Distribution of cholesteryl esters and other lipid in subcellular fractions of the adrenal gland of the pig

Total lipids from whole pig adrenal glands as well as from their mitochondria, microsomes, liposomes, and cell sap were extracted and fractionated first into neutral lipids and phospholipids. The highest percentage of neutral lipids was found in the cell sap, and the lowest in the microsomal fraction. Neutral lipids were subfractionated into cholesteryl esters, free cholesterol, triglycerides, and free fatty acids. Cholesteryl esters were distributed throughout the liposomes. Free fatty acids represented a substantial part of cell sap lipids, but were present also in the mitochondria, microsomes, and liposomes. Fatty acids of all fractions were analyzed by gas liquid chromatography. Free fatty acids and cholesteryl ester fatty acids from all cellular fractions were similar in composition and were characterized by considerable quantities of linoleic and arachidonic acid. Triglycerides were characterized by an increased percentage of palmitic and a low content of arachidonic acid. Phosphatidyl choline, phosphatidyl ethanolamine, diphosphatidyl glycerol, and sphingomyelin plus phosphatidyl inositol were isolated from the lipids by preparative thin layer chromatography, and their fatty acids analyzed by gas liquid chromatography. Phosphatidyl choline and phosphatidyl ethanolamine from mitochondria, microsomes, and cell sap were very similar in respect of their fatty acid composition. Sphingomyelin plus phosphatidyl inositol was characterized by a high content of C22:2omega6. Diphosphatidyl glycerol was present in mitochondria and in the cell sap.     

Analyse hoch ungesättigter Fettsäuremethylester

Analysis of Highly Unsaturated Methyl Esters of Fatty Acids The conditions for an optimum separation and identification of fatty acids having 0-6 double bonds were investigated. Thin-layer chromatographic separation of methyl esters of fatty acids on silver nitrate containing plates by a special technique in two different solvent systems enables a fractionation according to the degree of saturation as well as a resolution of highly unsaturated ω3 fatty acids from their ω6 positional isomers. Subsequent gas chromatography of individual fractions permits the examination of homologues with the same degree of saturation but different chain length.     

Lipid Composition of Garlic

Neutral, glyco- and phospholipids of garlic were resolved into their component fractions by thin layer chromatography. Neutral lipids contained considerable quantities of monoglycerides (18.5%), diglycerides (14.2%), sterols (16.3%) and triglycerides (41.5%) respectively. The phospholipid fraction was rich in phosphatidyl choline (23.5%), phosphatidyl ethanolamine (17.9%), lysophosphatidyl choline (11.8%) and lysophosphatidyl ethanolamine (8.2%). Digalactosyl diglyceride (10.1%), sterol glycoside (15.6%), cerebrosides (8.1%), acylsterol glycoside (38.6%) and monogalactosyl diglyceride (22.5%) were the major components of the glycolipids of garlic. Lauric, myristic, palmitic and linoleic acids constituted the major fatty acids of monoglycerides, diglycerides and free fatty acid fractions whereas palmitic, linoleic and linolenic acids were the major fatty acids of triglycerides. Palmitic and linoleic acids were the major fatty acids of garlic phospholipids. Except the acylsterol glycoside fraction glycolipids were rich in lauric, palmitic, linoleic and linolenic acids; palmitic acid was the only major fatty acid of acylsterol glycosides.     

Glycosphingolipids of human thyroid

Glycosphingolipids were isolated from total lipids of female and male human thyroids by alkaline hydrolysis, silicic acid, diethylaminoethyl-celluose and thin layer chromatography and analyzed by gas liquid chromatography. On the basis of their mobility in two dimensions on thin layer chromatography, IR analysis, and of sugar molar ratio, four neutral glycolipids, a sulfatide, and a hematoside fraction were identified. Glucosyl, plus galactosyl ceramide, and trihexosyl ceramide were the major fractions and accounted for 33% and 28% of total neutral glycolipids, respectively. Dihexosyl ceramide was a mixture of lactosyl and digalactosyl ceramide. The acidic lower phase glycolipids comprised ceramide galactosyl sulfate as the major component of male thyroids. Hematoside was identified tentatively as a minor component of the thyroids of both sexes. Major fatty acids of all neutral glycolipid fractions were 20∶0, 22∶0, 24∶0, and 24∶1; 24∶0 and 24∶1 for sulfatides. Low proportions of α-hydroxy fatty acids were identified. Total neutral glycosphingolipids of male thyroids were comparable in quantities with human liver but lower than kidneys, leucocytes, and platelets. Male thyroids comprised higher quantities of neutral glycosphingolipids (4.04±0.32 μmoles/g total lipid) as compared to females (2.34±0.21 μmoles/g total lipid), and much higher sulfatide than the females. These marked differences may suggest that the biosynthesis of the glycosphingolipids in the thyroid gland is under hormonal control. Similarities in glycosphingolipid composition of human thyroid and kidney are discussed in relation to a possible role played by glycolipids in ion transport, which is a common feature of the two organs.     

