Peroxynitrite (ONOO-), one of representative reactive nitrogen species with strong oxidative and nitrative properties, is known to be associated with various human diseases, such as Alzheimer's disease, drug-induced liver injury (DILI), inflammation, and cancer. Probing its fluctuations throughout diseases holds profound promise for advancing early diagnosis and enabling prompt intervention. In this work, we designed and synthesized a β-diketonate Eu3+ complex-based probe, [Eu(Cy-CDHH)3(terpy)], for the time-gated luminescence (TGL) detection of ONOO-. The probe, composed of a cyanine-dye-conjugated β-diketonate-Eu3+ coordination structure, is non-luminescent due to the intramolecular energy transfer from β-diketonate to cyanine-dye, which inhibits the energy transfer from β-diketonate to central Eu3+ ion. Upon reaction with ONOO-, the unsaturated CC bond of cyanine-dye is cleaved. This process leads to the recovery of the intense long-lived luminescence of the β-diketonate-Eu3+ complex (ϕ = 17.3 %, τ = 436 μs), showcasing characteristics of rapid response (within 10 s), high selectivity, low detection limit (17.4 nM), and low cytotoxicity. These features enable the probe to be used for the quantitative TGL detection of ONOO- in aqueous media as well as for the background-free TGL imaging of ONOO- in living cells under assorted stimuli. Furthermore, the probe was effectively implemented for imaging of ONOO- in livers of drug-induced liver injury mice, revealing the up-regulation of ONOO- levels in this disease and the therapeutic efficacy of glutathione (GSH) via precluding the onset of reactive oxygen/nitrogen species. This research paves a new way for the fabrication of lanthanide complex bioprobes, providing a useful tool for understanding the interconnection between ONOO- and disease-related physiological processes.