A Pulsed Laser Study of Excited States of Aromatic Molecules Absorbed in Globular Proteins

Pyrene and several derivatives of pyrene such as pyrene sulfonic acid, and pyrene butyric acid were incorporated into bovine serum albumin (BSA) in aqueous solution. The pyrene chromophore was subsequently excited by a pulse of uv light (λ = 3471 Å) from a Q switched frequency doubled ruby laser. The lifetime of the pyrene excited singlet and triplet states were monitored by time resolved spectrophotometry. Various molecules, such as O₂ and I^-, dissolved in the aqueous phase, diffused into the protein and quenched pyrene excited states. The rates of these reactions were followed under a variety of conditions such as pH and temperature and in the presence of inert additives. The rates of pyrene excited-state quenching were often considerably smaller than the rates observed in simple solutions. A comparison of the rates in the protein and homogeneous solutions gives information on the factors such as temperature, charge, and pH that control the movement of small molecules in and into BSA.