Identification of the pathway of α-oxidation of cerebronic acid in peroxisomes

Cerebronic acid (2-hydroxytetracosanoic acid), an α-hydroxy very long-chain fatty acid (VLCFA) and a component of cerebrosides and sulfatides, is unique to nervous tissues. Studies were carried out to identify the pathway and the subcellular site involved in the oxidation of cerebronic acid. The results from these studies revealed that cerebronic acid was catabolized by α-oxidation to CO₂ and tricosanoic acid (23:0). Studies with subcellular fractions indicated that cerebronic acid was α-oxidized in fractions having particulate bound catalase and enzyme systems for the β-oxidation of VLCFA (e.g., lignoceric acid), suggesting peroxisomes as the subcellular organelle responsible for α-oxidation of cerebronic acid. Etomoxir, an inhibitor of mitochondrial fatty acid oxidation, had no effect on cerebronic acid α-oxidation. Further, cerebronic acid oxidation was found to be dependent on the presence of NAD⁺ but not FAD, NADPH, ATP, Mg²⁺, or CoASH. Intraorganellar localization studies indicated that the enzyme system for the α-oxidation of cerebronic acid was associated with the peroxisomal limiting membranes. Studies on cultured fibroblasts from normal subjects and patients with peroxisomal disorders indicated an impairment of α-oxidation of cerebronic acid in cell lines that lack peroxisomes [e.g., Zellweger syndrome (ZS)]. On the other hand, α-oxidation of cerebronic acid was found to be normal in cell lines from X-linked adrenoleukodystrophy, adult Refsum disease, and rhizomelic chondrodysplasia punctata. Our results clearly demonstrate that α-oxidation of α-hydroxy VLCFA (cerebronic acid) is a peroxisomal function and that this oxidation is impaired in ZS. Furthermore, this α-oxidation enzyme system is distinct from the one for the α-oxidation of β-carbon branched-chain fatty acids (e.g., phytanic acid).     

Very long chain fatty acids in higher animals—A review

Fatty acids with greater than 22 carbon atoms (very long chain fatty acids, VLCFA) are present in small amounts in most animal tissues. Saturated and monoenoic VLCFA are major components of brain, while the polyenoic VLCFA occur in significant amounts in certain specialized animal tissues such at retina and spermatozoa. Biosynthesis of VLCFA occurs by carbon chain elongation of shorter chain fatty acid precursors while β-oxidation takes place, almost exclusively in peroxisomes. Mitochondria are unable to oxidize VLCFA because they lack a specific VLCFA coenzyme A synthetase, the first enzyme in the β-oxidation pathway. VLCFA accumulate in the tissues of patients with inherited abnormalities in peroxisomal assembly, and also in individuals with defects in enzymes catalyzing individual reactions along the β-oxidation pathway. It is believed that the accumulation of VLCFA in patient tissues contributes to the severe pathological changes which are a feature of these conditions. However, little is known of the role of VLCFA in normal cellular processes, and of the molecular basis for their contribution to the disease process. The present review provides an outline of the current knowledge of VLCFA including their biosynthesis, degradation, possible function and involvement in human disease.     

Molecular Species of Phospholipids with Very Long Chain Fatty Acids in Skin Fibroblasts of Zellweger Syndrome

The ratio of C_26:0/C_22:0 fatty acids in patient lipids is widely accepted as a critical clinical criterion of peroxisomal diseases, such as Zellweger syndrome and X-linked adrenoleukodystrophy (X-ALD). However, phospholipid molecular species with very long chain fatty acids (VLCFA) have not been precisely characterized. In the present study, the structures of such molecules in fibroblasts of Zellweger syndrome and X-ALD were examined using LC–ESI–MS/MS analysis. In fibroblasts from Zellweger patients, a large number of VLCFA-containing molecular species were detected in several phospholipid classes as well as neutral lipids, including triacylglycerol and cholesteryl esters. Among these lipids, phosphatidylcholine showed the most diversity in the structures of VLCFA-containing molecular species. Some VLCFA possessed longer carbon chains and/or larger number of double bonds than C_26:0-fatty acid (FA). Similar VLCFA were also found in other phospholipid classes, such as phosphatidylethanolamine and phosphatidylserine. In addition, VLCFA-containing phospholipid species showed some differences among fibroblasts from Zellweger patients. It appears that phospholipids with VLCFA, with or without double bonds, as well as C_26:0-FA might affect cellular functions, thus leading to the pathogenesis of peroxisomal diseases, such as Zellweger syndrome and X-ALD.     

