OPTIMIZING ASSAY AND EXTRACTION OF LIPOXYGENASE IN WHEAT-GERM

Using three-level seven-factor response surface methodology, wheat ger m lipoxygenase (LPO) assay conditions were standardized. The important parameters were concentration of the substrate (linoleic acid), and s urfactant (Tween 20), pH and temperature. The standardized LPO assay c onditions for the 1 mL reaction volume were : 450 muM linoleic acid, 1 29 muM Tween 20, 175 mM ethanol, 1 mM EDTA, pH 6.2 (phosphate buffer), ionic strength 100 mM and 40-degrees-C. LPO extraction conditions wer e standardized by sequential variation of parameters. Optimum conditio ns were a single extraction of defatted wheat germ flour at 2-5-degree s-C with magnetic stirring of an extractant acetate buffer pH 4.5, ion ic strength 100 mM, at buffer-to-solid ratio 10:1.