Quercetin Diminishes The Binding Of Hemoglobin To The Red Blood Cell Membrane

Hemoglobin (Hb) oxidation leads to the formation of hemichrome, which binds to the membrane and causes red blood cell removal by the reticuloendothelial system. In the present investigation, the effect of flavonoids on Hb oxidation and their binding to red blood cell (RBC) membranes were studied using tert-butyl hydroperoxide (tert-BOOH) to promote oxidative stress. The intrinsic antioxidant activity of RBC was able to prevent the binding of Hb to the membrane at tert-BOOH concentrations up to 0.4 mM. At higher concentrations, a brown pellet was observed and represented the appearance of membrane-bound oxidized Hb. Oxidations performed in membrane-free Hb solutions with an identical oxidative system showed less Hb oxidation. These observations suggest that erythrocyte membrane lipid peroxidation enhances the oxidative damage of Hb, increasing its binding to membranes. Quercetin partially protected Hb against oxidation by tert-BOOH and reduced the levels of the membrane bound hemichrome. Lipid peroxidation was also significantly suppressed by quercetin. Rutin and morin had little effect in preventing Hb binding to RBC membranes, indicating the importance of structure in the antioxidant properties of flavonoids. In the absence of oxidant, the peroxidation of erythrocyte membrane and isotonic hemolysis were protected by quercetin. These results suggest that quercetin displays a beneficial role on aging of RBC.415563Luna, E.J., Kidd, G.H., Branton, D., Identification by peptide analysis of the spectrin-binding protein in human erythrocytes (1979) J Biol Chem, 254, pp. 2526-2532Hargreaves, W., Giedd, K.N., Verkleij, A., Reassociation of ankyrin with band 3 in erythrocyte membranes and in lipid vesicles (1980) J Biol Chem, 255, pp. 11965-11972Bennett, V., Stenbuck, P.J., The membrane attachment protein for spectrin is associated with band 3 in human erythrocyte membrane (1979) Nature, 280, pp. 468-473Bennett, V., Stenbuck, P.J., Association between ankyrin and the cytoplasmic domain of band 3 isolated from the human erythrocyte membrane (1980) J Biol Chem, 255, pp. 6424-6432Shaklai, N., Yguerabide, J., Ranney, H.M., Classification and localization of hemoglobin binding sites on the red blood cell membrane (1977) Biochemistry, 16, pp. 5585-5592Salhany, J.M., Cordes, K.A., Gaines, E.D., Light-scattering measurements of hemoglobin binding to the erythrocyte membrane. Evidence for transmembrane effects related to a disulfonic stilbene binding to band 3 (1980) Biochemistry, 19, pp. 1447-1454Waugh, S.M., Low, P.S., Hemichrome binding to band 3: Nucleation of Heinz bodies on the erythrocyte membrane (1985) Biochemistry, 24, pp. 34-39Kiefer, C.R., Trainor, J.F., McKenney, J.B., Hemoglobin-spectrin complexes: Interference with spectrin tetramer assembly as a mechanism for compartmentalization of band 1 and band 2 complexes (1995) Blood, 86, pp. 366-371Bates, D., Winterbourn, C.C., Haemoglobin denaturation, lipid peroxidation and haemolysis in phenylhydrazine-induced anaemia (1984) Biochim Biophys Acta, 798, pp. 84-87Snyder, L.M., Fortier, N.L., Trainor, J., Effect of hydrogen peroxide exposure on normal human erythrocyte deformability, morphology, surface characteristics, and spectrin-hemoglobin cross-linking (1985) J Clin Invest, 76, pp. 1971-1977Fortier, N., Snyder, L.M., Garver, F., The relationship between in vivo generated hemoglobin skeletal protein complex and increased red cell membrane rigidity (1988) Blood, 71, pp. 1427-1431Itano, H.A., Hirota, K., Vedvick, T.S., Ligands and oxidants in ferrihemichrome formation and oxidative hemolysis (1977) Proc Natl Acad Sci USA, 74, pp. 2556-2560French, J.K., Winterboum, C.C., Carrell, R.W., Mechanism of oxyhaemoglobin breakdown on reaction with acetylphenylhydrazine (1978) Biochem J, 173, pp. 19-26Trotta, R.J., Sullivan, S.G., Stern, A., Lipid peroxidation and hemoglobin degradation in red blood cells exposed to t-butyl hydroperoxide. Dependence on glucose metabolism and hemoglobin status (1981) Biochim Biophys Acta, 678, pp. 230-237Van Der Zee, J., Van Steveninck, J., Koster, J.F., Inhibition of enzymes and oxidative damage of red blood cells induced by t-butylhydroxperoxide-derived radicals (1989) Biochim Biophys Acta, 980, pp. 175-180Slater, T., Free radical mechanisms in tissue injury (1984) Biochem J, 222, pp. 1-15Pryor, W., Oxy-radicals and related species: Their formation, lifetimes, and reactions (1986) Annu