Chromatographic analysis of carotenol fatty acid esters in Physalis alkekengi and Hippophae rhamnoides

The carotenol fatty acid esters of two potentially valuable sources of plant carotenoids, sepals of Physalis alkekengi (Chinese lantern) and fruits of Hippophae rhamnoides (sea buckthorn), were separated by column chromatography and identified by HPLC-DAD and HPLC-MS. A chemical and an enzymatic hydrolysis were employed to identify the parent carotenoids and to remove the lipid components. Zeaxanthin and beta-cryptoxanthin esters represented the main fraction in P. alkekengi sepals and an important one in H. rhamnoides fruits. Beta-Cryptoxanthin palmitate and zeaxanthin dipalmitate were identified as major compounds in both plants. In P. alkekengi, the carotenoids were mainly (> 90%) esterified with palmitic acid, and a high proportion (> 80%) of saturated medium chain fatty acids was found (by GC-MS) in the total lipid extract. Although the total lipid extract of H. rhamnoides contained significant amounts of unsaturated fatty acids, especially oleic and palmitoleic acids, the xanthophylls were mainly esterified with saturated fatty acids. The oleoresins of both species represent potential sources of carotenoid esters and can be used as food additives, cosmetic ingredients or nutraceuticals.     

Utilization of endogenous lipid by the isolated perfused rat heart

1. The lipids of the rat heart have been studied with regard to amount, classes present and fatty acid composition of free fatty acids, triglycerides and phospholipids. Myocardial lipid contained 300μmoles of total fatty acid/g. dry wt. of which only 2–4μmoles were free; the remainder was esterified, chiefly as phospholipid. Neutral esters, of which triglyceride was the principal form, made up 15% of the total fatty acids. 2. When normal hearts were perfused with a nutrient-free medium until exhaustion, the triglyceride concentration declined from 43 to 13μmoles/g. dry wt. The content of phospholipids, partial glycerides and cholesteryl esters did not change. When the lipids of the rat heart were labelled with [1-¹⁴C]palmitate before perfusion with non-nutrient medium, radioactivity disappeared from the triglyceride, diglyceride and free fatty acid fractions, but not from the phospholipid or other ester classes. 3. These experiments support the view that only a small fraction of the total cardiac lipid, principally triglycerides and to a smaller extent diglycerides, is available as a source of fuel in the absence of exogenous substrate.     

Adsorption of 3H-Fatty Acid Esters of Streptococcal Groups A and E Cell Wall Polysaccharide Antigens by Red Blood Cells and Their Effect on Hemagglutination

The streptococcal group A and E cell wall polysaccharide (PS) antigens were esterified under identical conditions with four fatty acid chlorides (lauroyl, myristoyl, palmitoyl, and stearoyl), varying from 12 to 18 carbon atoms. With group A PS, it was shown that the four resulting esters varied in their ability to sensitize red blood cells (RBC) to agglutination in the presence of specific antiserum. The most active was palmitoyl (16C) followed by myristoyl (14C). The least active was the lauroyl ester (12C). One-tenth as much palmitoyl ester was required as stearoyl group A PS ester. Such variation in the ability to sensitize RBC was not demonstrated with the group E esters, with the exception of the lauroyl ester which was the least active. Removal of N-acetylglucosamine from the esterified and the nonesterified group A PS by enzyme action resulted in a significant loss of serological activity of both antigens. No appreciable difference in the rate or total loss of activity was found in either case. It was demonstrated that both tritium-labeled stearic and palmitic acids and their respective PS esters were adsorbed in significant amounts to RBC. The results indicate that the esterified antigens were adsorbed to the RBC because of the presence of the fatty acid in the PS ester. Attempts to block the receptor sites on the red cell by presensitizing the cells with fatty acid were negative. Likewise, the adsorbed ester did not prevent the uptake of fatty acid at the levels tested. Tritium-labeled esterified group A PS and group E PS were used to show that the amount of antigen required to produce maximal agglutination was the same when cells from the same individual were used, whereas this was not the case when cells from different individuals were used. The amount of antigen required to produce maximal agglutination varied from one batch of sheep RBC to another. Once the optimal concentration of antigen was reached, any additional adsorption did not increase the titer of agglutination.     

