Maple syrup urine disease (MSUD) is a metabolic disorder caused by the deficiency of the activity of the mitochondrial enzyme complex branched-chain L-2-keto acid dehydrogenase. The metabolic block results in tissue and body fluid accumulation of the branched-chain amino acids leucine (Leu), isoleucine and valine, as well as of their respective alpha-keto acids. Neurological sequelae are usually present in MSUD, but the pathophysiologic mechanisms of neurotoxicity are still poorly known. It was previously demonstrated that Leu elicits oxidative stress in rat brain. In the present study we investigated the possible mechanisms involved in Leu-induced oxidative damage. We observed a significant attenuation of Leu-elicited increase of thiobarbituric acid-reactive substances (TBA-RS) measurement when cortical homogenates were incubated in the presence of the free radical scavengers ascorbic acid plus trolox, dithiothreitol, glutathione, and superoxide dismutase, suggesting a probable involvement of superoxide and hydroxyl radicals in this effect. In contrast, the use of Nomega-nitro-L-arginine methyl ester or catalase (CAT) did not affect TBA-RS values. We also demonstrated an inhibitory effect of Leu on the activities of the antioxidant enzymes CAT and gluthathione peroxidase, as well as a significant reduction in the membrane-protein thiol content from mitochondrial enriched preparations. Furthermore, dichlorofluorescein levels were increased although not significantly by Leu. Taken together, our present data indicate that an unbalance between free radical formation and inhibition of critical enzyme activities may explain the mechanisms involved in the Leu-induced oxidative damage.

5-Oxoproline (pyroglutamic acid) accumulates in glutathione synthetase deficiency, an inborn metabolic defect of the gamma-glutamyl cycle. This disorder is clinically characterized by hemolytic anemia, metabolic acidosis and severe neurological disorders. Considering that the mechanisms of brain damage in this disease are poorly known, in the present study we investigated whether oxidative stress is elicited by 5-oxoproline. The in vitro effect of (0.5-3.0 mM) 5-oxoproline was studied on various parameters of oxidative stress, such as total radical-trapping antioxidant potential, total antioxidant reactivity, chemiluminescence, thiobarbituric acid-reactive substances, sulfhydryl content, carbonyl content, and 2',7'-dichlorofluorescein fluorescence, as well as on the activities of the antioxidant enzymes catalase, superoxide dismutase and glutathione peroxidase in cerebral cortex and cerebellum of 14-day-old rats. Total radical-trapping antioxidant potential and total antioxidant reactivity were significantly reduced in both cerebral structures. Carbonyl content and 2',7'-dichlorofluorescein fluorescence were significantly enhanced, while sulfhydryl content was significantly diminished. In contrast, chemiluminescence and thiobarbituric acid-reactive substances were not affected by 5-oxoproline. The activities of catalase, superoxide dismutase and glutathione peroxidase were also not altered by 5-oxoproline. These results indicate that 5-oxoproline causes protein oxidation and reactive species production and decrease the non-enzymatic antioxidant defenses in rat brain, but does not cause lipid peroxidation. Taken together, it may be presumed that 5-oxoproline elicits oxidative stress that may represent a pathophysiological mechanism in the disorder in which this metabolite accumulates.