Cytochrome c degrading activity in rat liver mitochondria

Benzophenone can be used as an extrinsic triplet state probe, as its phosphorescence, a broad band centered at 445 nm, is readily observable in aqueous solution at room temperature. When bound covalently as an acyl enzyme at the active site of chymotrypsin, the benzophenone probe produces phosphorescence which is unusually resistant to quenching by O₂, $\text{trans}$-cinnamic acid, and H₃O⁺. Sodium 2-naphthalenesulfonate quenches the phosphorescence, probably indirectly. The quenching data indicate that the local protein structure at the enzyme active site provides a rigid and protective substrate environment, which is not penetrated by even the smallest triplet quenchers.