Deoxyribose degradation catalyzed by Fe(III)-EDTA: kinetic aspects and potential usefulness for submicromolar iron measurements.
Iron ions play a central role in .OH radicals formation and induction of oxidative stress in living organisms. Iron-catalyzed .OH radical formation degrades deoxyribose to thiobarbituric acid reactive substances (TBA-RS). This paper analyzes kinetic properties of the Fe(III)-EDTA-catalyzed deoxyribose degradation in the presence of ascorbate. The yield of TBA-RS formation in the presence of EDTA was 4-fold higher than in its absence, contrasting with results reported elsewhere, Cu(II)-EDTA and Fe(III)-citrate were unable to catalyze deoxyribose degradation. The dependence on deoxyribose concentration was fitted to a Lineweaver Burk-like plot and it was calculated that approximately 4.5 mM deoxyribose scavenged half of the .OH radicals formed. The data for Fe(III)-EDTA concentration dependence could also be fitted to a rectangular hyperbolic function. This function was linear up to 1 microM added FeCl3 and this property could be utilized as an assay for the estimation of submicromolar iron concentrations. Submicromolar concentrations of iron could induce measurable yields of TBA-RS. Differences of as little as 0.1 microM Fe(III)-EDTA could be reproducibly detected under optimum experimental conditions, above a consistent background absorbance that was equivalent to 0.35 +/- 0.05 microM Fe(III)-EDTA and represented contaminating iron in the reactants that could not be removed with Chelex-100. The low method determination limit makes the deoxyribose degradation reaction potentially useful as a new, highly sensitive and cost effective assay for iron quantification.
Institute of Biochemistry, Carleton University, Ottawa, Canada.
M
M
Hermes-Lima
E M
EM
Wang
H M
HM
Schulman
K B
KB
Storey
P
P
Ponka
D001205:Ascorbic Acid / Q000378:metabolism; D002374:Catalase / Q000494:pharmacology; D002384:Catalysis; D003855:Deoxyribose / Q000378:metabolism*; D004492:Edetic Acid / Q000494:pharmacology*; D005290:Ferric Compounds / Q000494:pharmacology*; D006863:Hydrogen-Ion Concentration; D017665:Hydroxyl Radical / Q000378:metabolism; D007501:Iron / Q000032:analysis*; D007700:Kinetics; D015203:Reproducibility of Results; D013482:Superoxide Dismutase / Q000494:pharmacology; D017392:Thiobarbituric Acid Reactive Substances / Q000032:analysis*
D005290:Ferric Compounds; D017392:Thiobarbituric Acid Reactive Substances; D017665:Hydroxyl Radical; D003855:Deoxyribose; D004492:Edetic Acid; D007501:Iron; D002374:Catalase; D013482:Superoxide Dismutase; C019179:Fe(III)-EDTA; D001205:Ascorbic Acid