Purification and identification of linoleic acid hydroperoxides generated by soybean seed lipoxygenases 2 and 3

It has been known that lipoxygenase (LOX) isozymes exhibit differences in product formation, but most product information to date is for LOX 1 among soybean (Glycine max) LOX isozymes. In this study, LOXs 2 and 3 were purified and used to generate hydroperoxide (HPOD) products in an in vitro system using linoleic acid as a substrate in the presence of either air or O2. The products were analyzed to determine their stereoisomeric characteristics. The control (no enzyme) showed only low levels of hydroperoxide production and no stereoisomeric specificity. LOX 2 shows high specificity in product formation, producing roughly 4 times more 13-HPOD than 9-HPOD, nearly all of which was 13-S(Z,E)-HPOD. LOX 3 produced more 9-HPOD than 13-HPOD at approximately a 2:1 ratio. No single stereoisomer was predominant among LOX 3 products. These results demonstrate that different isozymes of LOX have characteristic product profiles in in vitro reactions.