There is much evidence showing that a major cause of diseases in humans, such as iron sulfate poisoning and cancer from ultraviolet radiation, is free radicals present within the body.Iron is known to be very important in the oxidation of these free radicals in biological systems based on the fact that it acts as a catalyst in these reactions [1].A well known free radical, amongst others, is the hydroxyl radical which can cause oxidative damage to DNA.Nearly a century ago, it was reported that hydrogen peroxide was a strong oxidant when in the presence of ferrous ion.This combination, along with ferrous salt, is the Fenton’s reagent.Almost 40 years later, it was found that from the Fenton reagent reaction, the hydroxyl free radical is formed[2].This reaction is called the Fenton Reaction:
FeII+ H2O2→FeIII+HO- + HO. A second type of reaction can also occur producing the hydroxyl radical is known as the Haber-Weiss Reaction which is catalyzed by iron[1]:
O2.- + H2O2 → O2 + OH- + HO. These highly reactive species are now believed to be directly involved in oxygen toxicity, which leads to many clinical problems [2]. Many studies are involved in the production of hydroxyl radicals from organelles, cells, and possibly organs.Radicals with oxygen centers are of great interest, but their short lifetimes make these radicals very difficult to monitor [3].In addition, these free radicals are often in very low concentrations in biological system, which is too low to be detection by ESR [4].Due to these reasons, the involvement of free radicals in these systems has lacked direct evidence.To overcome these difficulties, the method of spin trapping is employed [5]. Spin trapping appears, from many studies, to be the only approach towards detecting free radicals, such as hydroxyl radicals, which are short lived and in low concentrations biological systems [4]
Spin Trapping was first used in 1969 and has gained acceptance over the years from studies involving the highly reactive, short lived free radicals.This technique has now been used to detect these species in organs and animals [6]. In general, spin trapping involves a nitrone or nitroso compound reacting with an unstable, targeted free radical to form a more stable free radical.This stabilization makes it more detectable and distinguishable by EPR spectroscopy [6].The nitrone and nitroso compounds are known as spin traps.Another common spin trap for the hydroxyl radical is DMPO (5,5-dimethyl-1-1-pyrroline N- oxide) [3].These spin traps form an accumulation of stable radicals called a spin adduct.The spin adducts accumulate to a high enough concentration that the radicals can be effeciantly detected by EPR/ESR spectroscopy[4]. EPR (electron paramagnetic resonance) detects the free radicals in biological systems.It is the process of absorption of microwave radiation by molecules with one or more unpaired electron spins with the absorption being shown in a spectrum. The yield of each spin adduct is determined by the height of the peak in the spectrum [1].
Figure 1.
Figure 1 displays two EPR spectra after spin trapping was done to detect the presence of free radicals in fungal media that had been incubated with asbestos.By comparing each spectrum, the presence of free radicals in shown as prominent peaks and their absence is a lack of peaks [7].
Works Cited
[1] Qian, S.Y. and G.R. Buettner.1999. Iron and Dioxygen Chemistry is an Important Route To Initiation of Biological Free Radical Oxidations: An Electron Paramagnetic Resonance Spin Trapping Study. Free Radical Biology & Medicine 26: 1447-1456.
[2]Yamazaki, I., and L.H. Piette. 1991. EPR spin-trapping study on the oxidizing species formed in the Reaction of the ferrous ion with hydrogen peroxide. J.Am.Chem.Soc. 113: 7588-7593.
[3]Buettner, G.R. and Mason, R.P. 2003. Spin-Trapping Methods for Detecting Superoxide and Hydroxyl Free Radicals In Vitro and In Vivo. Critical Reviews of Oxidative Stress and Aging: Advances in Basic Science, Diagnostics and Intervention 1: 27-38.
[4]Janzen, E.G., H.J. Stronks, C.M. Dubose, J.L. Poyer, and P.B. McCay. 1985. Chemistry and Biology of Spin-Trapping Radicals Associated with Halocarbon Metabolism in Vitro and in Vivo. Environmental Health Perspectives 64: 151-170.
[5]Janzen, E.G., D.E. Nutter, E.R. Davis, B.J. Blackburn, J.L. Poyer, and P.B. McCay. 1978. On Spin Trapping Hydroxyl and Hydroperoxyl Radicals. Can. J. Chem. 56: 2237-2242.
[6]Li, A.S.W., K.B. Cummings, H.P. Roethling, G.R. Buettner, and C.F. Chignell. 1988. A Spin-Trapping Database Implemented on the IBM PC/AT. Journal of Magnetic Resonance79: 140-142.
