A pH-Sensitive Spirocyclization Strategy for Constructing a Single Fluorescent Probe Simultaneous Two-Color Visualizing of Lipid Droplets and Lysosomes and Monitoring of Lipophagy

Lipid droplets (LDs) and lysosomes are crucial for maintaining intracellular homeostasis. But single fluorescent probes (SFPs) capable of simultaneous and discriminative visualizing of two organelles above and their interaction in living cells are still challenging due to the lack of rational design strategies. To break this bottleneck, herein, we develop a reliable strategy based on a pH-sensitive intramolecular spirocyclization. As a proof of concept, an SFP CMHCH, which possesses a switchable hemicyanine/spiro-oxazine moiety induced by pH, has been designed and synthesized. In acidic environments, the ring-open form CMHCH exhibits red-shift emission and low logP value, whereas the ring-closed form CMHC displays blue-shift emission and high logP value in neutral or basic environments. Thus, the distinct different hydrophilicity/hydrophobicity and absorption/emission properties of these two forms enable targeting LDs and lysosomes simultaneously and discriminatingly. Very importantly, the dynamic process of lipophagy can be directly monitored with CMHCH. The success of CMHCH indicated that the spirocyclization strategy is efficient for constructing SFPs to LDs and lysosomes.