Distribution of hexadecenoic,octadecenoic and octadecadienoic acid isomers in human tissue lipids

Thetrans 16∶1, 18∶1 and 18∶2 fatty acid composition of various human organ lipids was studied to determine if isomers accumulated in specific tissues. “Trans” isomers are defined as those fatty acids containing one or moretrans double bonds. Adipose, kidney, brain, heart and liver tissue lipids were analyzed. Gas chromatography with a 100-SP2560 capillary column was used to characterize the various positional and/or geometrical isomers. The distribution ofrans 16∶1 and 18∶1 isomers ranged from 0.3% in the brain to 4.0% in adipose tissue, whiletrans 18∶2 isomers ranged from 0.0% in the brain to 0.4% in adipose tissue. Notrans 18∶3 isomers were detected. Positional isomer ratios forcis 16∶1 (Δ9 vs Δ7) andcis 18∶1 (Δ11 vs Δ9) were also determined. Since these ratios are reproducible from one individual to the next, they might be useful for diagnosis of human metabolic disorders.     

Occurrence of octadecenoic fatty acid isomers from hydrogenated fats in human tissue lipid classes

The level oftrans-18∶1 isomers in several isolated lipid classes of human liver, heart, red blood cells and plasma was determined. Phospholipids contained substantially fewertrans-18∶1 isomers than triglycerides. The double bond distribution of thecis andtrans octadecenoate fraction of triglycerides and phosphatidylcholines from human liver and heart was determined. Whereas the double bond distribution of the triglycerides correlated closely with the pattern found in dietary hydrogenated vegetable oils, the phosphatidylcholine fraction showed evidence of selective incorporation or metabolism of specifictrans positional isomers. In general, isomers with double bonds near the methyl terminus were present at levels higher than expected from their relative abundance in the diet. Refinements in methodology needed to analyze octadecenoate double bond configuration and location in human tissues are presented.     

Trans fatty acid isomers in Canadian human milk

The fatty acid composition, totaltrans content (i.e., sum of all the fatty acids which may have one or moretrans double bonds) and geometric and positional isomer distribution of unsaturated fatty acids of 198 human milk samples collected in 1992 from nine provinces of Canada were determined using a combination of capillary gas-liquid chromatography and silver nitrate thin-layer chromatography. The mean totaltrans fatty acid content was 7.19±3.03% of the total milk fatty acids and ranged from 0.10 to 17.15%. Twenty-five of the 198 samples contained more than 10% totaltrans fatty acids, and thirteen samples contained less than 4%. Totaltrans isomers of linoleic acid were 0.89% of the total milk fatty acids with 18∶2Δ9c, 13t being the most prevalent isomer, followed by 18∶2Δ9c, 12t and 18∶2Δ9t, 12c. Using the totaltrans values in human milk determined in the present study, the intake of totaltrans fatty acids from various dietary sources by Canadian lactating women was estimated to be 10.6±3.7 g/person/d, and in some individuals, the intake could be as high as 20.3 g/d. The 18∶1trans isomer distribution differed from that of cow's milk fat but was remarkably similar to that in partially hydrogenated soybean and canola oils, suggesting that partially hydrogenated vegetable oils are the major source of thesetrans fatty acids.     

Polymorphism in single-acid triglycerides of positional and geometric isomers of octadecenoic acid

The polymorphism of 25 glycerol trioctadecenoates with double bonds ranging from Δ4-Δ17 was investigated by differential scanning calorimetry. Triglycerides withcis bonds in odd positions Δ7-Δ13 exhibited three intermediate melting (β′-) forms, but those withcis bonds in even positions, exceptcis Δ4, lacked β′-forms. Among thetrans compounds, only Δ11, 13, and 14 showed β′-forms. Thecis andtrans Δ5 triglycerides were unusual, because they readily assumed low melting (α-) forms that were not easily converted to high melting (β-) forms. β-Form mp of compounds in each series (cis ortrans) alternated depending upon double bond position; an even position correlated with high mp. Heats of fusion (ΔH_f) for β-forms, likewise, fluctuated with double bond position but nonuniformly;trans Δ6 had the highest ΔH_f (43 cal/g),cis Δ12 the lowest (21 cal/g).     

