The binding of iron by melanin is a potentially important phenomenon as detailed knowledge of this binding is essential for understanding the role of melanin and iron in the pathogenesis of oxidative damage in the substantia nigra. Electron paramagnetic resonance spectroscopy is one of the most useful approaches in the investigation of melanins and their interaction with iron. This study was undertaken to obtain systematic data on the effects of ferric iron on the electron paramagnetic resonance spectra of neuromelanin and synthetic models of neuromelanin. Data from the latter can potentially be used to infer as to the state of neuromelanin in the human substantia nigra. The results of this study indicate that the spectra at g = 4.3, attributable to Fe3+, provides a useful parameter for determining the amount of paramagnetic iron bound to melanin. These data together with the magnitude of the free radical signal from melanin provides an indication of the amount of iron bound to neuromelanin in intact human substantia nigra. After binding to melanin, the iron can change its location and/or state, which is indicated by the change in the microwave power saturation that occurs gradually after the binding of the iron. At least part of this process could occur at low temperatures (i.e., during storage at -15 degrees C).