Evaluation of inhibitory activity of aqueous, methanol and ethyl acetate extracts of Citrus aurantifolia leaf on crude xanthine oxidase isolated from cow milk

Lime (Citrus aurantifolia) is a citrus fruit from the Rutaceae family which is used in beverages, as food additives and in cosmetic industries. Citrus aurantifolia plant is used in the treatment of several complications including anemia, oxidative damages and inflammation. Xanthine oxidase is the enzyme that catalyzes the oxidation of hypoxanthine to xanthine and xanthine to uric acid. This study was aimed at determining the inhibitory activity of aqueous, methanol and ethyl acetate extract of Citrus aurantifolia leaves on xanthine oxidase isolated from cow milk. The inhibitory activities of the three extract at different concentrations (20, 40, 60, 80 and 100 µg/ml) was analyzed at 290 nm, pH 7.3 and at 25°C using spectrophotometric methods. Line Weaver-Burk plots were also used to determine enzyme kinetic constants. The aqueous extract of Citrus aurantifolia leaves has maximum velocity (Vmax); 0.05 U/ml and Michaelis-Menten constant (Km) value of 510.64 µM. The Vmax and Km value for the methanolic extract were 0.02 U/ml and 86.05 µM, while ethyl acetate extract has Vmax 0.03 U/ml and Km value of 87.84 µM. Compared to allopurinol (6.51±0.01 µg/mL), the IC50 of aqueous extract, methanol extract and ethyl acetate extracts were 14.68±0.13 µg/mL, 16.08±0.18 µg/mL and 19.13±0.25 µg/mL respectively. The study showed that the extracts can be used as a good source of natural xanthine oxidase inhibitors and this can also be used as alternative herbal drugs in the management and treatment of gout.