Chromatographic determination of optical configuration of 3-hydroxy fatty acids composing microbial surfactants

Abstract The chromatographic determination of the optical configuration of 3-hydroxy fatty acids of microbial surfactants was achieved in chiral high pressure liquid chromatography (HPLC) by injecting 3,5-dinitroaniline-derivatives of crude hydrolysates (less than 1 mg). Serrawettin W2, a surface-active cyclodepsipeptide of Serratia marcescens , was shown to contain d -3-hydroxydecanoic acid. Rubiwettin R1 and RG1, surface active glycolipid and linked fatty acids of Serratia rubidaea , were shown to contain d -3-hydroxytetradecanoic acid and d -3-hydroxydecanoic acid. The new method does not require purified sample or authentic optical isomers, and could be useful in the structural analysis of microbial lipids.     

Resolution and chromatographic configuration analysis of 2-hydroxy fatty acids

2-Hydroxyoctadecanoic acid was resolved into D and L isomers as salts of 1-phenylethylamine enantiomers The diastereomers of phenylethylamides of 2-hydroxy fatty acids and the corresponding derivatives with protected hydroxy group (acetyl, methyl, trifluoro-acetyl, trimethylsilyl) are well separated by thin-layer or gas-liquid chromatography. This allows a simple microanalysis of configuration and optical purity of 2-hydroxy fatty acids. With this method 2-hydroxy fatty acids from sphingomyelin of the honey-bee were shown to belong exclusively to the D series.     

Analysis of the stereochemistry of lipoxygenase-derived hydroxypolyenoic fatty acids by means of chiral phase high-pressure liquid chromatography

A chiral phase HPLC method was developed for the simultaneous determination of the positional and optical isomers of the lipoxygenase-derived hydroxypolyenoic fatty acids. With a Bakerbond chiral phase HPLC column (dinitrobenzoyl phenylglycine as chiral phase) the positional and optical isomers of the reduced dioxygenation products (by triphenylphosphine or borohydride) of linoleic acid and arachidonic acid were separated after methylation of the carboxylic groups. No cumbersome chemical derivatization such as conversion to a diastereomer was necessary. As compared with the methods used up till now chiral phase HPLC proved to be simpler and more sensitive. About 10 pmol of hydroxy fatty acids suffice for an analysis. The chiral phase HPLC can be used for the preparative separation of the optical antipodes of the lipoxygenase products. An optical purity of more than 90% can be reached in one preparative run. The method was applied to the determination of the stereochemistry of the dioxygenation products of polyenoic fatty acids formed by the lipoxygenases from soybeans, reticulocytes, pea seeds (isoenzyme I and II), tomato fruits, by the quasilipoxygenase activity of hemoglobin, and by the methylene blue-mediated photooxidation of arachidonic acid.     

Cellular Lipid and Fatty Acid Compositions of Burkholderia pseudomallei Strains Isolated from Human and Environment in Viet Nam

