Urea-based fractionation of seed oil samples containing fatty acids and acylglycerols of polyunsaturated and hydroxy fatty acids

The selectivity and efficiency of urea complex (UC) formation-based fractionation of free fatty acids (FFA) were examined. A rapid, simple, and inexpensive procedure recently developed for urea fractionation was applied to lipid mixtures containing various polyunsaturated and hydroxy FFA species. Urea treatment proved useful for isolating polyunsaturated FFA (PUFA) from FFA derived from fish, borage, and linseed oils by removal of saturated and monounsaturated FFA, but was not effective for isolating hydroxy FFA from the FFA derived from castor, Lesquerella, and Dimorphotheca oils. In situations where FFA within the crystalline or UC phase were rich in PUFA, the urea/FFA mole ratio of the UC was relatively higher, with lower recovery of FFA in this phase. The distribution of urea between the crystalline phase and the solvent was not significantly affected by the FFA composition of feed nor the overall ratio of FFA to urea. It was strongly dependent on the overall mass fraction of solvent. Phospholipids and mono-, di-, and triacylglycerols were poor templates for UC formation relative to FFA. Their inclusion in acylglycerol mixtures containing FFA reduced UC formation.     

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.     

Production of hydroxy fatty acids from unsaturated fatty acids byFlavobacterium sp. DS5 hydratase,a C-10 positional- andcis unsaturation-specific enzyme

A new microbial isolate,Flavobacterium sp. DS5, converted oleic and linoleic acids to their corresponding 10-keto-and 10-hydroxy fatty acids. The hydration enzyme seems to be specific to the C-10 position. Conversion products from α- and γ-linolenic acids were identified by gas chromatography/mass spectrometry, Fourier transform infrared, and nuclear magnetic resonance as 10-hydroxy-12(Z),15(Z)-octadecadienoic and 10-hydroxy-6(Z),12(Z)-octadecadienoic acids, respectively. Products from other 9(Z)-unsaturated fatty acids also were identified as their corresponding 10-hydroxy- and 10-keto-fatty acids.Trans unsaturated fatty acid was not converted. From these results, it is concluded that strain DS5 hydratase is indeed a C-10 positional-specific andcis-specific enzyme. DS5 hydratase prefers an 18-carbon monounsaturated fatty acid. Among the C₁₈ unsaturated fatty acids, an additional double bond at either side of the 9,10-position lowers the enzyme hydration activity. Because hydratases from other microbes also convert 9(Z)-unsaturated fatty acids to 10-hydroxy fatty acids, the C-10 positional specificity of microbial hydratases may be universal.     

Structure-activity relationship of unsaturated fatty acids as mosquito ovipositional repellents

Various straight-chain unsaturated fatty acids from C₁₄ to C₂₄ were evaluated for their ovipositional repellency against gravid females of the southern house mosquitoCulex quinquefasciatus Say, and the relationship between the structures of the fatty acids and their ovipositional repellency was determined. A double bond withZ configuration was prerequisite for an unsaturated fatty acid to be highly repellent;E isomers were less active or even inactive. No relationship was found between the repellency and the number of double bonds in the unsaturated fatty acids. In C₁₈ monounsaturated fatty acids, (Z)-9 acid was more active than (Z)-11 and (Z)-6 acids, indicating that a double bond at the 9 position rendered an acid highly repellent. Among (Z)-9-alkenoic acids of different chain lengths, the most repellent was C₁₈ acid which was also more active than (Z)-11-C₂₀, (Z)-13-C₂₂, and (Z)-15-C₂₄ acids. Oleic[(Z)-9-octadecenoic]acid, which met all these criteria, was the most ovipositionally repellent among the unsaturated fatty acids tested.Diptera: Culicidae.     

Monoethylenic fatty acids of a partially hydrogenated herring oil

A Canadian Atlantic herring oil hydrogenated for margarine use to an iodine value of 76 and melting point of 32.5 C was found to have 30% saturated acids and 66% monounsaturated fatty acids. The monounsaturated fatty acids could be analytically determined ascis andtrans isomers by open tubular gas liquid chromatography.Trans acids were 33% of the C₁₆ and C₁₈ monounsaturated acids, and 32 and 28%, respectively, of the C₂₀ and C₂₂ monounsaturated acids. After separation of geometric isomers by Florisil-silver nitrate chromatography the positional isomers in each class were determined by oxidative fission. The double bond positions of the originalcis fatty acids were largely retained in bothcis andtrans isomers, but additional isomers were observed, especially in thetrans fatty acids.     

