Preparation of deuterium-labelled methyl linoleate and its geometric isomers from natural seed oils

Multi-gram quantities of deuterium-labelled methyl linoleate (methyl cis-9,cis-12-octadecadienoate) and its geometric isomers are readily synthesized fromCrépis alpina (70–80% cis-9-octadecen-12-ynoic acid) andVernonia galamensis (70–80% 12,13-epoxy-cis-9-octadecenoic acid) seed oils. Methylcis- 9,cis- 12- andtrans- 9,cis- – octadecadienoate-12,13-d₂ were prepared by the Lindlar-catalyzed reduction (with D₂ gas) of methylcis- 9- and trans-9-octadecen-12-ynoates, respectively. Methyltrans- 9- octadecen-12-ynoate was synthesized by thep-toluene-sulfinic acid-catalyzed isomerization of the corresponding cis isomer. Methylcis- 9fiis- 12., trans- 9fiis- 12;cis- 9,trans- 12- andtrans- 9, frans-12-octadecadienoate-d₂, d₄ and d₆ were prepared by the Wittig coupling (with stereochemical control) of the appropriate d₂-, d₄- or de-alkyltriphenyl-phosphonium salt with methyl 12-oxo-cis-9- ortrans- 9- dodecenoate (prepared by the para-periodic acid cleavage of methyl 12,13-dihydroxy-cis-9- or trans-9-octadecenoate). Thecis dihydroxy ester was synthesized fromVernonia galamensis seed oil by acetolysis, saponification and then esterification. Thecis dihydroxy ester was isomerized by nitric acid/sodium nitrite to thetrans form and purified by silver resin chromatography. Isotopic purities ranged from 88% (for the d₆ isomers) to 99% (for the d₂ isomers). The mention of firm names or trade products does not imply that they are endorsed or recommended over other firms or similar products not mentioned.     

Hydrolysis of linoleate geometric isomers byGeotrichum candidum lipase

Lipase (EC 3.1.1.3) from the microorganismGeotrichum candidum preferentially hydrolyzescis-9 18∶1 andcis,cis-9,12 18∶2 from triacylglycerols, largely ignoring all other positional isomers ofcis 18∶1 as well astrans-9 18∶1. To obtain additional information about the specificity of the enzyme, two triacylglycerols were prepared and utilized as substrates. The lipase hydrolyzed 85%cis,cis-9,12 18∶2 and 15%trans,trans-9,12 18∶2 from the triacylglycerol, containing ca. 50% of each acid. From the triacylglycerol containing 46.3%cis,trans-9,12 18∶2 and 53.7%trans,cis-9,12 18∶2, 44.8 and 55.2% of the two acids were hydrolyzed. Therefore the enzyme discriminated against thetrans,trans isomer but not between thecis,trans andtrans,cis isomers. Scientific contribution No. 535, Agricultural Experiment Station, University of Connecticut. ARS, USDA.     

Separation of conjugated methyl octadecadienoate and trienoate geometric isomers by silver-resin column and preparative gas-liquid chromatography

Silver-resin column chromatography, preceded by preparative gas-liquid chromatography to remove unconjugated esters, has been used to separate conjugated methyl octadecadienoates intocis,cis-, cis,trans-, andtrans,trans- isomers, also methyl eleostearate into a- and β-isomers in 91 to 100% purity. 1 Presented at the AOCS Meeting, Cincinnati, October 1965. 2 No. Util. Res. Dev. Div., ARS, USDA.     

Autoxidation of methyl linolenate: Analysis of methyl hydroxylinolenate isomers by high performance liquid chromatography

The mixture of methyl hydroxylinolenates obtained by reduction of the hydroperoxide isomers formed by autoxidation of methyl linolenate was resolved by high performance liquid chromatography into eight major components. These are positional isomers with the hydroxyl group at positions 9, 12, 13, and 16. Two geometrical isomers of each positional isomer are present; these differ in the configuration of the conjugated double bonds (cis-trans andtrans-trans). Autoxidation of methyl linolenate is regioselective and favors the formation of positional isomers 9 and 16.     

