Cytochrome c oxidation determined by multiwavelength optical spectral analysis in the isolated perfused heart

摘要

Optical spectroscdpy has previously been used to investigate mitochondrial function in muscle, but spectral overlap of the absorbing species in muscle has limited the accuracy of these determinations. This report demonstrates a multispectral approach to determine cytochrome c redox state in the isolated perfused heart using partial least squares analysis. Optical spectra were acquired in the visible region (500 to 650 nm) from the surface of the isolated perfused guinea pig heart and interpreted using reference spectra obtained from in vitro solutions. Calibration spectra were acquired from separate light scattering solutions that contained cytochrome c or myoglobin in both oxidized (oxygenated) and reduced (deoxygenated) states. Cytochrome c redox state and myoglobin oxygen saturation were determined separately from each spectrum obtained from 16 guinea pig hearts during baseline perfusion with oxygenated buffer, during 90 seconds of ischemia and during recovery following the ischemic period. A brief delay between myoglobin deoxygenation and cytochrome c reduction was determined, demonstrating that both processes could be determined simultaneously from the same spectra. This approach may lead to improved determinations of mitochondrial function in the isolated perfused heart.