Rev Physiol, 48, pp. 657-667Yagi, K., Lipid peroxidation and human diseases (1987) Chem Phys Lipids, 45, pp. 337-351Hollman, P.C.H., Katan, M.B., Absorption, metabolism and health effects of dietary flavonoids in man (1997) Biomed Pharmacother, 51, pp. 305-310Van Aker, F.A.A., Schouten, O., Haenen, G.R.M.M., Flavonoids can replace α-tocopherol as an antioxidant (2000) FEBS Lett, 473, pp. 145-148Bors, W., Michel, C., Saran, M., Flavonoid antioxidants: Rate constants for reactions with oxygen radicals (1994) Methods Enzymol, 234, pp. 420-429Jovanovic, S.V., Steenken, S., Tosic, M., Flavonoids as antioxidants (1994) J Am Chem Soc, 116, pp. 4846-4851Ratty, A.K., Das, N.P., Effects of flavonoids on nonenzymatic lipid peroxidation: Structure-activity relationship (1988) Biochem Med Metab Biol, 39, pp. 69-79Laughton, M.J., Evans, P.J., Moroney, M.A., Inhibition of mammalian 5-lipooxygenase and cyclooxygenase by flavonoids and phenolic dietary additives. Relationship to antioxidant activity and to iron ion-reducing ability (1991) Biochem Pharmacol, 42, pp. 1673-1681Decharneux, T., Dubois, F., Beauloye, C., Effect of various flavonoids on lysosomes subjected to an oxidative or an osmotic stress (1992) Biochem Pharmacol, 44, pp. 1243-1248Korkina, L.G., Afanas'ev, I.B., Antioxidant and chelating properties of flavonoids (1997) Adv Pharmacol, 38, pp. 151-163Afanas'ev, I.B., Dorozhko, A.I., Brodskii, A.V., Chelating and free radical scavenging mechanisms of inhibitory action of rutin and quercetin in lipid peroxidation (1989) Biochem Pharmacol, 38, pp. 1763-1769Saija, A., Scalese, M., Lanza, M., Flavonoids as antioxidant agents: Importance of their interaction with biomembranes (1995) Free Radic Biol Med, 19, pp. 481-486Ferrali, M., Signorini, C., Caciotti, B., Protection against oxidative damage of erythrocyte membrane by the flavonoid quercetin and its relation to iron chelating activity (1997) FEBS Lett, 416, pp. 123-129Winterbourn, C.C., Oxidative reactions of hemoglobin (1990) Methods Enzymol, 186, pp. 265-272Stocks, J., Normandy, T.L., The autoxidation of human red cell lipids induced by hydrogen peroxide (1971) Br J Haematol, 20, pp. 95-111Stern, A., Red cell oxidative damage (1985) Oxidative Stress, pp. 75-90. , Sies H, ed. London: Academic PressPeisach, J., Blumberg, W.E., Rachmilewitz, E.A., The demonstration of ferrihemichrome intermediates in Heinz body formation following the reduction of oxyhemoglobin A by acetylphenylhydrazine (1975) Biochim Biophys Acta, 393, pp. 404-418Minetti, M., Mallozzi, C., Scorza, G., Role of oxygen and carbon radicals in hemoglobin oxidation (1993) Arch Biochem Biophys, 302, pp. 233-244Skaper, S.D., Fabris, M., Ferrari, V., Quercetin protects cutaneous tissue-associated cell types including sensory neurons from oxidative stress induced by glutathione depletion: Cooperative effects of ascorbic acid (1997) Free Radic Biol Med, 22, pp. 669-678Cao, G., Sofic, E., Prior, R.L., Antioxidant and prooxidant behavior of flavonoids: Structure-activity relationship (1997) Free Radic Biol Med, 22, pp. 749-760Hebbel, R.P., Eaton, J.W., Balasingam, M., Spontaneous oxygen radical generation by sickle erythrocytes (1982) J Clin Invest, 70, pp. 1253-1259Saija, A., Scalese, M., Lanza, M., Flavonoids as antioxidant agents: Importance of their interaction with biomembranes (1995) Free Radic Biol Med, 19, pp. 481-486Boyer, R.F., Clark, H.M., La Roche, A.P., Reduction and release of ferritin iron by plant phenolics (1988) J Inorg Biochem, 32, pp. 171-181Uri, N., (1961) Antoxidation and Antioxidants, pp. 133-169. , London: LundbergErben-Russ, M., Michel, C., Bors, W., Absolute rate constant of alkoxyl radical reactions in aqueous solution (1987) J Phys Chem, 91, pp. 2362-2365Jain, S.K., Hochstein, P., Polymerization of membrane components in aging red blood cells (1980) Biochem Biophys Res Commun, 92, pp. 247-254Barclay, L.R.C., Locke, S.J., MacNeil, J.M., Autoxidation of micelles and model membranes - Quantitative kinetic measuurements can be made by using either water or lipid-soluble initiators with water-soluble or lipid-soluble chain-breaking antioxidants (1984) J Am Chem Soc, 106, pp. 2479-2481Grinberg, L.N., Rachmilewitz, E.A., Newmark, H., Protective effects of rutin against hemoglobin oxidation (1994) Biochem Pharmacol, 48, pp. 643-649Hollman, P.C.H., Van Der Gaag, M., Mengelers, M.J.B., Absorption and disposition kinetics of the dietary antioxidant quercetin in man (1996) Free Radic Biol Med, 21, pp. 703-70