Liu_2025_Anal.Chim.Acta_1367_344332

BACKGROUND: As a serine hydrolase synthesized in hepatocytes, butyrylcholinesterase (BChE) serves dual physiological roles: a validated biomarker for hepatic injury and a potential detoxifier of organophosphorus compounds (OPs). However, current fluorescent probes for BChE detection remain constrained by suboptimal clinical applicability due to limitations in tissue penetration, interference resistance, and non-ideal interactions with biological substrates. Consequently, the development of near-infrared (NIR) probes that integrate high sensitivity, rapid response kinetics and biocompatibility has emerged as a critical priority in hepatopathy diagnostics. RESULTS: A novel near-infrared small-molecule fluorescent probe, LQBr-CP, was synthesized based on the novel indocyanine green IR-820 scaffold through rational structural design. Upon interaction with BChE, LQBr-CP undergoes specific ester bond hydrolysis to release the fluorophore LQBr-OH, resulting in a 55.6-fold fluorescence enhancement with absorption/emission wavelengths at 705 nm and 758 nm, respectively. This probe demonstrates exceptional optical properties and superior aqueous solubility. Furthermore, it exhibits a detection limit of 2.64 U/L for BChE with no cross-reactivity toward acetylcholinesterase (AChE) or other biomacromolecules. In cellular and murine models, LQBr-CP enabled real-time imaging of endogenous BChE and revealed significant downregulation of BChE expression during acetaminophen (APAP)-induced acute liver injury, consistent with histopathological and blood biochemical analyses. SIGNIFICANCE AND NOVELTY: As the first reported near-infrared "off-on" fluorescence probe derived from IR-820, LQBr-CP addresses the penetration depth limitations of conventional probes in vivo, providing a highly sensitive visual tool for early diagnosis and real-time monitoring of hepatic injury, while offering novel insights into organophosphorus compound detoxification. The selectivity mechanism of BChE targeting was elucidated through integrated molecular docking and experimental validation, establishing a new design strategy for enzyme activity-targeted probes. This study not only advances the application of fluorescent sensing technology in hepatopathy therapeutics but also lays the technical groundwork for clinical assessment and mechanistic investigation of hepatotoxicity.