| Autor: |
McCallum AM; School of Chemistry and Biochemistry and Petit Institute for Bioengineering and Bioscience, Georgia Institute of Technology, 901 Atlantic Drive, Atlanta, Georgia 30332, United States., Yu J; School of Chemistry and Biochemistry and Petit Institute for Bioengineering and Bioscience, Georgia Institute of Technology, 901 Atlantic Drive, Atlanta, Georgia 30332, United States., Sumalekshmy S; School of Chemistry and Biochemistry and Petit Institute for Bioengineering and Bioscience, Georgia Institute of Technology, 901 Atlantic Drive, Atlanta, Georgia 30332, United States., Hagwood A; School of Chemistry and Biochemistry and Petit Institute for Bioengineering and Bioscience, Georgia Institute of Technology, 901 Atlantic Drive, Atlanta, Georgia 30332, United States., Fahrni CJ; School of Chemistry and Biochemistry and Petit Institute for Bioengineering and Bioscience, Georgia Institute of Technology, 901 Atlantic Drive, Atlanta, Georgia 30332, United States. |
| Abstrakt: |
Push-pull fluorophores with donor-π-acceptor architectures are attractive scaffolds for the design of probes and labels for two-photon microscopy. Such fluorophores undergo a significant charge-delocalization in the excited state, which is essential for achieving a large two-photon absorption cross-section and brightness. The polarized excited state may, however, also facilitate excited-state proton transfer (ESPT) pathways that can interfere with the probe response. Herein, we employed steady-state and time-resolved spectroscopic studies to elucidate whether ESPT is responsible for the pH-dependent emission response of the Zn(II)-selective fluorescent probe chromis-1. Composed of a push-pull architecture with a pyridine ring as the acceptor, the chromis-1 fluorophore core acts as a photobase that promotes ESPT upon acidification. Although the p K a of the pyridine acceptor increases more than six orders of magnitude upon excitation, the photobasicity is not sufficient to deprotonate solvent water molecules under neutral conditions. Rather, the pH-dependent emission response is caused by the pendant bis-isonicotinic acid chelating group which upon protonation facilitates an excited-state intramolecular proton transfer to the pyridine acceptor. A simple permutation of the core pyridine nitrogen from the para- to the ortho-position relative to the thiazole substituent was sufficient to reduce the excited-state basicity by two orders of magnitude without compromising the two-photon excited brightness. These results highlight the importance of choosing the appropriate fluorophore core and chelating moiety for minimizing pH-dependent responses in the design of fluorescent probes for biological imaging. |
