Gas-Phase Fluorescence of Proflavine Reveals Two Close-Lying, Brightly Emitting States

Abstract

Surprising excitation-dependent, dual emission from a small organic model fluorophore is reported. Gas-phase fluorescence spectra of proflavine (a diaminoacridine) ions reveal two long-lived emitting states, with distinct bands separated by just 1700 cm −1 . The relative intensities of these two bands depend on the excitation wavelength. Time-dependent density functional theory (TD-DFT) calculations support the existence of two close-lying singlet electronic states, with excitation into S 2 predicted to be >1000-fold more likely than into S 1 . These data strongly suggest that internal conversion (IC) rates are suppressed relative to solvated proflavine, and that IC is competitive with intramolecular vibrational relaxation (IVR). This work offers an in-depth assessment of the gas-phase photophysics of a simple fluorophore that could open a new pathway to understanding dual emission in fluorophores.

Cryogenic Fluorescence Spectroscopy of Ionic Fluorones in Gaseous and Condensed Phases: New Light on Their Intrinsic Photophysics

“…Most of these gas-phase studies have considered fluorescence emission spectra, with only a handful attempting to record fluorescence excitation spectra. 15,23,24,27,34 More sophisticated applications of gas-phase fluorescence spectroscopy have considered conformational studies of larger biomolecules, such as peptides, oligonucleotides, and proteins, 12,28,35,36 by exploiting Forster (fluorescence) resonance energy transfer (FRET) between through-space-coupled fluorophores. 37 The above-mentioned studies were performed almost exclusively at room temperature; there are only a few reports of gas-phase fluorescence instruments incorporating cryogenic cooling of the ions (i.e., T < 120 K) through cooling of the ion trap and buffer gas with liquid nitrogen or attachment of the ion trap to a helium refrigerator.…”

“…These investigations often provide a comparison to spectral properties in solution and contribute toward establishing a foundation from which tailored fluorophores can be developed. Most of these gas-phase studies have considered fluorescence emission spectra, with only a handful attempting to record fluorescence excitation spectra. ,,,, More sophisticated applications of gas-phase fluorescence spectroscopy have considered conformational studies of larger biomolecules, such as peptides, oligonucleotides, and proteins, ,,, by exploiting Förster (fluorescence) resonance energy transfer (FRET) between through-space-coupled fluorophores …”

“…While each type of trap has advantages, 3D Paul traps are simple and approximate a point-like trapping volume, making them easily adapted for fluorescence experiments. The majority of gas-phase fluorescence studies on ions generated through ESI have considered organic dyes, particularly rhodamines, ,− ,− and other xanthene-based species − at T = 300 K due to their desirable benchmarking characteristics. These investigations often provide a comparison to spectral properties in solution and contribute toward establishing a foundation from which tailored fluorophores can be developed.…”

Robust Fluorescence Collection Module for Wide-Bore Ion Cyclotron Resonance Mass Spectrometers

“…As a vessel for optical spectroscopy, QITs benefit from higher ion density than ICRs as well as from collisional cooling from buffer gas in the trapping region. The experimental setup on our QIT yielded high-quality fluorescence data from the start, which has allowed continuous usage for more than a decade. − However, commercial FT-ICR instruments provide superior mass spectral manipulation and analysis: ultrahigh mass resolving power, better dynamic and mass ranges, and an array of tandem MS possibilities. These capabilities are key to exploiting the full potential of methods such as FRET to characterize the conformation and dynamics of mass-selected biomolecular ions in carefully controlled environments.…”

Environmentally Driven Symmetry Breaking Quenches Dual Fluorescence in Proflavine

“…The energy gap is found to be strongly dependent on the choice of DFT functional, with B3LYP predicting a significantly smaller splitting at the ground-state optimized geometry. 17 We have performed benchmark calculations of a single optimized ground-state geometry using the equation-of-motion coupledcluster singles and doubles (EOM-CCSD) method, 78−80 resulting in an S 1 −S 2 gap of 0.394 eV. Because the average S 1 −S 2 gap controls the location of the conical intersection between the two states in the Condon region, it is expected to have a strong influence on the system dynamics upon excitation.…”