Development of a mitochondria targetable ratiometric time-gated luminescence probe for biothiols based on lanthanide...

A mitochondria targetable ratiometric luminescence probe based on a mixture of Eu³⁺ and Tb³⁺ complexes, Mito-NSTTA-Eu³⁺/Tb³⁺, has been developed for the specific recognition and time-gated luminescence detection of biothiols in aqueous and living samples. This probe was synthesized by conjugating a mitochondria-anchoring motif, triphenylphosphonium, with a biothiol-responsive terpyridine derivative, [4'-(2,4-dinitrobenzenesulfonyloxy)-2,2':6',2''-terpyridine-6,6''-diyl] bis(methylenenitrilo) tetrakis(acetic acid), for coordinating with Eu³⁺ and Tb³⁺ ions. When the probe interacts with biothiols, the cleavage of the 2,4-dinitrobenzenesulfonyl moiety leads to a remarkable enhancement of the Tb³⁺ emission at 540 nm, while the Eu³⁺ emission at 610 nm is unchanged, which allows Mito-NSTTA-Eu³⁺/Tb³⁺ to be used as a ratiometric probe for the time-gated luminescence detection of biothiols, with the luminescence intensity ratio of I₅₄₀/I₆₁₀ as a signal. The triphenylphosphonium group enables the probe molecules to be specifically driven into the mitochondria after cell loading for responding to biothiols therein. Mito-NSTTA-Eu³⁺/Tb³⁺ showed high selectivity and sensitivity for the detection of biothiols under physiological conditions, with detection limits of 0.098 μM for glutathione (GSH) and 0.14 μM for cysteine (Cys). In cell imaging experiments, Mito-NSTTA-Eu³⁺/Tb³⁺ displayed an excellent mitochondria-localization feature, which allowed biothiols in the mitochondria of live cells to be successfully imaged under ratiometric and time-gated luminescence modes. In addition, biothiols in living Daphnia magna were also imaged using Mito-NSTTA-Eu³⁺/Tb³⁺ as a probe. All of the results suggested that Mito-NSTTA-Eu³⁺/Tb³⁺ could serve as a useful tool for monitoring biothiols in vitro and in vivo with high specificity and sensitivity.