Characterization of the Long-Range Internal Dynamics of Pyrene-Labeled Macromolecules by Pyrene Excimer Fluorescence

The long-range internal dynamics (LRID) of 74 pyrene-labeled macromolecules (PyLMs) were characterized in four solvents representing a broad range of dielectric constants equal to 2.4 (toluene), 7.6 (tetrahydrofuran, THF), 37.8 (N,N-dimethylformamide, DMF), and 46.7 (dimethyl sulfoxide, DMSO). The LRID of the PyLMs were quantified based on the parameters retrieved from the global model free analysis (MFA) of the time-resolved fluorescence (TRF) decays of the pyrene monomer and excimer. These parameters were combined to yield ⟨k⟩, the average rate constant of pyrene excimer formation in the PyLMs, and (IE/IM)ᵀᴿF(ffᵣₑₑ = 0), the ratio of excimer to monomer fluorescence intensity obtained in the absence of pyrene labels that do not form excimer. (IE/IM)ᵀᴿF(ffᵣₑₑ = 0) was found to increase linearly with increasing ⟨k⟩ values over 3 orders of magnitude in the four solvents studied with a slope that equaled the average lifetime of the pyrene excimer (τE). The ⟨k⟩ values obtained to build these master lines could be correlated back to the expected LRID of the macromolecules. The lowest ⟨k⟩ values were obtained for the end-labeled linear chains holding the pyrene groups far apart, whereas the branched macromolecules bringing the pyrene labels close to each other yielded the larger ⟨k⟩ values. Furthermore, the fact that these master curves were observed for so many different PyLMs in the four solvents covering such a broad range of solvent polarity suggests that this relationship represents a general physical phenomenon that applies to all PyLMs. Considering the importance of characterizing the LRID of macromolecules in solution, the (IE/IM)ᵀᴿF(ffᵣₑₑ = 0) vs ⟨k⟩ plots presented in this report can be viewed as calibration curves against which the LRID of any PyLM can now be compared. Thus, the 3 orders of magnitude range found for these master curves offers the scientific community an impressive analytical opportunity to gauge the LRID of macromolecules in solution.