The tryptophan fluorescence lifetime puzzle. A study of decay times in aqueous solution as a function of pH and buffer composition

Tryptophan fluorescence decay kinetics have been systematically investigated in aqueous solutions as a function of pH as well as in a variety of buffer solutions. Below pH 7.0, the decay appears to be double exponential with a subnanosecond component confirming the previous findings of Rayner and Szabo (3). In the low pH region, where the proton concentration becomes kinetically significant, tryptophan fluorescence is collisionally quenched by [H+] with diffusion controlled rate and no experimental evidence is found regarding the appearance at low pH of a new tryptophan molecular species, namely the cationic form. At pH ≥ 7.0, the decay becomes triple-exponential with the appearance of a long component whose contribution to the total emission intensity increases rapidly with increasing pH at the expense of the other two. Lifetimes and relative intensities of each decay component depend in a complex way on pH and on the buffer chemical composition.