Production of polyhydroxy fatty acids from linoleic acid by Clavibacter sp. ALA2

Hydroxy fatty acids are important industrial materials. We isolated a microbial culture, Clavibacter sp. ALA2, which converts linoleic acid to many polyhydroxy fatty acids. Structures of the products were determined as: 12,13,17-trihydroxy-9(Z)-octadecenoic (THOA, main product), 12-[5-ethyl-2-tetrahydrofuranyl]-7,12-dihydroxy-9(Z)-dodecenoic (ETDDA), and 12-[5-ethyl-2-tetrahydrofuranyl]-12-hydroxy-9(Z)-dodecenoic (ETHDA) acid. The yield of THOA was 25% and the relative amount of the products were THOA/ETDDA/ETHDA =9:1.3:1. The structures of the hydroxy unsaturated fatty acids resemble those of plant self-defense substances.     

A convenient preparative method of 3-hydroxy acids,and the reaction of 3-hydroxy acids with acidic materials

3-Hydroxy acids were synthesised in good yield from ketones and carboxylic acids using lithium naphthalenide in the presence of diethylamine. From cyclohexanone and propionic acid, 2-(1′-hydroxycyclohexan-1′-yl) propionic acid ( I ) was obtained in 98.3% yield. 3-Hydroxy acids were treated with various acidic materials to give 3,4-unsaturated carboxylic acids and γ-butyrolactones. From the reaction of ( I ) with p-toluenesulphonic acid or potassium bisulphate, 2-(cyclohexen-1′-yl) propionic acid ( II ) (a 3,4-unsaturated acid) was obtained (yield 98%), and with 97% sulphuric acid 2-(1′-hydroxycyclohexan-1′-yl) propionic acid lactone ( III ) was formed (93% yield). Several new γ-butyrolactones were obtained in good yield by this method.     

Cyclic fatty esters: Synthesis and characterization of methyl ω-(6-alkyl-3-cyclohexenyl) alkenoates

Diunsaturated C₁₈ cyclic fatty acid methyl esters of known structure and configuration were synthesized as model derivatives of cyclic fatty acids formed in heat-abused vegetable oils for characterization and further biological evaluation. The Wittig reaction was used to prepare 5 pure methyl esters: (a) 12-(3-cyclohexenyl)-11-dodecenoate, (b) 11-(6-methyl-3-cyclohexenyl)-10-undecenoate, (c) 10-(6-ethyl-3-cyclohexenyl)-9-decenoate, (d) 9-(6-propyl-3-cyclohexenyl)-8-nonenoate and (e) 8-(6-butyl-3-cyclohexenyl)-7-octenoate. Diels-Alder cycloaddition reactions between 1,3-butadiene and appropriate (E)-2-alkenals produced 3-cyclohexenal intermediates. The appropriate methyl ω-bromoesters and their triphenylphosphonium bromides were made and converted to their respective ylids with NaOCH₃ in DMF. The appropriate 3-cyclohexenals and phospho-ylids were reacted, and the desired cyclic ester products were isolated in crude yields of 30–83% as liquids and fractionally distilled. The crude cyclic esters were purified either by preparative TLC or by saponification-esterification. Double bonds in purified cyclic esters weretrans-isomerized and hydrogenated. Each derivative was characterized by IR,¹H-NMR,¹³C-NMR, capillary GLC and GC-MS. On the basis of these analyses, no positional isomers were detected, Z-unsaturated isomers were produced in better than 90% purity, and the alkyl and ester ring substituents were predominantlytrans to each other. Presented in part at the AOCS meeting, New Orleans, LA, May 1981.     

Derivatization of keto fatty acids: II. synthesis of long-chain enolacetates

Long-chain enolacetates were prepared by refluxing the keto fatty acids with acetic anhydride andp-toluene sulfonic acid (p-TSA). 10-Oxoundecanoic and 2-oxohexadecanoic acids gave mainly 10-acetoxy-10-undecenoic and 2-acetoxy-2-hexadecenoic acids, respec-tively. On the other harld, 12-oxooctadecanoic acid on similar treatment was assumed to yield an isomeric mixture of 12-acetoxy-12(11)-octadecenoic acids. The structure of the individual reaction products were established by spectral studies.     

