Time-resolved fluorescence spectroscopy response

At about the same time as this work was published, Znamenskiy and Kobrak simulated the absorption spectrum of betaine-30, a commonly used solvatochromic probe molecule, in [C4inim][PF6]. They investigated the interactions responsible for the solvatochromic shift. Because this shift is used experimentally to assess solvent polarity, the calculations can thus provide a direct window into the nature of polarity in ionic Hquid systems. To conduct the study, a single molecule of betaine-30 was immersed in a Hquid containing 200 ion pairs. Twelve independent 1-ns runs were then carried out, and from that the absorption spectrum was computed. They observe two distinct time scales one on subpicosecond time scales and one that is on the order of 100 ps. This result is consistent with previous simulation studies as well as time-resolved fluorescence spectroscopy experiments. Although the actual absorption spectra computed do not agree quantitatively with experimental results, the qualitative features do. 

The rotational reorientation times of the sample in several solvents at room temperature were measured by picosecond time-resolved fluorescence and absorption depolarization spectroscopy. Details of our experimental setups were described elsewhere. For the time-correlated single photon counting measurement of which the response time is a ut 40 ps, the sample solution was excited with a second harmonics of a femtosecond Ti sapphire laser (370 nm) and the fluorescence polarized parallel and perpendicular to the direction of the excitation pulse polarization as well as the magic angle one were monitored. The second harmonics of the rhodamine-640 dye laser (313 nm 10 ps FWHM) was used to raesisure the polarized transient absorption spectra. The synthesis of the sample is given elsewhere. All the solvents of spectro-grade were used without further purification. 

TNP-ATP complex obtained by the single-molecule time-resolved spectroscopy, together with a fluorescence decay curve of TNP-ATP obtained by a bulk measurement. Both curves were well fitted to biexponential functions. The instrument-response function in 195-ps fwhm is also displayed. (B) Representative fluorescence spectrums of two individual enzyme-TNP-ATP complexes showing different emission peaks. A fluorescence spectrum of TNP-ATP obtained from a bulk measurement is also displayed for comparison. All spectrums were normalized to unity at their maximum. (From Ref. 18.)... 

Heiftje GM, Vogelstein EE. A linear response theory approach to time-resolved fluorometry. In Wehry EL, ed. Modern fluorescence spectroscopy. New York Plenum Press, vol 4, 1981 25-50. 

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