[7]Daghino, S. F. Turci, M. Tomatis, A. Favier, S. Perotto, T. Douki, and B. Fubini. 2006. Soil Fungi Reduce The Iron Content and the DNA Damaging Effects of Asbestos Fibers. Environmental Science and Technology 2006: 5793-5798.
FeII+ H2O2→FeIII+HO- + HO.
A second type of reaction can also occur producing the hydroxyl radical is known as the Haber-Weiss Reaction which is catalyzed by iron[1]:
O2.- + H2O2 → O2 + OH- + HO.
These highly reactive species are now believed to be directly involved in oxygen toxicity, which leads to many clinical problems [2]. Many studies are involved in the production of hydroxyl radicals from organelles, cells, and possibly organs. Radicals with oxygen centers are of great interest, but their short lifetimes make these radicals very difficult to monitor [3]. In addition, these free radicals are often in very low concentrations in biological system, which is too low to be detection by ESR [4]. Due to these reasons, the involvement of free radicals in these systems has lacked direct evidence. To overcome these difficulties, the method of spin trapping is employed [5]. Spin trapping appears, from many studies, to be the only approach towards detecting free radicals, such as hydroxyl radicals, which are short lived and in low concentrations biological systems [4]
Spin Trapping was first used in 1969 and has gained acceptance over the years from studies involving the highly reactive, short lived free radicals. This technique has now been used to detect these species in organs and animals [6]. In general, spin trapping involves a nitrone or nitroso compound reacting with an unstable, targeted free radical to form a more stable free radical. This stabilization makes it more detectable and distinguishable by EPR spectroscopy [6]. The nitrone and nitroso compounds are known as spin traps. Another common spin trap for the hydroxyl radical is DMPO (5,5-dimethyl-1-1-pyrroline N- oxide) [3]. These spin traps form an accumulation of stable radicals called a spin adduct. The spin adducts accumulate to a high enough concentration that the radicals can be effeciantly detected by EPR/ESR spectroscopy[4]. EPR (electron paramagnetic resonance) detects the free radicals in biological systems. It is the process of absorption of microwave radiation by molecules with one or more unpaired electron spins with the absorption being shown in a spectrum. The yield of each spin adduct is determined by the height of the peak in the spectrum [1].
Figure 1.
Figure 1 displays two EPR spectra after spin trapping was done to detect the presence of free radicals in fungal media that had been incubated with asbestos. By comparing each spectrum, the presence of free radicals in shown as prominent peaks and their absence is a lack of peaks [7].
Works Cited
[1] Qian, S.Y. and G.R. Buettner.1999. Iron and Dioxygen Chemistry is an Important Route To Initiation of
Biological Free Radical Oxidations: An Electron Paramagnetic Resonance Spin Trapping Study.
Free Radical Biology & Medicine 26: 1447-1456.
[2]Yamazaki, I., and L.H. Piette. 1991. EPR spin-trapping study on the oxidizing species formed in the
Reaction of the ferrous ion with hydrogen peroxide. J.Am.Chem.Soc. 113: 7588-7593.
[3]Buettner, G.R. and Mason, R.P. 2003. Spin-Trapping Methods for Detecting Superoxide and Hydroxyl
Free Radicals In Vitro and In Vivo. Critical Reviews of Oxidative Stress and Aging: Advances in
Basic Science, Diagnostics and Intervention 1: 27-38.
[4]Janzen, E.G., H.J. Stronks, C.M. Dubose, J.L. Poyer, and P.B. McCay. 1985. Chemistry and Biology of
Spin-Trapping Radicals Associated with Halocarbon Metabolism in Vitro and in Vivo.
Environmental Health Perspectives 64: 151-170.
[5]Janzen, E.G., D.E. Nutter, E.R. Davis, B.J. Blackburn, J.L. Poyer, and P.B. McCay. 1978. On Spin Trapping
Hydroxyl and Hydroperoxyl Radicals. Can. J. Chem. 56: 2237-2242.
[6]Li, A.S.W., K.B. Cummings, H.P. Roethling, G.R. Buettner, and C.F. Chignell. 1988. A Spin-Trapping
Database Implemented on the IBM PC/AT. Journal of Magnetic Resonance79: 140-142.
[7]Daghino, S. F. Turci, M. Tomatis, A. Favier, S. Perotto, T. Douki, and B. Fubini. 2006. Soil Fungi Reduce
The Iron Content and the DNA Damaging Effects of Asbestos Fibers. Environmental Science and
Technology 2006: 5793-5798.