Tannous Rozan, Kopp Tal, Shabat Doron
School of Chemistry, Raymond and Beverly Sackler Faculty of Exact Sciences, Tel-Aviv University, Tel Aviv 69978, Israel.
JACS Au. 2025 May 23;5(6):2871-2883. doi: 10.1021/jacsau.5c00465. eCollection 2025 Jun 23.
Chemiluminescence imaging has emerged as a powerful alternative to fluorescence-based methods, offering significant advantages such as reduced background noise, elimination of autofluorescence, and prevention of photobleaching. These benefits are particularly critical for singlet oxygen detection, where the excitation light in fluorescence techniques can inadvertently generate singlet oxygen, compromising measurement accuracy. Despite this potential, the development of highly sensitive chemiluminescent probes for singlet oxygen detection under physiological conditions remains an urgent challenge. Here, we present a comprehensive structure-activity optimization of phenoxy-1,2-dioxetane precursors as probes for singlet oxygen detection in physiological environments. By systematically evaluating key parameterssteric hindrance at the oxidation site, the chemiexcitation rate of the luminophore, and total light emissionwe significantly increased the detection sensitivity of the singlet oxygen probe. Notably, a cyclobutyl-enolether probe (SOCL-CB) and a dimethyl-enolether probe (SOCL-DM) demonstrated 57-fold and 118-fold higher signal-to-noise (S/N) ratios, respectively, compared to the previously reported chemiluminescent adamantyl-enolether probe (SOCL-AD). The superior detection sensitivity of probe SOCL-DM was validated in an enzymatic model where singlet oxygen production was mediated by horseradish peroxidase. Remarkably, probe SOCL-DM detected singlet oxygen concentrations as low as 127 nM in this system, outperforming the previously reported probe SOCL-AD. These results establish probe SOCL-DM as the most sensitive chemiluminescent probe for singlet oxygen detection under physiological conditions reported to date. This study underscores the potential of chemiluminescent probes like SOCL-DM to facilitate real-time monitoring of singlet oxygen, providing invaluable tools for studying oxidative stress, elucidating cellular processes, and advancing diagnostic applications.
化学发光成像已成为基于荧光的方法的有力替代方案，具有显著优势，如降低背景噪声、消除自发荧光和防止光漂白。这些优点对于单线态氧检测尤为关键，在荧光技术中，激发光可能会意外产生单线态氧，从而影响测量准确性。尽管有这种潜力，但开发用于生理条件下单线态氧检测的高灵敏度化学发光探针仍然是一项紧迫的挑战。在此，我们对苯氧基-1,2-二氧杂环丁烷前体进行了全面的构效优化，以作为生理环境中单线态氧检测的探针。通过系统评估关键参数——氧化位点的空间位阻、发光体的化学激发速率和总发光量——我们显著提高了单线态氧探针的检测灵敏度。值得注意的是，与先前报道的化学发光金刚烷基烯醚探针（SOCL-AD）相比，环丁基烯醚探针（SOCL-CB）和二甲基烯醚探针（SOCL-DM）的信噪比分别提高了57倍和118倍。探针SOCL-DM的卓越检测灵敏度在一个酶促模型中得到验证，在该模型中，单线态氧的产生由辣根过氧化物酶介导。值得注意的是，在该系统中，探针SOCL-DM检测到低至127 nM的单线态氧浓度，优于先前报道的探针SOCL-AD。这些结果表明，探针SOCL-DM是迄今为止报道的生理条件下单线态氧检测最灵敏的化学发光探针。这项研究强调了像SOCL-DM这样的化学发光探针在促进单线态氧实时监测方面的潜力，为研究氧化应激、阐明细胞过程和推进诊断应用提供了宝贵工具。
