Dietary arachidonic acid and hepatic desaturation of fatty acids in obese zucker rats

The effect of low levels of dietary arachidonic acid (20:4n-6) on Δ6 desaturation of linoleic acid (18:2n-6) and α-linolenic acid (18:3n-3), and on Δ5 desaturation of dihomo-γ-linolenic acid (20:3n-6) were studied in liver microsomes of obese Zucker rats, in comparison with their lean littermates. Fatty acid composition of serum total lipids and of phospholipids from liver microsomes and from total heart and kidney was determined to see whether modifications of desaturation rate, if any, were reflected in the tissue fatty acid profiles. Animals fed for 12 wk on a balanced diet, containing 20:4n-6 and 18:2n-6, were compared to those fed 18:2n-6 only. The low amount of dietary 20:4n-6 greatly inhibited Δ6 desaturation of 18:2n-6 and Δ5 desaturation of 20:3n-6, whereas Δ6 desaturation of 18:3n-3 was slightly increased in obese rats. Inhibition of the biosynthesis of long-chain n-6 fatty acids by dietary arachidonic acid was only slightly reflected in the 20:4n-6 content of liver microsome phospholipids. On the contrary, the enrichment of serum total lipids and heart and kidney phospholipids in this fatty acid was pronounced, more in obese than in lean animals. Our results show that, although the desaturation rate of the n-6 fatty acids in liver microsomes was greatly decreased by the presence of arachidonic acid in the diet, the tissue phospholipid content in arachidonic acid was not depressed. The potentiality of synthesis of eicosanoids of the 2 family from this fatty acid is consequently not lower, especially in obese rats, in which certain tissues are deficient in arachidonic acid, in comparison with their lean littermates.     

Distribution of (n-9) and (n-7) Isomers of Monounsaturated Fatty Acids in Indian Mustard (Brassica juncea)

The seed oils of 19 Indian mustard varieties were analyzed for fatty acid composition using GC–MS, reporting the presence of n-7 isomers of C18:1, C20:1 and C22:1 fatty acids. n-7 isomers namely cis-vaccenic (C18:1), 11, eicosenoic(C20:1) and 13-cis-docosenoic (C22:1) acids ranged from 0.61 to 1.73, 1.04 to 1.69 and 0.58 to 1.17 %, respectively. The average values of C20:1(n-7) was highest (1.36) amongst the three acids. Nervonic acid was also reported in the range of 0.69 to 2.52 %. The ratios of (n-7)/(n-9) ranged from 6.22 to 14.62, 12.38 to 27.35 and 2.01 to 3.24 % for C18:1, C20:1 and C22:1 fatty acids, respectively. The variety RLM-619 had the lowest elongation ratio (ER) as 0.44 and the highest desaturation ratio (DR) as 0.41 indicating higher efficiency of the desaturation pathway than for other varieties. The oleic desaturation ratio (ODR) values varied from 0.68 (Basanti) to 0.75 (GM-2) with a mean value of 0.72 and linoleic desaturation ratio from 0.40 (Basanti) to 0.49 (Pusa Bold) with a mean value of 0.45. Palmitoleic acid showed positive correlation with C18:1(n-7), C20:1(n-7), C20:2, C22:0, C22:1(n-7), C24:1, (n-7)/(n-9) ratio of C18:1 and C22:1 and a positive trend with ER but a significant negative correlation with C18:3, DR and ODR. The results indicated that palmitoleic acid is an important intermediate component in the synthesis of long chain (n-7) fatty acids.     

