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Redox transitions in a film of detergent-purified bovine cytochrome bc[sub 1] complex were investigated by perfusion-induced attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopy. The technique provides a flexible method for generating redox-induced IR changes of components of bovine cytochrome bc[sub 1] complex at a high signal:noise ratio. These IR redox difference spectra arise from perturbations of prosthetic groups and surrounding protein. Visible difference spectra were recorded synchronously using a light beam reflected from the exposed prism surface and provided a quantitative means of determining the redox transitions that were occurring. IR and visible redox difference spectra of iron-sulfur protein/cytochrome c[sub 1], heme b[sub H], and heme b[sub L] were separated by selective reduction and/or oxidation that extends published data on the homologous bacterial enzyme. Several bands could be tentatively assigned to redox-sensitive modes of hemes and ubiquinone and changes in the surrounding protein by comparison with available data for bacterial bc[sub 1] complex, other related heme proteins, and model compounds. Some tentative assignments of further signals to specific amino acids are made on the basis of known crystal structures. [ABSTRACT FROM AUTHOR]