Chromatographic characterization of internal polar lipids from wool

Wool internal polar lipids were isolated and separated into different fractions based on polarity. Qualitative and quantitative analyses of the different fractions were performed by thin-layer chromatography and thin-layer chromatography coupled to flame-ionization detection, respectively. Cholesterol esters, free fatty acids, sterols, ceramides, glycosylceramides, and cholesterol sulfate were the main components, with ceramides being in the highest proportion. The fatty acid composition of ceramides and glycosylceramides was determined by gas chromatography/mass spectrometry. As for other keratinized tissues, long-chain fatty acids predominated in comparison to either free fatty acids or phospholipid-linked fatty acids; in both cases, stearic and lignoceric acids were the most abundant fatty acids, and a low amount of 18-methyleicosanoic acid was found. This work opens new avenues in the study of lipid rearrangement in more complex and realistic vesicle structures than conventional liposomes.     

Composition of fatty acids and structure of triglycerides in medium and low erucic acid rapeseed

Total triglycerides in medium (MEAR) and low (LEAR) erucic acid cultivars of rapeseed were fractionated by argentation chromatography into twelve and ten fractions, respectively. Gas liquid chromatography of the fatty acids in the triglyceride fractions and their 2-monoglycerides was used to evaluate the structural characteristics of the individual fractions. Fractionation occurred on the basis of degree of unsaturation, molecular weight and positional characteristics. The most mobile fractions contained 34–50% of saturated fatty acids while the less mobile had 59–65% of polyunsaturated fatty acids. In the medium erucic acid oil, long chain fatty acids (C20–C22) were found in all fractions, but four fractions of low erucic acid oil were essentially free of long chain acids. Two of these fractions in the latter oil, which represented 44% of the total triglycerides, were glycerol trioleate and dioleoyllinoleoylglycerol. The majority of the 2-positions were occupied by unsaturated C18 fatty acids, generally in the order of linoleic ≥linolenic> oleic acids. The saturated and long chain fatty acids occurred predominantly in the 1-and 3-positions. The various fractions of medium and low erucic acid oils were similar in structural composition except that eicosenoic and erucic acids substituted for oleic acid in some external positions. Erucic acid did not appear to substitute directly for oleic acid in the 2-position.     

Fatty acid composition of oleaster pulp and pit oils

Fatty acid compositions of oleaster pulp and pit oils were determined by gas chromatography in 4 samples of different varieties. Pit oils were highly unsaturated, containing >90% linoleic, oleic, and linolenic acids, as well as traces of palmitoleic acid. Saturated fatty acids consisted of palmitic and stearic acids with traces of arachidic acid. Pulp oils showed fatty acid compositions entirely different from that of pit oils. They contained 9 saturated fatty acids, C₁₂ to C₂₄, some of them with high quantities, up to 34.9%, of the total fatty acids. Unsaturated fatty acids, mainly oleic and linoleic, with low quantities of palmitoleic and linolenic acids composed about one-third of the total fatty acids.     

Fractional crystallization and gas chromatographie analysis of fatty acids as a means of detecting butterfat adulteration

A method has been devised which gives the distribution of saturated and unsaturated fatty acids of pure and adulterated cow and buffalo ghee with lard or margarine. It involves fractionation of pure and adulterated butterfat into fractions by fractional crystallization. The composition of the fatty acids liberated by the hydrolysis of each of the fractions was determined by gas chromatography. Adulteration of cow and buffalo ghee with various levels of lard or margarine caused significant changes in certain fatty acids, i.e., 22:0, 18:1, 18:0 and 16:0. It is possible to determine the extent of admixture of lard or margarine to either cow or buffalo ghee by applying a simple regression equation for certain fatty acids. This technique provides a basis for the detection of lipid adulteration.     

