Integration of Time-Gated Luminescence and ¹⁹F Magnetic Resonance in Lanthanide Complexes-Based Probes for Quantitative Imaging of Mitochondrial Superoxide Anions.

Abstract

Time-gated luminescence (TGL) bioimaging offers high-sensitivity biosensing with background-free characteristics, yet its limited tissue penetration constrains broad in vivo applications. To address this issue, we developed unique lanthanide complexes to act as modular probes that integrate the high-sensitivity of TGL and deep-tissue imaging capability of ¹⁹F magnetic resonance (MR). The probes are constructed from a multifunctional ligand, Mito-BOTTA, comprising a mitochondria-targeting 1-(2-aminoethyl)-4-methylpyridinium group, a superoxide anion (O₂ ^•-)-responsive 3,5-bis(trifluoromethyl)benzenesulfonyl (TFBS) moiety, and a terpyridine polyacid-derived scaffold for coordination to Eu³⁺, Tb³⁺, or Gd³⁺. The coordination with Eu³⁺/Tb³⁺-mixture produces a ratiometric TGL probe Mito-BOTTA-Eu³⁺/Tb³⁺ that, upon O₂ ^•--triggered cleavage of the TFBS moiety, exhibits a pronounced increase in Tb³⁺ emission at 538 nm, accompanied by a decrease in Eu³⁺ emission at 608 nm. This reciprocal spectral response enables precise and autofluorescence-free ratiometric TGL imaging of mitochondrial O₂ ^•- using the I₅₃₈/I₆₀₈ ratio as a quantitative readout. In parallel, the complexation with Gd³⁺ yields Mito-BOTTA-Gd³⁺, where the paramagnetic Gd³⁺ ion quenches the ¹⁹F MR signal. The cleavage of TFBS in the presence of O₂ ^•- releases the fluorinated TFBS moiety, which restores the ¹⁹F MR signal, enabling a complementary "turn-on" MR detection for O₂ ^•-. Both probes demonstrated excellent mitochondrial localization, low cytotoxicity, and robust responsiveness to O₂ ^•- in HepG2 cells as well as in murine models of alcohol- and CCl₄-induced acute liver injuries, which suggested the potential of the probes for dual-mode TGL and ¹⁹F MR bioimaging of O₂ ^•-, offering a promising approach for the diagnostics of oxidative stress-related diseases.