Near-infrared Fluorescent Probes for Sensitive Detection of NADH in Live Cells

该项目旨在开发用于精确和定量的水溶性近红外荧光探针 烟酰胺掺入对活细胞线粒体NAD(P)H浓度变化的影响 作为电子受体的不同吸电子荧光团的一部分，如四苯乙烯 (TPE)、二氰亚甲基-4H-色烯、香豆素、萘酰亚胺、罗丹明、罗多尔和BODIPY。我们会 研制比例式近红外荧光探针克服基于强度的系统误差 近红外BODIPY供体引入近红外罗丹明和罗多尔的荧光探针 分别是接受者。为了显著提高水的溶解性和生物相容性，我们将引入 支链寡聚(乙二醇基)残基连接到探针。它具有很强的吸电子性能 荧光团，如二氰亚甲基-4H-色烯、香豆素、萘酰亚胺、罗丹明、罗多尔和 BODIPY染料将促进NAD(P)H将烟酰胺部分快速还原为电子供体 二氢烟酰胺部分，并显著打开探针荧光。我们将使用计算 化学指导如何设计有效的荧光探针与研究光学关系 探针的性质及其化学结构，例如HOMO和LUMO水平，pKa值， 吸收和发射波长、荧光量子产率和探针之间的氧化还原反应 和NAD(P)H，并评估了从给体到受体的通过键能量转移效率 NAD(P)H存在下的比率荧光探针我们将研究底物特异性的影响 正常细胞和癌细胞中葡萄糖和丙酮酸/乳酸失衡对糖酵解途径中NADH水平的影响。 我们还将研究GSH/GSSG(谷胱甘肽/谷胱甘肽二硫化物)和 细胞氧化还原调节因子NAD/NADH在维持关键的氧化还原平衡中的作用 不同的代谢过程。最后，我们将研究nadh在氧化中独特的重要作用。 线粒体的磷酸化，并监测有丝分裂过程中NADH浓度的变化 由饥饿和药物治疗引起的。

The project aims to develop water-soluble near-infrared fluorescent probes for the precise and quantitative analysis of NAD(P)H concentration changes in mitochondria of live cells by incorporating the nicotinamide moiety into different electron-withdrawing fluorophores as electron acceptors such as tetraphenylethylene (TPE), dicyanomethylene-4H-chromene, coumarin, naphthalimide, rhodamine, rhodol, and BODIPY. We will develop ratiometric near-infrared fluorescent probes to overcome systematic errors of intensity-based fluorescent probes by introducing near-infrared BODIPY donors to near-infrared rhodamine and rhodol acceptors, respectively. In order to significantly enhance water solubility and biocompatibility, we will introduce branched oligo(ethylene glycol) residues to the probes. The strong electron-withdrawing properties of the fluorophores such as dicyanomethylene-4H-chromene, coumarin, naphthalimide, rhodamine, rhodol, and BODIPY dyes will facilitate a quick reduction of the nicotinamide moiety by NAD(P)H into an electron-donating dihydronicotinamide moiety and significantly turn on the probe fluorescence. We will use computation chemistry to guide how to design effective fluorescent probes and study the relationship of the optical properties of the probes and their chemical structures such as HOMO and LUMO levels, pKa values, absorption and emission wavelengths, fluorescence quantum yields, and redox reaction between the probes and NAD(P)H, and assess the through-bond energy transfer efficiency from the donors to the acceptors of ratiometric fluorescent probes in the presence of NAD(P)H. We will investigate the effects of substrate-specific glucose and pyruvate/lactate imbalance on NADH levels in the glycolysis pathway in normal and cancer cells. We will also investigate the co-relationship between GSH/GSSG (glutathione/glutathione disulfide) and the cellular redox regulator NAD+/NADH in maintaining a critical redox balance for proper functioning in the different metabolic processes. Finally, we will investigate the unique significant role of NADH in oxidative phosphorylation in mitochondria, and monitor NADH concentration changes during the mitophagy process caused by starvation and drug treatment.