Viet nam is known as an endemic area of melioidosis but its etiologic agent originated in Viet nam was not extensively studied. For the first time, we analyzed the cellular lipid and fatty acid compositions of 15 Vietnamese isolates of Burkholderia pseudomallei, 10 from humans and 5 from the environment. Cellular lipid compositions were analyzed by two-dimensional thin-layer chromatography on silica gel G plates. Cellular fatty acid methyl esters were analyzed by gas chromatography (GC) and gas chromatography/mass spectrometry (GC/MS). The major lipids in all the isolates were phosphatidylglycerol (PG), two forms of phosphatidylethanolamine (PE-1 and PE-2), and two forms of ornithine-containing lipid (OL-1 and OL-2). PE-1 contained non-hydroxy fatty acids at both sn-1 and ?2 positions, while PE-2 possessed 2-hydroxy fatty acids and non-hydroxy fatty acids in a ratio of 1: 1. Since snake venom phospholipase A₂ digestion of PE-2 liberated 2-hydroxy fatty acids, it was confirmed that these acids are at the sn-2 position of glycerol moiety. In both OL-1 and OL-2, amide-linked fatty acid was 3-hydroxy palmitic acid (3-OH-C16: 0), while ester-linked fatty acids were non-hydroxy acids in OL-1 and 2-hydroxy acids in OL-2. The total cellular fatty acid compositions of the test strains were characterized by the presence of 2-hydroxy palmitic (2-OH-C16: 0), 2-hydroxy hexadecenoic (2-OH-C16: 1), 2-hydroxy octadecenoic (2-OH-C18: 1), 2-hydroxy methylene octadecanoic (2-OH-C19CPA), 3-hydroxy myristic (3-OH-C14: 0) and 3-hydroxy palmitic (3-OH-C16: 0) acids. There were significant differences in the concentration of hexadecenoic (C16: 1), methylene hexadecanoic (C17CPA), octadecenoic (C18: 1) and methylene octadecanoic (C19CPA) acids among the Vietnamese isolates of B. pseudomallei. However, no significant difference was observed in cellular lipid and fatty acid components between strains of human and environmental origins.     

Structural components of sphingophosphonolipids from the ciliated protozoan, Tetrahymena pyriformis WH-14.

1. Two sphingophosphonolipids were isolated from the lipids of the ciliated protozoan, Tetrahymena pyriformis WH-14. They were ceramide N-methyl-2-aminoethylphosphonate (CMAEP) and ceramide 2-aminoethylphosphonate (CAEP), in yields of 0.05 mg/g and 1.74 mg/g dry cells, respectively. 2. Two chromatographically distinguishable CAEP species were found, a slow-moving major component and a minor component which moved faster; the slow-moving one contained primarily hydroxy fatty acids, while in the other one nonhydroxy fatty acids were predominant. However, their long-chain base constituents were similar. 3. The major fatty acids of CAEP were 2-hydroxy acids with carbon numbers of 16 to 19, which were almost exclusively iso-types. The fatty acids of CMAEP consisted mainly of palmitic, iso-octadecanoic, and 2-hydroxy iso-heptadecanoic acids. 4. The long-chain bases were dominated by C16, C17, and C19 iso-4-sphingenine homologs.     

FATTY ACIDS AND HYDROXY FATTY ACIDS IN THREE SPECIES OF FRESHWATER EUSTIGMATOPHYTES

The monocarboxylic fatty acids and hydroxy fatty acids of three species of freshwater microalgae—Vischeria punctata Vischer, Vischeria helvetica (Vischer et Pascher) Taylor, and Eustigmatos vischeri (Hulbert) Taylor, all from the class Eustigmatophyceae— were examined. Each species displayed a very similar distribution of fatty acids, the most abundant of which were 20:5n-3, 16:0, and 16:1n-7; C₁₈ polyunsaturated fatty acids were minor components. These fatty acid distributions closely resemble those found in marine eustigmatophytes but are quite distinct from those found in most other algal classes. These microalgae also contain long-chain saturated and unsaturated monohydroxy fatty acids. Two distinct types of hydroxy fatty acids were found: a series of saturated α-hydroxy acids ranging from C₂₄ to C₃₀ with a shorter series of monounsaturated α-hydroxy acids ranging from C₂₆ to C₃₀ together with a series of saturated β-hydroxy acids ranging from C₂₆ to C₃₀. The latter have not previously been reported in either marine or freshwater microalgae, although C₃₀ to C₃₄ midchain (ω-18)-hydroxy fatty acids have been identified in hydrolyzed extracts from marine eustigmatophytes of the genus Nannochloropsis, and C₂₂ to C₂₆ saturated and monounsaturated α-hydroxy fatty acids have been found in three marine chlorophytes. These findings have provided a more complete picture of the lipid distributions within this little studied group of microalgae as well as a range of unusual compounds that might prove useful chemotaxonomic markers. The functions of the hydroxy fatty acids are not known, but a link to the formation of the lipid precursors of highly aliphatic biopolymers is suggested.     