δ8 desaturationin vivo of deuterated eicosatrienoic acid by mouse liver

In vitro evidence has been reported for an alternate pathway that involves δ8 desaturation of n-6 and n-3 polyunsaturated fatty acids (PUFA). The present study was designed to allow detection of δ8 desaturationin vivo and to provide an estimation of the relative contribution of δ8 desaturation to thein vivo synthesis of n-3 fatty acids. Male adult ICR mice were fed a semisynthetic fat-free diet for eight days, and then the diets were supplemented for three days with deuterated 11,14,17-eicosatrienoic acid (20:3-d₈) labeled at the 3,3,4,4,8,8,9,9 carbon positions. Analysis of liver total lipid by gas chromatography/mass spectroscopy indicates that the total deuterated fatty acids contained 22.3% 20:3n-3-d₈ and 28.9% of metabolites formed by elongation and δ5 desaturation of 20:3n-3-d₈. Deuterated metabolites resulting from retroconversion to 18:3-d₆ and subsequent metabolism by classical pathways represented 35.3% of the total deuterated fatty acids. The retroconversion product (18:4n-3-d6) of 20:4n-3-d₆ and/or-d₈ was 9.0% of the total. A minor percentage (4.4%) of the products identified (20:4n-3-d₆, 20:5n-3-d₆, 22:5n-3-d₆, 22:6n-3-d₅ and 24:6n-3-d₅) were formed by δ8 desaturation. This study provides the first clear evidence of δ8 desaturationin vivo in the mouse liver. Whether δ8 desaturation would have a greater importancein vivo when the δ6 desaturase pathway is disrupted remains to be determined.     

Role of essential fatty acids in the function of the developing nervous system

The basis for n-3 fatty acid essentiality in humans includes not only biochemical evidence but functional measures associated with n-3 deficiency in human and nonhuman primates. Functional development of the retina and the occipital cortex are affected by α-linolenic acid deficiency and by a lack of docosahexaenoic acid (DHA) in preterm infant formulas and, as reported more recently, in term diets. Functional effects of n-3 supply on sleep-wake cycles and heart rate rhythms support the need for dietary n-3 fatty acids during early development. Our results indicate that n-3 long-chain polyunsaturated fatty acids should be considered provisionally essential for infant nutrition. DHA may also be required by individuals with inherited metabolic defects in elongation and desaturation activity, such as patients with peroxisomal disorders and some forms of retinitis pigmentosa.     

Norflurazon—An inhibitor of essential fatty acid desaturation in isolated liver cells

Norflurazon is a herbicide known to inhibit carotene biosynthesis and linolenic acid biosynthesis in plants. In the present work, the effect of norflurazon on the metabolism of essential fatty acids was studied in isolated rat liver cells and in rat liver microsomes, incubated with [1-¹⁴C] labeled linolenic acid (18∶3, n−3), dihomogammalinolenic acid (20∶3, n−6) and eicosapentaenoic acid (20∶5, n−3). Norflurazon (0.1 mM, 1.0 mM) was found to inhibit essential fatty acid desaturation. The Δ6 desaturation is inhibited more efficiently than the Δ5 and Δ4 desaturation. The chain elongation of essential C₁₈ fatty acids to their C₂₀ and C₂₂ homoglogs was not inhibited by norflurazon.     

Aktuelle Beurteilung ernährungswissenschaftlicher Aspekte der Linolensäuren

Current Views of Nutritional Aspects of Linolenic Acids The linolenic (n-3) type of fatty acid has many metabolic and chemical properties similar to those of the linoleic (n-6) type. Activation to CoA esters and esterification to lipids occurs relatively independent of the n-3 or n-6 structure. Both types of dietary polyunsaturated fatty acid appear able to reduce serum cholesterol levels and diminish some risk factors associated with cardiovascular disease. Nevertheless, the presence of the added terminal double bond in the n-3 fatty acids diminishes their reactivity in physiological conditions with the cyclooxygenase enzyme that forms prostaglandins. Also the formation of leukotriene derivatives from 20:5n-3 appears decreased relative those from 20:4n-6. This lower reactivity of n-3 acids also is associated with a lower ability to relieve the symptoms of essential fatty acid deficiency in experimental animals. The n-6 acids are thus defined as the essential fatty acids that physiologically form the active eicosanoids. Because villagers traditionally ingesting maritime foods have decreased incidence of cardiovascular disease and thrombotic deaths, current attention is directed to the dietary balance of n-3 and n-6 fatty acids. Recent concepts of platelet function and dietary lipids in coronary thrombosis, vasospasm and angina (Herz 5, 34–41 [1980]) suggest that some dietary balance between the n-3 and n-6 polyunsaturated fatty acids may be beneficial in decreasing the development of coronary artery disease and thromboembolisms.     

