Linoleic reactions lipoxygenase

Liquid C h rom atography/M ass Spectrometry. Increased use of liquid chromatography/mass spectrometry (lc/ms) for structural identification and trace analysis has become apparent. Thermospray lc/ms has been used to identify by-products in phenyl isocyanate precolumn derivatization reactions (74). Five compounds resulting from the reaction of phenylisocyanate and the reaction medium were identified two from a reaction between phenyl isocyanate and methanol, two from the reaction between phenyl isocyanate and water, and one from the polymerization of phenyl isocyanate. There were also two reports of derivatization to enhance either the response or structural information from thermospray lc/ms for linoleic acid lipoxygenase metabolites (75) and for cortisol (76). 

Garssen, G.J., Vliegenthart, J.F.G. and Boldingh, J. (1972). The origin and structures of dimeric fetty acids from the anaerobic reaction between soya-bean lipoxygenase, linoleic acid and its hydroperoxide. Biochem. J. 130, 435-442. 

Sekiya, J. Aoshima, H., Kajiwara, T., Togo, T. and Hatanaka, A. (1977). Purification and some properties of potato tuber lipoxygenase and detection of linoleic acid radical in the enzyme reaction. Agric. Biol. Chem. 41, 827-832. 

We previously described [25] the function of soybean lipoxygenase-1 in a biphasic system (modified Lewis cell) composed of an aqueous phase (borate buffer) and octane. The substrate of the reaction is linoleic acid (LA) and the main product is hydro-peroxyoctadecadienoic acid (LIP). The system involves two phenomena LA transfer from the organic to the aqueous phase and lipoxygenase kinetics in the aqueous medium. 

As mentioned earlier, oxidation of LDL is initiated by free radical attack at the diallylic positions of unsaturated fatty acids. For example, copper- or endothelial cell-initiated LDL oxidation resulted in a large formation of monohydroxy derivatives of linoleic and arachi-donic acids at the early stage of the reaction [175], During the reaction, the amount of these products is diminished, and monohydroxy derivatives of oleic acid appeared. Thus, monohydroxy derivatives of unsaturated acids are the major products of the oxidation of human LDL. Breuer et al. [176] measured cholesterol oxidation products (oxysterols) formed during copper- or soybean lipoxygenase-initiated LDL oxidation. They identified chlolcst-5-cnc-3(3, 4a-diol, cholest-5-ene-3(3, 4(3-diol, and cholestane-3 3, 5a, 6a-triol, which are present in human atherosclerotic plaques. 

Schnurr et al. [22] showed that rabbit 15-LOX oxidized beef heart submitochondrial particles to form phospholipid-bound hydroperoxy- and keto-polyenoic fatty acids and induced the oxidative modification of membrane proteins. It was also found that the total oxygen uptake significantly exceeded the formation of oxygenated polyenoic acids supposedly due to the formation of hydroxyl radicals by the reaction of ubiquinone with lipid 15-LOX-derived hydroperoxides. However, it is impossible to agree with this proposal because it is known for a long time [23] that quinones cannot catalyze the formation of hydroxyl radicals by the Fenton reaction. Oxidation of intracellular unsaturated acids (for example, linoleic and arachidonic acids) by lipoxygenases can be suppressed by fatty acid binding proteins [24]. 

Lipoxygenase [EC 1.13.11.12] catalyzes the reaction of linoleate with dioxygen to produce (9Z,11 )-(135 )-13-hydroperoxyoctadeca-9,ll-dienoate. This iron-depen-dent enzyme can also oxidize other methylene-interrupted polyunsaturated fatty acids. See also specific enzyme... 

