PROOXIDANT EFFECTS OF CROSS-LINKED HEMOGLOBINS EXPLORED USING LIPOSOME AND CYTOCHROME-C-OXIDASE VESICLE MODEL MEMBRANES

The therapeutic use of cell-free haemoglobin as a blood substitute has been hampered by toxicological effects. A model asolectin (phosphatid ylcholine/phosphatidylethanolamine) liposome system was utilized to st udy the pro-oxidant efficiency of several chemically modified haemoglo bins on biological membranes. Lipid peroxidation, resulting from the i nteractions between haemoglobin and liposomes, was measured by conjuga ted diene formation and the maximal rates of oxygen uptake. Spectral c hanges gave insight into the occurrence of the ferryl iron species. Th e residual reactivity of oxidatively damaged haemoglobins with ligands during incubation with liposomes was assessed from rapid kinetic carb on monoxide-binding experiments. Liposomes in which cytochrome c oxida se was embedded show both haemoglobin and the enzyme to be oxidatively damaged during incubation. The functional state of cytochrome c oxida se was monitored in the presence and absence of a free radical scaveng er. Once in contact, both unmodified and modified haemoglobins trigger ed and maintained severe radical-mediated membrane damage. Differences in the pro-oxidant activities among haemoglobins may be explained by either the differential population of their ferryl intermediates or di sparate dimerization and transfer of haem into the membrane with subse quent haem degradation. This study may contribute to a better understa nding of the molecular determinants of haemoglobin interactions with a variety of biological membranes.