Submitted by Dr. Tamer Fouad, M.D.
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Free radicals are a chemical species that possess an
unpaired electron in the outer shell of the molecule. |
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Measurement of free radicals
This increasing interest in the role of
free radicals in the pathogenesis of disease has led to an increased need for
techniques to measure free radicals and their reactions in vivo and, most
importantly, in the clinical situation.
Several problems arise when considering
measurement of free radicals (Cheesman
and Slater, 1993): First the ultra-short half-life of these radicals
(usually measured in microseconds). Second any free radicals produced in vivo
react at or close to their source of formation. Hence, the methods devised to
quantitate free radicals are more or less indirect (Slater,
1984; Pryor and Godber, 1991). A third problem is that many of these end
products are in themselves reactive although to a lesser degree. Last but not
least, the only samples that are readily available for clinical practice are
blood, urine and expired breath. It is completely impossible for a free radical
that is produced in an internal tissue, having a half-life measured in
microseconds to diffuse to the blood. In order to overcome these problems the
following methods are used:
Electron spin resonance and radical trapping:
The only analytical technique that
directly measures free radicals is electron spin resonance spectrometry.
However, it is relatively insensitive and requires steady-state concentrations
of free radicals in the micromolecular range; thus it is very limited for use in
vivo. Spin trapping involves the addition of a compound known as a spin trap,
that reacts rapidly with free radicals to form radical-adducts that are very
much more stable and longer lived than the original species (Cheesman
and Slater, 1993).
An alternative technique for trapping
free radicals produced by cells, organelles and perfused organs is by measuring
the non-radical products produced when free radicals attack aromatic compounds.
These substances are not produced normally hence their presence is evidence of
hydroxyl radical production (Grootveld
and Halliwell, 1986). These methods like ESR spin trapping can only produce
semi-quantitative data.
Measurement of DNA damage:
Free radical species such as ?OH can
produce irreversible DNA modification of DNA bases; theoretically a large number
of products are possible but relatively few have been detected in biological
systems.
Among these the major products of ?OH
with thymine is thymine glycol and guanine is 8-hydroxy-guanine. These DNA
products are eliminated by repair enzymes (excision enzymes & glycolases)
and are excreted in urine either as the free base or as the nucleoside
derivatives, thymidine glycol and 8-hydroxydeoxyguanosine. The latter products
can be used as an index of radical attacks against DNA (Cathcart
et al. 1984). This method is limited by several considerations including the
obscurity of the tissue of origin of the products.
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