Fatty Acids of Extractable and Bound Lipids of Rhodomicrobium vannielii

Cells of Rhodomicrobium vannielii grown at 29 C in a lactate-containing medium were extracted at room temperature with organic solvents. The extractable fraction contained the bulk of the simple lipid (1.87% of cell dry weight) and complex lipids (phospholipids, 4.2%; sulfolipid, 0.01%), coenzyme Q (0.09%), and pigments (carotenoids 1.2%; bacteriochlorophyll, 1.9%). The cell residue contained the bound lipids (nonpolar fatty acid fraction, 1.86%; polar hydroxy fatty acids, 0.49%). The residue also contained poly-β-hydroxybutyric acid (0.2%), which was extracted in boiling chloroform. In both the simple and complex lipids, vaccenic acid (11-octadecenoic acid) was the largest single component (approximately 90% in each fraction). The fatty acids of the bound lipid contained 35% vaccenic acid, even- and odd-numbered saturated and unsaturated straight-chain fatty acids, cyclopropane-, branched-, and α- and β-hydroxy fatty acids. The extractable lipids contained only straight-chain saturated and unsaturated even-numbered fatty acids. Nearly 60% of hydroxy fatty acid fraction was α-hydroxydodecanoic acid (24%) and β-hydroxydodecanoic acid (34.5%). Coenzyme Q was crystallized and identified as Q₉ on the basis of melting point and chromatographic properties. Q₁₀ had been previously reported.     

Quantification of ergosterol and 3-hydroxy fatty acids in settled house dust by gas chromatography-mass spectrometry: comparison with fungal culture and determination of endotoxin by a Limulus amebocyte lysate assay.

Ergosterol and 3-hydroxy fatty acids, chemical markers for fungal biomass and the endotoxin of gram-negative bacteria, respectively, may be useful in studies of health effects of organic dusts, including domestic house dust. This paper reports a method for the combined determination of ergosterol and 3-hydroxy fatty acids in a single dust sample and a comparison of these chemical biomarkers determined by gas chromatography-mass spectrometry with results from fungal culture and Limulus assay. Analyses of replicate house dust samples resulted in correlations of 0.91 (ergosterol in six replicates; P < 0.01) and 0.94 (3-hydroxy fatty acids in nine replicates; P < 0.001). The amounts of ergosterol (range, 2 to 16.5 ng/mg of dust) correlated with those of total culturable fungi (range, 6 to 1,400 CFU/mg of dust) in 17 samples, (r = 0.65; P < 0.005). The amounts of endotoxin (range, 11 to 243 endotoxin units/mg of dust) measured with a modified chromogenic Limulus assay correlated with those of lipopolysaccharide (LPS) determined from 3-hydroxy fatty acid analysis of 15 samples. The correlation coefficient depended on the chain lengths of 3-hydroxy acids used to compute the LPS content. The correlation was high (r = 0.88 +/- 0.01; P < 0.001) when fatty acid chains of 10 to 14 carbon atoms were included; the correlation was much lower when hydroxy acids of 16- or 18-carbon chains were included. In conclusion, the results of the described extraction and analysis procedure for ergosterol and 3-hydroxy fatty acids are reproducible, and the results can be correlated with fungal culture and endotoxin activity of organic dust samples.     

Chemotaxonomy of planktonic cyanobacteria based on non-polar and 3-hydroxy fatty acid composition

