Reacction of Guava Lipoxygenase with Arachidonic Acid

• Main Author: 沈明璁
• Other Authors: 羅初英
• Format: Others
• Language: zh-TW
• Published: 1995
• Online Access: http://ndltd.ncl.edu.tw/handle/57695372864803334235

碩士 === 國立成功大學 === 化學學系 === 83 === Lipoxygenases (EC 1.13.11.12; abbrev.LOX) are a group of closely related enzymes that appear to be widely distributed in plants and mammalian tissues. This enzyme can catalyze the oxygenation ofpolyunsaturated fatty acids (PUFA) containing a cis, cis-l,4-pentadiene system. The nomenclature of the lipoxygenases is based upon the site of insertion of the oxygen molecule onto the fatty acid molecule. In the case of arachidonic acid (abbrev. AA), they have been classfied as 5-, 8-, 9-, 11-, 12-, and 15- lipoxygenase. The reaction of LOX with arachidonic acid can lead to hydroper- oxyeicosatetraenoic acids (abbrev. HPETEs) or leukotrienes (abbrev. LTs) in mammals. These eicosanoids are known to have various physiologic effects such as leukocytes agglomeration, vasoconstriction and hypersensi- tive reactions. When LOX reacts with polyunsaturated fatty acids such as linoleic acid and linolenic acid in plants, hydroperoxides of the unsaturated fatty acids are produced. These compounds play important roles in plant growth, development, senecence and defence against insects and pathogens. Thus, investigations on the various sources of LOX and their metabolites can not only result in great academic importance but also has significant economic value and human well-being. In this study, we employed guava (Psidium guajava L.) as a source of lipoxygenase. Guava LOX was extrated under potassium phosphate buffer (pH 6.0) and isolated by ammonium sulfate (65%) precipitation. This partially purified enzyme was subjectd to incubation (0℃) with AA in potassium phosphate buffer (pH 6.0). The products were extrated with chloform : methanoi (3:1, v/v) and purified with High Perfomance Liquid Chromatography (HPLC) to yield a sole product : 15-hydroperoxyei- cosatetafinoic acid (15-HPETE), in 72.7% yield. The identity of 15-HPETE was characterized by triphenylphosphine reduction to its corresponding hydroeicosatetraenoic acid (HETE), further analyzed with HPLC, and identified by GC/MS after proper derivatizations.