The Neutral Lipids of Paramecium tetraurelia: Changes with Culture Age and the Detection of Steryl Esters in Ciliary Membranes1

Neutral lipids, particularly triglycerides, accounted for the major decrease in the total lipid content in Paramecium cells that occurs with culture age. Sterols, triglycerides, and steryl esters were the major classes of neutral lipids in cells and isolated cilia. Free as well as high concentrations of esterified sterols were detected in purified ciliary membrane preparations. Stigmasterol and 7-dehydrostigmasterol were the major components of both free and esterified sterols of cells and cilia; however, when cholesterol was present in the growth medium, it was desaturated to 7-dehydrocholesterol and incorporated into cellular and ciliary lipids. Free fatty acids from cells and triglycerides from cells and cilia were low in polyunsaturated fatty acids and reflected the composition of fatty acids in the culture medium. An exception was the reduced concentration of stearate in triglycerides from whole cells. Greater than 50% of triglyceride fatty acids from cilia were saturated. The fatty acid compositions of cellular triglycerides and ciliary steryl esters did not change with culture age, but those of cellular steryl esters and ciliary triglycerides did change. In comparison with phospholipids, these neutral lipid fatty acid compositional changes were smaller. The sensitivity of these stigmasterol-containing cells to polyene antibiotics indicated that they were killed by nystatin > filipin > amphotericin B. The unexpected finding of high concentrations of steryl esters in ciliary membrane preparations is discussed.     

Differential distribution and metabolism of arachidonic acid and docosahexaenoic acid by human placental choriocarcinoma (BeWo) cells

The time course of incorporation of [¹⁴C]arachidonic acid and [³H]docosahexaenoic acid into various lipid fractions in placental choriocarcinoma (BeWo) cells was investigated. BeWo cells were found to rapidly incorporate exogenous [¹⁴C]arachidonic acid and [³H] docosahexaenoic acid into the total cellular lipid pool. The extent of docosahexaenoic acid esterification was more rapid than for arachidonic acid, although this difference abated with time to leave only a small percentage of the fatty acids in their unesterified form. Furthermore, uptake was found to be saturable. In the cellular lipids these fatty acids were mainly esterified into the phospholipid (PL) and the triacyglycerol (TAG) fractions. Smaller amounts were also detected in the diacylglycerol and cholesterol ester fractions. Almost 60% of the total amount of [3H]Docosahexaenoic acid taken up by the cells was esterified into TAG whereas 37% was in PL fractions. For arachidonic acid the reverse was true, 60% of the total uptake was incorporated into PL fractions whereas less than 35% was in TAG. Marked differences were also found in the distribution of the fatty acids into individual phospholipid classes. The higher incorporation of docosahexaenoic acid and arachidonic acid was found in PC and PE, respectively. The greater cellular uptake of docosahexaenoic acid and its preferential incorporation in TAG suggests that both uptake and transport modes of this fatty acid by the placenta to fetus is different from that of arachidonic acid.     

Lipid analysis and fatty acid profiles of individual arterial atherosclerotic plaques

A protocol for the analysis of the lipid profile of microsamples of aortic tissue was developed. Lipid extraction was from intact tissue using acetone and chloroform/methanol (2/1, v/v). The extract was analyzed for total lipid, esterified cholesterol, cholesterol, triacylglycerol, and phospholipid. The extract was then processed to separate cholesteryl esters, triacylglycerol, and phospholipid which were hydrolyzed and the fatty acid composition was determined by GLC of pentafluorobenzyl ester derivatives. A lipid profile could be obtained on samples with a wet weight of 5 mg.     

