New branched-chain fatty acids from the senegalese gorgonianLeptogorgia piccola (white and yellow morphs)

Fatty acids from total lipids of the gorgonianLeptogorgia piccola (white and yellow morphs), collected from the same area at two different periods with regard to the average water temperature, were studied. More than fifty fatty acids were identified as methyl esters andN-acyl pyrrolidides by gas chromatography and gas chromatography/mass spectrometry. Three new, branched-chain unsaturated fatty acids were identified in addition to the unusual 7-methyl-6-hexadecenoic acid, namely 10-methyl-6-hexadecenoic, 7,9-dimethyl-6-hexadecenoic, and 10-methyl-6,9-heptadecadienoic acids. Also 6,9-heptadecadienoic acid was identified. The fatty acid patterns of specimens harvested in colder waters were quite different from those harvested in warmer waters in that the former contained high amounts of methylene-interrupted polyunsaturated acids, including tetracosapolyenoic acids, especially 6,9,12,15,18–24∶5 (up to 15.8% of the total acid mixture) and 6,9,12,15,18,21–24∶6 (up to 5.3%). Arachidonic acid was, nevertheless, a major component in all the fatty acid mixtures studied (13.6–20.5%). Based on gas chromatography/Fourier transform infrared experiments, the double bonds were assigned the (Z) configuration. Several fatty aldehydes and their dimethyl acetals were also detected, of which the most abundant was octadecanal.     

Volatile components from tristearin heated in air

Tristearin was heated to 192 C in air, and its volatile oxidation products were collected directly on a cooled (−60 C) gas liquid chromatography column. Subsequently, the volatile products were separated by temperature programing up to 250 C and identified by mass spectrometry. Methyl ketones and aldehydes were the major degradation products along with minor amounts of monobasic acids, n-hydrocarbons, primary alcohols, and γ-lactones. Qualitative results indicated all the fatty acid methylene carbon atoms are susceptible to oxidation. Quantities of aldehydes and ketones were found to be in excess of their taste threshold concentrations, suggesting thermally oxidized saturated fatty acids may be precursors of some odors and flavors associated with heated lipids. Presented at the AOCS Fall Meeting, Philadelphia, September 1974. ARS, USDA.     

GLC and TLC analysis of isopropylidene derivatives of isomeric polyhydroxy acids derived from positional and geometrical isomers of unsaturated fatty acids

Gas-liquid chromatography (GLC) and thin-layer chromatography (TLC) were used to investigate the isomeric positional geometrical isopropylidene derivatives of nine isomeric dihydroxy esters, four isomeric methyl 9,10-12-trihydroxystearates, and eight isomeric methyl 9,10-12,13-tetrahydroxystearates prepared from unsaturated fatty acids. The isopropylidenes derived fromcis andtrans monounsaturated fatty acids were easily separated on both polar and nonpolar columns. Positional isopropylidenes derived from positional isomers of monounsaturated fatty acids were not separated on either liquid phase but were resolved by TLC. Four of the eight isomeric isopropylidenes derived from the four geometrical isomers of linoleic acid were resolved on the polar column; the other four isomers eluted as a single peak. The four isomeric isopropylidene-trifluoroacetate derivatives derived from ricinoleic and ricinelaidic acids were also resolved on the polar column. GLC analyses were carried out with liquid phases of ethylene glycol succinate methyl silicone polymer (EGSS-X) and methyl silicone polymer (SE-30) packed columns. Isopropylidenes, in addition to their applicability for the resolution of polyhydroxy acid mixtures, are particularly useful for the determination of double bond positions and geometrical configurations of fatty acids without cleavage. Under contract with the U. S. Atomic Energy Commission.     

Structure of bovine milk fat triglycerides

The triglycerides of bovine milk fat globules were isolated and separated into short, medium and long chain lengths by thin-layer chromatography. The molecular weight distribution and the fatty acid composition of the component triglycerides was then separately determined by gas chromatography following argentation-thin-layer and preparative gas chromatography. Some 38 triglyceride types (28% of total), of which there could be up to 6 isomers, were specifically identified and quantitatively estimated. The quantitative estimates for the rest of the milk fat triglycerides were limited to much more complex glyceride groups. The results confirm the earlier claim that butyric and caproic acids occur in milk fat almost exclusively in combination with medium and long chain fatty acids. Presented in part at the AOCS Meeting, Philadelphia, October, 1966.     

