Free radical scavenging activity of silibinin in nitrite-induced hemoglobin oxidation and membrane fragility models

Free radical formation in heme proteins is recognized as a factor in mediating the toxicity of many drugs. Xenobiotics and drug therapy-related toxicity, due to oxidative modification of hemoglobin (Hb), has been attributed in part to the uncontrolled oxidative reactions. A variety of antioxidant strategies to ameliorate potential oxidative damage in vivo have been suggested. The present study was designed to evaluate the dose-response relationship of the free radical scavenging properties of silibinin dihemisuccinate (SDH) in nitrite-induced Hb oxidation in vitro and in vivo. Different concentrations of SDH were added, before and after different intervals of inducing Hb oxidation in erythrocytes lysate, and formation of methemoglobin (MetHb) was monitored spectrophotometrically; the same approach was utilized to evaluate the effect of the same doses of SDH on the integrity of erythrocytes after induction of hemolysis. Moreover, the most effective dose of SDH was administered in rats before challenge with toxic dose of sodium nitrite, and MetHb formation was monitored as mentioned before. The results showed that in both in vitro and in vivo models, SDH successfully attenuates Hb oxidation after challenge with sodium nitrite; this protective effect was not related to the stage of the catalytic stage of Hb oxidation, though the effect was more prominent when the compound was administered before nitrite. In conclusion, SDH can effectively, in concentration-dependent pattern, attenuate sodium nitrite-induced Hb oxidation and maintain integrity of red blood cells both in vitro and in vivo.