Separation and quantitation of dolichyl esters by high-performance liquid chromatography

An HPLC procedure for the isolation and quantitation of total and individual dolichyl esters in tissues has been developed. The purified lipid extracts are subjected to sequential reversed-phase, straight-phase, and reversed-phase HPLC, which yield complete resolution and high recovery of the individual dolichyl esters. The isoprenoid distribution in the esterified fraction was similar to that of the free alcohol fraction in liver, kidney, and spleen. All fatty acids present in the total fraction were also recovered in all the individual polyisoprenoids. Dolichyl esters thus appear to differ from other lipid esters in tissues in containing a broader range of fatty acids.     

Oxidative degradation of model lipids representative for main paper pulp lipophilic extractives by the laccase–mediator system

Different model lipids-alkanes, fatty alcohols, fatty acids, resin acids, free sterols, sterol esters, and triglycerides-were treated with Pycnoporus cinnabarinus laccase in the presence of 1-hydroxybenzotriazole as mediator, and the products were analyzed by gas chromatography. The laccase alone decreased the concentration of some unsaturated lipids. However, the most extensive lipid modification was obtained with the laccase-mediator system. Unsaturated lipids were largely oxidized and the dominant products detected were epoxy and hydroxy fatty acids from fatty acids and free and esterified 7-ketosterols and steroid ketones from sterols and sterol esters. The former compounds suggested unsaturated lipid attack via the corresponding hydroperoxides. The enzymatic reaction on sterol esters largely depended on the nature of the fatty acyl moiety, i.e., oxidation of saturated fatty acid esters started at the sterol moiety, whereas the initial attack of unsaturated fatty acid esters was produced on the fatty acid double bonds. In contrast, saturated lipids were not modified, although some of them decreased when the laccase-mediator reactions were carried out in the presence of unsaturated lipids suggesting participation of lipid peroxidation radicals. These results are discussed in the context of enzymatic control of pitch to explain the removal of lipid mixtures during laccase-mediator treatment of different pulp types.     

Yeast sterol esters and their relationship to the growth of yeast.

Variation in the percentage of sterols esterified to long-chain fatty acids during cellular growth has been examined. Under all conditions, a constant percentage of sterol esters was maintained during exponential growth. This maintenance level was found to vary with different growth conditions. A sharp increase in the rate of esterification was observed upon entry of the culture into the stationary growth phase. The minor cellular sterol components were found to accumulate after this period of rapid sterol ester synthesis, with a relative decrease in the size of the ergosterol pool. Evidence is presented that sterol esters of ergosterol precursors are unable to be metabolized to ergosterol. Once esterified, the fatty acids do not appear to be scavenged during starvation conditions.     

Lipid and sterol composition of the pollen of the west african oilpalm, Elaeis guineensis

The lipid classes, fatty acid methyl esters and the sterols of oilpalm pollen were analysed. The neutral lipid fraction consisted of triglycerides, esterified and free sterols and trace amounts of hydrocarbons. Monogalactosyl and digalactosyl diglycerides, phosphatidyl choline, phosphatidyl inositol and phosphatidyl ethanolamine represented the polar lipids. The major fatty acids were linoleic, palmitic and linolenic acids together with small to trace amounts of oleic, stearic, arachidic, myristic, lauric, palmitoleic and margaric acids. Unsaturated fatty acids predominated over saturated ones in the ratio of 3:2. The 4-desmethyl sterols were the major phytosterols in the free form but they constituted a lower proportion of the sterols in the esterified state. 28-Isofucosterol was isolated and characterized as the principal sterol.     