Fatty acid specificity in the inhibition of cell proliferation and its relationship to lipid peroxidation and prostaglandin biosynthesis

Primary cultures of smooth muscle cells were established from the medial layer of guinea pig aorta. Cells at passage level 4 were treated with different series of fatty acids belonging to the n-9, n-6 and n-3 families. Lipid peroxidation was measured by the thiobarbituric acid assay and prostaglandin biosynthesis was measured by the radioimmunoassay of PGE and 6-keto-PGF_1α. Cell proliferation was estimated from the total cell number of cultures seeded at low density. 18∶1(n-9) did not form lipid peroxides and this fatty acid stimulated cell proliferation. All fatty acids which generated lipid peroxides inhibited cell proliferation, but inhibition was correlated with the degree of lipid peroxidation only in the n-9 fatty acid family. 22∶4(n-6) and 22∶6(n-3) inhibited prostaglandin biosynthesis. 18∶2(n-6), 18∶2(n-9), 18∶3(n-3), 20∶2(n-9), 20∶3(n-3) and 20∶5(n-3) had no effect on prostaglandin biosynthesis. 18∶3(n-6), 20∶3(n-6) and 20∶4(n-6) generated prostaglandins. 20∶3(n-9) generated metabolites with prostaglandin immunoreactivity. The inhibition of cell proliferation did not correlate with enhanced or inhibited prostaglandin synthesis. The inhibition of cell proliferation was related to the structures of the different polyunsaturated fatty acid families decreasing in the order n-9>n-6>n-3. Eicosatrienoic acids were the most effective inhibitors of cell proliferation in each fatty acid family and 20∶3(n-9) was the most potent eicosatrienoic acid. These data show that specific as yet unrecognized products of fatty acid metabolism are responsible for the inhibition of cell proliferation. Fatty acids are designated by the number of carbon atoms: number of double bonds and the position of the first double bond from the methyl terminus of the acyl chain is noted in parenthesis: 18∶1(n-9), 9-octadecenoic acid; 18∶2(n-9), 6,9-octadecadienoic acid; 18∶2(n-6), 9,12-octadecadienoic acid; 18∶3(n-6), 6,9,12-octadecatrienoic acid, 18∶3(n-3), 9,12,15-octadecatrienoic acid; 20∶2(n-9), 8,11-eicosadienoic acid; 20∶3(n-9), 5,8,11-eicosatrienoic acid; 20∶3(n-6), 8,11,-14-eicosatrienoic acid, 20∶4(n-6), 5,8,11,14-eicosatetraenoic acid; 20∶5(n-3), 5,8,11,14,17-eicosapentaenoic acid; 22∶4-(n-6), 7,10,13,16-docosatetraenoic acid, 22∶6(n-3), 4,7,10,13,16,19-docosahexaenoic acid. Presented at the 73rd AOCS annual meeting, Toronto, Canada, May 1982.     

Comparative effect of glucagon,dibutyryl cyclic AMP,and epinephrine on the desaturation and elongation of linoleic acid by rat liver microsomes

The effect of glucagon, dibutyryl cyclic adenosine 3′,5′-monophosphate, and epinephrine on the biosynthesis of polyunsaturated fatty acids of the linoleic acid family was studied. The incubations were performed with rat liver microsomes and labeled linoleic acid under desaturating and elongating conditions. Under desaturating conditions, linoleic acid was converted to γ-linolenic acid, whereas under elongating conditions it was converted to 20∶2ω6. Glucagon, dibutyryl cyclic AMP, and epinephrine decreased the oxidative desaturation of linoleic acid to γ-linolenic acid while the elongating reaction was not modified in the experimental conditions tested. Consequently, the results support the hypothesis that the oxidative desaturation of linoleic acid to γ-linolenic acid is the main controllable step in the biosynthesis of polyunsaturated fatty acids of the linoleic acid family in the microsomes.     

Cessation of polyunsaturated fatty acid formation in four selected filamentous fungi when grown on plant oils

Four fungi,Conidiobolus nanodes, Entomophthora exitalis, Mortierella isabellina, andMucor circinelloides, were grown on various oils (triolein, sesame, safflower, linseed, and oil fromM. isabellina) and produced lipids in which the fatty acids were predominantly the same as those of the original staring substrate. Only in the first two cases was there evidence of a small amount of chain elongation and of fatty acid desaturation taking place. The extent of this was only about 10% of that seen in glucose-grown cells. The apparent repression of the fatty acid desaturases and elongases was not reversed by growing cells on glucose and oils as mixed substrates—the fatty acid profiles were the same as when the fungi had grown in oils alone. Neither was the cessation of polyunsaturated fatty acid synthesis due to the presence of nonoil components (NOC) in the oil. Only the NOC from sesame oil affected one single conversion, that of 20∶3n-3 to 20∶4n–6. We conclude that fatty acid desaturase and elongase systems are repressed either partially or completely in a filamentous fungi grown on triacylglycerol oils.     