Variation in fatty acid composition of the different acyl lipids in seed oils from fourSesamum species

Seeds from different collections of cultivatedSesamum indicum Linn. and three related wild species [specifically,S. alatum Thonn.,S. radiatum Schum and Thonn. andS. angustifolium (Oliv.) Engl.] were studied for their oil content and fatty acid composition of the total lipids. The wild seeds contained less oil (ca. 30%) than the cultivated seeds (ca. 50%). Lipids from all four species were comparable in their total fatty acid composition, with palmitic (8.2–12.7%), stearic (5.6–9.1%), oleic (33.4–46.9%) and linoleic acid (33.2–48.4%) as the major acids. The total lipids from selected samples were fractionated by thin-layer chromatography into five fractions: triacylglycerols (TAG; 80.3–88.9%), diacylglycerols (DAG; 6.5–10.4%), free fatty acids (FFA; 1.2–5.1%), polar lipids (PL; 2.3–3.5%) and steryl esters (SE; 0.3–0.6%). Compared to the TAG, the four other fractions (viz, DAG, FFA, PL and SE) were generally characterized by higher percentages of saturated acids, notably palmitic and stearic acids, and lower percentages of linoleic and oleic acids in all species. Slightly higher percentages of long-chain fatty acids (20∶0, 20∶1, 22∶0 and 24∶0) were observed for lipid classes other than TAG in all four species. Based on the fatty acid composition of the total lipids and of the different acyl lipid classes, it seems thatS. radiatum andS. angustifolium are more related to each other than they are to the other two species.     

Biotransformation of saturated monohydroxyl fatty acids to 2-tetrahydrofuranyl acetic acid derivatives: Mechanism of formations and the biological activity of 5-n-hexyl-tetrahydrofuran-2-acetic acid

Transformation of 12-hydroxyoctadecanoic acid (12-HOA) to 5-n-hexyl-tetrahydrofuran-2-acetic acid (5-HTFA) byBacillus lentus NRRL B-14864 (B-14864) was carried out in the presence or absence of oligomycin, 2-bromooctanoic acid (2-BA), or sodium azide. In addition, several saturated and monounsaturated monohydroxyfatty acids, saturated monooxofatty acids, and monounsaturated fatty acid were used as substrates for transformation reactions by B-14864 or corynebacterium FUI-2. Methyl esters of the transformation products were analyzed by gas chromatography and gas chromatography/mass spectroscopy. Various γ-lactones and tetrahydrofuran fatty acid derivatives were conversion products when saturated monohydroxyfatty acids were used as substrates; the production of 5-HTFA from 12-HOA by B-14864 cells was completely inhibited in the presence of high concentration of oligomycin, 2-BA, or sodium azide; and fatty acid β-oxidation metabolic intermediates, 6-hydroxydodecanoic, 4-oxododecanoic, and 4-oxodecanoic acids were products when 12-HOA, 10-oxo-, and 12-oxooctadecanoic acids were used as substrates. Our results suggest that the production of 5-HTFA from 12-HOA by B-14864 was through the fatty acid β-oxidation pathway. Three-day-old driedfruit beetle pupae were topically treated with 5-HTFA to test for juvenile hormone activity, and 5-HTFA was found to possess juvenile hormone-like activity in pure form but not when it was diluted to 10%. Presented in part at the 37th West Central States Biochemistry Conference Annual Meeting, Columbia, Missouri, Oct. 1994.     

Rapid determination of double-bond positions in monoenoic fatty acids by periodate-permanganate oxidation

Tetramethylammonium hydroxide has been used in the extraction and pyrolysis methylation of the carboxylic acids produced by periodate-permanganate oxidation of monounsaturated fatty acid methyl esters. This modification of the von Rudloff procedure allows rapid determination of double-bond positions and analysis of mixtures of positional isomers of monoenoic fatty acids.     

Enzymatic synthesis of steryl esters of polyunsaturated fatty acids

Steryl esters of long-chain fatty acids have water-holding properties, and polyunsaturated fatty acids (PUFA) have various physiological functions. Because steryl ester of PUFA can be expected to have both features, we attempted to synthesize steryl esters of PUFA by enzymatic methods. Among lipases used, Pseudomonas lipase was the most effective for the synthesis of cholesteryl docosahexaenoate. When a mixture of cholesterol/docosahexaenoic acid (3:1, mol/mol), 30% water, and 3000 units/g of lipase was stirred at 40°C for 24 h, the esterification extent attained 89.5%. Under the same reaction conditions, cholesterol, cholestanol, and sitosterol were also esterified efficiently with docosahexaenoic, eicosapentaenoic, arachidonic, and γ-linolenic acids.     

