A Colorimetric Method for the Determination of Lipoxygenase Activity Suitable for Use in a High Throughput Assay Format

A rapid and sensitive method for measuring the activity of lipoxygenase is described. The assay is based on the principle that under acidic conditions a lipid hydroperoxide can oxidize Fe2+ to Fe3+ which then oxidizes xylenol orange to form a product that absorbs strongly in the visible region (see Z-Y. Jiang, A. C. S. Woollard, and S. P. Wolff, Lipids 26, 853-856, 1991). This methodology was modified to measure lipoxygenase activity and the system was optimized using platelet 12-lipoxygenase. This assay is suitable for use in a 96-well microtiter plate and may be utilized as a high throughput screen for the identification of novel lipoxygenase inhibitors.