Twenty-eight axenio planktonic cyanobacterial strains (10 Microcystis, three Oscillatoria, one Spirulina, one Aphanizomenon, 13 Anabaena) were investigated for their fatty acid composition by measurement of non-polar and hydroxy fatty acids. No 2-hydroxy fatty acids were detected in any strain, but 3-hydroxy fatty acids were detected in minor quantities in 24 strains. The highest portion of total fatty acids were non-polar fatty acids. Qualitative and quantitative analyses of 3-hydroxy fatty acids showed no taxonomic value in these strains, while the type of non-polar fatty acid composition was shown to be consistent within Microcystis and Anabaena strains, distinguishing them as type 4, characterized by the presence of 18:4, and type 2, characterized by 18:3 (α) of the Kenyon-Murata system. Two Oscillatoria agardhii Gomont strains were also included in the type 2 group due to the presence of 18: 3 (α), but the difference in characteristics of 16:2 and 16:3 between O. agardhii and Anabaena further divided type 2 into two subgroups: type 2A for Anabaena and type 2B for O. agardhii. A simplified unweighted pair group method with arithmetic averages (UPGMA) dendrogram demonstrated that the classification of 28 strains (Microcystis spp., Anabaena spp., Aphanizomenon flos-aquae (Lemmermann) Ralfs f. gracile (Lemmermann) Elenkin, O. agardhii and Spirullnasubsalsa Oersted ex Gomont based on numerical analysis of non-polar fatty acids corresponded to morphological species criteria, suggesting that non-polar fatty acid composition is a valuable chemical marker in the taxonomy of planktonic cyanobacteria. However, the fatty acid composition in Oscillatoria raciborskii is similar to that of Microcystis and very different from that of O. agardhii, suggesting its special position in Oscillatoria and the chemical diversity in the genus Oscillatoria.     

Hydroxy fatty acids in Bacteroides species: D-(--)-3-hydroxy-15-methylhexadecanoate and its homologs.

Acid hydrolysates of 140 strains, representing 11 species of the genus Bacteroides, were analyzed by capillary gas-liquid chromatography for total cellular fatty acid. All samples contained components which appeared to be hydroxy fatty acid. The relative amount and chain length distribution of the hydroxy fatty acids, as well as the nonhydroxy fatty acids, varied according to species. To characterize the presumed hydroxy acids, a composite of some 40 of these samples was analyzed by thin-layer and capillary gas-liquid chromatography, mass spectrometry, infrared spectrophotometry, and polarimetry. The hydroxy acids were shown to be of the D-(--)-3-hydroxy acid family. The predominant component was the iso-branched D-(--)-3-hydroxy-15-methylhexadecanoic acid. Lesser amounts of the iso-branched 15-carbon, straight-chain 16-carbon, and anteiso-branched 17-carbon acids were also found.     

Absolute configuration of 3-hydroxy acids formed by Stenotrophomonas maltophilia: application of multidimensional gas chromatography and circular dichroism spectroscopy.

The soil bacterium Stenotrophomonas maltophilia was found to transform various long-chain fatty acids selectively into 3-hydroxy fatty acids of shorter chain length. Their chiral evaluation was performed by multidimensional gas chromatography (MDGC) on modified cyclodextrin phase comparing the enantiodistribution of 1,3-diol formed without loss of stereochemical information from a representative microbial product with those of synthetic (3RS)- and (3S)-1,3-diols. Enantiomeric excesses of 84-98% (R) were determined for the microbially produced 3-hydroxy acids. In addition, the CD exciton chirality method was applied to determine their absolute configuration. Derivatization with 9-anthryldiazomethane and 2-naphthoylimidazole led to the required bichromophoric structures. Their CD spectra displayed a positive first Cotton effect around 254 nm and a negative second Cotton effect around 237 nm, which confirmed the (R)-configuration of the bacterial products.     

Substrate specificity of <Emphasis Type="Italic">Stenotrophomonas nitritireducens</Emphasis> in the hydroxylation of unsaturated fatty acid

An isolated bacterium that converted unsaturated fatty acids to hydroxy fatty acids was identified as Stenotrophomonas nitritireducens by API analysis, cellular fatty acids compositions, sequencing the full 16S ribosomal ribonucleic acid, and evaluating its nitrite reduction ability. S. nitritireducens has unique regio-specificity for C16 and C18 cis-9 unsaturated fatty acids. These fatty acids are converted to their 10-hydroxy fatty acids without detectable byproducts. Among the cis-9-unsaturated fatty acids, S. nitritireducens showed the highest specificity for linoleic acid. The cells converted 20 mM linoleic acid to 13.5 mM 10-hydroxy-12(Z)-octadecenoic acid at 30°C and pH 7.5 with a yield of 67.5% (mol/mol).     