Composition of muscle tissue lipids of silver carp and bighead carp

Lipids have a complex role in the nutritional value of food. Some polyunsaturated fatty acids, characterized as essential, are extremely important for human health. This is primarily related to α-linolenic acid (18:3n-3), eicosapentaenoic acid (20:5n-3) and docosahexaenoic acid (22:6n-3). Content of polyunsaturated n-3 fatty acids is usually much higher in lipids of marine fish than in freshwater fish. Previous investigations have shown that muscle tissue of silver carp and bighead carp from fish farms may be a rich source of essential fatty acids. Because of that, the objective of this work was to examine contents and composition of fatty acids and total lipids in the muscle tissue of silver and bighead carp, with the aim to find out whether there are significant differences in this respect between the two species and to what extent the harvest season can influence the composition of lipids in these freshwater fish. This study showed that there is no significant difference either in the content of polyunsaturated n-3 and n-6 fatty acids, or in the n-6/n-3 fatty acid ratio in these two fish species. The lipids of both the silver and bighead carp from the spring harvest have significantly higher contents of the n-3 acids and a significantly lower n-6/n-3 ratio than fish from the autumn harvest.     

Biosynthesis of Long Chain Polyunsaturated Fatty Acids in the Marine Ichthyosporean Sphaeroforma arctica

Sphaeroforma arctica is a unique, recently discovered marine protist belonging to a group falling close to the yeast/animal border. S. arctica is found in cold environments, and accordingly has a fatty acid composition containing a high proportion of very long chain polyunsaturated fatty acids, including the ω3 polyunsaturated fatty acids eicosapentaenoic acid (EPA) and docosahexanoic acid (DHA). Two elongases and five desaturases, representing the complete set of enzymes necessary for the synthesis of DHA from oleic acid, were isolated from this species and characterized in yeast. One elongase showed high conversion rates on a wide range of 18 and 20 carbon substrates, and was capable of sequential elongation reactions. The second elongase had a strong preference for the 20-carbon fatty acids EPA and arachidonic acid, with over 80 % of EPA converted to docosapentaenoic acid (DPA) in the heterologous yeast host. The isolation of a Δ8-desaturase, along with the detection of eicosadienoic acid in S. arctica cultures indicated that this species uses the alternate Δ8-pathway for the synthesis of long-chain polyunsaturated fatty acids. S. arctica also carried a Δ4-desaturase that proved to be very active in the production of DHA from DPA. Finally, a long chain acyl-CoA synthetase from S. arctica improved DHA uptake in the heterologous yeast host and led to an improvement in desaturation and elongation efficiencies.     

Significance of docosahexaenoic acid for rainbow trout (Oncorhynchus mykiss) larvae

N-3 fatty acids are essential for rainbow trout (Oncorhynchus mykiss). Our investigations specify the fatty acid requirements of rainbow trout larvae. One group of larvae was fed with Artemia ssp. rich in linolenic acid (18:3 n-3) and octadecatetraenoic acid (18:4 n-3), while the other group received a commercial fish feed containing high levels of long-chain polyunsaturated fatty acids (PUFA), mainly docosahexaenoic acid (22:6 n-3). After two and four weeks of feeding, the growth and fatty acid pattern of the larvae were determined. The fatty acid composition of the diet is reflected in the triglyceride composition of the fish, but there is no sign of conversion of 18:3 n-3 and 18:4 n-3 into long-chain polyunsaturated fatty acids. This is caused by limited chain elongation rather than by low enzyme activity of desaturase. In the triglycerides of the larvae, high levels of 18:4 n-3 were determined. This fatty acid was not transformed into the corresponding long-chain polyunsaturated fatty acids. Deficiency of 22:6 n-3 resulted in reduced larval growth. Therefore, it can be stated that docosahexaenoic acid is essential for rainbow trout larvae.     

