Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
10738026 | Free Radical Biology and Medicine | 2011 | 11 Pages |
Abstract
Acetoacetate (AA) and 2-methylacetoacetate (MAA) are accumulated in metabolic disorders such as diabetes and isoleucinemia. Here we examine the mechanism of AA and MAA aerobic oxidation initiated by myoglobin (Mb)/H2O2. We propose a chemiluminescent route involving a dioxetanone intermediate whose thermolysis yields triplet α-dicarbonyl species (methylglyoxal and diacetyl). The observed ultraweak chemiluminescence increased linearly on raising the concentration of either Mb (10-500 μM) or AA (10-100 mM). Oxygen uptake studies revealed that MAA is almost a 100-fold more reactive than AA. EPR spin-trapping studies with MNP/MAA revealed the intermediacy of an α-carbon-centered radical and acetyl radical. The latter radical, probably derived from triplet diacetyl, is totally suppressed by sorbate, a well-known quencher of triplet carbonyls. Furthermore, an EPR signal assignable to MNP-AA
- adduct was observed and confirmed by isotope effects. Oxygen consumption and α-dicarbonyl yield were shown to be dependent on AA or MAA concentrations (1-50 mM) and on H2O2 or tert-butOOH added to the Mb-containing reaction mixtures. That ferrylMb is involved in a peroxidase cycle acting on the substrates is suggested by the reaction pH profiles and immunospin-trapping experiments. The generation of radicals and triplet dicarbonyl products by Mb/H2O2/β-ketoacids may contribute to the adverse health effects of ketogenic unbalance.
- adduct was observed and confirmed by isotope effects. Oxygen consumption and α-dicarbonyl yield were shown to be dependent on AA or MAA concentrations (1-50 mM) and on H2O2 or tert-butOOH added to the Mb-containing reaction mixtures. That ferrylMb is involved in a peroxidase cycle acting on the substrates is suggested by the reaction pH profiles and immunospin-trapping experiments. The generation of radicals and triplet dicarbonyl products by Mb/H2O2/β-ketoacids may contribute to the adverse health effects of ketogenic unbalance.
Keywords
nuclear factor κβAdvanced glycated end productsMNPDMPO5,5′-dimethyl-1-pyrroline N-oxideacetoacetateRAGEMAAAGEsHRPGSHCATo-PhenylenediamineopdELISAEnzyme-linked immunosorbent assayElectron paramagnetic resonanceEPRtumor necrosis factor αDiacetylFree radicalsSODSuperoxide dismutaseTNF-αMethylglyoxalMyoglobintert-butyl hydroperoxideHorseradish peroxidaseCatalasereduced glutathioneKetosishigh performance liquid chromatographyHPLC
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Authors
Douglas Ganini, Marcelo Christoff, Marilyn Ehrenshaft, Maria B. Kadiiska, Ronald P. Mason, Etelvino J.H. Bechara,