Diesters of alkane diols and 2-hydroxy fatty acids: identification and discrimination of isomers with the aid of NMR spectroscopy.

High resolution nuclear magnetic resonance spectroscopy has been shown to be extremely useful for the identification and discrimination of naturally occurring diesters of 1,2- and 2,3-alkanediols as well as for fatty alkyl esters of acylated 2-hydroxy fatty acids. A comparison of 220 MHz spectra of 1,2 and erythro- 2,3-alkanediol diesters exhibits the following distinguishing features: (1) two non-equivalent methylene protons from the glycol group of 1,2-alkanediol diesters resonate at 3.87 ppm and 4.17 ppm respectively while these resonances are completely absent in the spectrum of 2,3-isomer; (2) methylene protons adjacent to esther carbonyl groups appear as two overlapping triplets at 2.22 ppm in 1,2-alkanediol diesters while the corresponding protons in the 2,3-isomer are displayed as two partially overlapping triplets centered at 2.15 ppm and 2.2 ppm respectively; and (3) methyl protons adjacent to glycol group in 2,3-isomer appear as downfield doublet at 1.13 ppm; this downfield doublet is not shown by 1,2-alkanediol diesters. Erythro- and threo-2,3-alkanediol diesters have also been distinguished from each other; two alpha-methylenes in erythro isomers appear as partially overlapping triplets while these protons in threo isomer display an apparent quartet centered at 2.22 ppm. Fatty alkyl esters of acylated 2-hydroxy fatty acids display a triplet at 4.79 for 2-position methylene proton, a distinguishing feature not shown by diacyl alkanediols. A distinction between diester lipids and other classes of neutral lipids has also been achieved by the study of nuclear magnetic resonance spectra, particularly in the region of 3-6 ppm.     

Unusual wax esters and glycolipids: Biocatalytical formation and physico-chemical characterization

By aid of lipases, e.g. of Mucor michei, in n-hexane wax esters were produced from usual primary fatty alcohols and unusual hydroxy fatty acids (in part of microbial origin). Thus, (S)-17-hydroxyoctadecanoic acid dodecyl ester and (R)-3-hydroxy decanoic acid dodecyl ester were formed. In measurements of the film pressure using a LANGMUIR film balance the monolayers of both compounds indicated good stability compared to the non-hydroxy wax esters. Glycolipids de novo produced by microorganisms did not show suitable wetting properties, but they were able to lower ze surface tension of water to a higher extent than the unusual waxes.     

Photochemical Reactions of Methyl trans-dihydrojasmonate

The occurrence of positional isomers of minor C₂₄ unsaturated fatty acids in female gonad lipids of the limpet Cellana grata was clarified by gas chromatography–mass spectrometry of the combination of their 4,4-dimethyloxazoline and picolinyl ester derivatives. In this study, in addition to 5,9-24:2, 9,15-24:2, 5,9,15-24:3, and 5,9,17-24:3, previously identified, 24:4n-6, 24:5n-3, and four novel nonmethylene-interrupted fatty acids, 9,17-24:2, 9,15,18-24:3, 5,9,15,18-24:4, and 5,9,15,18,21-24:5, were newly recognized. All C₂₄ unsaturated fatty acids detected were present only in triacylglycerols.     