Δ6 Desaturase mRNA Abundance in HepG2 Cells Is Suppressed by Unsaturated Fatty Acids

The effect of unsaturated fatty acids on the abundance of Δ6 desaturase (D6D) mRNA and the fatty acid composition of HepG2 cell membranes was examined. Supplementation of HepG2 cells with oleic acid (18:1n-9, OA), linoleic acid (18:2n-6, LA), α-linolenic acid (18:3n-3, ALA), arachidonic acid (20:4n-6, AA) or eicosapentaenoic acid (20:5n-3, EPA) reduced D6D mRNA abundance by 39 ± 6.6, 40 ± 2.2, 31 ± 5.2, 55 ± 4.8, and 52 ± 5.0%, respectively, compared with control cells. Despite the reduction in D6D mRNA abundance, the level of D6D conversion products (20:3n-9, EPA and AA) in OA, ALA and LA supplemented cells, respectively, was elevated above that in control cells. Our results suggest that although unsaturated fatty acids decrease the abundance of D6D mRNA by as much as 50%, the conversion of polyunsaturated fatty acids and accumulation of long chain polyunsaturated fatty acids (LCPUFA) in HepG2 cell phospholipids continues to occur.     

Fatty Acid Composition of the Brain,Retina, Liver and Adipose Tissue of the Grey Mouse Lemur (<Emphasis Type="Italic">Microcebus murinus</Emphasis>, Primate)

The particular interest in supplementing human foods with n-3 fatty acids has arisen from the findings that this series of polyunsaturated fatty acids (PUFA) have an impact on neuronal functions. Indeed vertebrates, including humans, preferentially use docosahexaenoic acid (DHA, 22:6n-3) over other long-chain n-3 PUFA for the genesis of their neuronal and retinal membranes. The grey mouse lemur is a nocturnal prosimian primate originating from Madagascar. The increased use of this omnivorous primate in nutritional studies (chronic caloric restriction, n-3 fatty acids supplementation), justifies the interest of determining their fatty acids body composition. In the present study, we report the fatty acid composition in lipid classes from the main target tissues (brain, retina, liver and adipose tissue) of six adult mouse lemurs raised under laboratory nutritional conditions. Among the main findings, n-6-docosapentaenoic acid (n-6-DPA; 22:5n-6) is very low in the brain cortex and retina, whereas there is a very high accumulation of docosahexaenoic acid (DHA, 22:6n-3) in the neural tissues compared to liver and plasma. In particular, DHA accounts for about one half of the total fatty acids in the retina ethanolamine glycerophospholipids. This high concentration clearly indicates that DHA is efficiently transferred from blood lipids to the outer segment of the mouse lemur retina. We conclude that the mouse lemur n-3 PUFA metabolism efficiently drives DHA to neural tissues, through the blood-brain barrier and the blood-retina barrier.     

Characteristics of the Fatty Acid Composition of a Deep-Sea Vent Gastropod, <Emphasis Type="Italic">Ifremeria nautilei</Emphasis>

Neutral and polar lipids in the soft parts of a gastropod species, Ifremeria nautilei, collected from deep-sea hydrothermal vents, were examined to assess the trophic relationships in hydrothermal vents. The vent gastropod obtains many of its lipids from symbiotic chemosynthetic microorganisms. The major polyunsaturated fatty acids (PUFA) both in the triacylglycerols and phospholipids of the gastropod consist of a limited number of n-3 and n-6 PUFA: arachidonic acid (20:4n-6), icosapentaenoic acid (20:5n-3), and docosapentaenoic acid (22:5n-3), without docosahexaenoic acid (DHA, 22:6n-3). Noticeable levels of various n-6 PUFA, such as 18:2n-6,9, 20:2n-6,9, 20:3n-6,9,12, and 20:3n-6,9,15 with significant levels of 16:1n-6 and 18:1n-6 indicate the biosynthetic characteristic of the endosymbionts. The lack of DHA in all specimens suggests a limitation of its lipid biosynthesis ability with its symbionts. This finding with regard to the lipids is unusual for a marine animal in the grazing or detrital food chain because many marine animal lipids evidently contain high levels of DHA with low levels of n-6 fatty acids. Such contradictory findings lead to some new insights into the absence of a biosynthetic pathway for DHA in I. nautilei, and provide evidence that DHA in this species is dispensable. Similar to herbivorous gastropods, the lack of DHA with significant levels of n-6 PUFA in this species also indicates its selective assimilation of specific microorganisms, such as chemosynthetic bacteria in hydrothermal vents, because significant levels of DHA were found in carnivorous mollusk lipids.     