Fatty acid and lipid analysis of the house cricket,Acheta domesticus

The fatty acid content and composition of the house cricket Acheta domesticus have been investigated in entire insects at different developmental stages and in selected organs of male and female adults. We have also determined the fatty acid composition of the various lipid classes within extracts of the organs of adult female insects. Fatty acids were analysed by capillary gas chromatography or mass spectrometry as their methyl esters (FAMEs) after direct transesterification of insect material or separated lipid classes.The major esterified fatty acids in all extracts were palmitate (C_16:0), stearate (C_18:0), oleate (C_18:1) and linoleate (C_18:2). Levels of esterified fatty acid varied considerably between organs but the fatty acid compositions showed only small variations. The levels of polyunsaturated fatty acids of the C₁₈ series were considerably higher in phospholipid fractions than in other lipid classes. Triacylglycerols formed the major lipid class in ovaries, fat-body and newly-laid eggs, whereas diacylglycerols and phospholipid predominate in the haemolymph. Triacylglycerols, phospholipids, diacylglycerols and free fatty acids were all found in significant amounts in the gut tissue.     

Lipid composition of the electron transport membrane of Haemophilus parainfluenzae

The principal lipids associated with the electron transport membrane of Haemophilus parainfluenzae are phosphatidylethanolamine (78%), phosphatidylmonomethylethanolamine (0.4%), phosphatidylglycerol (18%), phosphatidylcholine (0.4%), phosphatidylserine (0.4%), phosphatidic acid (0.2%), and cardiolipin (3.0%). Phospholipids account for 98.4% of the extractible fatty acids. There are no glycolipids, plasmalogens, alkyl ethers, or lipo amino acid esters in the membrane lipids. Glycerol phosphate esters derived from the phospholipids by mild alkaline methanolysis were identified by their staining reactions, mobility on paper and ion-exchange column chromatography, and by the molar glycerol to phosphate ratios. Eleven diacyl phospholipids can be separated by two-dimensional thin-layer chromatography. Each lipid served as a substrate for phospholipase D, and had a fatty acid to phosphate ratio of 2:1. Each separated diacyl phospholipid was deacylated and the glycerol phosphate ester was identified by paper chromatography in four solvent systems. Of the 11 separated phospholipids, 3 were phosphatidylethanolamines, 2 were phosphatidylserines, and 2 were phosphatidylglycerols. Phosphatidylcholine, cardiolipin, and phosphatidic acid were found at a single location. Phosphatidylmonomethylethanolamine was found with the major phosphatidylethanolamine. Three distinct classes of phospholipids are separable according to their relative fatty acid compositions. (i) The trace lipids consist of two phosphatidylethanolamines, two phosphatidylserines, phosphatidylcholine, phosphatidic acid, and a phosphatidylglycerol. Each lipid represents less than 0.3% of the total lipid phosphate. These lipids are characterized by high proportions of the short (C(10) to C(14)) and long (C(19) to C(22)) fatty acids with practically no palmitoleic acid. (ii) The major phospholipids (93% of the lipid phosphate) are phosphatidylethanolamine, phosphatidylmonomethylethanolamine, and phosphatidylglycerol. These lipids contain a low proportion of the short (C(19)) fatty acids. Palmitic and palmitoleic acids represent over 80% of the total fatty acids. (iii) The fatty acid composition of the cardiolipin is intermediate between the other two classes. Both palmitoleic and the longer fatty acids represent a significant proportion of the total fatty acid.     

Occurrence of steryl and wax esters in Dicranum elongatum

The steryl and wax esters of the frozen subarctic moss Dicranum elongatum Schleich contained fatty acids 39.8 mg per gram dry green tissue. The content decreased with increasing age of the moss shoots, but no great changes were found in the fatty acid pattern of the esters. The major part of the steryl and wax ester fraction of the green shoots was made up of esterified sterols (85%), and the rest (15%) of esterified aliphatic alcohols. No great changes were found in their relative proportions with increased age of the shoots. Some changes were evident in the pattern of individual esterified sterols, however. The proportion of cycloartenol was lower in the older parts than in the green part, and the proportion of campesterol, stigmasterol, and β-sitosterol were lower in the green part. The major esterified aliphatic alcohols were 1-octadecanol, phytol and geranylgeraniol. The proportion of geranylgeraniol was highest in the green part and that of phytol in the older parts. The main alcohol of the surface lipids was 1-octadecanol.     

