Highly Stable Electrochemical Probe with Bidentate Thiols for Ratiometric Monitoring of Endogenous Polysulfide in Living Mouse Brains

The lack of reliable approaches for real-time measurement and quantification of polysulfides (H₂S_n) in vivo greatly limits the exploration of their potential roles in brain functions. Herein, an electrochemical probe, 4-(5-(1,2-dithiolan-3-yl)pentanamido)-1,2-phenylene bis(2-fluoro-5-nitrobenzoate) (FP2), was rationally designed and created for determination of H₂S_n. The bis-electrophilic groups of FP2 could specifically recognize two −SH groups in H₂S_n and trigger the generation of an electroactive pyrocatechol moiety, resulting in a well-defined faradic current signal at ∼0.24 V (vs Ag/AgCl). Meanwhile, bidentate thiols were designed as anchoring sites to greatly improve the assembled stability of FP2 at the Au surface, which efficiently defended the interference of glutathione (GSH) with a current decrease of less than 5.2% even after long-term measurements in 5 mM GSH for 3 h. In addition, a stable inner reference molecule with dithiols, α-lipoic acid ferrocenylamide (FcBT), was synthesized to construct a ratiometric electrochemical strategy for in vivo determination of H₂S_n through one-step coassembling with FP2 via double S–Au bonds. The present ratiometric strategy demonstrated high selectivity for real-time tracking of H₂S_n in a linear range of 0.25–20 μM. Eventually, the developed microelectrode with high selectivity, accuracy, and stability was employed for in vivo assaying of H₂S_n in mouse brains with ischemia.