Rationally engineering recognition group to construct a two-photon reaction-based fluorescent probe for rapid and selective sensing of cysteine

Abstract

It is highly requested to rationally design fluorescent probes with desired analytical performances for the applications in sensing and imaging through molecular engineering strategy. The reaction-based fluorescent probes for highly selective sensing of cysteine (Cys) are mainly based on Cys-participated addition-cyclization with acrylates, cyclization with aldehydes, coordination displacement, Michael addition reaction, and cleavage reaction. Cys-triggered reaction with “O” ether bond has also been used to construct the reaction-based fluorescent probe based on the substitution of the ether with the nucleophilic thiolate of Cys. However, many of the developed probes still suffer from long response time, interfering from homocysteine (Hcy) and glutathione (GSH), high background fluorescence, and lack of two-photon absorption (TPA) properties. Herein, we successfully design a Cys-sensitive two-photon fluorescent probe (F-BTD) using a donor-acceptor-donor (D-A-D) type π-extended benzothiadiazole framework as the fluorophore, while nitrobenzofuran (NBD) as the recognition unit. The proposed F-BTD probe displays some advantages including rapid response, high selectivity, low background, and two-photon imaging capability. The F-BTD probe is applied to two-photon fluorescence imaging of endogenous and exogenous Cys in HeLa cells with satisfactory results. For comparing, commonly used recognition groups of biothiol including 2,4-dinitrobezensulfonyl and 2,4-dinitrophenyl are also used to construct S-BTD and N-BTD probes, respectively. The responsive mechanism of F-BTD to Cys is studied in detail through the kinetic studies and transition-state analysis. This study may provide an example to design fluorescent probe with desired analytical performances by taking recognition group into consideration as an important index.