The distribution of lipids and sterols in cell types from the marine spongePseudaxinyssa sp

The sponge Pseudaxinyssa sp., unique in sterol and fatty acid composition, was cellularly dissected into fractions enriched in each of the major cell types present in the sponge: microbial symbionts (cyanobacteria), small sponge cells (pinacocytes and choanocytes), and large sponge cells (archeocytes and cyanophytes). Three phototrophic microbial symbionts were also isolated from the cell fractions and grown in culture. An unsymmetrical distribution of fatty acids and sterols was observed for the sponge cells: small cells contained larger quantities of long chain fatty acids (greater than C24) and smaller quantities of sterols than were present in the larger sponge cells. Moreover, the rare sterols 24-isopropylcholesterol predominated in the smaller sponge cells, whereas its 22-dehydro analog predominated in the larger sponge cells. Long chain fatty acids and sterols were not detected in the cultured microbial symbionts. This constitutes the first report of lipid variability according to cell type for this most primitive group of Metazoa.     

Recent advances in the analytical chemistry of fatty acids and derivatives

Recent advances in the analysis of industrial fatty acids and their derivatives almost always involve complex instrumentation. One of the most important developments in the analysis of fatty acids and their derivatives was the application of gas chromatography (GC). The result has been so effective that the time-consuming fractional distillation and detailed analysis of fractions previously employed are rarely used. Even though the so-called GLC technique has now been applied for over twenty years, new advances continue to be made in this area. Perhaps the most potentially valuable new development is the coupling of GLC with mass spectrometry (GLC-MS). Two newer chromatographic methods which have great potential in the field of fatty material analysis are thin layer chromatography (TLC) and high performance liquid chromatography (HPLC). The former is applicable to difficult separations; e.g., separation of broad lipid classes, in microquantities. The latter has already been applied with some success to the separation of individual component triglycerides of fats and oils and fatty acids using reverse phase HPLC. Instrumentation techniques include supportive methods which are frequently used in conjunction with other methods. Among these, the techniques of nuclear magnetic resonance (NMR) and infrared absorption (IR) are the most prominent.¹³C NMR is useful in defining fine structure of fatty acids, particularly with respect to branching. X-ray diffraction is used to study polymorphism in fatty acids.     

Study of color stability of tall oil fatty acids through isolation and characterization of minor constituents

Minor constituents in high quality tall oil fatty acids have been isolated successfully by liquid column chromatography, using silicic acid as the adsorbent. The minor constituents contained two types of compounds: those which were noneffective and those which were effective in causing the darkening of tall oil fatty acids during heating. The former consisted oftrans-3,5-dimethoxystilbene and rosin acids. The latter was separated into numerous fractions by the combination of chemical methods, silicic acid column chromatography, and low temperature fractional crystallization. The fractions were characterized by functional group analyses, chemical reactions, and UV and IR spectrometric methods. Most of the fractions contained two-three times as much oxygen in the molecule as the original sample and were highly oxidized fatty acids. They had mol wt ranging 300–551 and contained double bonds, carbonyl, ester, peroxide, and hydroxyl groups. The effect of these minor constituents upon the color stability of tall oil fatty acids during heating was postulated as being due to the hydroxyl groups located in the α-position to the double bond in the molecule.     

Hydrierung von Lipiden als Hilfsmittel zur Identifizierung und Quantifizierung von Phosphatiden aus Membransystemen des Herzmuskels