Fatty acid composition of meat from ruminants, with special emphasis on trans fatty acids

The fatty acid composition was determined in 39 samples of beef, 20 samples of veal, and 34 samples of lamb, representative of the supply of ruminant meat in Denmark. Five cuts of beef and veal and three cuts of lamb with increasing fat content were selected, and analysis of the fatty acid methyl esters was performed by gas-liquid chromatography (GLC) on a polar 50-m capillary column CP Sil 88 with flame-ionization detection. Lamb had the highest content of saturated fatty acids (52.8±1.8 g/100 g fatty acids), higher than beef and veal (45.3±3.1 and 45.4±0.8 g/100 g fatty acids, respectively). Cis monounsaturated fatty acids were 49.2±3.1, 44.9±1.8, and 37.7±1.7, and polyunsaturated fatty acids were 3.3±0.7, 5.8±2.0, and 5.0±0.1 g/100 g fatty acids in beef, veal, and lamb, respectively. Beef contained 2.1±0.8 g trans C_18:1 per 100 g fatty acids, about half that found in veal (4.0±1.2 g/100 g fatty acids) and lamb (4.5±0.6 g/100 g fatty acids). Trans C_16:1 was 0.24±0.01, 0.14±0.02, and 0.79±0.02 g/100 g fatty acids in beef, veal, and lamb, respectively. Only small variations in trans and other fatty acids could be demonstrated between cuts. The overlap between cis and trans C_18:1 by capillary GLC was verified by argentation-thin-layer chromatography followed by GLC, on three samples of veal and three samples of lamb. In veal 1.0 g, and in lamb 1.4 g trans C_18:1 per 100 g fatty acids were hidden under the cis C_18:1 peak. The mean intake of trans fatty acids from ruminant meat is estimated at 0.2 g/d.     

Perineal scent gland of wild and domestic guinea pigs

Wild and domestic male guinea pigs (Cavia aperea andCavia porcellus) prefer the perineal secretion from males of the same species to that of males of the other species. Gas chromatographic-mass spectroscopic analyses of the volatile components of the secretions show complex mixtures comprised primarily of fatty acids, alcohols, and ketones. Inter-species differences in the composition of the volatiles are evident. The possible role of bacteria in odor production is discussed.Camille and Henry Dreyfus Teacher-Scholar, 1978–1983.     

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.     

Liver lipids and fatty acids of the sting ray Dasyatis bleekeri (Blyth)

The sting ray, Dasyatis bleekeri (Blyth), has been studied for lipids and fatty acids of its liver. The neutral lipids identified were hydrocarbons, wax esters, steryl esters, 1-O-alkyl-2,3-diacylglycerols, triacylglycerols, and sterols. Neutral lipids were predominant (91.8%), major components being triacylglycerols (92.7%). Polyenoic fatty acids of n-3 series, viz. eicosapentaenoic acid and docosahexaenoic acid, were high in the phospholipid and neutral lipid fractions. Cholesterol was the major component (67.9%) in the steryl ester fraction. Glyceryl ethers, with chainlengths up to 30 carbons, were recorded with unsaturated, anteiso, iso, and normal chains. In wax ester alcohols, up to 32-carbon chains were recorded. Hydrocarbons were up to 36-carbon chains with anteiso, iso, and normal chains. Among branched chain hydrocarbons, pristane was the major component (6.7%) and squalene was present at the level of 3.5%. Chimyl and batyl alcohol backbones were the major components found in 1-O-alkyl-diacylglycerols.     

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.     

Identification of cyclopentenyl fatty acids by 1H and 13C nuclear magnetic resonance

Gorlic, chaulmoogric and hydnocarpic fatty acids, specific to the seed oil of the genus Hydnocarpus sp. (Flacourtiaceae), are determined only with difficulty by gas chromatography. These fatty acids were isolated in their methyl ester form by a combination of different chromatographic techniques (thin-layer chromatography/Ag⁺ and high-pressure liquid chromatography). The proton and carbon nuclear magnetic resonance analysis of these fatty acid methyl esters showed some characteristic signals of the cyclopentenyl ring. The presence of these signals in the proton and/or carbon nuclear magnetic resonance spectrum of an oil thus will allow us to confirm the presence of these cyclopentenyl fatty acids in lipids.     

The enrichment of n−3 polyunsaturated fatty acids using aminopropyl solid phase extraction columns

A rapid, simple and reliable method is described for the preparation of concentrates of methyl or ethyl esters of n−3 polyunsaturated fatty acids by solid phase extraction using aminopropyl bonded silica columns. After applying mixtures of fatty acid esters in hexane, saturated and monounsaturated fatty acid esters are preferentially eluted with hexane whereas polyunsaturated fatty acids (PUFA) can subsequently be eluted with dichloromethane. Concentrates containing 80–90% n−3 PUFA can thus be obtained using fish oil fatty acids esters as a starting material.     

