FREE OXYGEN AND THIYL RADICALS
Several reactive oxygen species (ROS) and one thiyl radical (RS.) are known. Among them, the most frequently studied are given below.
1 - Superoxide radical (O2.- )
This ROS is formed when oxygen takes up one electron and as leaks
in the mitochondrial electron transport but its formation is easily increased when
exogenous components (redox cycling compounds) are present. Its first production site is
the internal mitochondrial membrane (NADH ubiquinone reductase and ubiquinone cytochrome C
reductase). This species is reduced and forms hydrogen peroxide (H2O2).
The production of superoxide radicals at the membrane level (NADPH oxidase) is initiated
in specialized cells (oxidative burst) with phagocytic functions (macrophages) and
contributes to their bactericid action. The flavin cytosolic enzyme xanthine oxidase found
in quite all tissues and in milkfat globules generates superoxide radicals from
hypoxanthine and oxygen and is supposed to be at the origin of vascular pathologies.
While superoxide radical can be directly toxic (Fridovich I, Arch Biochem
Biophys 1986, 247, 1), it has a limited reactivity with lipids, raising
questions about its true toxicity. Thus, its action is frequently considered to
result from a secondary production of the far-more reactive .OH
species by the iron-catalyzed Haber-Weiss reaction. As that production may be
limited in vivo, it was proposed that nitric oxide reacting with O2.-
generates secondary cytotoxic species (peroxinitrite
anion).
2 - Hydrogen peroxide (H2O2)
Hydrogen peroxide is mainly produced by enzymatic reactions. These
enzymes are located in microsomes, peroxysomes and mitochondria. Even in normoxia
conditions, the hydrogen peroxide production in relatively important and leads to a
constant cellular concentration between 10-9 and 10-7 M.
In plant and animal cells, superoxide dismutase is able to produce H2O2
by dismutation of O2.- , thus contributing to the
lowering of oxidative reactions. The natural combination of dismutase and catalase
contributes to remove H2O2 and thus has a true cellular antioxidant
activity. H2O2 is also able to diffuse easily through cellular
membranes.
3 - Hydroxyl radical (.OH)
In the presence of Fe2+, H2O2 produces the very active species .OH by the Fenton reaction (described in 1894):
Fe2+ + H2O2 ----> Fe3+ + .OH + OH-
This iron-catalyzed decomposition of oxygen peroxide is considered
the most prevalent reaction in biological systems and the source of various deleterious
lipid peroxidation products.
It must be noticed that an important part of hydroxyl radicals is also produced
(with .NO2)
by the decay of peroxinitrite or peroxynitrous acid
Another reaction involving myeloperoxidase and Cl- ions
represent an important .OH production process in neutrophils during
phagocytosis.
Nitric oxide is produced in various types of cells and
is well studied
in vascular endothelium. While this species is not too reactive (poorly oxidizing
function), even antioxidant under physiological concentrations (up to
100 nM), it reacts rapidly with oxygen to yield nitrogen dioxide (.NO2)
which in turn may react with .NO to
yield nitrogen trioxide (N2O3).
The rapid reaction of O2.-,
produced in different pathological states, with .NO gives the extremely reactive peroxinitrite
(ONOO-) which mediates oxidation,
nitrosation, and nitration reactions. In alkaline solutions, ONOO-
is stable but decays rapidly once protonated into peroxynitrous acid. Its decomposition forms .OH
and nitrogen dioxide (NO2.),
radicals important in the formation of acid rain. The high rate
and broad distribution of .NO
production, combined with its facile reactions with oxygen radical species,
assure that it plays a central role in regulating oxidant reactions.
Multiple mechanisms account for the nitration of lipids by .NO-derived
species (O’Donnell V B et al., Circ Res 2001, 88, 12). In acidic conditions, protonation of NO2- to HNO2
can mediate the nitration of polyunsaturated fatty acids and lipid
hydroperoxides giving nitrated lipid products whose structure and function are incompletely characterized.
5 - Singlet oxygen (1O2)
This chemical form of oxygen is not a true radical but is reported
to be an important ROS in reactions related to ultraviolet exposition (UVA, 320-400 nm).
Its toxicity is reinforced when appropriate photoexcitable compounds (sensitizers) are
present with molecular oxygen. Several natural sensitizers are known to catalyzed
oxidative reactions such as tetrapyrroles (bilirubin), flavins, chlorophyll, hemoproteins
and reduced pyridine nucleotides (NADH). Some of these sensitizers are also found in foods
and cosmetics. Some others are used for therapeutic purposes (anticancer treatments) and
are sensitive to visible light. The presence of metals contributes to increase the
production of singlet oxygen, as well as anion superoxide, and thus accelerates the
oxidation of unsaturated lipids generating hydroperoxides.
It has been suggested that singlet O2 may be formed during the degradation of
lipid peroxides and thus may cause the production of other peroxide molecules. This
singlet O2 formation may account for the chemiluminescence observed during
lipid peroxidation.
6 - Ozone (O3)
This natural compound present in the higher atmosphere and in the
lower atmosphere of our polluted cities is a major pollutant formed by photochemical
reactions between hydrocarbons and nitrogen oxides. Ozone is not a free radical but, as
singlet oxygen, may produce them, stimulates lipid peroxidation and thus induces damages
at the lipid and protein levels in vivo mainly in airways.
The exact chemistry of ozone-mediated stimulation of peroxidation is not entirely
known. Ozone may add on across a double bond and decomposes to form a free radical. The
proposed mechanism is given below.
Aliphatic thiols (RSH)
are contained in living organisms in high concentrations. Typical levels of
intracellular of glutathione are about of 5 to 10 mM. Furthermore, the level of
protein SH may exceed that of GSH. The thiolate specie RS– is one of the most
reactive functional groups found in proteins. It can react as a nucleophile and
attack a disulfide bond. In the absence of
oxygen, a thiyl radical was shown to induced cis/trans-isomerization of linoleic
acid and led to several isomers. (Schwinn J, Int J
Radiat Biol 1998, 74, 359). A review of the effects of thiyl radicals on
lipid structures and metabolisms may be consulted (Ferreri
C et al., Cell Mol Life Sci 2005, 62, 834).
Thiol compounds (RSH) are frequently oxidized in the presence of
iron or copper ions:
RSH + Cu2+ ----> RS. + Cu+ + H+
These thiyl radicals have strong reactivity in combining with O2 :
RS. + O2 ---> RSO2.
Furthermore, they are able to oxidize NADH into NAD. , ascorbic acid and to generate various free radicals (.OH and O2.-). These thiyl radicals may also be formed by homolytic fission of disulfide bonds in proteins.
8 - Carbon-centered radicals
The formation of these reactive free radical is observed in cells treated with CCl4 . The action of the cytochrome P450 system generates the trichloromethyl radical (.CCl3) which is able to react with oxygen to give several peroxyl radicals (i.e. .O2CCl3).
