Submitted to: Lipids
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/20/2001
Publication Date: 5/20/2002
Citation: NUNEZ, A., FOGLIA, T.A., PIAZZA, G.J. PURIFICATION OF LIPOXYGENASE FROM CHLORELLA: PRODUCTION OF THE 9 AND 13-HYDROPEROXIDE DERIVATES OF LINOLEIC ACID. LIPIDS. 2002. v. 37. p. 1027-1032. Interpretive Summary: Fats and oils are abundantly produced in the United States, but their domestic food usage has remained stagnant. Accordingly, new outlets for domestic fats and oils need to be identified, particularly in non-food uses, which also would decrease our need to import non-domestic oils. Fatty acids common to most domestic fats and oils can be converted to derivatives that can act as substitutes for currently used industrial products that are presently derived from such imported oils as castor oil and petroleum. In this study we have identified and characterized a micro- algae enzyme system that can convert the unsaturated fatty acids commonly found in domestic oils, e.g., soy and corn oils, into products that can be substituted for products that are widely used in industrial applications, such as surfactants and polymers. The development of this biotechnological approach of producing these materials from domestic oils has the potential of not only reducing our dependence on imported oils but of providing a more benign environmental process for producing these materials.
Technical Abstract: Oxygenation of linoleic acid by the enzyme lipoxygenase (LOX) present in the micro-alga Chlorella pyrenoidosa is reported to produce the corresponding 13- and 9-hydroperoxide derivatives of linoleic acid (13-HPOD and 9-HPOD, respectively). Previous work with this micro-alga indicated that partially purified LOX present in the 30-45% and 45-80% saturated ammonium sufate precipitate produced both HPOD isomers in different ratios It was not clear, however, if the observed activity in the two isolates represented the presence of one or more isozymes. In the present work, LOX activity isolated from the intracellular fraction of C. pyrenoidosa and fusca was purified by 35-80% saturated ammonium sufate precipitation, ion exchange and hydrophobic interaction chromatography to apparent homogeneity. SDS-PAGE analysis of the purified LOX, however, showed two protein bands in equal amounts with apparent molecular weights of 34 and 47 7kDa, respectively. Purification of the enzyme by size exclusion chromatography, indicated a single monomeric LOX with a molecular weight of approximately 47 kDa. This isolated protein produced both 13-HPOD and 9- HPOD from linoleic acid in equal amounts. This HPOD product ratio was not altered by pH in the range of 6 to 9, with the optimal activity of LOX at pH 7.8.