A selective fluorescence-on reaction of spiro form fluorescein hydrazide with Cu(II).

The spiro form fluorescein hydrazide (1) bearing a cleavable active bond is characterized as a highly selective and sensitive fluorescence probe for Cu2+. In 0.01 M Tris-HCl buffer (pH 7.2), the probe displays a highly selective fluorescence-on response to Cu2+ only, instead of other common metal ions. The reaction mechanism has been explored and proposed as that the hydrazide group of 1 recognizes and binds Cu2+, and the subsequent complexation of Cu2+ promotes hydrolytic cleavage of the amide bond, causing the release of fluorophore (fluorescein) and thereby the retrievement of fluorescence. Conditions for fluorescence measurement were optimized, generating a calibration curve that is linear over the concentration range of 0.1-10 microM Cu2+. The detection limit for Cu2+ is 64 nM based on 11 blank determinations (k=3), also showing a highly sensitive feature. The fluorescence-on reaction described here has been used for the determination of trace Cu2+ in real biological fluids with satisfactory results.