Isomeric fatty acids: Evaluating status and implications for maternal and child health

“Isomeric fatty acids” is a term that refers to the trans- and positional isomers formed during hydrogenation of naturally occurring oils. The purposes of this paper are as follows: (i) to summarize potential exposure of infants to isomeric fatty acids by reviewing estimates of isomeric fatty acids in the maternal diet, in human milk, and in infant formula/infant foods, and (ii) to evaluate the evidence for adverse effects of isomeric fatty acids on infant development with respect to growth and essential fatty acid status. Estimates of the intake of trans-fatty acids vary widely both within and across populations. Current estimates of trans-fatty acids in the North American population are 4–11% of total fatty acids or 3–13 g/(person·d), whereas in Mediterranean countries in which olive oil is the primary fat and in Far Eastern countries in which little commercially hydrogenated fat is consumed, per capita consumption of trans-fatty acids is <1–2 g/d. The trans-fatty acid content of human milk reflects the cross-cultural variation in the maternal diet, with trans-fatty acids in human milk samples ranging from 6 to 7% in North America to <0.5% in Hong Kong. Trans-fatty acids are transferred from the maternal diet through the placenta to the developing fetus or through milk to the breast-fed infant. In some studies, plasma trans-fatty acids are inversely related to birth weight and head circumference. The hypothesis that dietary trans-fatty acids could inhibit biosynthesis of long-chain polyunsaturated fatty acids with 20 and 22 carbon atoms and thus affect infant development is supported by studies demonstrating an inverse correlation of plasma trans-fatty acids with n−3 and n−6 long-chain polyunsaturated fatty acids in infants. However, no such relationship has been observed in human milk. A definitive answer concerning a potentially adverse effect of dietary trans-fatty acids on infant development awaits future studies.     

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.     

Fatty acid synthesis from [2-14C]acetate in normal and peroxisome-deficient (Zellweger) fibroblasts

The incorporation of [2-¹⁴C]acetate into the lipids of normal and peroxisome-deficient (Zellweger's syndrome) skin fibroblasts was examined. Most of the label was incorporated into triacyglycerol fatty acids in normal as well as Zellweger's syndrome cells. Triacylglycerols and cholesteryl esters in Zellweger's syndrome cells contained increased levels of labelled saturated and monounsaturated very long-chain fatty acids (VLCFA, that is fatty acids with more than 22 carbon atoms), in particular hexacosanoic (26∶0) and hexacosaenoic (26∶1) acids. As traces of labelled VLCFA with up to 32 carbon atoms were detected in triacylglycerols even in control cells it is probable that these fatty acids are formed naturally during the elongation process. Our data suggest that peroxisomes are involved in the chain shortening of the saturated and monounsaturated VLCFA.     

Metabolism of very long chain polyunsaturated fatty acids in isolated rat germ cells

Which cell type is responsible for the high levels of very long chain polyunsaturated fatty acids in testis and whether this fatty acid pattern is a result of a local synthesis are not presently known. In this study, fatty acid conversion from 20∶4n−6 to 22∶5n−6 and from 20∶5n−3 to 22∶6n−3 was investigated in isolated rat germ cells incubated with [1-¹⁴C]-labeled fatty acids. The germ cells elongated the fatty acids from 20- to 22-carbon atoms and from 22- to 24-carbon atoms but had a low Δ6 desaturation activity. Thus, little [¹⁴C]22∶5n−6 and [¹⁴C]22∶6n−3 were synthesized. When Sertoli cells were incubated with [1-¹⁴C]20∶5n−3 for 24 h, an active fatty acid elongation and desaturation were observed. In vivo germ cells normally have a higher content of 22∶5n−6 or 22∶6n−3 than Sertoli cells. An eventual transport of essential fatty acids from Sertoli cells to germ cells was thus studied. Different co-culture systems were used in which germ cells were on one side of a filter and Sertoli cells on the opposite side. When isolated pachytene spermatocytes or round spermatids were added to the opposite side of a semipermeable filter, approximately 1 nmol [¹⁴C]-22∶6n−3 crossed the filter. Little of this was esterified in the germ cells. Similarly, in using [1-¹⁴C]20∶4n−6 in identical experiments, very little [¹⁴C]22∶5n−6 was esterified in germ cells on the opposite side of the filter. Although the very active synthesis of 22∶5n−6 and 22∶6n−3 observed in Sertoli cells suggests a transport of these compounds to germ cells, this was not experimentally determined.     

