EXTRACTION, CHARACTERIZATION AND INHIBITION SENSITIVITY OF ASCORBATE PEROXIDASE FROM KIWANO (CUCUMIS METULIFERUS)

Abstract

The aim of the present study is to extract and characterize ascorbate peroxidase (APX) from kiwano (Cucumis metuliferus) and determine the effect of various inhibitors and metal ions on the enzyme activity. The enzyme activity was measured by the decrease in absorbance at 285 nm during 1 min with the spectrophotometric method. The optimum pH and temperature were found to be 6.2 and 30 degrees C respectively. Km and Vmax of APX extracted from kiwano were 0.2848 mM, 0.0035 U min-1 for aspartic acid (AsA), and 0.0649 mM, 0.0077 U min-1 for H2O2, respectively. APX extracted from kiwano was strongly inhibited by 2-nitrobenzoic acid (1.48% residual activity, 1 mM), KCN (13.3% residual activity, 1 mM), and NaN3 (25.9% residual activity, 1 mM), but less inhibited by L-cysteine, 2-mercaptoethanol, iodoacetamide, and EDTA. Analysis was performed for determining the effect of various metal ions, of which Fe2+ and Ba2+ activated the activity of APX at the concentrations of 1, 5 and 10 mM and inhibited completely by Cu2+ and Sn2+ at above 5 mM. APX extracted from kiwano lost its activity to some extent due to interacting with the rest of the metal ions. In this study, it was reported, for the first time, the extraction and biochemical characterization of the APX extracted from kiwano, which could be useful in potential applications.