---

Background: Biochemical studies have shown that iron produces reactive oxygen species via Haber-Weiss and Fenton reactions. The goal of this study is to examine the role of iron in oxidation of human hemoglobin and its structural changes in erythrocytes. Materials and Methods: In this experimental study, blood samples from healthy subjects were incubated aerobically with the iron containing metal catalyzed oxidation (MCO) system in the presence of 0.036, 0.7, 0.14, 0.28, 0.57, 1.14, 2.28, 4.55, 9.09, and 18.18 micromole of iron. Structural changes in Hb were followed by spectrophotometric analysis from 300 to 650 nm. In addition, carbonyl assay was performed for estimation of protein oxidation in globin chains. Results: Based on the results, oxy-Hb decreased up to 68% in iron-treated erythrocytes. Decrease in the absorbance ratio (A577, A542 wavelength) indicated the conversion of oxy-Hb to met-Hb. Also, met-Hb concentration was 4.7 fold of hemichrome. After 24 hours of incubation, oxyHb concentration decreased up to 50% and metHb concentration increased up to 85%. Moreover, increase in iron concentration resulted in significant carbonyl formation in hemoglobin. Conclusion: These findings indicate that Hb oxidation instead of its oxygenation leads to anemia and hypoxia. The findings of this study may be directly applicable to oxidation states during hemolytic diseases and iron treatment.

---

Background: The aim of this study is to assess antioxidative and pro-oxidative efficacy of ascorbate on serum albumin during iron-induced oxidative stress. Materials and Methods: In this experimental study, albumin was placed in the oxidative system containing iron ions and different concentrations of ascorbate. To monitor albumin degradation, sodium dodecyl sulfate polyacrylamide gel electrophoresis was performed according to Laemmli procedure. Oxidative modification of albumin was demonstrated using a method for determination of carbonyl groups by 2,4-dinitrophenylhydrazine. Results: By applying the carbonyl assay, ascorbate showed a dual effect: initial pro-oxidative effect on albumin changed to an antioxidant one in a dose-dependent manner. Our findings showed prooxidant effects for ascorbate in low concentrations (0-100 µM) and antioxidant effects in higher concentrations (100-300 µM). Also, electrophoretic pattern of plasma proteins showed significant protein aggregations in the range of 35 to 45 kDa of MW and protein degradations in the range of 115 to 180 kDa. Conclusion: Ascorbate can produce reactive oxygen species and can also inhibit the production of these oxidants in the presence of iron ions as well. These findings may be directly applicable to oxidative states during the administration of ascorbate and may be important in preventing oxidative modifications of proteins in blood circulation and other biological fluids.

---

Background: The presence of oxidant agents yields higher levels of free radical reaction products in erythrocyte membrane proteins and serum proteins. The aim of this study is to investigate the oxidative modifications of recombinant human coagulation factor VIII (rHFVIII) by spectrophotometric analysis. Materials and Methods: In this experimental study, rHFVIII was incubated aerobically with vitamin C and ferro ions in metal catalyzed oxidation (MCO) system for 4 to 28 hr. Carbonyl assay was used as an index of protein oxidation. For this purpose, 2,4dinitrophenyl hydrazine (DNPH) was used. Reaction of this reagent with carbonyl groups produces dinitrophenylhydrazone derivatives that their concentration was estimated by spectrophotometry. Results: Carbonyl groups in rHFVIII changed in the presence of vitamin C and ferro ions. Dose-dependent effects of vitamin C showed a decrease in carbonyl groups of rHFVIII whileferro ions increased oxidation and carbonyl group formation in this protein. Conclusion: These findings indicate that changes in carbonyl groups in rHFVIII are related to the generation of reactive oxygen species. Also, antioxidant mechanisms are activated in a dose- and time-dependent manner.