THE CHARACTERIZATION OF SPHINGOLIPIDS OF OLIGODENDROGLIA FROM CALF BRAIN

Abstract— Purified oligodendroglia isolated from bovine brain white matter were found to contain, in addition to galactosylceramide, sulfatide and sphingomyelin, significant quantities of glucosylcerai-mide, dihexosylceramide and esterified galactosylceramide. These sphingolipids were isolated and quan-titated and their fatty acid and long chain base patterns compared with those from sphingolipids isolated from bovine myelin, white matter and gray matter.
The minor glycosphingolipids, glucosylceramide, dihexosylceramide and esterified galactosylceramide, constituted a higher percentage of glial lipids than of myelin lipids. Glucosylceramide accounted for 12% of the total glial monohexosylceramide fraction and 0.8% of total lipids; dihexosylceramide was 0.9% of total glial lipids. Both of these lipids had small quantities of α-hydroxy fatty acids. The unsubstituted fatty acids of glucosylceramide were mostly short chain (16 and 18 carbons) and were different from those of the dihexosylceramides which were a mixture of short and long chain. The hydroxy acids of each of these lipids were, however, similar and resembled those of galactosylceramide.
The fatty acid patterns of galactosylceramide, sulfatide and sphingomyelin from glial cells resembled those of the corresponding lipids from myelin and white matter. The amide-linked acids of esterified galactosylceramide contained both unsubstituted and α-hydroxy chains. Their patterns were not identical to those of galactosylceramide, but were similar in all brain fractions.
With the exception of sphingomyelin and dihexosylceramide, which contained small amounts of C₂₀-sphingosine, all sphingolipids analyzed contained mostly sphingosine and dihydrosphingosine.
We conclude that the distribution of sphingolipids in the oligodendroglia is characteristic, but the lipophilic residues of these lipids are not cell-specific.     

The lipid composition of the electric organ of the ray, Torpedo marmorata, with specific reference to sulfatides and Na+-K+-ATPase.

The lipids from the electric organ of the ray, Torpedo marmorata, have been isolated and characterized. The major lipids were cholesterol, choline phospholipids, ethanolamine phospholipids, and sphingomyelins. The major fatty acids of ethanolamine phospholipids were 18:1, 18:0, 22:6, and 20:4. More than 50% of the acids in choline phospholipids were 16:0. The sphingomyelins consisted of five major ceramide species, all with sphingosine and the fatty acids 14:0, 15:0, 16:0, 22:1, and 24:1. The fatty acid 15:0 was mostly branched (n-2), a fatty acid earlier identified in sphingomyelins of the rectal gland of spiny dogfish. All long-chain bases were dihydroxy bases with a small percentage of branched chains. Sulfatides (cerebroside sulfate) made up the largest glycolipid fraction. The polar moiety wase galactose-3-sulfate. The fatty acids were normal and 2-hydroxy; the homologue 24:1 was the most abundant in both types of fatty acids. Most fatty acids were higher homologues of mono-unsaturated acids, but normal 18:0 fatty acid was also found. The long-chain bases were both dihydroxy and trihydroxy, with very small amounts of branched chains. The two major ceramide species of sulfatides were sphingosine combined with normal and hydroxy 24:1 fatty acids, respectively. Smaller amounts of trihydroxy base (18:0) were found linked to hydroxy 24:1 fatty acid, but not to its normal homologue. The cerebrosides contained the two major species mentioned above but lacked the trihydroxy base-hydroxy fatty acid species. The ratio of the activity of Na+-K+-dependent ATPase (EC 3.6.1.3) and the concentration of sulfatides was similar to ratios found for other tissues with normal and increased Na+ and K+ transporting capacity. The significance of this finding is discussed.     

Influence of Exogenous Natural Oils on the ω-1 and ω-2 Hydroxy Fatty Acid Moiety of Sophorose Lipid Produced by Candida bombicola

Candida bombicola can synthesize monohydroxy fatty acid as a moiety of sophorose lipids. The hydroxy fatty acids contained in a major lactone were identified by GC-MS, after culturing with natural oils such as coconut, rapeseed, olive, and soybean oils. Hydroxy fatty acids of C18 and C16 were always synthesized, but differences were observed among the oils regarding the positions of hydroxyl groups, unsaturation, and composition of the fatty acids. A new C17 hydroxy acid was found without addition of oil.     

