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

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 I-540/I-610 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 M for glutathione (GSH) and 0.14 M 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.