Cholesterol alterations in young dystrophic mice

Cholesterol and cholesteryl ester concentrations and cholesteryl ester fatty acid substituents have been measured during the first 10 weeks of life in tissues of normal and dystrophic mice. In normal Swiss and 129ReJ(+/?) mice the concentrations of both cholesterol and cholesteryl esters remain essentially constant in liver, increase in brain and fall sharply in both thigh (mixed fiber type muscles) and chest muscles (predominantly slow oxidative muscles) over this period. In all cases the concentration of free cholesterol exceeds that of esterified cholesterol. In dystrophic mice, similar patterns are found in brain and liver. In both thigh and chest muscles, however, the developmental pattern is significantly different. After an initial decrease the concentrations of cholesterol and cholesteryl esters increase rapidly with the largest increase occurring in the concentration of cholesteryl esters which by 10 weeks of age exceeds the concentration of cholesterol in chest muscle. During the same period the pattern of esterified fatty acids changes gradually in dystrophic tissues towards an increasing ratio of unsaturated/saturated fatty acids. By 10 weeks of age this ratio is significantly higher in dystrophic tissues than normal in all tissues tested.     

Incorporation of lipoxygenase products into cholesteryl esters by acyl-CoA:cholesterol acyltransferase in cholesterol-rich macrophages.

Macrophages which were incubated with acetylated low-density lipoproteins, resulting in cholesteryl ester accumulation, incorporated the monohydroxyeicosatetraenoic acids (5-, 15-, and 12-HETEs) into cholesteryl esters. The esterification of these hydroxy fatty acids to cholesterol by total membrane preparations of cholesterol-rich macrophages was dependent on the synthesis of the fatty acyl-CoA derivative, and was catalysed by acyl-CoA:cholesterol acyltransferase (ACAT). Stimulation of membrane ACAT activity by 25-hydroxycholesterol increased the synthesis of cholesteryl 12-HETE by 40%. In contrast, inhibiting ACAT activity by progesterone and compound 58-035 decreased cholesteryl 12-HETE production by 60% and 90% respectively. Although 5-, 15- and 12-HETE were esterified to cholesterol by ACAT, these monohydroxy fatty acids were less optimal as substrates compared with oleic acid or arachidonic acid. The hydrolysis and release of 12-HETE and the other monohydroxyeicosatetraenoic acids from intracellular cholesteryl esters and phospholipids occurred at a faster rate than for the more conventional fatty acids, oleate and arachidonate. Cholesteryl esters which contain hydroxy fatty acids therefore provide only a transient storage for lipoxygenase products, as these fatty acids are released into the medium as readily as hydroxy fatty acids found in phospholipids and triacylglycerols. The data provide evidence, for the first time, of an ACAT-dependent esterification of the lipoxygenase products 5-, 15- and 12-HETEs to cholesterol in the macrophage-derived foam cell. The channelling of these monohydroxy fatty acids to cholesteryl esters provides a mechanism which can alter the amount of lipoxygenase products incorporated into cellular phospholipids, thus averting deleterious changes to cell membranes. ACAT, by catalysing the esterification of monohydroxyeicosatetraenoic acids to cholesterol, could play a key role in regulating the amount of lipoxygenase products in the pericellular space of the cholesterol-enriched macrophage.     

Effect of light on lipid composition of Ricinus cell cultures

The changes of lipid composition were determined in callus cultures from Ricinus communis endosperm upon transfer from darkness into light. Culture in light induced chlorophyll synthesis and formation of differentiated chloroplasts. In light-grown cultures the major lipid classes were phospho- and glycolipids, dark-grown cultures were rich in triacylglycerol. The major fatty acids were linolenic acid and palmitic acid in both cultures. In the green cultures linolenic acid was predominantly esterified in glycolipids whereas in the dark-grown cultures this fatty acid was the major component of phospholipids. Ricinoleic acid was not found.Abbreviations PC phosphatidylcholine - PE phosphatidylethanolamine - MGD monogalactosyldigylceride - DGD digalactosyldiglyceride - SE steryl esters - NL neutral lipids     

