REACTION OF MYOGLOBIN WITH HYDROGEN-PEROXIDE FORMS A PEROXYL RADICAL WHICH OXIDIZES SUBSTRATES

Evidence is presented that the radical observed upon reaction of myogl obin with hydrogen peroxide is a peroxyl radical. Simulation of this s pectrum gives principal values for the g tenser of g(x) = 2.0357, g(y) = 2.0082, and g(z) = 2.0016, which are consistent with those of a per oxyl radical. Use of molecular oxygen isotopically labeled with O-17 c onfirmed that the radical observed was a peroxyl radical. Removal of o xygen from the incubation by use of glucose and glucose oxidase reveal ed two radicals, one at g(iso) = 2.0028 and the other at g(iso) = 2.00 73. Addition of various amounts of the spin trap 5,5-dimethyl-1-pyrrol ine N-oxide revealed that the spin trap and oxygen compete for the sam e radical site. Four model substrates, glutathione, styrene, arachidon ic acid and linoleic acid, were individually added to both the aerobic and anoxic systems. Glutathione reacted with the peroxyl radical, red ucing its intensity by 98%, and entirely eliminated the g(iso) = 2.002 8 line from the spectrum of the anoxic incubation. Styrene, arachidoni c acid and linoleic acid reacted with the peroxyl radical, reducing it s amplitude by 84, 57, and 35%, respectively, but did not decrease the amplitude of either radical species in the anoxic incubation. The g(i so) = 2.0028 species detected in the anoxic incubation appears to be t he original radical site to which molecular oxygen binds to form the p eroxyl radical. This myoglobin-derived peroxyl radical species is resp onsible for the advent of lipid peroxidation as proposed in ischemia/r epurfusion injury, as well as other reactions, as exemplified by the O -2-dependent epoxidation of styrene.