Photophysics of a Live-Cell-Marker, Red Silicon-Substituted Xanthene Dye
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- 作者：Luis Crovetto ; Angel Orte ; Jose M. Paredes ; Sandra Resa ; Javier Valverde ; Fabio Castello ; Delia Miguel ; Juan M. Cuerva ; Eva M. Talavera ; Jose M. Alvarez-Pez
- 刊名：Journal of Physical Chemistry A
- 出版年：2015
- 出版时间：November 5, 2015
- 年：2015
- 卷：119
- 期：44
- 页码：10854-10862
- 全文大小：454K
- ISSN：1520-5215
文摘
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 silicon-substituted xanthene, 7-hydroxy-5,5-dimethyl-10-(o-tolyl)dibenzo[b,e]silin-3(5H)-one (2-Me TM), was explored by means absorption, steady-state, and time-resolved fluorescence. First, the near-neutral pH, ground-state acidity constant of the dye, pKN-A, was determined by absorbance and steady-state fluorescence at very low buffer concentrations. Next, we determined whether the addition of phosphate buffer promoted the excited-state proton-transfer (ESPT) reaction among 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, the values of the excited-state rate constants were determined by global analysis of the fluorescence decay traces recorded as a function of pH, and buffer concentration. 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.