On the chemical reactivity of tricyanofuran(TCF)-based near-infrared fluorescent redox probes – Effects of glutathione on the probe response and product fluorescence

Abstract

Recent research towards the development of the redox probes for in vivo applications focuses on near-infrared fluorescent sensors and tricyanofuran-based fluorophores gained popularity thanks to their favorable spectral properties. The tricyanofuran-based boronate probe (TCF-BA) has been proposed for specific fluorescent detection of selected biological oxidants in vitro and in vivo. Here, we report the detailed chemical reactivity of TCF-BA toward hydrogen peroxide, hypochlorite, and peroxynitrite in the presence and absence of glutathione, a major small molecule biothiol present intracellularly at millimolar concentrations. We demonstrate that, at the physiologically relevant concentration of glutathione, the TCF-BA probe forms an adduct, resulting in decreased reactivity of the probe toward the oxidants tested. Only peroxynitrite efficiently oxidizes TCF-BA in the presence of GSH. Furthermore, the fluorescent phenolic oxidation product, TCF-OH, also reacts with glutathione, which results in a decreased fluorescence intensity. This observation suggests that the results reported with TCF-based probes may be affected by the changes in intracellular glutathione, in addition to the desired analyte. We also report a modified probe (TCF-BA-2) derived from 1-naphthalene boronic acid, which has similar reactivity toward peroxynitrite. Although the TCF-BA-2 probe also reacts with glutathione, the absorption spectrum of its oxidation product, TCF–OH–2, is not influenced by glutathione and, therefore, can be applied for real-time monitoring of peroxynitrite formation in biological systems.