Abstract
Publisher Summary This chapter discusses the preparation of 15-L-hydroperoxy- 5, 8,11,13- eicosatetraenoic acid (15-HPETE) by soybean lipoxygenase. In preparation, a slurry of silicic acid in ethyl acetate in petroleum ether is prepared. The slurry is poured into glass column. Prior to application of the lipoxygenase extract to the column, the fraction of ethyl acetate in petroleum ether in the extract has to be reduced from 50% to 3%. A trace amount of [ 3 H]arachidonic acid, added as an internal standard to demonstrate the separation of fatty acid from its hydroperoxy counterpart, is eluted quantitatively with 3% ethyl acetate between fractions 1 and 20. 15-HPETE acid is eluted from the column between fractions 25 and 32 with 8% ethyl acetate in petroleum ether. A minor component, unresolved from 15-HPETE, emerges between fractions 33 and 60. TLC analysis of an aliquot of the combined fractions 25- 32, representing 18μmol ±3, showed 15-HPETE as a single component with an Rf value of 0.36.
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