Influence of partially hydrogenated vegetable and marine oils on lipid metabolism in rat liver and heart

Partially hydrogenated marine oils containing 18∶1-, 20∶1- and 22∶1-isomers and partially hydrogenated peanut oil containing 18∶1-isomers were fed as 24–28 wt % of the diet with or without supplement of linoleic acid. Reference groups were fed peanut, soybean, or rapeseed oils with low or high erucic acid content. Dietary monoene isomers reduced the conversion of linoleic acid into arachidonic acid and the deposition of the latter in liver and heart phosphatidylcholine. This effect was more pronounced for the partially hydrogenated marine oils than for the partially hydrogenated peanut oil. The content oftrans fatty acids in liver phospholipids was similar in groups fed partially hydrogenated fats. The distribution of various phospholipids in heart and liver was unaffected by the dietary fat. The decrease in deposition of arachidonic acid in rats fed partially hydrogenated marine oils was shown in vitro to be a consequence of lower Δ6-desaturase activity rather than an increase in the peroxisomal β-oxidation of arachidonic acid. The lower amounts of arachidonic acid deposited may be a result of competition in the Δ6-desaturation not only from the C22-and C20-monoenoic fatty acids originally present in the partially hydrogenated marine oil, but also from C18- and C16-monoenes produced by peroxisomal β-oxidation of the long-chain fatty acids. Part of this work was presented at the ISF-AOCS Congress, New York City, 1980.     

Antagonistic effects ofMyxococcus xanthus on fungi: II. Isolation and characterization of inhibitory lipid factors

The chemical composition of the lipophilic excretion ofMyxococcus xanthus inhibitory to the germination of fungal spores and growth has been investigated. The inhibitory effect was attributed to a mixture of fungistatically acting fatty acids and a component of antibiotic character. The fatty acid mixture has been fully characterized and found to constitute a mixture of saturated (68%) and unsaturated (32%) structures in the C₁₃–C₁₇ range. The major part is methylbranched of theiso-type, with 13-methyltetradecanoic acid being the main component (33% of the total). The fungistatic activity of the fatty acid mixture on spore germination is attributed to the structures withiso-configuration. The presence of unsaturation is of minor importance. Observed morphological changes of the spores and hyphae in the presence ofiso-fatty acids suggest that they act on the plasma membrane.     

Identification and Ruminal Outflow of Long-Chain Fatty Acid Biohydrogenation Intermediates in Cows Fed Diets Containing Fish Oil

The abundance of 20- to 24-carbon fatty acids in omasal digesta of cows fed grass silage-based diets supplemented with 0 (Control) and 250 g/day of fish oil (FO) was examined to investigate the fate of long-chain unsaturated fatty acids in the rumen. Complimentary argentation thin-layer chromatography and gas-chromatography mass-spectrometry analysis of fatty acid methyl esters and corresponding 4,4-dimethyloxazoline derivatives prepared from fish oil and omasal digesta enabled the structure of novel 20- to 22-carbon fatty acids to be elucidated. Compared with the Control, the FO treatment resulted in the formation and accumulation of 27 novel 20- and 22-carbon biohydrogenation intermediates containing at least one trans double bond and the appearance of cis-14 20:1, 20:2n-3, 21:4n-3 and 22:3n-6 not contained in fish oil. No conjugated ≥20-carbon fatty acids were detected in Control or FO digesta. In conclusion, fish oil in the diet results in the formation of numerous long-chain biohydrogenation intermediates in the rumen of lactating cows. Comparison of the intake and flow of 20-, 21- and 22-carbon fatty acids at the omasum in cows fed the Control and FO treatments suggests that the first committed steps of 20:5n-3, 21:5n-3 and 22:6n-3 hydrogenation in the rumen involve the reduction and/or isomerisation of double bonds closest to the carboxyl group.     

Effect of very long chain fatty acids on peroxisomal β-oxidation in primary rat hepatocyte cultures

Previous studies have demonstrated that certain high fat diets can induce peroxisomal fatty acid β-oxidation in rodent liver and that this may be due to their content oftrans 22∶1 fatty acids. In this study we have examined the effects ofcis andtrans 22∶1 fatty acids (erucic and brassidic) and oleic acid (18∶1) on palmitoyl-CoA oxidation, carnitine acetyltransferase and carnitine palmitoyltransferase activities in primary rat hepatocyte cultures. Brassidic and erucic acid and, to a lesser extent, oleic acid were cytotoxic to rat hepatocytes. However, at a concentration of 0.1 mM, brassidic acid produced small increases in palmitoyl-CoA oxidation and carnitine acetyltransferase activities in hepatocytes cultured 70 hr. Treatment of cells with 0.1 and 0.3 mM of either erucic or oleic acid had no effect on any of the enzymes measured.     

