Epoxy and hydroxy fatty acids

In plant tissues, various enzymes convert the hydroperoxides produced by LOX to other products, some of which are important as flavor compounds. These enzymes include hydroperoxide lyase, which catalyzes the formation of aldehydes and oxo acids hydroperoxide-dependent peroxygenase and epoxygenase, which catalyze the formation of epoxy and hydroxy fatty acids, and hydroperoxide isomerase, which catalyzes the formation of epoxyhydroxy fatty acids and trihydroxy fatty acids. LOX produces flavor volatiles similar to those produced during autoxidation, although the relative proportions of the products may vary widely, depending on the specificity of the enzyme and the reaction conditions. 

Thomceus S, Carlsson AS, Stymne S. (2001) Distribution of fatty acids in polar and neutral hp-ids during seed development in Arabidopsis thaliana genetically engineered to produce acetylenic, epoxy and hydroxy fatty acids. Plant Science 161 997-1003. 

The seeds of Acacia arabica were analyzed for composition and nutritional properties. The seed contains 5.2% oil and is rich in linoleic (39.2%) and oleic (32.8%) acids. Trace levels of epoxy and hydroxy fatty acids were also detected. When animals were fed 10% seed oil in their diet, they showed poor growth performance and low feed efficiency ratios. The digestibility of the seed oil was 90% compared to 94% for peanut oil. Because of the low oil content of the seed and inferior nutritive value of the seed oil, the seed of A. arabica was not considered a prime candidate for commercial exploitation as a source of dietary fat (Maity and Mandel, 1990). 

Li, R.Z., Yu, K.S. and Hildebrand, D.F. 2010. DGATl, DGAT2 and PDATexpression in seeds and other tissues of epoxy and hydroxy fatty acid accumulating plants. Lipids. 45 145-157. 

It is generally accepted that a set of desaturases which is located in the plastids introduces double bonds into the acyl moieties bound as thioesters to acyl carrier protein or as oxoesters in monogalactosyl diacylglycerols. Both saturated and unsaturated fatty acids are translocated as acyl-CoAs from the plastids to the endoplasmic reticulum. Acyl chains of these activated fatty acids are elongated or incorporated into phosphatidylchohnes and other polar lipids II (cf. Fig. 1), and partly modified, for example, by desaturation. Further modification of acyl moieties leads to hydroxy-, epoxy-, and cyclic fatty acids. 

Fig. 3.32. Degradation of linoleic acid hydroperoxides to hydroxy-, epoxy- and oxo-fatty acids. The postulated reaction sequence explains the formation of identified products. Only segments of the structures are presented (according to Gardner, 1985)...

The cytochrome P-450 pathway enzymes (not shown on Figure 2) are monooxygenases, whereas COX is a dioxygenase (Estabrook et al, 1982). Unlike COX and lipoxygenases, cytochrome P-450 enzymes require several cofactors to metabolize fatty acids, including P-450 reductase and NADPH (Fitzpatrick and Murphy, 1989). In the presence of these cofactors and molecular oxygen, cytochrome P-450 can serve as the catalyst for the biotransformation of EPA to a variety of oxygenated metabolites (Fitzpatrick and Murphy, 1989). These include epoxides and hydroxy fatty acids such as epoxy-EPA (EEP) and HEPE (Kiss et al, 1998). 

The aliphatic components of SOM, derived from various sources, tend to persist in soil (Almendros et al. 1998 Lichtfouse et al. 1998a Lichtfouse et al. 1998b Mosle et al. 1999 Poirier et al. 2000). The principal source of aliphatic materials in soil is plant cuticular materials, especially cutin, an insoluble polyester of cross-linked hydroxy-fatty acids and hydroxy epoxy-fatty acids (Kolattukudy 2001). Some plant cuticles also contain an acid and base hydrolysis-resistant biopolymer, comprised of aliphatic chains attached to aromatic cores known as cutan (Tegelaar et al. 1989 McKinney et al. 1996 Chefetz 2003 Sachleben et al. 2004). 

Conventional breeding has developed oilseed rape (Brassica napus) cultivars that can accumulate long-chain fatty acids such as C20 1 and C22 l, however the ability to accumulate short-chain fatty acids is limited. Similarly the ability to accumulate industrially useful hydroxy fatty acids and epoxy fatty acids is also limited with conventional breeding methods. Due to its close relationship to the crucifer Arabidopsis and its associated characterised genome, and the relative ease with which genes can be inserted into Brassica species, oilseed rape is seen as a key target species for genetic manipulation. 