Formation of monohydroxy derivatives of arachidonic acid, linoleic acid, and oleic acid during oxidation of low density lipoprotein by copper ions and endothelial cells.

An important event in the formation of atherosclerotic lesions is the uptake of modified low density lipoprotein (LDL) by macrophages via scavenger receptors. Modification of LDL, which results in its recognition by these receptors, can be initiated by peroxidation of LDL lipids. The first step in this process is the formation of monohydroperoxy derivatives of fatty acids, which are subsequently degraded to the corresponding monohydroxy compounds, or to a variety of secondary oxidation products. In order to understand this process more completely, we have developed a mass spectrometric procedure to measure the amounts of specific hydroperoxy/hydroxy fatty acids formed by oxidation of the major unsaturated fatty acids in human LDL, oleic acid, linoleic acid, and arachidonic acid. Oxidation of human LDL in the presence of a relatively strong stimulus (20 microM CuSO4) resulted in very large increases in the amounts of the major monohydroxy derivatives of linoleic acid (9- and 13-hydroxy derivatives) and arachidonic acid (5-, 8-, 9-, 11-, 12-, and 15-hydroxy derivatives) in LDL lipids in the early stages of the reaction. After 20 h, the amounts of these products declined due to substrate depletion, but large amounts of monohydroxy derivatives of oleic acid (8-, 10-, and 11-hydroxy derivatives) were detected. Although thiobarbituric acid-reactive substances clearly increased under these conditions, the changes were not nearly so dramatic as those observed for monohydroxy fatty acids. Oxidation of LDL in the presence of endothelial cells, a much milder stimulus, resulted in 2.5- to 3-fold increases in the amounts of monohydroxy derivatives of linoleic and arachidonic acids, as well as thiobarbituric acid-reactive substances, with more modest increases in the amounts of hydroxylated derivatives of oleic acid. There was little positional specificity in the oxidation of the above fatty acids in the presence of either stimulus, suggesting that the formation of these products proceeds primarily by lipid peroxidation, rather than by catalysis by lipoxygenases. However, an important role for lipoxygenases in the initiation of these reactions cannot be excluded. In conclusion, oxidation of LDL in the presence of copper ions or endothelial cells results in the formation of a large number of monohydroxy derivatives of oleic, linoleic, and arachidonic acids. The relative amounts of products formed from each of these fatty acids depends on the strength of the stimulus as well as the incubation time.     

Stereochemical aspects of synthetic and naturally occurring 2-hydroxy fatty acids. Their absolute configurations and assays of optical purity

d- and l-Cerebronic acid⁵, ⁶ methyl esters were obtained from a dl-mixture by converting the racemic ester to the l-acetylmandelates and separating the diastereomers by thin-layer chromatography; the diasteromers were then decomposed with mild acid methanolysis. The optical purities of the d- and l-enantiomers were determined as described below and found to be 82.5% and 87.1%, respectively. Absolute configurations of the enantiomers were confirmed by their ORD spectra; specific rotations at the peak of the anomalous curves at 223 nm of the d- and l-isomers were ?989 ° and +1120 °, respectively. Considering their optical purities, the actual maximum rotations of these enantiomers were calculated as ?1520 ° and +1509 °, respectively. The conditions for the reaction of l-acetylmandelyl chloride and 2-hydroxy fatty acid esters were modified to be suitable for the optical assay of 2-hydroxy fatty acid methyl esters. The method was also shown to be applicable to the assay of 3-hydroxy isomers and possibly to other positional isomers. The absolute configuration of 2-hydroxy fatty acid methyl esters obtained from calf brain cerebrosides and from yeast cerebrins were determined to be d(?), confirming previous assignments. Methyl cerebronate obtained from its less soluble strychnine salt was found to be the l(+)-enantiomer, contrary to a previous assignment. Methyl d-cerebronate was hydrolyzed with tetrahydrofuran-HCl to obtain D-cerebronic acid without racemization.