Creating new water-soluble Iso- and n-fatty acid arginate hydrochloride with antimicrobial properties

Typically, short- and long-chain lipids from oils exhibit different antimicrobial activities and therefore have been used in agriculture and aquaculture, biomedical therapeutic and antibacterial fields. However, these fatty acids have limitations in terms of bioactive efficacy, thermostability and aqueous solubility. In this study, water-soluble iso-fatty acid arginate hydrochloride derivatives with antimicrobial properties were produced by introducing branched (iso-) chain and other linear- (n-) chain fatty acids to the “arginine” amino acid molecule. The two-step synthetic route was straightforward and provided an efficient 88% and 76% product yields for ethyl n-oleoyl arginate hydrochloride and ethyl iso-oleoyl arginate hydrochloride, respectively. ATR-FT-IR, NMR, and LC-MS-Q-TOF techniques were used to thoroughly characterize and confirm the products. These arginate products had strong antimicrobial activities against Listeria innucua, a Gram-positive bacterium with minimum inhibitory concentrations and minimum bactericidal concentrations ranging from 1.8 µg mL⁻¹ to 29.1 µg mL⁻¹. Therefore, the study demonstrated the development of a novel class of antimicrobial compounds from iso-fatty acids and arginates.     

Metabolism of meadowfoam oil fatty acids in mice

Meadowfoam oil is unusual because over 95% of the fatty acids are 20- and 22-carbon aliphatic acids withcis double bonds located principally at the 5- and/or 13-position. Since little information is available on the metabolism of the 5c−20∶1 and 5c,13c−22∶2 fatty acids, an exploratory study in mice was conducted to investigate the metabolism of purified samples of the free fatty acids isolated from meadowfoam oil, and to determine the effect of meadowfoam oil on weight gain and tissue lipid composition. Mice fed diets containing 5% by wt of the purified 5c−20∶1 or 5c,13c−22∶2 for 6 days exhibited no apparent physiological problems. Total liver lipids from mice fed the purified fatty acid diets contained mean values of 2.0% 5c−20∶1 and 2.1% 5c,13c−22∶2; total heart lipids contained 1.7% 5c−20∶1 and 10.7% 5c,13c−22∶2. Liver total phospholipids from mice fed a 5% meadowfoam oil diet for 19 wk contained 1.4% 5c−20∶1 and 1.9% 5c,13c−22∶2. There was no evidence of desaturation, elongation or retroconversion. Weight gain for mice fed the meadowfoam oil diet for 19 wk was similar to mice fed corn oil, and was higher than for mice fed hydrogenated cottonseed oil. Considering the high 5c−20∶1 and 5c,13c−22∶2 content of the diets, the percentages of these fatty acids in mouse tissue lipids from both the short- and long-term studies were low. Weight gain was surprisingly good since the meadowfoam oil diet was essential fatty acid-deficient. Results of this initial investigation suggest that the 5c−20∶1 and 5c,13c−22∶2 fatty acids were utilized primarily for energy. In the short-term study, these fatty acids did not produce toxic effects or cause metabolic problems. The mention of firm names or trade products does not imply that they are endorsed or recommended by the U.S. Department of Agriculture over other firms or similar products not mentioned.     

Total and peroxisomal oxidation of various saturated and unsaturated fatty acids in rat liver,heart and m. quadriceps

Rates of total and peroxisomal fatty acid oxidation were estimated from the production of¹⁴C-labeled CO₂ and acid-soluble products from differently labeled [¹⁴C]fatty acids, in the absence and presence of antimycinrotenone, in homogenates of liver, heart and m. quadriceps. Total and peroxisomal oxidation rates of palmitic, oleic and linoleic acid were 3–4 times higher than those of arachidonic and adrenic acid which had higher oxidation rates than those of lignoceric and erucic acid. The peroxisomal contribution to the oxidation of the last fatty acids was similar to or higher than that of palmitic acid. For all fatty acids tested in these tissues, the mitochondrial contribution to β-oxidation was higher than the peroxisomal contribution. Production of¹⁴CO₂ and¹⁴C-labeled, acid-soluble metabolites from [13-¹⁴]arachidonic acid indicated that polyunsaturated fatty acids can be chain-shortened beyond their double bonds in m. quadriceps and heart as well as in liver. Although 2,4-dienoyl-CoA reductase requires NADPH, addition of this coenzyme did not influence arachidonic acid oxidation. Arachidonic acid oxidation was inhibited by palmitic acid in mitochondria and peroxisomes, but arachidonic acid had only a slight effect on palmitic acid oxidation.     