Holmes et al. (1998) performed two enzymatic reactions, the lipase-catalyzed hydrolysis of y>-nitrophenol butyrate and lipoxygenase-catalyzed peroxidation of linoleic acid, in w/c microemulsions stabilized by a fluorinated two-chained sulfosuccinate surfactant (di-HCF4). The activity of both enzymes in the w/c microemulsion environment was found to be essentially equivalent to that in a water/heptane microemulsion stabilized by Aerosol OT, a surfactant with the same headgroup as di-HCF4. The buffer 2-(A-morpholino)ethanesulfonic acid (MES) was used to fix the pH in the range 5-6. 

Molecular oxygen, as distinct from reactions involving radicals or singlet oxygen, is directly inserted into free fatty acids by lipoxygenase (LOX) enzymes. Lipoxygenases, both regio- and stereospecific enzymes, react on the 1,4-pentadienyl moieties such as those of linoleic and a-linolenic acids. 

Hamberg and Samuelsson (111) had tried a number of polyunsaturated fatty acids earlier and concluded that the structural requirement for lipoxygenase attack was a cis,ci5-l,4-pentadiene group whose methylene carbon was at position a>-8. In other words the requirement was a distal double bond in position w-6. These authors also examined the modified products of the reaction by mass spectroscopy and found that in all cases O2 was inserted at position w-6 except for linoleic acid. In... 

Formation of Secondary Products and Lipohydroperoxide Destruction. As early as 1945 Holman and Burr (132) found that crude soybean lipoxygenase acting on a number of substrates produced carbonyl-containing material in addition to diene. Holman, as noted above (107), used his crystalline enzyme and found that it was difficult to establish a correspondence between O2 consumption and diene conjugation. The diene concentration always tended to be too low. Privett et al. 123) found that the reaction products varied with enzyme concentration and method of addition. Vioque and Holman 133) identified 9-keto-ll,13- and 13-keto-9,ll-octadecadienoate with the usual hydroperoxides in a reaction carried out with linoleic acid and a relatively large amount of crude soybean lipoxygenase at pH 9. 

Garssen et al. 134) observed that soybean lipoxygenase, presumably lipoxygenase-1, acting at pH 9.0 under anaerobic conditions and in the presence of linoleic acid and 13-hydroperoxy octadeca-ci5-9-trans-ll-dienoic acid i.e., the major product formed in the hydroperoxidation of linoleic acid by this enzyme) gives 13-keto-octadeca-9,ll-dienoic acid and the split products, pentane and 13-keto-trideca.-cis trans)-9-trans-ll-dienoic acid. The D-9-hydroperoxy compound cannot substitute for the L-13-hydroperoxide 135). Dimers are also formed under these conditions. These reactions do not occur under aerobic conditions. Two possible pathways for the anaerobic reaction suggested by these workers are shown in Figures 1 and 2. 

A. Arachidonic acid is produced from linoleic acid (an essential fatty acid) by a series of elongation and desaturation reactions. Arachidonic acid is stored in membrane phospholipids, released, and oxidized by a cyclooxygenase (which is inhibited by aspirin) in the first step in the synthesis of prostaglandins, prostacyclins, and thromboxanes. Leukotrienes require a lipoxygenase, rather than a cyclooxygenase, for their synthesis from arachidonic acid. 

Recently, Veldink et al.10s) reported the observation of chemiluminescence during the oxygenation of linoleic acid by lipoxygenase. This phenomenon is observed only during the aerobic phase of the reaction and is quenched by superoxide dismutase. This suggests the involvement of superoxide in this process, which is not unreasonable considering the participation of ferrous iron and free radicals in the enzyme reaction. 

The hypothesis that HTPLO and SLO were able to mediate the N-demethyl-ation of selected phenothiazines and insecticides in the presence of polyunsaturated linoleic acid (LA) was proposed by Hover and Kulkarni [163]. This N-demethylation reaction might be limited by the incubation time, pH of the medium and concentration of the enzyme and substrate. The reaction was followed by measuring the formaldehyde production. The results confirmed that the polyunsaturated free fatty acids could support the N-demethylation of xenobiotics via the lipoxygenase pathway. 

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