Trans octadecenoic fatty acid (TFA) isomers in German foods and bakery goods

In the present study, 122 food samples from the German food market were analysed for their C18:1 trans fatty acid (TFA) content and profile. A particular focus of the survey were baked and fried foods. TFA analysis was performed by means of silver ion SPE (Ag⁺‐SPE) in combination with high‐resolution GC (HRGC‐FID). Overall, 51 bakery product samples were analysed of which 25 samples were prepacked bakery products purchased from local retail stores and 26 samples of unpacked bakery products purchased from local bakery shops. In addition, 14 French fries samples obtained from small local fast food restaurants as well as from internationally operating fast food chains, 27 potato and tortillas chips, 15 instant soups as well as 15 dry culinary sauces were analysed. The highest amounts of C18:1 TFA isomers were found in deep‐fried bakery products. Prepacked branded cookies and biscuits on the other hand contained only negligible C18:1 TFA amounts. Regarding their C18:1 trans isomer profile most deep‐fried bakery products exhibited a Gaussian‐distributed isomer profile. The analysed prepacked croissants, cookies and biscuits contained predominantly ruminant TFA (TFA) as suggested by the presence of vaccenic acid (C18:1 trans 11), which was the major C18:1 TFA isomer in these products. All non‐bakery samples (n = 71) contained less than 3 g C18:1 TFA per 100 g fat. In conclusion, TFA still occur in considerable amounts in a few German food products, especially in some deep‐fried bakery products (‘Berliner’ type of doughnuts). Practical applications: Trans fatty acids, in particular the trans octadecenoic fatty acid isomers (C18:1), are generally considered from the nutritional point of view as undesirable food components due to their negative health effects. Tremendous efforts have been made by major food processors in order to decrease or even eliminate the presence of TFA in some foodstuffs (e.g. in margarines in European countries). However, some food processors of other food sectors are still applying oils and fats containing partially hydrogenated vegetable oils, whereas others within the same food category have already switched their processing conditions and/or raw materials towards TFA alternatives. Therefore, actual TFA data of foodstuffs determined by means of state‐of‐the‐art analytical procedures (Ag⁺‐SPE in combination with GC‐FID) is necessary to detect areas of further improvement in the food supply chain and to provide data for an update of dietary TFA intake.     

Maternal dietary alpine butter intake affects human milk: Fatty acids and conjugated linoleic acid isomers

Consumption of CLA by lactating women affects the composition of their milk, but the pattern of the different CLA isomers is still unknown. We determined the effects of short maternal supplementation with CLA-rich Alpine butter on the occurrence of FA and CLA isomers in human milk. In an open randomized controlled study with a two-period cross-over design, milk FA and CLA isomer concentrations were measured on postpartum days >-20 in two parallel groups of lactating women before, during, and after consumption of defined quantities of Alpine butter or margarine with comparable fat content (10 d of butter followed by 10 d of margarine for one group, and vice versa in the other). In the 16 women who completed the study (8/group), Alpine butter supplementation, increased the C₁₆ and C₁₈ FA, the sum of saturated FA, the 18∶1 trans FA, and the trans FA with CLA. The CLA isomer 18∶2 c9, t11 increased by 19.7%. Significant increases were also found for the isomers t9,t111, t7,c9,t11,c13, and t8,c10 18∶2. The remaining nine of the total 14 detectable isomers showed no changes, and concentrations were <5 mg/100g fat. A breastfeeding mother can therefore modulate the FA/CLA supply of her child by consuming Alpine butter. Further studies will show whether human milk containing this FA and CLA isomer pattern acts as a functional food for newborns.     

