Difference spectra of mitochondria

These difference spectra of rat liver mitochondria were recorded in a double-beam spectrophotometer.

The black line shows the difference spectrum of fully reduced versus fully oxidized mitochondria.

Mitochondria reduced with substrate under anaerobic conditions were in the sample chamber, and oxygenated mitochondria in the reference chamber.

The peaks and shoulders refer to NADH, flavin, and the a and g absorption bands, as indicated, for cytochromes a, a3, b, and c.

The red line shows the effect of adding a respiratory inhibitor, antimycin A, which blocks electron flow from cytochrome b to c1.

The inhibitor causes all of the carriers beyond cytochrome b to become fully oxidized, while NADH, flavin, and cytochrome b are in the reduced state. Note the expanded absorbance scale beyond 500 nm.