Multiple Factors Regulate the Spirocyclization Equilibrium of Si-Rhodamines.

Si-rhodamine has been extensively used in super-resolution fluorescence imaging in recent years. Its equilibrium between ring-closed nonfluorescent spirolactones and ring-opened fluorescent zwitterions endows Si-rhodamine with excellent fluorogenicity, membrane permeability, and photostability. In this paper, the equilibrium of Si-rhodamine between lactones and zwitterions was revealed to be greatly affected by various environmental factors, including molecular aggregation, solvent polarity, pH, metal ions, irradiation, and temperature. These environmental sensitivities make Si-rhodamine useful as a hydrochromic material, a fluorescent sensor array for metal ions or solvents, and a photoactivatable switch. Importantly, these results indicate that using Si-rhodamine as a fluorogenic probe or a blinking fluorophore in single-molecule localization super-resolution microscopy requires caution on possible false signals caused by its environmental sensitivity.