Hydrogenation of Lipids for Identification and Quantification of Phosphatides from Pellicle Systems of Cardiac Muscle. It was the aim of our research to show that hydrogenation of lipids is an auxiliary technique in phospholipid analysis of cardiac membranes. This is of interest if a preliminary overview on lipid fractions containing highly unsaturated fatty acids is needed. The fatty acids and the diglycerides from phospholipids were hydrogenated according to the procedure described by Appelqvist (A simple and convenient procedure for the hydrogenation of lipids on the micro- and nanomole scale, J. Lipid Res. 13 (1972), 146) with platinum oxide as a catalyst. The lipids (fatty acid methyl esters or acetylated diglycerides) were taken to dryness in a test-tube under nitrogen and flushed with hydrogen. The catalyst, suspended in methanol was injected through a septum. For identification purposes thin-layer chromatography on silica gel and on silica gel impregnated with silver nitrate was combined with gas chromatography before and after hydrogenation. After hydrogenation the fatty acid profile is much simpler and the fatty acid methyl esters can easily be differentiated from dimethyl acetals, as the latter are more volatile. Diacylglycerides and alkenylacylglycerides were also separated by thin-layer chromatography in individual subclasses before they were analysed by gaschromatography. Hydrogenating the lipids makes it possible to circumvent in part difficulties which arise often with polyunsaturated fatty acids. As the chain length of C20 and C22 are mainly represented by C20:4 , the arachidonic acid and C22:6 the docosahexaenoic acid, both fatty acids can be assessed after hydrogenation. The fatty acid profile of phosphatidylcholine and phophatidylethanolamine of cardiac muscle from rat, guinea pig and pig was determined. Each sample was analysed before and after hydrogenation. The fatty acids with the same chain length were summed up and compared to the corresponding chain length after hydrogenation.     

Lipids of the pawpaw fruit: Asimina triloba

The fatty acid composition and structure of pawpaw fruit (Asimina triloba) triglycerides were examined and found to contain fatty acids ranging from C₆ to C₂₀. Octanoate represented 20% of the fatty acids while other medium-chain fatty acids were present in low amounts. Analysis of the intact triglycerides by high-temperature gas-liquid chromatography gave an unusual three-cycle carbon number distribution. Analysis of triglyceride fractions separated according to degree of unsaturation suggested that one octanoate was paired with diglyceride species containing long-chain fatty acids. Determination of the double-bond positions of monoene fatty acids revealed cis Δ9 and cis Δ11 hexadecenoate and cis Δ9, cis Δ11, and cis Δ13 octadecenoate isomers were present in significant quantities. Octanoate and positional monoene fatty acid isomers were found only in the fruit lipids and not in the seed lipids. Phenacyl esters of fatty acids were found to be useful derivatives for structure determination using multiple types of analyses.     

Detection of cow milk admixture to buffalo milk

A method has been devised that gives the distribution of various fatty acids of pure and adulterated buffalo milk with cow milk. Gas chromatography (GC) was used for the qualitative and quantitative determination of fatty acids of authentic buffalo milk, cow milk and buffalo milk adulterated with cow milk. The milk fat was separated by fractional crystallization at -20 C into 2 fractions, i.e., semisolid and mother liquor. The concentration of fatty acids in the mother liquor changed significantly for 14:0, 16:0 and 18:1 as adulteration levels were increased. The fatty acids of the semisolid fractions change in the proportion of 16:0, 18:0 and 18:1 when cow milk is mixed with buffalo milk. By applying simple regression equations for these acids, adulteration of buffalo milk with 5% cow milk could be distinguished.     

Analysis of estolides in technical hydroxylated fatty acids from plant oils

Gel permeation chromatography of hydroxylated fatty acids (HOFA), prepared from various plant oils by a novel technical process, showed the presence of considerable amounts of estolides formed by intermolecular esterification of the HOFA. Thin-layer chromatographic fractionation followed by gas chromatography of the fractions revealed that the nonpolar estolides contain predominantly saturated fatty acids esterified tothero-9, 10-dihydroxy octadecanoic acid or dihydroxy tetrahydrofuran octadecanoic acids, e.g., 9,12-dihydroxy-10, 13-epoxy octadecanoic acid and 10,13-dihydroxy-9, 12-epoxy octadecanoic acid. The fractions of polar estolides consist mainly of intermolecular esters of the above dihydroxy fatty acids.     

Gas chromatography-mass spectrometry methods for structural analysis of fatty acids

Procedures for structural analysis of fatty acids are reviewed. The emphasis is on methods that involve gas chromatography-mass spectrometry and, in particular, the use of picolinyl ester and dimethyloxazoline derivatives. These should be considered as complementing each other, not simply as alternatives. However, additional derivatization procedures can be of value, including hydrogenation and deuteration, and preparation of dimethyl disulfide and 4-methyl-1,2,4-triazoline-3,5-dione adducts. Sometimes complex mixtures must be separated into simpler fractions prior to analysis by gas chromatography-mass spectrometry. Silver ion and reversed-phase high-performance liquid chromatography are then of special value. In particular, a novel application of the latter technique, involving a base-deactivated stationary phase and acetonitrile as mobile phase, is described that is suited to the separation of fatty acids in the form of picolinyl ester and dimethyloxazoline derivatives, as well as methyl esters.     