Lewis Acid Induced Additions to Unsaturated Fatty Compounds III: Alkylaluminium Halide Induced Friedel-Crafts Acylations of Unsaturated Fatty Compounds

Alkylaluminium halide induced Friedel-Crafts acylations of unsaturated fatty compounds with acyl chlorides give the corresponding β,γ-unsaturated ketones. Acylation of oleic acid and 10-undecenoic acid with acetyl chloride yields (E)-9(10)-acetyloctadec-10(8)-enoic acid and 12-oxo-tridec-9-enoic acid, respectively. Acylation of 10-undecenoic acid with heptanoyl chloride followed by NaBH₄-reduction gives ricinelaidic acid. The very interesting β,γ-unsaturated ketodicarboxylic acid (E)-9(10)-(1-oxo-3-carboxy-propyl)octadec-10(8)-enoic acid is obtained by acylation of oleic acid with succinic anhydride.     

New additives derived from fatty acids for water-based cutting fluids

A variety of derivatives from long chain fatty acids were prepared, and corrosion and lubricity tests for these products as water-based cutting fluid additives were carried out. We have found that triethanolamine salts of 11-(carboxymethylthio)-undecanoic acid (II), 10,11-dichloromethylene undecanoic acid (IV) and 12-isovaleroyloxy stearic acid (VI) showed effective rust-inhibiting and anti-wear properties for water-based cutting fluids.     

Oxidative cyclization of unsaturated fatty acids with lead tetraacetate

The reaction of 10-undecenoic acid with lead tetraacetate (LTA) in the presence of acetic acid and anhydrous potassium acetate gave a eutectic mixture of 10(11)-acetoxy-11(10)-methylundecanoic acid (25%), 10,11-diacetoxyundecanoic acid (15%) and 5-(ω-carboxyoctyl) γ-butyrolactone (45%). Similar reaction withcis-9-octadecenoic acid yielded a eutectic mixture of 9(10)-acetoxy-10(9)-methyloctadecanoic acid (20%) and 4(5)-(ω-carboxyheptyl)-5-(4)-octyl γ-butyrolactone (40%). In the reaction withtrans-2-enoic acids (C₁₆ and C₁₈) products obtained were: 1,2-diacetoxypenta- and heptadecane (27%), 3-acetoxyhexa- and octadecanoic acid (31%) and 2-(2-penta- and heptadecyl) succinic anhydride (23%), respectively. The structures of individual compounds were characterized by spectral methods.     

Produkte der thermischen En-Reaktion von ungesättigten Fettstoffen und Maleinsäureanhydrid

Products of the Thermal Ene-Reaction of Unsaturated Fatty Compounds and Maleic Anhydride The thermal ene reaction of the methylates of unsaturated fatty acids 10-undecenoic acid, oleic acid and (E)-10-eicosendioic acid with maleic anhydride was carried out at 190°C. The mono addition products were isolated and their stereochemistry was deduced from ¹HNMR- and ¹³C-NMR-spectroscopy.     

Diels-alder adducts from safflower fatty acids: III. Acrylic and related acid dienophiles

The Diels-Alder adducts from alkali conjugated and elaidinized safflower fatty acids with acrylic, methacrylic, crotonic and cinnamic acids and their esters as dienophiles were obtained in 40–64% yields. Low yields of 16–18% were obtained when 10-undecenoic acid and its ester were used. Most of the adducts could be estimated quantitatively in the reaction products by column chromatography or gas liquid chromatography (GLC). For the estimation of the adducts from crotonic and 10-undecenoic acids a combination of column chromatography and GLC techniques was employed. Partitioning between hexane and 90% methanol, partial esterification with alkali washing, and splitting and fractional distillation under reduced pressure were methods used to obtain the adducts in 90–95% purity.     

Fatty acid distribution in the phospholipids ofFrancisella tularensis

Francisella tularensis, LVS (live vaccine strain) grown in a chemically defined medium was found to have a lipid content of 21% by dry weight. The two major phospholipids were identified as phosphatidylethanolamine (PE; 76%) and phosphatidylglycerol (PG; 24%) by thin layer chromatographic analysis, staining characteristics and quantitative chemical analyses of fatty acid, phosphate and glycerol constituents. PE contained a high proportion of 24∶0 fatty acid, with lesser amounts of 24∶1, 22∶0 and 10∶0. The major fatty acids of PG were 18∶1 and 22∶0. Hydroxy fatty acids, which are prominent components ofF. tularensis, were conspicuously lacking in these phospholipids; it is therefore concluded that hydroxy fatty acids are constituents of other structures of the organism.     

