Phenothiazine-Based Fluorescence-Quenched Photoacoustic Contrast Agents for Molecular Imaging

Photoacoustic imaging (PAI), renowned for its high spatial resolution and deep tissue penetration, holds great promise for disease diagnosis and molecular detection. However, the performance of commercially available PAI contrast agent is often hindered by intrinsic fluorescence, which reduces photoacoustic (PA) signal intensity and contrast, thereby limiting their applications. Herein, we rationally designed and synthesized a class of PAI contrast agents based on a phenothiazine core. By precise molecular engineering, we introduced a twisted intramolecular charge transfer (TICT) effect to suppress fluorescence, minimize radiative decay, and facilitate nonradiative energy dissipation, thereby enhancing photothermal conversion efficiency and significantly improving PA signal intensity and imaging contrast-to-noise ratio both <i>in vitro</i> and <i>in vivo</i>. Moreover, the structure facilitates the convenient incorporation of derivatization sites, allowing for further structural expansion and modification. Building upon this platform, we developed a hypochlorous acid (HOCl)-responsive PA probe, DHU-PAOCl-1, to further explore the biomedical imaging potential of these designed contrast agents. DHU-PAOCl-1 exhibited high selectivity and pH stability, with a detection limit of 11.88 nM. In a dermatitis animal model, DHU-PAOCl-1 enabled precise in situ visualization of endogenously generated HOCl at inflammatory sites, offering a powerful imaging tool for the early diagnosis and real-time monitoring of inflammation-related diseases. Collectively, this study provides an effective molecular design strategy for high-performance PAI contrast agents and expands the application potential of PAI technology in molecular diagnostics and therapeutic monitoring.