Determination of cyclopropenoid fatty acids inSterculia seed oils from senegal

Seed oils ofSterculia tomentosa andS. tragacantha (Sterculiaceae) were found to contain malvalic (5.8 and 5.1%), sterculic (11.3 and 30.2%) and dihydrosterculic (0.9 and 0.5%) acids. The total amount of these two cyclopropenoid fatty acids was established by¹H nuclear magnetic resonance and their cooccurrence by gas chromatography. Besides these unusual compounds, the main common fatty acids were palmitic (20 and 24%), oleic (21 and 15%) and linoleic (30 and 16%) acids.     

Influence of moderate amounts of trans fatty acids on the formation of polyunsaturated fatty acids

The effect of trans fatty acids from partially hydrogenated soybean oil and butterfat on the formation of polyunsaturated fatty acids was investigated. Five groups of rats were fed diets that contained 20 wt% fat. The content of linoleic acid was adjusted to 10 wt% of the dietary fats in all diets, whereas the amount of trans fatty acids from partially hydrogenated soybean oil (PHSBO) was varied from 4.5 to 15 wt% in three of the five diets. The fourth group received trans fatty acids from butterfat (BF), while the control group was fed palm oil without trans fatty acids. Trans fatty acids in the diet were portionally reflected in rat liver and heart phosphatidylethanolamine (PE), phosphatidylcholine (PC), phosphatidylinositol, and phosphatidylserine. Incorporation in the sn-1 position was compensated by a decrease in saturated fatty acids. Trans fatty acids were not detected in diphosphatidylglycerol. Compared to the presence in the dietary fats, 8t- and 10t-18:1 were discriminated against in the incorporation in PE and PC from liver and heart, whereas 9t- and 12t-18:1 were preferred. The formation of 20:4n-6 was not influenced by 4.5 wt% trans fatty acids (from PHSBO) but apparently was by 10 wt% in liver. In contrast, even a content of 2.5 wt% trans fatty acids from BF reduced the formation of 20:4n-6. The inhibitory effect of trans isomers on linoleic acid conversion was reflected less in heart than in liver and less for PE than for PC. Groups with trans fatty acids showed increased 22:6n-3 and 22:5n-3 deposition in liver and heart PE and PC.     

Alteration of steryl ester content and positional distribution of fatty acids in triacylglycerols by chemical and enzymatic interesterification of plant oils

Steryl ester content of refined and interesterified corn, soybean, and rapeseed oils has been measured via clean-up on a short silica gel column, followed by high performance liquid chromatography with evaporative light-scattering mass detector. Chemical interesterification, catalyzed by sodium methoxide, led to random positional distribution of fatty acids in triacylglycerols and some increase in the steryl ester content of all three oils. Enzymatic interesterification, catalyzed by the immobilized lipase from Rhizomucor miehei (Lipozyme), resulted in a distinct reduction in steryl ester content, but essentially no alteration in positional distribution of fatty acids in triacylglycerols occurred. Formation of steryl esters during chemical and enzymatic interesterification was also examined by radioactive tracer technique with [4-¹⁴C]β-sitosterol added as marker to refined rapeseed oil and measurement of the radioactive steryl esters formed. Chemical interesterification of rapeseed oil resulted in moderate formation (10% of total radioactivity) of radioactive β-sitosteryl esters. Enzymatic interesterification of the oil, catalyzed by Lipozyme, led to little formation of radioactive β-sitosteryl esters, whereas with the lipase from Candida cylindracea high proportions (>90% of total radioactivity) of ¹⁴C-labeled β-sitosteryl esters were formed. Part of doctoral thesis of Roseli Ap. Ferrari to be submitted to Faculdade de Engenharia de Alimentos, Universidade de Campinas, Campinas, Brazil.     

Estolides from meadowfoam oil fatty acids and other monounsaturated fatty acids

The formation of estolides was detected during the studies on dimerization of meadowfoam oil fatty acids. By adjusting the reaction conditions, it was possible to produce monoestolides with little dimer or trimer formations. Estolides have potential use in lubricant, cosmetic and ink formulations and in plasticizers. This paper reports the conditions for production of estolides from mixed meadow-foam fatty acids, commercial oleic acid, high-oleic sun-flower oil fatty acids,cis-5,cis-13-docosadienoic acid, petroselinic acid and linoleic acid.