Dibutylated Si-fluorescein: enhanced hydrophobicity for fluorogenic labeling in vivo

Fluorescein derivatives, characterized by a xanthene backbone, represent a prominent class of organic molecular dyes. Through silicon atom substitution, silicon-substituted fluoresceins (SiFs) exhibit bathochromic emission spectra with high quantum yields. However, the anionic configuration significantly restricts their application in vivo. Herein, we reported dibutylated silicon fluorescein, SiFC4 and HSiFC4. With increased hydrophobicity by dibutylation, SiFC4 tends to convert into the neutral lactone form and enables its aggregation (150 nm in diameter) with substantially reduced background fluorescence at physiological pH. Notably, HSiFC4 demonstrates 4.3-fold enhanced fluorogenicity in HaloTag labeling systems and robust fluorescence response (4.1-fold) in living cell imaging compared to its dimethyl derivative, HSiFC1. This approach underscores the potential of alkylation in modulating the properties of fluorophores, offering a promising strategy for advanced applications in the future.