Extraction, purification and characterization of Lipoxygenase from African oil bean (Pentaclethra macrophylla Benth.) seed.

Lipoxygenase (linoleate:oxygen oxidoreductase, LOX) was extracted from African oil bean seed and purified in three purification steps using ammonium sulphate precipitation, ion exchange chromatography on DEAE-Sephadex A50 column and gel filtration chromatography on Sephadex G 200. The optimum pH and temperature were investigated while the pH and temperature stability were also examined. The effect of metal ions on LOX was also investigated. The molecular weight and kinetic parameters were determined. The single protein band on SDS-PAGE suggested that the enzyme was homogenous. The molecular weight was found to be 77.54 kDa. A purification fold of 32.7 and 10.7% yield of the enzyme was achieved. The optimum temperature of the purified lipoxygenase was 40 °C and the enzyme was thermally stable at 40 °C and rapidly lost activity at 50 °C and above. The purified enzyme had an optimum pH of 6.0 and was stable at pH 5.0 and 6.0. The kinetic parameters K m and V max of the purified lipoxygenase were 0.1 mM and 94.40 μmol/min, respectively. The activity of the enzyme was enhanced in the presence of Ca²⁺ and Mg²⁺ while Zn²⁺, Cu²⁺ and Fe²⁺ significantly reduced the enzyme activity. Total inactivation of the lipoxygenase activity was observed at alkaline environment. This study confirmed lipoxygenase activity in African oil bean seed and showed properties that could be explored in improving its utilization in industrial food processing. [Display omitted] • Lipoxygenase was extracted from dormant seeds of African oil bean and purified. • The purified enzyme had temperature and pH optima of 40 °C and pH 6.0, and total inactivation in alkaline environment. • The kinetic parameters Km and Vmax of the purified lipoxygenase were 0.1 mM and 94.40 μmol/min, respectively. • Ca²⁺ and Mg²⁺ enhanced activity of the enzyme, while Zn²⁺, Cu²⁺ and Fe²⁺ inhibited the activity. • The African oil bean lipoxygenase could be exploited in industrial processes. [ABSTRACT FROM AUTHOR]