H2O2是一种常见的生物活性小分子, 其在细胞增殖, 炎症反应, 细胞信号传输等过程中都起着重要的作用。 细胞粘度是一项重要的生理微环境参数, 其为细胞是否正常运转的关键指标。 研究还表明, 一些重大的生理疾病与细胞组织的粘度及过氧化氢浓度异常均有着密切的关联, 如阿尔茨海默病、 癌症。 因此开发能有效同时检测H2O2及粘度的分析方法对揭示相关生理病理机制及重大疾病的诊断有着重要的意义。 研制了一种能有效检测H2O2与粘度的双响应型荧光探针(1)。 由于存在扭曲分子内电荷转移(TICT)的猝灭效应, 探针(1)几乎不具有荧光发射特性。 而随着环境体系粘度的增高, TICT效应被抑制, 探针(1)呈现出了强的荧光发射, 且发射峰位于近红外波段处(680 nm)。 随着粘度从1.996 cp增至851.8 cp, 探针的荧光强度增幅达到了85倍。 探针还能对H2O2产生灵敏的荧光响应, 发射峰位置在590 nm处。 响应原理为H2O2能与探针中的苯硼酸基团反应使探针脱除掉与吡啶氮原子相连的亚甲基苯硼酸, 从而减弱了探针分子中的TICT效应以及分子内电荷转移(ICT)效应, 因此探针溶液荧光发射显著增强且探针溶液的紫外光谱发生了明显的蓝移(吸收峰从540 nm移至460 nm), 对应溶液颜色从红色变为黄色。 荧光光谱测试表明探针(1)对H2O2的检测具有高的选择性与灵敏度, 探针体系在590 nm处的荧光强度与H2O2的浓度呈良好的线性关系, 线性范围为0～25 μmol·L-1, 根据IUPAC的3σ法计算得出检测限为0.34 μmol·L-1。 此外细胞荧光成像实验表明探针(1)具有较好的生物相容性及细胞膜通透性, 且能实现细胞内H2O2的成像分析。

Hydrogen peroxide (H2O2) is an important biological molecule and plays vital roles in cell growth, immune responses, and cell signaling pathways. Cellular viscosity is a significant physiological parameter and also indicates the normal or abnormal functions of cells. Moreover, both abnormal levels of H2O2 and cellular viscosity are found to be highly related to some major diseases, such as Alzheimer’s disease and cancers. Therefore, the development of effective analytical tools for simultaneously detecting H2O2 and cellular viscosity is of great significance to elucidating some critical physiological and pathological mechanisms, as well as the diagnosis of some relevant diseases. In this work, we developed a dual-responsive fluorescent probe (1) for viscosity and H2O2. Probe (1) is almost non-fluorescent due to the quenching effect arisen from the twisted intramolecular charge transfer (TICT) process within the probe. While the probe exhibited strong near-infrared fluorescence (~680 nm) in solution with high viscosities, which can be attributed the restricted TICT process. The turn-on fluorescence reached 85 folds with the solution viscosity increased from 1.996 cp to 851.8 cp. Furthermore, probe (1) also can sensitively response to H2O2 with the evolution of a new emission band at about 590 nm. H2O2 can effectively react with the phenylboronic acid moiety of probe (1) and result in the conversion of pyridinium unit to pyridine, which could attenuate the ICT (intramolecular charge transfer) and TICT effects of the probe, and thus lead to a dramatic increase in the fluorescence intensity as well as a blue-shift in absorption profile (from 540 to 460 nm) with the observed color of the solution changed from purple-red to yellow. Fluorescence measurements indicated that probe (1) is highly sensitive and selective for H2O2. The fluorescence intensity of the probe assay at 590 nm was found to vary linearly with the concentration of H2O2 in the range of 0～25 μmol·L-1, the detection limit was calculated to be 0.34 μmol·L-1 (3σ). Furthermore, cellular imaging experiment confirmed that probe (1) is highly biocompatible and cell-membrane permeable, and can be utilized for monitoring H2O2 in living cells.