Turnover of fatty acyl-CoA oxidase in the liver of rats fed on a partially hydrogenated marine oil

The change in turnover of fatty acyl-CoA oxidase (FAO), the rate-limiting enzyme of the peroxisomal β-oxidation system, was investigated in rats fed a 30% (w/w) partially hydrogenated marine oil (PHMO) diet. The FAO activity increased five-fold after two weeks of PHMO feeding, and decreased after withdrawal of the diet. Based onin vivo experiments using L-[4,5-³H]leucine and an immunoprecipitation technique, the increase in the activity of FAO could be accounted for by a 1.6-fold higher rate of FAO synthesis and a 3.4-fold slower rate of FAO degradation as compared to controls. In the same PHMO-fed rats, the rates of synthesis and degradation of carnitine palmitoyltransferase were 1.8-fold higher and 2.0-fold slower, respectively, as compared to controls. The results indicate that the observed increase in the activity of the enzymes of peroxisomal β-oxidation is mainly due to a reduced rate of FAO degradation in the liver of rats fed the PHMO diet.     

Activities of liver mitochondrial and peroxisomal fatty acid oxidation enzymes in rats fedtrans fat

The effect oftrans fat on the activities of liver mitochondrial and peroxisomal fatty acid oxidation enzymes was examined in various strains of rats. When Wistar and Sprague-Dawley rats were fed for 30 days diets containing either olive oil or partially hydrogenated corn oil as a source ofcis-ortrans-octadecenoate, respectively, the activities of various enzymes of mitochondrial and peroxisomal β-oxidation measured withcis- andtrans-9-octadecenoic acid as substratese showed little dietary fatdependent change. In Fischer 344 rats, feedingtrans fat for 15 mo increased only moderately various enzymes of β-oxidation except for carnitine acyltransferase. The rate of mitochondrial ketogenesis and the activity of carnitine acyltransferase measured withtrans-9-octadecenoic acid as a substrate were about half those with thecis-counterpart. Peroxisomes oxidizedtrans-9-octadecenoyl-CoA at a rate comparable to thecis-counterpart. It was concluded from this study and previous ones that the difference in the geometry of dietary fatty acid had only a marginal effect in modulating the hepatic fatty acid oxidation system, in spite of marked differences in the metabolic behavior ofcis-andtrans fatty acid in cell-free preparations and perfused liver.     

Fatty acid metabolism in renal ischemia

The increase in free fatty acids in the ischemic tissue is a consistent observation and these free fatty acids are considered, to play a role in the cellular toxicity. To elucidate the cause of higher levels of free fatty acids in ischemic tissue, we examined the catabolism of fatty acids. The β-oxidation of lignoceric (24∶0), palmitic (16∶0) and octanoic (8∶0) acids and the peroxidation of fatty acids were measured at different times of renal ischemia in whole kidney homogenate. The enzymatic activities for the oxidation of fatty acids decreased with the increase in ischemia time. However, the lipid peroxide levels increased 2.5-fold of control with ischemic injury. Sixty min of ischemia reduced the rate of oxidation of octanoic, palmitic and lignoceric acids by 57, 59 and 69%, respectively. Almost similar loss of fatty acid oxidation activity was observed in the peroxisomes and mitochondria. These data suggest that loss of mitochondrial and peroxisomal fatty acid β-oxidation enzyme activities from ischemic injury may be one of the factors responsible for the higher levels of free fatty acids.     

