Study of tetra-substituted amino aluminum phthalocyanine as a new red-region substrate for the fluorometric determination of peroxidase and hydrogen peroxide

This is a first report on a new promising red-region fluorescence substrate, tetra-substituted amino aluminum phthalocyanine, which displays an excitation maximum at 610 nm and an emission maximum at 678 nm in strongly acidic medium. It has been synthesized and applied to the determination of traces of hydrogen peroxide and horseradish peroxidase (HRP). The steady-state catalytic rate depends upon the enzyme and substrate concentrations, and the Michaelis-Menten parameters K_m and V_max are measured to be 2.82 × 10^-6moll^-1 and 6.0 × 10^-9moll^-1s^-1, respectively. Under optimum conditions, the calibration graph has a linear range of 0.0 to 3.94 × 10^-11moll^-1 HRP and 0.0 to 2.0 × 10^-7moll^-1 hydrogen peroxide, with 3σ detection limits of 5.9 × 10^-13moll^-1 HRP and 1.4 × 10^-9moll^-1 H₂O₂. By coupling with a glucose oxidase-catalyzed reaction, glucose in human serum has been quantified and the results are in good agreement with those reported by a hospital laboratory. The proposed method can greatly decrease the interference that results from background fluorescence or scattered light and has a high analytical sensitivity.