Coumarin- and Rhodamine-Fused Deep Red Fluorescent Dyes: Synthesis, Photophysical Properties, and Bioimaging in Vitro

A series of deep red fluorescent
dyes (CR1 to CR3) was developed via introduction
of a coumarin moiety into
the rhodamine molecular skeleton. The novel dyes possessed the individual
advantages of coumarin and rhodamine derivatives, and the emission
wavelength was extended to the deep red region (>650 nm) due to
the
extension of fused-ring conjugate structure simultaneously. To illustrate
its value, we designed and conveniently synthesized a series of novel
deep red bioimaging dyes (CR1E to CR3E)
by esterification of CR1 to CR3, which could
selectively stain mitochondria. They were superior to the MitoTrackers
for mitochondrial staining in terms of large Stokes shift, excellent
contrast for imaging, high photostability, and low cytotoxicity. Furthermore,
the fluorescence of the coumarin moiety and rhodamine-like fluorophore
could be switched like classical rhodamine. Thus, they could be used
as an effective platform in constructing fluorescence sensors. Based
on this fact, we constructed a novel ratiometric sensor (CR1S) for Hg2+ with good selectivity that could be successfully
applied to the imaging of Hg2+ in living A549 cells. This
design strategy is straightforward and adaptable to various deep red
dyes and sensing platforms by simply introducing different fluorophores.

本研究通过将香豆素（coumarin）基团引入罗丹明（rhodamine）分子骨架，成功开发了一系列深红色荧光染料（编号CR1至CR3）。该类新型染料兼具香豆素与罗丹明衍生物的双重优势，且因稠环共轭结构的同步延伸，其发射波长红移至深红色区域（>650 nm）。
为验证该类染料的应用价值，我们通过对CR1至CR3进行酯化反应，简便合成了一系列新型深红色生物成像染料（CR1E至CR3E），该系列染料可选择性靶向线粒体。相较于商用线粒体追踪探针MitoTracker，此类染料在大斯托克斯位移（Stokes shift）、优异成像对比度、高光稳定性与低细胞毒性等方面均表现更优。
此外，其香豆素基团与类罗丹明荧光发色团的荧光行为可仿照经典罗丹明进行切换，因此可作为构建荧光传感器的有效平台。基于此策略，我们构建了一种对Hg²+具有良好选择性的新型比率型荧光传感器CR1S，并成功将其应用于活A549细胞内Hg²+的成像检测。
该设计策略简洁通用，仅需引入不同荧光基团即可适配各类深红色染料与传感平台。