Gut Microbial Fatty Acid Metabolites Reduce Triacylglycerol Levels in Hepatocytes

Hydroxy and oxo fatty acids were recently found to be produced as intermediates during gut microbial fatty acid metabolism. Lactobacillus plantarum produces these fatty acids from unsaturated fatty acids such as linoleic acid. In this study, we investigated the effects of these gut microbial fatty acid metabolites on the lipogenesis in liver cells. We screened their effect on sterol regulatory element binding protein‐1c (SREBP‐1c) expression in HepG2 cells treated with a synthetic liver X receptor α (LXRα) agonist (T0901317). The results showed that 10‐hydroxy‐12(Z)‐octadecenoic acid (18:1) (HYA), 10‐hydroxy‐6(Z),12(Z)‐octadecadienoic acid (18:2) (γHYA), 10‐oxo‐12(Z)‐18:1 (KetoA), and 10‐oxo‐6(Z),12(Z)‐18:2 (γKetoA) significantly decreased SREBP‐1c mRNA expression induced by T0901317. These fatty acids also downregulated the mRNA expression of lipogenic genes by suppressing LXRα activity and inhibiting SREBP‐1 maturation. Oral administration of KetoA, which effectively reduced triacylglycerol accumulation and acetyl‐CoA carboxylase 2 (ACC2) expression in HepG2 cells, for 2 weeks significantly decreased Srebp‐1c, Scd‐1, and Acc2 expression in the liver of mice fed a high‐sucrose diet. Our findings suggest that the hypolipidemic effect of the fatty acid metabolites produced by L. plantarum can be exploited in the treatment of cardiovascular diseases or dyslipidemia.     

Periodate-permanganate oxidations for determining location and amount of unsaturation in monounsaturated fatty acids

Conclusions Periodate-permanganate oxidation of three samples of oleic acid considered to be of high purity has given reproducible but less than theoretical amounts of total dibasic acids. Recovery approximated 92% with about 90.3% of the expected azelaic acid. Recovery of other dibasic acids indicated that about 1.5% of the total unsaturation of these samples of oleic acid was present in positions 8 or 10 in the fatty acid molecule. Oxidation of elaidic acid produced from one of the oleic acids has given total dibasic acid yield of about 96%, with a smaller amount of its total unsaturation in the same position as in the parent oleic acid. Oxidation of high purity 9,10-dihydroxystearic acid has given essentially quantitative yield of total dibasic acids. The method described should be useful in determining the composition of similar unsaturated positional isomers. Controls showing the effect of the method on azelaic acid and on a mixture of azelaic acid with pelargonic have shown essentially quantitative recovery of azelaic acid. Failure to establish quantitative recovery on oxidation of oleic acid must be caused by some unknown factor during oxidationper se. The experimental technique described was satisfactory for quantitative studies of the type undertaken. Oxidation of moderate-purity, mono-unsaturated fatty acids, such as erucic, 10-undecenoic, and vaccenic acid, has given mixed dibasic acids corresponding to the respective positions of unsaturation. The data indicate that the method described shows the position of minor unsaturation within about 1%. Presented at meeting of American Oil Chemists' Society, Chicago, Ill., September 24–26, 1956.     

Identification of nine acetylenic fatty acids, 9-hydroxystearic acid and 9,10-epoxystearic acid in the seed oil ofJodina rhombifolia hook et arn. (Santalaceae)

In addition to some usual fatty acids, the seed oil ofJodina rhombifolia (Santalaceae) contains nine acetylenic fatty acids [9-octadecynoic acid (stearolic acid) (1.1%),trans-10-heptadecen-8-ynoic acid (pyrulic acid) (20.1%), 7-hydroxy-trans-10-heptadecen-8-ynoic acid (2.3%),trans-10,16-heptadecadien-8-ynoic acid (0.7%), 7-hydroxy-trans-10,16-heptadecadien-8-ynoic acid (0.1%),trans-11-octadecen-9-ynoic acid (ximenynic acid) (20.3%), 8-hydroxy-trans-11-octadecen-9-ynoic acid (12.2%),trans-11,17-octadecadien-9-ynoic acid (1.5%), 8-hydroxy-trans-11,17-octadecadien-9-ynoic acid (1.3%), 9-hydroxystearic acid (<0.1%) and 9,10-epoxystearic acid (0.7%)]. The fatty acids have been analyzed by gas chromatography/mass spectrometry of their methyl ester and 4,4-dimethyloxazoline derivatives. The hydroxy fatty acid methyl esters have been examined also as trimethyl-silyl ethers. Furthermore, the fatty acid methyl esters (FAME) have been fractionated according to their polarity (FAME-A: nonhydroxy; FAME-B: hydroxy fatty acids) and to their degree of unsaturation (FAME-A1/A2; FAME-B1/B2) by preparative thin-layer chromatography and argentation chromatography, respectively. All of these fractions have been analyzed by ultraviolet and infrared spectroscopy, and the fractions FAME-A and FAME-B have been analyzed further by nuclear magnetic resonance (¹H,¹³C, 2D H/C, attached proton test) spectroscopy and gas chromatography/mass spectrometry. This work is dedicated to the 65th birthday of Prof. Dr. K. Pfeilsticker, Institut of Food Science, University Bonn (Germany).     