Gas-liquid chromatographic analysis of cyclopropenoid fatty acids

A method is described for the analysis of cyclopropenoid fatty acids in oils. The method consists of reacting the methyl esters of the cyclopropenoid fatty acids with silver nitrate in methanol to form ether and ketone derivatives. The derivatives formed from the cyclopropenoid fatty acids are separated from the methyl esters of the normal fatty acids by gas-liquid chromatography on a 15% diethylene glycol succinate column. The method is applicable to oils containing from 0.01% to 100% of cyclopropenoid fatty acids. The derivatives of oils containing lew levels of cyclopropenoids are separated from the normal methyl esters by alumina chromatography prior to gas-liquid chromatography. Studies on the quantitative aspects of the derivative formation, alumina chromatography, and gas-liquid chromatography are reported. Analyses for total cyclopropenoid fatty acid content of cottonseed oil andSterculia foetida oil by the gas-liquid chromatographic and hydrobromic acid titration procedures showed good agreement. Replicate analyses of a sample ofSterculia foetida oil for malvalic and sterculic acid gave coefficients of variation of 6.04% and 1.17%, respectively.     

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.     

Analysis of seed oils containing cyclopentenyl fatty acids by combined chromatographic procedures

The fatty acids of seed oils of the Flacourtiaceae,Hydnocarpus anthelmintica, Caloncoba echinata andTaraktogenus kurzii, have been examined by a combination of capillary gas chromatography, silver ion high performance liquid chromatography and gas chromatography-mass spectrometry. In addition to the common range of cyclopentenyl fatty acids found in such oils, 13-cyclopent-2-enyltridec-4-enoic acid was a major component ofH. anthelmintica and was identified by mass spectrometry as its picolinyl ester and dimethyldisulphide adduct. It has not previously been found in nature. In the other seed oils, the isolated double bond in the corresponding fatty acid was in position 6, as expected. Similarly,cis-4-hexadecenoic acid and C₁₆ and C₁₈ cyclopentyl fatty acids were identified for the first time inH. anthelmintica. Iso- andanteiso-methylbranched fatty acids were present in trace amounts.     

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.     

Improved gas chromatographic analysis of fatty and resin acid mixtures with special reference to tall oil

A gas liquid chromatography system for the analysis of complex mixtures of fatty and resin acids has been developed. On 30–40 m long, 0.3 mm ID glass capillary columns coated with 1,4-butanediol succinate (BDS) and attaining over 90,000 effective theoretical plates, all main fatty and resin acids in wood extractives and various tall oil products can be separated and quantitatively determined without need of any prefractionation of the acids. Also the levopimaric acid is well separated. Retention values, including their temperature dependence, on columns coated with BDS or SE-30 are given for 49 significant fatty and resin acids. Applications on wood extractives, sulfate soaps, and crude and distilled tall oil are presented and discussed.     

Phospholipid and fatty acid composition of Bulgarian nut oils

The phospholipid and fatty acid composition of three Bulgarian nut oils were investigated. Phospholipids were separated by Folch′s method and two-directional thin-layer chromatography. Their content was determined spectrophotometrically. Phospholipids were present at levels of 0.8% in almond oil, 2.8% in hazelnut oil, and 0.9% in walnut oil. Phosphatidylcholine (18—50%), phosphatidylinositol (18—45%), and phosphatidylethanolamine (8—16%) were found to be the major components. Small amounts of phosphatidylserine, phosphatidic acids, phosphatidylglycerols, lysophosphatidylcholine, and lysophosphatidylethanolamine were also detected. The fatty acid composition of glyceride oils and of the four main phospholipids, namely phosphatidylcholine, phosphatidylethanolamine, phosphatidylinositol and phosphatidic acids was identified by capillary gas chromatography of their methyl esters. The predominant fatty acid present in almond and hazelnut oils was linoleic (83.2% and 80.8%, respectively). Oleic acid (18.7%), linoleic acid (48.5%), and linolenic acid (15.8%) were the major components in walnut oil. Higher quantities of saturated fatty acids (27.8—81.2%) were found to be in the phospholipids than in the corresponding oils (9.5—16.7%).     

The fatty acids of the spongeDysidea fragilis from the black sea

The fatty acid composition of the sponge,Dysidea fragilis, from the Black Sea has been determined by analytical gas chromatography, silver ion high-performance liquid chromatography and gas chromatography/mass spectrometry. More than a hundred different fatty acids were identified, of which many were similar to those in sponges from tropical seas. On the other hand, some of the fatty acids identified have not been found previously in sponges or other marine sources and perhaps are new to science. These include 13-methyl-tetradec-4-enoic and 14-methyl-hexadec-6-enoic acids, together with demospongic acids,i.e. 5,9,17-tetracosatrienoic, 5,9,17-pentacosatrienoic and 5,9,19-pentacosatrienoic acids. From the elution behavior on silver nitrate chromatography, all the double bonds were of thecis-configuration.     