Peroxisomal metabolism of adrenic acid; No Δ4 desaturase detected in rat liver peroxisomes

The existence of a peroxisomal Δ4 desaturation of 22:4n-6 and 22:5n-3 to yield, respectively, 22:5n-6 and 22:6n-3 has been questioned. An alternative pathway has been formulated to include microsomal chain elongation and Δ6 desaturation and peroxisomal chain shortening. We incubated [1-¹⁴C]adrenic acid (22:4n-6) in a system for desaturation (i.e., in the presence of NADH) with purified rat liver peroxisomes. The fatty acids were separated as methyl derivatives by high-performance liquid chromatography. Four ultraviolet-absorbing product peaks appeared, three of which contained radioactivity, which we investigated as methyl, trimethylsilyl, and oxazoline derivatives on gas chromatography-mass spectrometry. In addition to adrenic and arachidonic acids, the product peaks were trans-enoyl, hydroxy, and keto derivatives of adrenic acid: the three first steps of β-oxidation cycle. This indicated that the NAD-dependent dehydrogenase step in the peroxisomal β-oxidation cycle of adrenic acid was inhibited due to a high concentration of added NADH. Incubation in the presence of NAD instead of NADH reduced radioactivity in the peaks that corresponded to intermediates, while radioactivity in the acid-soluble fraction increased considerably, consistent with a complete β-oxidation cycle of adrenic to arachidonic acid. There were no indications of Δ4 desaturation in purified peroxisomes incubated in a standard desaturation system. Instead, adrenic acid as substrate underwent β-oxidation.     

Effects of Cigarette Smoke on Cell Viability,Linoleic Acid Metabolism and Cholesterol Synthesis,in THP−1 Cells

Cigarette smoke (CS) contains thousands of substances, mainly free radicals that have as a target the polyunsaturated fatty acids (PUFA). Long chain PUFA are produced through elongation and desaturation reactions from their precursors; the desaturation reactions are catalyzed by different enzymes: the conversion of 18:2n-6 (linoleic acid, LA) to 18:3n-6 by Delta6 desaturase, while that of 20:3n-6 to 20:4n-6 by Delta5 desaturase. The aim of this work is to evaluate the effect of serum exposed to cigarette smoke (SE-FBS) on (1) cell viability and proliferation, (2) [1-(14)C] LA conversion and desaturase activities in THP-1 cells, a monocytic cell line. In THP-1, CS inhibits cell proliferation dose-dependently, by producing a modification in the cell cycle with a reduced number of cells in synthesis and mitosis phases at higher concentrations. CS also decreases [1-(14)C] LA conversion to its derivatives in a concentration-dependent manner, inhibiting the activities of Delta6 and mainly Delta5 desaturase. In addition, CS does not modify the incorporation of LA into various lipid classes but it reduces cholesterol synthesis from radiolabelled acetate, and increases free fatty acid, TG and CE levels. In conclusion, CS affects lipid metabolism, inhibiting LA conversion and desaturase activities. CS also shifts the "de novo" lipid synthesis from free cholesterol to TG and CE, where LA is preferentially esterified.     

Exogenous Arachidonate Restores the Dimethoate-Induced Inhibition of Steroidogenesis in Rat Interstitial Cells

