A highly sensitive two-photon fluorescent probe for glutathione with near-infrared emission at 719 nm and intracellular glutathione imaging.

Abstract

A near-infrared turn-on two-photon fluorescent probe ST-BODIPY for glutathione-specific detection was designed and synthesized by attaching triphenylamine to BODIPY skeleton through the Knoevenagel condensation to prolong the maximum emission wavelength to the NIR region. And 2,4-dinitrobenzenesulfonyl group (DNBS), as the fluorescence quencher and thiol recognition moiety, was modified in 8 position of BODIPY. In the presence of GSH, the probe afforded an "off-on" signal response with a significant NIR fluorescence enhancement centered at 719 nm accompanying by quantum yield increased to 0.44, which was ascribed to the glutathione-induced SNAr (aromatic substitution) reaction. Surprisingly, we found that the probe could discriminate GSH from other biothiols including Cys and Hcy upon the addition of intracellular concentrations of them. Time-dependence also demonstrated that the probe could distinguish GSH from Cys and Hcy under physiological environment. The limit of detection (LOD) for GSH was calculated as 25.46 nM from the titration experiments, which is lower than most previously reported GSH-selective probes. Under the Ti:sapphire pulsed laser's 800 nm irradiation, ST-BODIPY toward GSH generated an "off-on" signal response with a significant enhancement of fluorescence emission at 719 nm after treatment with GSH. Besides, the 2PA cross section value (σ2) was calculated to be 410 GM, suggesting that it could not only function well as an excellent two-photon fluorescent probe for the detection of intracellular GSH, but also be applied for two-photon imaging with high sensitivity in living cells. Moreover, ST-BODIPY probe has been successfully employed for monitoring exogenous and endogenous GSH in MCF-7 cells with satisfying results, perhaps it was feasible for detecting abnormal contents of GSH in a biological system and accomplishing the goal of maintaining normal human activities.