Positional isomers formed during the hydrogenation of methyl linoleate under various conditions

Journal of the American Oil Chemists' Society -     

Volatile products from mild oxidation of methyl linoleate. Analysis by combined mass spectrometry-gas chromatography

The volatile products from autoxidation of methyl linoleate have been analyzed by combined mass spectrometry-gas chromatography. The principal components were pentanal, hexanal, amyl formate, methyl octanoate, and substituted dioxolanes. Minor components included esters, alcohols, ketones, aldehydes, hydrocarbons, and acetals. Certain unsaturated carbonyl compounds, previously reported, were not detected. A laboratory of the W. Utiliz. Res. Devel. Div., ARS, USDA.     

甲苯磺酰氯异构体的气相色谱测定法

介绍了用气相色谱法分别对甲苯磺酰氯异构体及其水分进行的分析实验,得出了最佳操作条件,并以两次内加法进行了水分测定。     

Gas-liquid chromatography of the positional isomers of methyl nonynoate

A mixture of all the positional isomers of methyl nonynoate is poorly resolved by gasliquid chromatography on packed polar (diethylene glycol succinate) and packed nonpolar (Apiezon L) columns. Better resolution is obtained on capillary columns, with the polar liquid phase giving baseline separations between all the isomers except 9:T4 and 9:T5. The nonynoic esters are eluted later than methyl nonanoate on either liquid phase. The isomers with the triple bond near the center of the molecule come off first, and the elution time increases as the triple bond moves toward either end of the chain. Methyl 8-nonynoate, the only isomer with a terminal triple bond, does not follow this pattern. Presented at the AOCS meeting in Cincinnati, 1965.     

毛细管气相色谱法分析甲基丙烯酸甲酯中的阻聚剂

本文采用毛细管气相色谱法测定甲基丙烯酸甲酯中的阻聚剂--2,4-二甲基-6-叔丁基苯酚,操作简便,结果准确.实验选择了最佳汽化温度、柱温和载气流速.外标法定量,回收率可达99%～102%,变异系数为0.6%.     

The geometric isomers of conjugated octadecadienoates from dehydrated methyl ricinoleate

The dehydration of methyl ricinoleate by heatingin vacuo in the presence of KHSO₄ resulted in the formation of the following conjugated octadecadienoates expressed as a percentage of the final product:cis, trans (trans, cis), 14.3;cis, cis, 11.2;trans, trans, 7.3. The isomers contained the double bonds predominantly in the 9,11 position but the possible presence of traces of 8,10 and other conjugated isomers is not excluded. Using urea “inclusion” fractionation and low temp crystallization from acetone methyl,cis-9,cis-11-octadecadienoate was isolated. The methyl esters of commercially dehydrated castor oil fatty acids on the other hand, contained the following percentages of conjugated octadecadienoate isomers:cis, trans (trans, cis), 20.3;cis, cis, 8.0;trans, trans, 5.4. From these mixtures conc ofcis, trans (trans, cis)- andtrans, trans-octadecadienoates were prepared by fractional distillation and low temp crystallization. It was found that the conjugated octadecadienoates consisted of mixtures of positional isomers with double bonds mainly in the 8,10 and 9,11 positions with lesser amounts in the 7,9 and 10,12 positions.     

High performance liquid chromatography and glass capillary gas chromatography of geometric and positional isomers of long chain monounsaturated fatty acids

Positional and geometrical isomers of monounsaturated long chain fatty acids were analyzed by the combination of high performance liquid chromatography (HPLC) and glass capillary gas chromatography (GC). A preparative group separation ofcis andtrans isomers of the monounsaturated fatty acid methyl esters was achieved according to chain length by reversed-phase HPLC, and using a highly sensitive interference refractive index detector. After collection of the different fractions containingcis andtrans forms of the monounsaturated fatty acid methyl esters, the fractions were analyzed for their content of positional isomers using glass capillary GC with Silar-5 CP as stationary phase. The preparative step in the HPLC was also used analytically for the determination of the ratio between thecis andtrans monounsaturated fatty acids. A comparison was made between the results obtained with the HPLC technique and the results of a GLC technique with a packed OV-275 column. There was a good correlation between the 2 techniques with a tendency to highertrans values with the HPLC technique (4%). It was shown with reference substances that 18∶1ω6-cis to ω11-cis and 18∶1ω5-trans to ω12-trans, the most common monounsaturated fatty acid isomers in partially hydrogenated vegetable oils, could be almost quantitatively recovered in the HPLC step. Most of the individual positional isomers of monounsaturated fatty acids of varying chain length could be separated and determined in the glass capillary GC step with the exception of those isomers containing the double bond in a relatively high ω-position. The relative standard deviation of the technique as determined with reference substances was better than 4%. The described technique was applied to the analysis of the isomeric monounsaturated fatty acid content in partially hydrogenated vegetable and marine oils, and about 5 samples a day could be executed. Part of this work has been presented at the ISF/AOCS World Congress, New York (1980)JAOCS 58, (4), 1981, abstr. no. 184.     

