reactive oxygen species are intimately involved with the redox regulation of cell functions, as seems likely from current evidence, it may be easier to understand why attempts to change antioxidant balance in aging experiments have failed. The cell will adapt to maintain its redox balance. Indeed, transgenic animals over‐expressing antioxidants show some abnormalities of function. There must therefore be a highly complex interrelationship between dietary, constitutive, and inducible antioxidants within the body, under genetic control. The challenge for the new century is to be able to understand these relationships, and how to manipulate them to our advantage to prevent and treat disease.
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Potentially damaging species (reactive oxygen, nitrogen and chlorine species) arise as by-products of metabolism and as physiological mediators and signalling molecules. Levels of these species are controlled by the antioxidant defence system. Several components of this system are micronutrients (e.g. vitamins C and E) or are dependent upon dietary micronutrients (e.g. CuZn and Mn superoxide dismutase). The antioxidant defences act as a coordinated system where deficiencies in one component may affect the efficiency of the others. Oxidative stress may be an important factor in infection if micronutrients are deficient.