Phosphorus derivatives of fatty acids. The addition of dialkyl phosphonates to unsaturated acids and the synthesis of some 11-dialkylphosphonoundecanoamides

A series of dialkylphosphonoundecanoic acids and 9(10)-dialkylphosphonostearic acids was prepared by the addition of dialkylphosphonates to 10-undecenoic acid and oleic acid under free radical conditions in 58–66% yield. The phosphonates used were: dimethyl, diethyl, di-n-butyl, di-n-hexyl and di-2-ethylhexyl. All of the products are colorless, odorless, thermally stable liquids, insoluble in water and soluble in organic solvents. The dialkylphosphonoundecanoic acids were converted into the corresponding acid chlorides by reaction with oxalyl chloride. The acid chlorides were then converted into the corresponding amides by addition of ammonia. All of the amides are white waxy solids except the 2-ethylhexyl derivative which is a high-boiling liquid. Two of the dialkylphosphonoundecanoic acids were converted into the N-decyl substituted amides by heating withn-decylamine. The N-n-decyl-11-dialkylphosphonoundecanoamides are white waxy solids. Presented at the AOCS Meeting, New Orleans, 1964.     

Analysis of oil and meal fromLesquerella fendleri seed

Attention is being focused onLesquerella species as a source of hydroxy acids to replace imported castor oil. Genetic and agronomic improvement and utilization of the seed oil and meal are being studies. We have conducted laboratory experiments to extract oil fromL. fendleri seed in preparation for extracting large quantities of seed.L. fendleri is a member of the Cruciferae family, and when seeds are crushed glucosinolates release isothiocyanates by the action of a thioglucosidase enzyme system. Therefore, our experiments included moist heat treatment of whole seeds to inactivate this enzyme. The seed was then flaked in a Wolf mill, and the flakes were exhaustively extracted with hexane. The oil was degummed and bleached, and then analyzed for hydroxyl (103), saponification (174), and iodine values (107), and for unsaponifiables (1.5%), FFA (1.13%) and P (10 ppm) contents. Hydroxy fatty acids, 55% lesquerolic (14-hydroxy-cis-11-eicosenoic) and 3% auricolic (14-hydroxy-cis-11,cis-17-eicosadienoic), and total fatty acid distribution were determined by gas chromatography of the methyl esters. The defatted meal was analyzed for residual oil (1%), protein (29.8%), non-protein nitrogen (0.7%), ash (6.45%). crude fiber (12.9%), and for distribution of amino acids. DefattedL. fendleri meal has an excellent distribution of amino acids, including favorable levels of lysine, methionine and threonine compared with soybean meal.     

Two-step enzymatic reaction for the synthesis of pure structured triacylglycerides

Structured triacylglycerides with medium-chain fatty acids (caprylic acid) in sn1- and sn3-positions and a long-chain unsaturated fatty acid (oleic or linoleic acid) in the sn2-position of glycerol (MLM) were synthesized by lipase catalysis in a two-step process. First, pure 2-monoacylglycerides (2-MG) were synthesized by alcoholysis of triacylglycerides (triolein, trilinolein, or peanut oil) in organic solvents with 1,3-regiospecific lipases (from Rhizomucor miehei, Rhizopus delemar, and Rhizopus javanicus). The 2-MG were purified by crystallization and obtained in up to 71.8% yield. These 2-MG were esterified in a second reaction with caprylic acid in n-hexane to form almost pure MLM. For 2-MG obtained from peanut oil, the final product contained more than 90% caprylic acid in the sn1- and sn3-positions, whereas the sn2-position was composed of 98.5% unsaturated long-chain fatty acids. Reaction conditions for both steps were optimized with respect to source and immobilization of lipase, water activity, and solvent.     

Epoxides of unsaturated fatty acids as anti-rust additives in water-based cutting fluids

Epoxides from a variety of unsaturated fatty acids were prepared, and corrosion tests for these products as water-based cutting fluid additives were performed. The triethanolamine salt of an epoxide of undecylenic acid showed very effective rust-inhibiting properties in a water-based cutting fluid.     