The Determination of Fatty Acid Primary Amides by Capillary Gas Chromatography

A method has been developed for the determination of 12 primary amides of long-chain fatty acids by capillary column gas chromatography. The method uses no derivatization or sample preparation other than extraction of the sample. A variety of commercial amidecontaining materials have been analyzed successfully. The amides, along with other soluble materials, are first separated from the host using refluxing 2-propanol containing an internal standard. The fatty acid amides are then identified and measured by gas chromatography over a programmed temperature range of 200 to 260 C. The chromatograms obtained show sharp peaks, unique retention times and acceptable reproducibility for quantitation.Cis- trans isomers of several of the fatty acid amides were tested and found to be resolved under the conditions employed.     

The effect of temperature on the fatty acid and phospholipid composition ofCephalosporium falciforme andCephalosporium kiliense

The effect of temperature on the lipid composition ofCephalosporium falciforme andCephalosporium kiliense, causative agents of maduromycosis, was investigated. The fungi were grown at 28.5 C and 37 C in a chemically defined medium. The lipids were solvent extracted, purified on Sephadex, and separated into their component classes by silicic acid column chromatography. Five lipid classes were found: (a) sterol esters, (b) triacylglycerides, (c) free fatty acids, (d) sterols, and (e) phospholipids. Fatty acids were analyzed by gas liquid chromatography and phospholipids by thin layer chromatography. Temperature induced changes of varying degrees occurred in both the fatty acid phospholipid fractions of each organism.     

Metabolism of arachidonate in rat testis: Characterization of 26–30 carbon polyenoic acids

Fatty acid methyl esters of long-chain polyenoic fatty acids (LCPA) from rat testis injected with [1-¹⁴C] arachidonate were analyzed and separated by reversed-phase high performance liquid chromatography (RP-HPLC). Earlier, all previously identified LCPA were prepared in high purity along with 4 previously unidentified fatty acids, which were further characterized by capillary gas chromatography (GC), catalytic hydrogenation and alkaline isomerization. Unidentified fatty acids proved to be 26∶4, 26∶5, 28∶5 and 30∶5. All of these LCPA incorporated¹⁴C from arachidonate (20∶4) to specific activities that were comparable to that of 20∶4 and previously identified metabolites of 20∶4 and much greater than specific activities of 18∶1n−9 or 22∶6n−3. LCPA were analyzed on a capillary GC system capable of resolving knowncis-trans and positional isomers of the n−3, n−6, n−7 and n−9 families of unsaturated fatty acids. Log plots of isothermal retention times and normal plots of temperature programmed retention times were linear (r=0.999) in carbon number when values for known and previously unidentified LCPA of 4 or 5 double bonds, respectively, were coplotted. Thus, the newly identified fatty acids belong to the n−6 family of fatty acids synthesized from arachidonic acid.     

Lipid metabolism ofAgaricus bisporus (Lange) sing.: I. Analysis of sporophore and mycelial lipids

The lipid components of four strains ofAgricus bisporus (Lange) Sing., the cultivated mushroom, were analyzed. Both sporophore and mycelial samples were obtained from beds in normal production. A method for obtaining mycelium free of compost was developed. Neutral lipids were separated from polar lipids by silicic acid column chromatography. Each fraction was separated by thin layer chromatography. Fatty acid methyl esters were analyzed by gas liquid chromatography and mass spectrometry. Sporophore extracts contained free sterol, free fatty acid, triglycerides, phosphatidyl choline and phosphatidyl ethanolamine. High amounts of linoleic acid were found in both neutral and polar lipid fractions. Mycelial extracts contained free fatty acids, triglycerides, phosphatidylcholine and phosphatidyl ethanolamine. No free sterol could be detected. Linoleic acid was also present in large amounts. Paper 3798 in the Journal Series of The Pennsylvania Agricultural Experiment Station.     

Analysis of hydroxylated fatty acids from plant oils

A novel process has been described recently for the preparation of hydroxylated fatty acids (HOFA) and HOFA methyl esters from plant oils. HOFA methyl esters prepared from conventional and alternative plant oils were characterized by various chromatographic methods (thin-layer chromatography, high-performance liquid chromatography, and gas chromatography) and gas chromatography-mass spectrometry as well as¹H and¹³C nuclear magnetic resonance spectroscopy. HOFA methyl esters obtained fromEuphorbia lathyris seed oil, low-erucic acid rapeseed oil, and sunflower oil contain as major constituents methylthreo-9,10-dihydroxy octadecanoate (derived from oleic acid) and methyl dihydroxy tetrahydrofuran octadecanoates, e.g., methyl 9,12-dihydroxy-10,13-epoxy octadecanoates and methyl 10,13-dihydroxy-9,12-epoxy octadecanoates (derived from linoleic acid). Other constituents detected in the products include methyl esters of saturated fatty acids (not epoxidized/derivatized) and traces of methyl esters of epoxy fatty acids (not hydrolyzed). The products that contain high levels of monomeric HOFA may find wide application in a variety of technical products.     

