Synthesis of coumarin derivatives as fluorescent probes for membrane and cell dynamics studies

• Three coumarin-derived fluorescent probes (FM1, FM2 and FM3) were synthesized.
• FM1, FM2 and FM3 localize preferentially to the plasma membranes of living cells.
• The coumarin moiety of all three probes penetrates to the glycerol level of membranes.
• The aliphatic chains of FM1 and FM3 align with the alkyl chains of the phospholipid.
• The aliphatic chain of FM2 lies parallel to the membrane surface.

Three coumarin-derived fluorescent probes, 3-acetyl-7-[(6-bromohexyl)oxy]-2H-chromen-2-one (FM1), 7-[(6-bromohexyl)oxy]-4-methyl-2H-chromen-2-one (FM2) and ethyl 2-{7-[(6-bromohexyl)oxy]-2-oxo-2H-chromen-4-yl}acetate (FM3), are described, with their photophysical constants. The compounds were tested in preliminary studies employing epifluorescence microscopy demonstrating that they allow the imaging of human neuroblastoma SH-SY5Y cell membranes. The structure of FM3 was confirmed by X-ray crystallographic analysis. Molecular dynamics (MD) simulations were used to characterize the localization and interactions of the studied compounds with a lipid bilayer model of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC).

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