One-pot synthesis of functionalised coumarin fluorophores enables rapid access to live cell bioorthogonal labelling and microenvironmental sensing agents

Coumarins are robust and versatile organic scaffolds, with broad applications in fluorescence imaging. Herein, we report a one-pot methodology to functionalised coumarins from commercially available and readily accessible intermediates. Using this streamlined approach, we synthesised a library of 7-aminocoumarins with diverse substituents and varied photophysical properties in good yields and on gram scale. To demonstrate the versatility of this methodology, we developed two functional imaging agents: CouPyC6, a mitochondrially targeted viscosity sensor for fluorescence intensity and lifetime imaging, and CouTz, the first zero-distance tetrazinyl-coumarin which can participate in fluorogenic IEDDA reactions with a range of dienophiles. The latter enables wash free, real-time bioorthogonal imaging of labelled proteins and glycan targets on the surface of live cells, offering simple access to functional coumarins for advanced imaging applications.