Analysis of north atlantic and baltic fish oil triacylglycerols by high-performance liquid chromatography with a silver ion column

Triacylglycerols from Atlantic herring (Clupea harengus), sandeel (Ammodytes sp.) and Baltic herring (Clupea harengus membras) have been fractionated by silver ion high-performance liquid chromatography. An ion exchange column loaded with silver ions was the stationary phase, and a gradient in the mobile phase from 1,2-dichloroethane/dichloromethane (1∶1, v/v) to acetone and then to acetone/acetonitrile (2∶1, v/v) was used to effect the separation with light-scattering (i.e., mass) detection. Fractions were collected via a streamsplitter, and fatty acid methyl esters were prepared by transesterification in the presence of an internal standard for identification and quantification by gas liquid chromatography. Triacylglycerols were separated according to the number of double bonds in the fatty acyl residues. Resolution was excellent at first, when the least unsaturated molecules eluted (trisaturated to dimonoene-monodiene fractions). Base-line resolution could no longer be achieved when molecules containing trienoic or more highly-unsaturated fatty acids began to elute because of overlapping components. Nonetheless, some valuable separations of species containing two saturated and/or monoenoic fatty acids and one polyenoic fatty acid were achieved. Double bond indices (average number of double bonds in each triacylglycerol molecule) were calculated to estimate the separations possible. Fractions containing at least 11–14 double bonds per molecule were obtained.     

Phospholipids of three xerophytic cucurbit seed oils

The composition of seed phospholipids was determined in three species of xerophytic cucurbits,Cucurbita digitata Gray,C. foetidissima HBK andApodanthera undulata Gray. The phospholipid fractions were isolated using silicic acid chromatography and quantitated by colorimetric analysis. The component phospholipids were separated using thin layer chromatography. All three species contained phosphatidylcholine, phosphatidyl-ethanolamine and phosphatidylinositol as their major component phospholipids. Analysis by gas liquid chromatography of fatty acids in total phospholipid samples revealed linoleic acid as the major component and myristic acid in significant amounts in each species. Small amounts of conjugated unsaturated fatty acids in the phospholipids of each species were determined by ultraviolet spectrometry. Close similarities in the composition of specific phospholipids were found in all species. This is Paper No. 3908 of the Arizona Agricultural Experiment Station.     

Proportions of C18∶1n−7 and C18∶1n−9 fatty acids in canola seedcoat surface and internal lipids

Lipids of canola seedcoats (Brassica napus L. andB. rapa L.) were prepared by surface washing and by complete extraction of seed coats with toluene. The major fatty acyl-containing triacylglycerols, wax esters and free fatty acids were separated by thin-layer chromatography prior to transesterification and analysis by gas-liquid chromatography. The proportion of C18∶1n−7 to C18∶1n−9 was higher in the extracted lipids than in the surface-washed lipids for all three classes.     

Novel branched-chain fatty acids in certain fish oils

Methyl-branched fatty acids, which are usually minor components (≤0.1%) in fish oils, were concentrated in the non-urea-complexing fraction along with polyunsaturated fatty acids during the enrichment of omega-3 fatty acids from certain fish oils via the urea complexation process. The methyl-branched fatty acids in the omega-3 polyunsaturated fatty acid concentrates, which were prepared from three fish body oils, were characterized by gas chromatography and gas chromatography/mass spectrometry. Among the major branched-chain fatty acids expected and identified were the known isoprenoid acids—mainly 4,8,12-trimethyltridecanoic, pristanic, and phytanic—and the well-known iso and anteiso structures. Two novel phytol-derived multimethyl-branched fatty acids, 2,2,6,10,14-pentamethylpentadecanoic and 2,3,7,11,15-pentamethylhexadecanoic, were identified in redfish (Sebastes sp.) oil. These two fatty acids were absent in oils from menhaden (Brevoortia tyrannus) and Pacific salmon (mixed, but mostly from sockeye,Oncorhynchus nerka). The major branched-chain fatty acid in the salmon oil, 7-methyl-7-hexadecenoic acid, was also present to a moderate extent in menhaden oil. A novel vicinal dimethyl-branched fatty acid, 7,8-dimethyl-7-hexadecenoic was detected in all of the fish oils examined, but was most important in the salmon oil. Three monomethyl-branched fatty acids, 11-methyltetradecanoic acid, and 11- and 13-methylhexadecanoic, hitherto undescribed in fish lipids, were also detected in salmon, redfish and menhaden oils. Presented in part at the First Annual Meeting of the Amer. Oil. Chem. Soc. Canadian Section, Guelph, Ontario, Oct. 8–9, 1986.