cis andtrans Analysis of fatty esters by gas chromatography: Octadecenoate and octadecadienoate isomers

A gas chromatographic method has been developed for quantitative determination of thecis andtrans percentages in octadecenoate and octadecodienoate esters. To separatecis- andtrans-monoene and diene isomers on a packed GC column, the fatty esters were stereospecifically epoxipized with peracetic acid. A simple and quantitative epoxidation procedure allows thecis- andtrans-epoxyoctadecanoates to be analyzed without prior isolation from the reaction mixture. No positional or geometric isomerization of the double bond occurred during epoxidation. Synthetic mixtures containingcis- andtrans-6,-9 and-12 octadecenoate isomers were completely separated intocis andtrans fractionstrans-15-Octadecenoate was the only isomer investigated that partially interfered in the analysis. Diene mixtures containingrans,trans-, cis,trans- andcis,-cis-9,12-octadecadienoates were also successfully analyzed by gas liquid chromatography (GLC) after epoxidation with peracetic acid. Each diene isomer formed two pairs of diepoxy diastereomers, some of which could be separated. Onecis,cis-diepoxyoctadecanoate diastereomer peak overlapped thecis,trans-diepoxyoctadecanoate peaks. The totalcis,-cis-diepoxyoctadecanoate percentages were calculated by using the ratio of the twocis,cis-diepoxyoctadecanoate diastereomers. Other positional octadecadienoate isomers were also epoxidized and analyzed by GLC. The large number of possible octadecadienoate isomers requires that some preeiminary fractionation be made before GC analysis is practical for diene isomers. Presented at the AOCS Meeting, Atlantic, City, October 1971 N. Market. Nutr. Res. Div., ARS, USDA.     

Proportion of geometrical hydroperoxide isomers generated by radical oxidation of methyl linoleate in homogeneous solution and in aqueous emulsion

The proportion of geometrical hydroperoxide isomers generated by aerobic oxidation of methyl linoleate (18∶2 Me) in either aqueous emulsion consisting of Tris-HCl buffer (pH 7.4) or in a homogeneous dichloromethane solution was determined to understand the mechanism of lipid oxidation in different reaction systems. Four geometrical isomers were generated after oxidation of 18∶2 Me in dichloromethane: methyl 13-hydroperoxy-cis-9,trans-11-octadecadienoate, methyl 13-hydroperoxy-trans-9,trans-11-octadecadienoate, methyl 9-hydroperoxy-trans-10,cis-12-octadecadienoate, and methyl 9-hydroperoxy-trans-10,trans-12-octadecadienoate in the ratios of 1∶4∶1∶4, respectively. The ratios between each isomer did not change until the peroxide value (PV) increased to 58 meq/kg. Oxidation of 18∶2 Me in aqueous emulsion yielded the same geometrical isomers of hydroperoxide. However, the ratios were different: 3∶2∶3∶2 until the PV increased to 110 meq/kg. Predominant (60%) formation of trans,trans hydroperoxide isomers was obtained in the oxidation of a mixture of 18∶2 Me and methyl laurate (12∶0 Me). These results are interpreted to reflect the importance of the concentration of hydrogen atom-donating equivalents to the kinetic preference for different products. The high effective concentration of hydrogen donors in the oxidation of 18∶2 Me in emulsions favored the formation of the less stable cis,trans isomers. The lower concentration of hydrogen donor in the dichloromethane solution effectively slowed hydrogen donation and led to the strong preference for the more stable trans,trans isomers. This interpretation was further tested by preparing emulsions of 18∶2 Me and 12∶0 Me to dilute concentration of hydrogen-donating species using the nonhydrogen-donating 12∶0 Me. Consistent with the proposed hypothesis, the proportion of trans,trans isomers increased as a result of 12∶0 Me addition.     

