USE OF A CONTINUOUS ASSAY OF OXYGEN-CONSUMPTION TO EVALUATE THE PHARMACOLOGY OF 5-LIPOXYGENASE INHIBITORS

A variety of assay systems have been utilized to evaluate the inhibiti on of the key enzyme in leukotriene (LT) biosynthesis, 5-lipoxygenase (5-LO). We have developed an assay utilizing a cytosolic preparation o f 5-LO from rat basophilic leukemia (RBL-1) cells. Enzyme activity was monitored by continuous measurement of oxygen consumption. High perfo rmance liquid chromatography (HPLC) analysis of products showed exclus ive generation of 5-LO products. The assay proved useful for the evalu ation of a variety of chemical classes of lipoxygenase inhibitors and clearly differentiated those compounds which extended the lag phase (e .g. A-64077) as opposed to the propagation phase of the enzyme activit y (e.g. SK & F 105561). The data generated were in reasonable agreemen t with results from the assay of isolated human monocyte 5-LO and, wit h the exception of compounds which appear to have a significant effect on 5-LO translocation (e.g. MK-886 and Wy-49 232), inhibition of LT p roduction by intact monocytes. This assay system proved to be a conven ient and informative method to analyze inhibition of 5-LO activity.