Bifunctional coumarin derivatives in solution and solid phase synthesis offluorogenic enzyme substrates

The use of the fluorescent bifunctional compounds 7-amino-4-coumarinyl-acet ic acid 1, 7-hydroxy-4-coumarinyl-acetic acid 2 and ethyl 7-amino-4-coumari nyl-acetate 3 in solution and solid phase synthesis of fluorogenic enzyme s ubstrates was examined. The intramolecularly quenched fluorogenic substrate N-(7-amino-4-coumarinyl-acetyl)-L-phenylalanyl-p-nitroanilide 5, and the f luorogenic one ethyl 7-(glutaryl-L-phenylalanilamido)-4-coumarinyl-acetate 8, both suitable for chymotrypsin and/or chymotrypsin like enzymes determin ation, were prepared in solution. The substrates 7-oleyloxy-4-coumarinyl-ac etic acid 13 and 7-palmitoyloxy-4-coumarinyl-acetic acid 14, suitable for t he enzymatic study of lipases, were prepared by solid phase technique using 2-chloro-chlorotrityl-resin. The study of the fluorescence properties of t he fluorophores 1, 2, 3, and substrates 5, 8, 13, 14 showed that the examin ed bifunctional coumarin derivatives are suitable markers for solution and solid phase synthesis of fluorogenic enzyme substrates.