A practical CO₂-mediated synthesis of 5,6-carboxylated silicon-rhodamines for targeted probe development

Silicon-rhodamine (SiR) dyes are among the most important fluorophores for super-resolution imaging owing to their far-red emission, high photostability, and tunable lactone-zwitterion equilibrium. However, their broader application in targeted probe development has been limited by the challenging synthesis of 5,6-carboxylated SiR derivatives, which are essential intermediates for bioconjugation. Here, we report a practical CO₂-mediated strategy for the synthesis of 5,6-carboxylated SiRs from brominated SiR precursors via lithium-halogen exchange and direct carboxylation. Using n-BuLi and readily available CO₂, this method delivers carboxylated SiR derivatives in 60-93% yields while avoiding the use of t-BuLi, toxic CO, and expensive palladium catalysts. In addition, the crude carboxylation mixtures can be directly subjected to amide coupling without chromatographic purification, enabling one-pot access to HaloTag-targeted SiR probes. The resulting probes were successfully applied to long-term live-cell super-resolution imaging, allowing visualization of filopodial dynamics and mitochondrial remodeling. This work establishes a concise and efficient route to functionalized SiR dyes and facilitates the rapid development of targeted SiR probes.