Oxidative stress is believed to play a role in the pathogenesis o
f several diseases, includingdiabetes and inborn errors o
f metabolism. The types o
f oxidative damage observed in thesepathologies have been attributed to the excessive production o
f reactive intermediates relatingto the accumulation o
f toxic metabolites. The production o
f extremely oxidizing peroxynitritecan also be high in these pathologies. We study here the oxidation initiated by peroxynitriteo
f the ethyl esters o
f acetoacetate (EAA) and 2-methylacetoacetate (EMAA), metabolites thataccumulate in diabetes and isoleucinemia, respectively. Oxygen consumption studies havecon
firmed that peroxynitrite promotes the aerobic oxidation o
f EAA and EMAA in phosphatebu
ffer. These reactions were accompanied by ultraweak light emission, which probably arises
from triplet carbonyl products
formed by thermolysis o
f dioxetane intermediates. The kineticso
f oxygen uptake and chemiluminescence by EAA and EMAA was strongly a
ffected by thephosphate ion, known to catalyze carbonyl enolization and nucleophilic additions to carbonyls.The reaction pH pro
files obtained by oxygen consumption and chemiluminescence measurements indicated that the peroxynitrite anion was the initiator o
f EAA and EMAA aerobicoxidation. EPR spin-trapping studies with the spin traps 3,5-dibromo-4-nitrosobenzenesul
fonicacid and 2-methyl-2-nitrosopropane showed the intermediacy o
f methyl and a carbon-centeredradical (
f">CH
2COR) in the oxidation o
f EAA by peroxynitrite. In the case o
f EMAA, a tertiarycarbon-centered radical (
f">EMAA) and an acyl radical were detected, the latter probably resulting
from the cleavage o
f a triplet carbonyl product. Superstoichiometric
formation o
f acetate
fromboth substrates con
firmed the occurrence o
f oxygen-dependent chain reactions, here proposedto be initiated by one-electron abstraction
from the enolic
form o
f the substrates. The
freeradicals and electronically excited species generated in the oxidation o
f EAA and EMAA mayhelp shed
further light on the molecular basis o
f these diseases.