Quantitative effects of dietary polyunsaturated fats on the composition of fatty acids in rat tissues

A method combining data on fatty acid composition into subsets is used to illustrate general relative competitive selectivities in the metabolic and transport events that maintain fatty acid compositions in tissue lipids and to minimize differences among tissues or species in the amount of individual fatty acids. Fatty acid compositions of triglycerides and phospholipids in several tissues of the rat were maintained with simple relationships between the exogenous n−3 and n−6 dietary polyunsaturated fatty acids and the endogenous n−7 and n−9 types of fatty acid. The general pattern of fatty acids in triglycerides was similar for liver, plasma and adipose tissue, averaging about 30% as saturated acids, 67% as 16- and 18-carbon unsaturated acids and only about 2% as 20- and 22-carbon highly unsaturated acids. The tissues maintained a linear relationship between the amount of 18-carbon polyunsaturated fatty acids in the diet and in the tissue triglycerides, with the proportionality constant for 18∶3n−3 being 60% of that for 18∶2n−6. The total phospholipids of liver, plasma and red blood cells maintained about 45% of the fatty acids in the form of saturated fatty acids and 20–30% as 20- and 22-carbon highly unsaturated fatty acids irrespective of very different proportions of n−3, n−6 and n−9 types of fatty acids. In all three tissues, the 20-carbon highly unsaturated fatty acids of the n−3, n−6 and n−9 type were maintained in a competitive hyperbolic relationship with apparent EC₅₀ values for dietary 18∶2n−6 and 18∶3n−3 near 0.1% of dietary calories. The consistent quantitative relationships described in this study illustrate an underlying principle of competition among fatty acids for a limited number of esterification sites. This approach may be useful in predicting the influence of diet upon tissue levels of the substrates and antagonists of eicosanoid biosynthesis.     

Sponge fatty acids. 3. Occurrence of series of n−7 monoenoic andiso-5,9 dienoic long-chain fatty acids in the phospholipids of the marine spongeCinachyrella aff.schulzei keller

The fatty acid composition of phospholipids from the New Caledonian spongeCinachyrella aff.schulzei Keller was studied. More than 60 fatty acids were identified as methyl esters andN-acyl pyrrolidides by gas chromatography and gas chromatography/mass spectrometry. Two isoprenoid fatty acids also were shown to be present, namely 4,8,12-trimethyltridecanoic and 5,9,13-trimethyltetradecanoic acids. The unusual 6-tetradecenoic, 6-pentadecenoic, 12-nonadecenoic and 26-methylheptacosanoic (iso-28∶0) acids were found for the first time in sponge phospholipids. A series of six n−7 monoenoic long-chain fatty acids (C₂₃ to C₂₈) were identified, including the rare 16-tricosenoic, 18-pentacosenoic and 21-octacosenoic acids. Fifteen fatty acids possessing the typical 5,9 dienoic moiety accounted for 30% of the total fatty acid mixture. Two new fatty acids were identified, namely 5(Z)-octacosenoic and 27-methyl-5(Z),9(Z)-octacosadienoic (iso-5,9-29∶2). Based on gas chromatography/Fourier transform infrared experiments, the double bonds were assigned the (Z) configuration. For part 2 of this series, see Reference 1.     

Differential alterations of ethanolamine and choline phosphoglyceride metabolism by clofibrate and retinoic acid in human fibroblasts are not mediated by phorbol ester-sensitive protein kinase C

Peroxisomal proliferators and retinoids have been reported to interact to regulate lipid metabolism, particularly β-oxidation of fatty acids. Based on postulated interactions of these agents at the levels of receptors and response elements, we examined whether interactions exist between the peroxisomal proliferator, clofibrate (CLF), and retinoic acid (RA) in modulation of phospholipid turnover in cultured human skin fibroblasts. Treatment of cultured cells with either 25 μM CLF or 1 μM RA alone decreased [¹⁴C]ethanolamine incorporation into ethanolamine phosphoglycerides (EPG) by 20–30%, and simultaneous exposure to both agents resulted in additive inhibition. By contrast, [³H]choline incorporation into phospholipid was stimulated 5–30% by incubation with either agent; when CLF and RA were administered together, the stimulatory effects were additive. Different types of pulse-chase studies examining effects on uptake, biosynthesis, and degradation of labelled phospholipids indicated stimulation of EPG degradation and inhibition of phosphatidylcholine degradation by CLF; no effect on catabolism of either phospholipid was observed with RA. Combinations of modifiers of protein kinase activity [4β-12-O-tetradecanoylphorbol-13-acetate (β-TPA), 1-(5-isoquino-linesulfonyl)-2-methylpiperazine dihydrochloride,N-(2′-guanidinoethyl)-5-isoquinolinesulfonamide hydrochloride,bis-indolylmaleimide, staurosporine] indicated that β-TPA-responsive protein kinases were not involved. Accordingly, CLF and RA regulate biosynthesis and degradation of ethanolamine and choline phosphoglycerides in cultured skin fibroblasts by different mechanisms that do not involve classical protein kinase C (PKC) isoforms, even though turnover of phospholipids generating lipid activators of PKC occurs.