Homolytic decomposition of linoleic acid hydroperoxide: Identification of fatty acid products

An isomeric mixture of linoleic acid hydroperoxides, 13-hydroperoxy-cis-9,trans-11-octadecadienoic acid (79%) and 9-hydroperoxy-cis-12,trans-10-octadecadienoic acid (21%), was decomposed homolytically by Fe(II) in an ethanol-water solution. In one series of experiments, the hydroperoxides were decomposed by catalytic concentrations of Fe(II). The 10⁻⁵ M Fe(III) used to initiate the decomposition was kept reduced as Fe(II) by a high concentration of cysteine added to the reaction in molar excess of the hydroperoxides. Nine different monomeric (no detectable dimeric) fatty acids were identified from the reaction. Analyses of these fatty acids revealed that they were mixtures of positional isomers identified as follows: (I) 13-oxo-trans,trans-(andcis,trans-) 9,11-octadecadienoic and 9-oxo-trans,trans- (andcis,trans-) 10,12-octadecadienoic acids; (II) 13-oxo-trans-9,10-epoxy-trans-11-octadecenoic and 9-oxo-trans-12, 13-epoxy-trans-10-octadecenoic acids; (III) 13-oxo-cis-9,10-epoxy-trans-11-octadecenoic and 9-oxo-cis-12, 13-epoxy-trans-10-octadecenoic acids; (IV) 13-hydroxy-9,11-octadecadienoic and 9-hydroxy-10,12-octadecadienoic acids; (V) 11-hydroxy-trans-12, 13-epoxy-cis-9-octadecenoic and 11-hydroxy-trans-9, 10-epoxy-cis-12-octadecenoic acids; (VI) 11-hydroxy-trans-12, 13-epoxy-trans-9-octadecenoic and 11-hydroxy-trans-9,10-epoxy-trans-12-octadecenoic acids; (VII) 13-oxo-9-hydroxy-trans-10-octadecenoic acids; (VIII) isomeric mixtures of 9, 12, 13-dihydroxyethoxy-trans-10-octadecenoic and 9, 10, 13-dihydroxyethoxy-trans-11-octadecenoic acids; and (IX) 9, 12, 13-trihydroxy-trans-10-octadecenoic and 9, 10, 13-trihydroxy-trans-11-octadecenoic acids. In another experiment, equimolar amounts of Fe(II) and hydroperoxide were reacted in the absence of cysteine. A large proportion of dimeric fatty acids and a smaller amount of monomeric fatty acids resulted. The monomeric fatty acids were examined by gas liquid chromatography-mass spectroscopy. Spectra indicated that the monomers were largely similar to those produced by the Fe(III)-cysteine reaction. Presented in part at the American Chemical Society Meeting, Los Angeles, March 1974. ARS, USDA.     

Cyclopropanation of unsaturated fatty acid methyl esters using diazomethane and palladium (II) acetate

Cyclopropanation of methyl esters of 10-undecenoic, oleic, elaidic, erucic,trans-2-docosenoic and a mixture of hydnocarpic and chaulmoogric acids was effected using diazomethane in the presence of palladium (II) acetate as catalyst. The products were isolated by silver ion thin-layer chromatography and characterized by infrared, proton nuclear magnetic resonance and mass spectrometric techniques. Terminally unsaturated, α, β-unsaturated and cyclopentene fatty acid esters were found to be more reactive than elaidate which in turn was more reactive than oleate. The reaction proceeds stereospecifically under milder conditions in a shorter time than with diiodomethane and zinc-copper couple.     

Synthesis of 6-hydroxy δ-lactones and 5,6-dihydroxy eicosanoic/docosanoic acids from meadowfoam fatty acids via a lipase-mediated self-epoxidation

Meadowfoam fatty acids were reacted with hydrogen peroxide in a lipase-catalyzed autocatalytic system, forming a mixture of 5,6-epoxyeicosanoic, 13,14-epoxydocosanoic, 5,6-epoxydocosanoic, and 5,6-13,14-diepoxydocosanoic acids in 98% yield. The 5,6-epoxy acids were cyclized to 6-hydroxy δ-eicosanoic/docosanoic lactones by sulfuric acid catalysis in high yield (99%). 5,6-Dihydroxy acids could be obtained from 6-hydroxy δ-lactones by a simple alkaline work-up procedure. Meadowfoam fatty acids were converted (77% yield) in a one-pot reaction to 6-hydroxy δ-lactones by in situ performic acid epoxidation and subsequent addition of sulfuric acid.