Lipofuscin-like fluorophores can result from reactions between oxidized ascorbic acid and glutamine. Carbonyl-protein cross-linking may represent a common reaction in oxygen radical and glycosylation-related ageing processes

Abstract

Ascorbic acid (AsA), after being oxidized in 0.1 M phosphate (pH 7.0) buffer under the catalytic influence of adventitious iron, reacted with glutamine (Gln) derivatives with the formation of stable fluorophores showing lipofuscin-like blue (350/430 nm) fluorescence. The fluorescence was reversibly quenched by acidity and enhanced by alkaline conditions, and the fluorescence intensity was directly proportional to the Gln and AsA concentrations. Addition of H 2O 2 considerably increased the velocity of the fluorescence formation. Incubation of AsA/Gln in 0.1 M phosphate buffer at pH 5.0 gave a slower fluorophore formation as compared with incubation at pH 7.0. The iron chelators DTPA and desferrioxamine inhibited the fluorophore development by preventing the iron catalyzed AsA oxidation. This was in contrast to the effects of the chelators ADP and EDTA which did not show such preventive activity. The fluorophores produced by the AsA/Gln reaction are thought to be Schiff bases formed secondary to Maillard reactions involving oxidized AsA. Considering that ascorbic and dehydroascorbic acid are active and common reductones [1], the oxidation-enhanced carbonyl-protein cross-linking is suggested to be an important chemical reaction which may take place during ageing and be involved in lipofuscinogenesis.