Figure 9.16 Chemical structure of cutin, a biopolyester mainly composed of interester-ified hydroxy and epoxy-hydroxy fatty acids with a chain length of 16 and/or 18 carbons (Ci6 and C[s class). Also, the chemical strcuture of the aliphatic monomers of suberin, derived from the general fatty acid biosynthetic pathway, namely from palmitic (16 0), stearic (18 0), and oleic acids.

Strain ALA2 converted y-linolenic acid (18 3 all ds-6,9,12) to several products including 12,17 13,17-diepoxy-6,9-octadecadienoic acid 12,13,17-trihydroxy-6(Z),9(Z)-octadecadienoic acid and 12-hydroxy-13,16-epoxy-6(Z),9(Z)-octadecadienoic acid as depicted in the top half of Figure 16.2 (Hosokawa et al, 2003a). Strain ALA2 also converted the substrate arachidonic acid (20 4 all civ-5,8,11,13) to cyclic and trihydroxyl fatty acids as reported previously (Hosokawa et al., 2003a). These reactions resulted in compounds 14,19 15,19-diepoxy-5(Z),8(Z),ll(Z)-eicosatrienoic acid 14-hydroxy-15,18-epoxy-5(Z),... 

Generation of aldehydes is discussed to be achieved by action of lyases and hydroperoxidases [198,217,218] epoxygenases are claimed to transfer IffODEs to epoxy hydroxy fatty acids [217,219] which are... 

Many of the biological actions of essential fatty acids are the result of their metabolic products, the eicosanoids. These are oxidized derivatives of AA and include prostaglandins and thromboxanes that are formed via the cyclo-oxygenase (COX) pathway, as well as hydroxy fatty acids and leukotrienes that arise by means of the lipoxygenase pathway. Another series of AA-derived products, the epoxy fatty acids, are produced by the cytochrome P450 epoxygenase pathway. The AA that serves as precursor for these reactions... 

Fig. 6. Structures of common cutin and suberin monomers, and ranges of typical composition values. Non-substituted fatty acids are not represented. There are overlaps in some classes of monomers (e.g. some monomers are epoxy hydroxy-fatty acids, of epoxy dicarboxylic acids).

The biochemical reaction catalyzed by epoxygenase in plants combines the common oilseed fatty acids, linoleic or linolenic acids, with O2, forming only H2O and epoxy fatty acids as products (CO2 and H2O are utilized to make linoleic or linolenic acids). A considerable market currently exists for epoxy fatty acids, particularly for resins, epoxy coatings, and plasticizers. The U.S. plasticizer market is estimated to be about 2 billion pounds per year (Hammond 1992). Presently, most of this is derived from petroleum. In addition, there is industrial interest in use of epoxy fatty acids in durable paints, resins, adhesives, insecticides and insect repellants, crop oil concentrates, and the formulation of carriers for slow-release pesticides and herbicides (Perdue 1989, Ayorinde et al. 1993). Also, epoxy fatty acids can readily and economically be converted to hydroxy and dihydroxy fatty acids and their derivatives, which are useful starting materials for the production of plastics as well as for detergents, lubricants, and lubricant additives. Such renewable derived lubricant and lubricant additives should facilitate use of plant/biomass-derived fuels. Examples of plastics that can be produced from hydroxy fatty acids are polyurethanes and polyesters (Weber et al. 1994). As commercial oilseeds are developed that accumulate epoxy fatty acids in the seed oil, it is likely that other valuable products would be developed to use this as an industrial chemical feedstock in the future. 

The walnut oil used in our study contained 26 6 mg naturally occurring hydroxy fatty acids. After the meal, plasma concentration of these unlabeled hydroxy fatty acids increased by 100% (from 0.54 0.07 to 1.09 0.24 imol/L, P < 0.05). One should realize that the calculation of absorption of the [U- C]-labeled hydroxy- or epoxy-fatty acids is based on the assumption that hydroxy TG are metabolized, transported in chylomicrons, and incorporated into VLDL as any other long-chain fatty acid (37). This may not be true. Less oxidized linoleic acid was incorporated into chylomicrons in CaCo-2 epithelial cells in culture (40). Thus, we may have underestimated the extent of absorption. Nevertheless, an important conclusion is that the more fat that is oxidized, the less there is to be absorbed. 

Silver ion TLC has also been used for separation of a variety of substituted unsaturated fatty acids such as epoxy, hydroxy and halohydroxy fatty acids, and these have been reviewed by Morris and Nichols (1972). Wax esters and steryl esters exhibit similar polarities and do not separate on column chromatography or normal silica TLC. Kiosseoglou and Boskou (1990) have separated the wax esters from steryl esters by using argentation TLC. They have used silica gel plates impregnated with 10% silver nitrate, with developing solvent hexane/chloroform (7 3 vol./vol.). 

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