Detection of cow's milk admixture to buffalo's milk

Gas chromatography was used for the qualitative and quantitative determination of fatty acids of authentic buffalo's milk, cow's milk and buffalo's milk adulterated with cow's milk. The milk fat was separated by fractional crystallization at ?20°C into two distinct fractions, i. e., semi-solid and mother liquor. The concentration of the fatty acids in the mother liquor changed significantly for 14:0, 16:0 and 18:1 as adulteration levels were increased. The fatty acids of the semi-solid fractions changes in the proportion of 16:0, 18:0 and 18:1 when cow's milk is mixed with buffalo's milk. By applying a simple regression equation for these acids, adulteration of buffalo's milk with 5% cow's milk could be distinguished.     

Fatty acid compositions of lipid fractions from vegetative cells and mature sorocarps of the cellular slime moldDictyostelium discoideum

A wild-type strain ofDictyostelium discoideum was grown uponAerobacter aerogenes. Fatty acid compositions of lipid fractions and of total lipids obtained from vegetative amoebae and mature sorocarps were compared. Fatty acids isolated from vegetative cells were found to include large quantities of 17- and 19-carbon cyclopropane fatty acids while straight-chain, saturated fatty acids represented only 10% (w/w) of total fatty acids. These cyclopropane fatty acids appear to be derived from ingested bacteria and are preferentially incorporated into neutral lipids of the slime mold. Development of amoebae to mature sorocarps is accompanied by a substantial decrease in cyclopropane fatty acid content and a concomitant increase in unsaturated fatty acids, mostly as octadeca-5,11-dienoic acid. The †-22 stigmastenyl ester fraction is the richest source of this acid. Fully 65% of the fatty acids in this fraction are the octadecadienoate.     

A comparison of physical and chemical properties of milk fat fractions obtained by two processing technologies

Milk fat fractions from supercritical carbon dioxide (SC-CO₂) extraction were compared with commercial melt crystallization (MC) fractions for their physical and chemical properties. The fractions were analyzed for fatty acids, triacylglycerols, cholesterol, total carotenoid content, and volatile compounds. The fractions were also evaluated for solid fat content (SFC) by pulsed nuclear magnetic resonance and thermal profiles by differential scanning calorimeter (DSC). The distribution of fatty acids and triacylglycerols in the fractions depended on the fractionation technique used. SC-CO₂ separated fractions based on molecular weight rather than on melting point, which is the driving force for the MC process. The differences among the fractions were quantified from their SFC and DSC curves. Triacylglycerol profiles by high-performance liquid chromatography showed that the SC-CO₂ fractions were distinctly different from each other and from MC fractions. The SC-CO₂ solid fraction (super stearin) was the most unique. It had a high concentration of long-chain, unsaturated fatty acid-containing triacylglycerols in a narrow range of high molecular weight, indicating a homogeneity of this fraction that has not been attainable by other techniques. It was also enriched in β-carotene and was devoid of volatile compounds. As compared to liquid MC fractions, the liquid SC-CO₂ fraction had a high concentration of low-melting triacylglycerols and was enriched in volatile compounds. With SC-CO₂, it is thus possible to simultaneously fractionate and produce a flavor-rich concentrate at no extra processing cost.     

Detection of trace fatty acids in fats and oils by urea fractionation and gas-liquid chromatography

The detection of trace fatty acids (<0.1%) in a fat or oil by gas-liquid chromatography is possible when the methyl esters are fractionated with urea to provide a number of less complex fractions. Identification and estimation of trace fatty acids is simplified by quantitative removal of other fatty acids having similar gas chromatographic retention times. A detailed knowledge of the order in which inclusion compounds are formed was obtained by fractionating a complex mixture of marine and vegetable fatty acids. In addition, lanolin was fractionated to determine the preferential order in which saturated, branched chain (iso-, anteiso-) and hydroxy acids form inclusion compounds. Using urea fractionation and gas chromatography, 52 trace fatty acids were tentatively identified in butter, 30 in lard, and 26 in walnut oil. Presented at the AOCS Meeting in New Orleans, 1964.