Facile synthesis of green-light and large Stokes-shift emitting coumarins for bioconjugation

Bright and hydrosoluble fluorescent dyes with large Stokes shifts are widely used as molecular probes and light-emitting markers in chemistry and biology. In this report, one-step synthesis of a bright green-emitting coumarin (Φ = 0.52, λem = 505 nm) was achieved from available reagents under neat conditions. The novel dye 1 showed very large Stokes shift (Δλ = 138 nm) due to strong intramolecular charge transfer (ICT) effect, which was confirmed by density functional theory (DFT) calculations. Dye 1 was hydrosoluble and displayed strong emission at a wide range of pH from 3 to 8. Importantly, the secondary amine in 1 made it extremely convenient for futher bioconjugation. Representative derivatives were showcased for various successful applications of fluorogenic enzyme assay, biolabelling and mitochondrial staining.