Room-Temperature Solution Fluorescence Excitation Correlation Spectroscopy

J Phys Chem Lett. 2024 Aug 22;15(33):8533-8539. doi: 10.1021/acs.jpclett.4c01798. Epub 2024 Aug 12.

Abstract

Single-molecule fluorescence spectroscopy is a powerful tool for investigating the physical properties of individual molecules, yet elucidating the fast fluctuation dynamics of freely diffusing single molecules in solution at room temperature, where a variety of chemical and biological processes occur, remains challenging. In this study, we report on fluorescence excitation correlation spectroscopy of room-temperature solutions, which enables the study of spontaneous fluctuation of the excitation spectrum with microsecond time resolution. By employing Fourier transform spectroscopy with broadband femtosecond pulses and time-correlated single-photon counting, we achieved fluorescence excitation spectroscopy of a room-temperature solution at the single-molecule level. Building upon this single-molecule measurement, we obtained an excitation wavelength-resolved fluorescence autocorrelation function in the microsecond to millisecond range, demonstrating the potential of this method to elucidate fast, spontaneous, time-dependent changes of excitation spectra in statistically equilibrated systems. With further development, this method will allow the study of spectral exchange associated with transitions between sub-ensembles of solution-phase molecules with unprecedented time resolution.