The present work studies the potential restorative effect of polyunsaturated fatty acids (PUFA, 5 μM/24 h) on the dimethoate (DMT)-induced inhibition of testosterone biosynthesis in Leydig cells isolated from rat testes. Various fatty acids (FA) from the n-6 (18:2, 20:3, 20:4, 22:4 and 22:5) and n-3 (18.3, 20:5, 22:5, 22:6) series were assayed in Leydig cells, alone (as delipidated BSA complexes) and in combination with DMT (1 ppm). The n-6 FA stimulated lipid peroxidation (LPO) and inhibited the activities of steroidogenic enzymes (3β- and 17β-hydroxysteroid dehydrogenases). The n-3 FA exerted an anti-oxidant effect, decreasing the production of thiobarbituric-acid reactive substances (TBARS) and inhibiting phospholipase A(2) activity. The biosynthesis of testosterone in DMT-treated cultures was completely normalized by ARA (20:4n-6) and partially restored by the addition of 20:3n-6, increasing ARA content inside the mitochondria. The other FA assayed failed to restore androgenesis. COX-2 protein and prostaglandin F2α and E2 production were stimulated by 20:3n-6, ARA, 18:3n-3 and 20:5 n-3. COX-2 protein decreased upon addition of 22:5n-3 and 22:6n-3. StAR protein was increased by ARA and partially increased by 20:3n-6, likely due to its metabolic conversion into ARA. Both FA increased the mitochondrial cholesterol pool available for testosterone biosynthesis. The rate of androgenesis is likely the result of various regulatory factors acting concomitantly on the physiology of Leydig cells.     

Isolation of a Δ5 Desaturase Gene from <Emphasis Type="Italic">Euglena gracilis</Emphasis> and Functional Dissection of Its HPGG and HDASH Motifs

Delta (Δ) 5 desaturase is a key enzyme for the biosynthesis of health-beneficial long chain polyunsaturated fatty acids such as arachidonic acid (ARA, C20:4n-6), eicosapentaenoic acid (C20:5n-3) and docosahexaenoic acid (C22:6n-3) via the “desaturation and elongation” pathways. A full length Δ5 desaturase gene from Euglena gracilis (EgΔ5D) was isolated by cloning the products of polymerase chain reaction with degenerate oligonucleotides as primers, followed by 5′ and 3′ rapid amplification of cDNA ends. The whole coding region of EgΔ5D was 1,350 nucleotides in length and encoded a polypeptide of 449 amino acids. BlastP search showed that EgΔ5D has about 39 % identity with a Δ5 desaturase of Phaeodactylum tricornutum. In a genetically modified dihomo-gamma-linoleic acid (DGLA, C20:3n-6) producing Yarrowia lipolytica strain, EgΔ5D had strong Δ5 desaturase activity with DGLA to ARA conversion of more than 24 %. Functional dissection of its HPGG and HDASH motifs demonstrated that both motifs were important, but not necessary in the exact form as encoded for the enzyme activity of EgΔ5D. A double mutant EgΔ5D-34G158G with altered sequences within both HPGG and HDASH motifs was generated and exhibited Δ5 desaturase activity similar to the wild type EgΔ5D. Codon optimization of the N-terminal region of EgΔ5D-34G158G and substitution of the arginine with serine at residue 347 improved substrate conversion to 27.6 %.     

Characterization of wax esters in the roe oil of amber fish,Seriola aureovittata

The wax esters of the roe oil of the amber fish,Seriola aureovittata, have been resolved by high-performance liquid chromatography (HPLC) in the silver-ion mode. Each of the fractions collected was transmethylated, and the fatty acids and alcohols were identified by gas chromatography/mass spectrometry (GC/MS) as the picolinyl esters and nicotinates, respectively. Their compositions were determined by GC. The fatty acid composition is complex, and the main components are C18:1n-9 (35.5 mol%), C22:6n-3 (20.3 mol%), and C16:1n-7 (10.7 mol%), while fatty alcohols are limited to saturated (C16:0, 60.3 mol%; C18:0, 15.3 mol%; C14:0, 5.1 mol%) and monoenoic alcohols (C18:1n-9, 6.5 mol%; C16:1n-7, 4.5 mol%) with traces (<0.1 mol%) of polyunsaturated fatty alcohols such as C20:3n-3, C20:4n-6, C20:5n-3, and C22:5n-3. Silver-ion HPLC exhibited excellent resolution in which fractions were resolved on the basis of the number and configuration of double bonds as well as the distribution pattern between the acid and alcohol moieties of the molecules with a given number of double bonds. The main wax ester fraction are those of monoenoic acid-saturated alcohol species, hexaenoic acid/saturated alcohol species, and pentaenoic acid/saturated alcohol species. Appreciable specificity was observed in the esterification of fatty acids with alcohols, and surprisingly, no saturated acid-monoenoic alcohol species were detected.