John M.C. Gutteridge

Biologically relevant metal ion-dependent hydroxyl radical generation. An update

Transition metal ions, especially iron, appear to be important mediators of oxidative damage in vivo. Iron(II) reacts with H2O2 to give more‐reactive radicals. On the basis of ESR spin‐trapping data with DMPO, supported by aromatic hydroxylation studies and patterns of DNA base modification, it is concluded that hydroxyl radical (OH•) is likely to be the major damaging species formed in Fenton Systems under biologically‐relevant conditions (which include iron concentrations no higher than the micromolar range). Although reactive oxo‐iron species (such as ferryl and perferryl) may also be important, chemical evidence for their formation and identity in biologically relevant Fenton systems is currently lacking. Studies at alkaline pH values show that iron(IV) and iron(V) species are highly oxidizing under those reaction conditions, with a pattern of reactivity different from that of OH•.

Biological origin of free radicals, and mechanisms of antioxidant protection.

Reduced intermediates of molecular oxygen, such as superoxide and hydrogen peroxide, are ubiquitous inorganic products of normal aerobic metabolism. Certain cells, such as phagocytes, have evolved to use superoxide and hydrogen peroxide for purposeful chemistry beneficial to the host, but most cells require antioxidant protection against excessive production of these intermediates. Superoxide and hydrogen peroxide are themselves poorly reactive in aqueous solution, and unable to directly damage DNA, lipids and proteins. Excessive generation, however, of superoxide and hydrogen peroxide invariably accompanies molecular damage. Substantial evidence suggests that conversion of these poorly reactive intermediates of oxygen to highly reactive forms requires the participation of transition metal ions, particularly iron. Iron ions react with hydrogen peroxide (Fenton chemistry) to generate hydroxyl radicals that can damage all organic molecules.

Peroxynitrite releases copper from caeruloplasmin: implications for atherosclerosis

Peroxynitrite may be formed in the vasculature by the reaction of puperoxide with nitric oxide. When the blue copper‐containing protein, caeruloplasmin, is incubated with peroxynitrite, copper is released, and ferroxidase activity and the blue colouration are lost. When plasma from normal subjects is incubated with peroxynitrite, the oxidant reacts with numerous plasma constituents but is still able to release copper from caeruloplasmin. As the ferroxidase activity of caeruloplasmin is lost in plasma in the presence of peroxynitrite, a second ferroxidase activity associated with peroxidised lipids, and not inhibited by azide, is formed.

Hydroxyl Radicals, Iron, Oxidative Stress, and Neurodegenerationa

One hundred years ago H. J. H. Fenton published his seminal paper “Oxidation of Tartaric Acid in presence of Iron”:’ Ferrous sulphate, hydrogen peroxide, tartaric acid, and sodium hydroxide, when mixed, yield a beautiful violet colour.z The reaction was shown to be driven by the mixing of a ferrous salt (Fez+) with hydrogen peroxide (HzO,), and the oxidation product of tartaric acid was later identified as a hydroxylacetaldehyde dimer. Although Fenton suggested that the iron was acting catalytically, the chemistry involved was not addressed until Haber and Weiss in the 1930s’ proposed that a ferrous salt reacting with H202 could yield hydroxyl radicals (.OH)

Ferrous Ions Detected in Iron-Overloaded Cord Blood Plasma From Preterm and Term Babies: Implications for Oxidative Stress

Redox active iron chelatable to bleomycin is often present in the plasma of cord blood samples taken from preterm and term babies. The low caeruloplasmin and high ascorbate levels in plasma at birth may allow this iron to exist in the reduced ferrous state. In support of this postulate thirteen cord blood samples showing the presence of low molecular mass iron were able to degrade DNA in the presence of bleomycin and plasma.

Hydroxyl radical formation from the auto-reduction of a ferric citrate complex

When a ferric citrate complex is prepared from citric acid and ferric chloride, and the pH value left unchanged, a reduction of the iron moiety takes place. Within several hours a substantial yield of ferrous ions can be detected in the solution. When placed in a phosphate buffer pH 7.0 with a suitable detector molecule, oxidative damage to the detector molecule can be observed. Thus, deoxyribose is degraded with the release of thiobarbituric acid-reactive material and benzoate is hydroxylated to form fluorescent dihydroxy products. Damage can be prevented by scavengers of the hydroxyl radical such as mannitol, formate the thiourea, by catalase and by the protein caeruloplasmin, suggesting that Fenton chemistry occurs leading to the formation of hydroxyl radicals.