Epoxidation of methyl linoleate. II. The two isomers of methyl 9, 10: 12, 13-diepoxystearate

Epoxidation of methyl linoleate results in the formation of two stereoisomeric methyl 9,10:12,13-diepoxystearates. The previously well-known isomer, mp 32C, represents only 60% of the total diepoxides formed. The newly isolated form is liquid at room temperature and is measured incompletely by standard titrimetric oxirane analysis. The isomers differ by the relative positions of their oxirane oxygen atoms. The oxygens of the solid are located on opposite sides and those of the liquid are on the same side of the molecular plane. E. Utiliz. Res. Devel. Div., ARS, USDA.     

Thermal decomposition of individual positional isomers of methyl linoleate hydroperoxide: Evidence of carbon-oxygen bond scission

The methyl ester of individual positional isomers of linoleate hydroperoxide were prepared by an enzymic oxidation of linoleate. On injection onto a gas chromatographic column they were thermally decomposed and the resulting volatile components analyzed. The major (67–80% yield on a molar basis) cleavage products were found to be hexanal, methyl octanoate, 2,4-decadienal isomers, and methyl 9-oxononanoate. Both the 9 and 13 isomers of linoleate hydroperoxide gave rise to these same four compounds, an observation suggesting carbon-oxygen scission in their decomposition. This was confirmed by using very pure individual isomers obtained by high performance liquid chromatography. The involvement of an isomerization reaction of the hydroperoxides is discussed.     

Autoxidation of methyl linoleate initiated by the ozonide of allylbenzene

Allylbenzene ozonide (ABO), a model for polyunsaturated fatty acid (PUFA) ozonides, initiates the autoxidation of methyl linoleate (18∶2 ME) at 37°C under 760 torr of oxygen. This process is inhibited by d-α-tocopherol (α-T) and 2,6-di-ert-butyl-4-methylphenol (BHT). The autoxidation was followed by the appearance of conjugated diene (CD), as well as by oxygen-uptake. The rates of autoxidation are proportional to the square root of ABO concentration, implying that the usual free radical autoxidation rate law is obeyed. Activation parameters for the thermal decomposition of ABO were determined under N₂ in the presence of radical scavengers and found to be E_a=28.2 ±0.3 kcal mol⁻¹ and log A=13.6±0.2; k_d (37°C) is calculated to be (5.1±0.3)×10⁻⁷ sec⁻¹. Autoxidation data are also reported for ozonides of 18∶2 ME and methyl oleate (18∶1 ME).     

Catalytic epoxidation of methyl linoleate

The epoxidation of methyl linoleate was examined using transition metal complexes as catalysts. With a catalytic amount of methyltrioxorhenium (4 mol%) and pyridine, methyl linoleate was completely epoxidized by aqueous H₂O₂ within 4 h. Longer reaction times (6 h) were needed with 1 mol% catalyst loading. Manganese tetraphenylporphyrin chloride was found to catalyze the partial epoxidation of methyl linoleate. A monoepoxidized species was obtained as the major product (63%) after 20 h.     

Autoxidation of methyl linoleate. Separation and analysis of isomeric mixtures of methyl linoleate hydroperoxides and methyl hydroxylinoleates

The mixture of conjugated diene hydroperoxide isomers obtained from autoxidation of methyl linoleate was separated by high performance liquid chromatography (HPLC). Four major isomers were obtained from adsorption chromatography and identified as the 9 and 13 positional isomers having thetrans-trans andcis-trans configurations. The latter geometrical isomers have thetrans double bond adjacent to the hydroperoxide group. The hydroxy compounds (methyl hydroxylinoleates) obtained from the hydroperoxides by NaBH₄ reduction were similarly separated but with improved resolution. This is the first instance of the complete separation of these compounds and provides a rapid method for their analysis. Unlike adsorption chromatography, reversed-phase chromatography separates the mixtures only according to the geometrical isomerism of the double bonds and not according to the position of the hydroxy or hydroperoxide function.     

Effect of temperature on the separation of some Δ9-cis-trans isomers of methyl esters of fatty acids by open tubular gas chromatography

The influence of temperature on the gas chromatographic separation ofcis-trans isomers of the methyl esters of some monounsaturated fatty acids was studied on capillary columns coated with Apiezon L, BDS and DEGS. As far as methyl oleate and methyl elaidate are concerned, the separation is better at lower temperatures on Apiezon L (180–210 C) and at higher temperatures on polyester phases (BDS, DEGS; 150–180 C). The influence of temperature on the separation ofcis-trans isomers on the three stationary phases under study is explained by the higher values of δECL/δt forcis isomers. The variation of the equivalent carbon chain length with temperature can be used for the identification ofcis-trans isomers in natural mixtures.