Cereal Chem 68:115-122   |  VIEW ARTICLE

Purification and Characterization of Lipoxygenase Isozymes from Wheat Germ.

K. Shiiba, Y. Negishi, K. Okada, and S. Nagao. Copyright 1991 by the American Association of Cereal Chemists, Inc. 

To investigate the mechanism by which lipoxygenase improves the rheological properties of wheat flour doughs, wheat lipoxygenase was extracted first with acetate buffer from wheat germ and fractionated by ammonium sulfate. The lipoxygenase fraction was further purified by diethylaminoethyl (SEAE)-Sepharose chromatography and then separated into three major isozymes (L-1, L-2, and L-3) and a minor isozyme (L- a) by carboxymethyl-Sepharose chromatography. Each isozyme was purified by gel filtration and DEAE- Sepharose chromatography. Sodium dodecyl sulfate-polyacrylamide gel chromatography of these purified isozymes exhibited a single band with almost the same retardation factor. These purified lipoxygenase isozymes were characterized by enzymatic properties (pH activity profile, thermal sensitivity, substrate specificity, effect of metal ions), amino acid composition, and their effects on wheat flour. After reaction with each purified lipoxygenase isozyme, the treated flour exhibited higher foaming activity and lower contents of the sulfhydryl group than did the control flour, which received no enzyme treatment. The treated flour also showed a shorter dough development time and less resistance after peak time on a mixogram than did the control flour. The flour treated with the L-3 isozyme exhibited the highest foaming activity and bread-making quality.