THE COMPOSITION OF LIPIDS IN CEREBROSPINAL FLUID OF CHILDREN AND ADULTS

Abstract— The proportions of esterified cholesterol and phosphatidyl ethanolamine in lipids of cerebrospinal fluid (CSF) from children were found to be lower than the corresponcling values for adult CSF. The fatty acid patterns of the cholesterol ester, triglyceride + non-esterified fatty acids and phospholipid fractions all displayed low proportions of linoleate; palmitate and oleate were the principal acids present. The fatty acid composition of these lipid classes for CSF derived from children was similar to that from adult subjects. Degradation of CSF lecithin by snake-venom phospholipase A₂ revealed the saturated acids to be located predominantly in the 1-position with the unsaturated ones mainly in the 2-position.     

Hexane soluble non-volatiles in flowering to fruiting stages of Fatsia japonica

The n-hexane soluble non-volatile fraction of the acetone extracts from the flower buds, the flowers and the immature and the mature fruits of Fatsia japonica were all found to contain fatty acids, fatty acid methyl esters, squalene, β-amyrin and sterols. At all the stages between budding to the mature fruit, the major fatty acids were palmitic and linoleic acids and the major phytosterol was stigmasterol. In addition steryl and β-amyrenyl esters were found in the flowers and the immature and the mature fruits, but these esters were not present in the flower buds. Sitosteryl ester was the major constituent of the steryl ester fraction in the fruiting stages. Phytol was found in only the flowering stage and triglycerides in only the mature fruits. The variations in the lipid constituents is discussed in relation to the stages from budding to the mature fruit.     

CHANGES IN CEREBRAL CORTICAL LIPIDS IN COBALT-INDUCED EPILEPSY

Abstract– In control rats and in rats rendered epileptic by insertion of cobalt slivers into the cerebral cortex, total free fatty acids, free cholesterol, esterified cholesterol, triglycerides and phospholipids were measured in normal and lesion areas of cerebral cortex. The cortical lipid profile of the adult rat resembled that of the whole brain of very young rats rather than that of adult whole brain, with the principal differences from whole adult brain being lower total lipid content, increased proportions of phosphatidyl choline in the phospholipid fraction, and higher levels of cholesterol esters. Cobalt-induced epilepsy was associated with significant changes in cerebral cortical lipids in the area of the lesion and in the non-necrotic tissue adjacent to the lesion. The total lipid in the area of the lesion decreased sharply as a result of reductions in free cholesterol and total phospholipids. The levels of cholesterol esters and triglycerides increased in the area of the lesion, and cholesterol esters were also increased in the adjacent tissue. In addition there were decreases in the proportion of phosphatidyl ethanolamine in the phospholipids from the lesion site and adjacent tissue and decreases in the proportions of oleic, arachidonic and nervonic acids (unsaturated acids), and an increase in the proportions of lignoceric acid in the phospholipids. In the site of the lesion only, we observed a decrease in phospholipid palmitic acid and an appreciable increase in the proportions of an unidentified long-chained fatty acid.     

Cellular distribution and linkage of D-(-)-3-hydroxy fatty acids in Bacteroides species.

Two strains of Bacteroides asaccharolyticus and two strains of Bacteroides fragilis were analyzed for total fatty acid, total lipid fatty acid, and total bound fatty acid profiles. Extracted lipids and defatted cell residues were subjected to sequential alkaline and acid methanolyses to distinguish ester- and amide-linked fatty acids in each fraction. In the lipid fractions, all the ester-linked fatty acids were nonhydroxylated, whereas all of the amide-linked fatty acids were hydroxylated. In the nonextractable fractions, both hydroxy and nonhydroxy fatty acids were found in both ester and amide linkage, although hydroxy acids predominated. The fatty acid profiles of the bound fractions differed widely from those of the lipid fractions. Bound fatty acid represented approximately 10% of the total cellular fatty acids.