Characterization of coupled ground state and excited state equilibria by fluorescence spectral deconvolution.
Fluorescence probes with multiparametric response based on the relative variation in the intensities of several emission bands are of great general utility. An accurate interpretation of the system requires the determination of the number, positions and intensities of the spectral components. We have developed a new algorithm for spectral deconvolution that is applicable to fluorescence probes exhibiting a two-state ground-state equilibrium and a two-state excited-state reaction. Three distinct fluorescence emission bands are resolved, with a distribution of intensities that is excitation-wavelength-dependent. The deconvolution of the spectrum into individual components is based on their representation as asymmetric Siano-Metzler log-normal functions. The application of the algorithm to the solvation response of a 3-hydroxychromone (3HC) derivative that exhibits an H-bonding-dependent excited-state intramolecular proton transfer (ESIPT) reaction allowed the separation of the spectral signatures characteristic of polarity and hydrogen bonding. This example demonstrates the ability of the method to characterize two potentially uncorrelated parameters characterizing dye environment and interactions.
Caarls
Wouter
W
Laboratory of Cellular Dynamics, Max Planck Institute for Biophysical Chemistry, Am Fassberg 11, D37077, Göttingen, Germany.
Soledad Celej
M
M
Demchenko
Alexander P
AP
Jovin
Thomas M
TM
D000465:Algorithms; D005456:Fluorescent Dyes / Q000737:chemistry; D006860:Hydrogen Bonding; D011522:Protons; D013050:Spectrometry, Fluorescence / Q000706:statistics & numerical data*
D005456:Fluorescent Dyes; D011522:Protons