Blood cardioplegia increases plasma iron overload and thiol levels during cardiopulmonary bypass

BACKGROUNDnCardiopulmonary bypass and crossclamping of the ascending aorta introduce two well-characterized phases of oxidative stress, namely, the extracorporeal circulation of blood and the reoxygenation of ischemic tissue. A feature of both forms of stress is the release of reactive and damaging oxygen species.nnnMETHODSnForty-seven patients undergoing aortic valve replacement received either cold crystalloid, cold blood, or warm blood cardioplegia. Plasma thiol levels were measured in all groups before and during bypass. All cardiopulmonary bypass patients had, before going onto bypass, low plasma thiol levels (3.80 +/- 0.22 nmol/mg protein) compared with normal healthy controls (5.48 +/- 0.14 nmol/mg protein).nnnRESULTSnThiol values remained low throughout bypass in patients receiving cold crystalloid cardioplegia, but rose in patients receiving cold blood cardioplegia, and rose even more in patients receiving warm blood cardioplegia to reach normal plasma values. During cardiopulmonary bypass it has previously been reported that plasma transferrin can become fully saturated with iron and cause transient iron overload. Two patients (13%) receiving cold crystalloid cardioplegia went into plasma iron overload, whereas 18% receiving cold blood and 27% receiving warm blood cardioplegia showed plasma iron overload.nnnCONCLUSIONSnWe suggest that blood cardioplegia provides an additional source of thiols as well as a source of reactive iron. However, the reactive iron and thiol-containing molecules have the potential to interact and exacerbate oxidative stress, already a feature of bypass. Control of reactive iron by chelation may be strongly indicated when blood cardioplegia is used.

Sequential Oxidative Damage, and Changes in Iron-Binding and Iron-Oxidising Plasma Antioxidants During Cardiopulmonary Bypass Surgery

Cardiopulmonary bypass patients undergoing heart valve replacement surgery appear to be under oxidative stress, when compared with normal healthy controls, by showing increased levels of protein and lipid damage. During bypass surgery two further episodes of oxidative stress occur. The first is seen when patients are placed on extracorporeal blood circulation and oxygenation which results in a rise in lipid peroxides and thiobarbituric acid-reactive substances. The second phase of oxidative stress occurs during reperfusion of the myocardium following removal of the aortic cross clamp. Coincident with evidence of increased oxidative damage to lipids during these latter phases of oxidative stress were decreases in plasma iron-binding and iron-oxidising antioxidant activities.

4-Hydroxy-2-Nonenal Levels Increase in the Plasma of Patients with Adult Respiratory Distress Syndrome as Linoleic Acid Appears to Fall

Gas chromatograph-mass spectrometry has been applied to the analysis of plasma linoleic acid and one of its oxidation products, 4-hydroxy-2-nonenal (HNE), in adult patients with the acute respiratory distress syndrome (ARDS). Peak areas of total ion chromatograms showed there to be negative correlations between loss of linoleic acid and formation of HNE (measured by selective ion monitoring) in 7 out 10 patients studied. When HNE was quantitated by selective ion monitoring, with reference to a pure standard of HNE and an internal standard of nonanoic acid, ARDS patients showed significantly increased levels of HNE (0.412 +/- 0.023 nmol/ml) compared with normal healthy controls (0.205 +/- 0.018 nmol/ml).

DNA base damage by β-lactam, tetracycline, bacitracin and rifamycin antibacterial antibiotics

Several antibacterial antibiotics have been shown to participate with transition metal ions in chemical reactions leading to the formation of reactive oxygen species. An important host defence mechanism for dealing with invading bacteria involves the production of reactive oxygen species, such as superoxide, hydrogen peroxide and hypochlorous acid, by phagocytic cells. The production of reactive oxygens by redox cycling antibacterial antibiotics has led us to suggest that a phagomimetic contribution may also be made in vivo. Here we show that four structurally different antibacterial antibiotics, in the presence of added copper salt, bring about oxidative modification to bases in DNA detected using gas chromatography-mass spectrometry. The drug most damaging to DNA was rifamycin SV which was more active than a reference mixture of hydrogen peroxide and ascorbic acid.