Photophysics of a Live-Cell-Marker, Red Silicon-Substituted Xanthene Dye Crovetto González, Luis Orte Gutiérrez, Ángel Paredes, José M. Resa, Sandra Valverde Pozo, Javier Castello, Fabio Miguel, Delia Cuerva, Juan M. Talavera Rodríguez, Eva María Álvarez Pez, José María This document is the Accepted Manuscript version of a Published Work that appeared in final form in Journal of Physical Chemistry A, copyright © American Chemical Society after peer review and technical editing by the publisher. To access the final edited and published work see https://pubs.acs.org/doi/10.1021/acs.jpca.5b07898. Dyes with near-red emission are of great interest because of their undoubted advantages for use as probes in living cells. In-depth knowledge of their photophysics is essential for employment of such dyes. In this article, the photophysical behavior of a new siliconsubstituted xanthene, 7-hydroxy-5,5-dimethyl-10-(o-tolyl)dibenzo[b,e]silin-3(5H)-one (2-Me TM), was explored using absorption, steady-state, and time-resolved fluorescence. First, the near-neutral pH, ground state acidity constant of the dye, pK*N-A, was determined by both absorbance and steady-state fluorescence methods in the presence of very low buffer concentrations. Next, we determined whether the addition of phosphate buffer promoted the excited-state proton exchange reaction between the neutral and anion form of 2-Me TM in aqueous solutions at near-neutral pH. For this analysis, both the steady-state fluorescence method and time-resolved emission spectroscopy (TRES) were employed. The TRES experiments demonstrated a remarkably favored conversion of the neutral form to the anion form. Then, fluorescence decay traces recorded as a function of buffer concentration and pH were globally analyzed to determine the values of the excited-state rate constants. The revealed kinetic parameters were consistent with the TRES results, exhibiting a higher rate constant for deprotonation than for protonation, which implies an unusual low value of the excited-state acidity constant pK*N-A and therefore an enhanced photoacid behavior of the neutral form. Finally, we determined whether 2-Me TM could be used as a sensor inside live cells by measuring the intensity profile of the probe in different cellular compartments of HeLa 229 cells. 2019-03-07T11:15:08Z 2019-03-07T11:15:08Z 2015 info:eu-repo/semantics/article Crovetto, L. [et al.] Photophysics of a Live-Cell-Marker, Red Silicon-Substituted Xanthene Dye. J. Phys. Chem. A 2015, 119, 44, 10854-10862 https://pubs.acs.org/doi/10.1021/acs.jpca.5b07898 http://hdl.handle.net/10481/54892 10.1021/acs.jpca.5b07898 eng info:eu-repo/semantics/openAccess American Chemical Society