Trans- and cis-octadecenoic acid isomers in the hump and milk lipids from Camelus dromedarius

The distribution profiles of individual trans- as well as cis-18∶1 isomers from the fat prepared from the hump adipose tissue and the milk from Camelus dromedarius (the single-humped Arabian species) are described. Gas-liquid chromatography on two capillary columns with different polarities and lengths were used for this purpose in combination with argentation thin-layer chromatography. A comparison of the profiles established is made with that of true ruminant fats. In the fats from the dromedarius as well as from true ruminants, the trans-18∶1 isomers have their ethylenic bonds in all positions between Δ4 and Δ16. The prominent trans isomer is the 11–18∶1 (vaccenic) acid in all species, and the complete distribution profiles are quite similar. Concerning the cis isomers, the prominent isomer is oleic acid, followed by cis-vaccenic acid, as in true ruminant fats. Other cis isomers encompass the Δ6–8 and the Δ12 to Δ15 isomers. Camelidae (suborder Tylopoda) and Bovidae (suborder Ruminantia) have evolved independently since the Eocene, that is for approximately 50 million years. Despite this considerable period, and the profound differences in anatomy, morphology, physiology, ecological and dietary habits between the extant species of these suborders, the rumen microflora has continued to synthesize the same trans- and cis-octadecenoic acid isomers, in comparable proportions, at least as deduced from their composition profiles. We conclude that the trans-18∶1 acid profile is not intrinsically species-dependent, but it can be affected by the nature and the proportions of dietary unsaturated fatty acids that themselves depend on the feed, and that may be species-specific.     

Gas-liquid chromatographic analysis of geometrical and positional octadecenoic and octadecadienoic acid isomers produced by catalytic hydrogenation of linoleic acid on Ir/Al2O3

Catalytic hydrogenation of linoleic acid was studied on Ir/Al₂O₃. A detailed analysis of geometrical and positional isomers of octadecenoic acid (18:1) in the products was performed by capillary gas-liquid chromatography with a new capillary column coated with isocyanopropyl trisilphenylene siloxane (TC-70). Well-resolved peaks of 10 species of 18:1 were observed in the product. In addition to monoenoic acid isomers, four species of trans-dienoic isomers and conjugated dienoic isomers were found. From the distribution of 18:1 isomers, it was found that the double bond closer to the methyl end (Δ12) showed higher reactivity than that closer to the carboxyl end (Δ9) for hydrogenation. Because cis-8 18:1 and trans-8 18:1 were not observed but cis-10 18:1 and trans-10 18:1 were observed in the products, the double-bond Δ9 did not migrate to the carboxyl end but migrated to the methyl end. On the other hand, the Δ12 bond migrated to both methyl and carboxyl ends. From the distribution of 18:1 isomers in the reaction pathway, the hydrogenation of linoleic acid proceeds via half-hydrogenation states. Cis-18:1 isomers were produced predominantly in the initial stage of the reaction, while trans-18:1 isomers were produced during progress of the reaction. The cis/trans and positional isomerization took place by readsorption of 18:1 produced by the partial hydrogenation of linoleic acid.     

Synthesis of octadecynoic acids and [1-14C] labeled isomers of octadecenoic acids

Geometric and positional isomers of [1-¹⁴C] octadecenoic acids have been synthesized by modifications of published procedures. Positional isomers of octadecynoic acids also have been synthesized to obtain the geometric and positional isomers of the unlabeled octadecenoic acid analogs. The syntheses were accomplished by coupling a haloalkyl compound with a substituted acetylene using n-butyl lithium in hexamethylphosphoramide. The coupled product, either a 17-or 18-carbon acetylenic alcohol, could be semihydrogenated and chain extended to afford a carboxy labeled derivative, could be partially hydrogenated and chain extended to afford a carboxyl labeledcis-ortrans-octadecenoic acid in the former case. In the latter case, octadecynoic,cis-octadecenoic ortrans-octadecenoic acids could be obtained by the appropriate reactions. The methods used in this study enabled the synthesis of¹⁴C-labeled fatty acids in generally higher yields and by simpler reactions than were previously possible.     

Trans-geometrical and positional isomers of linoleic acid including conjugated linoleic acid (CLA) in German milk and vegetable fats

A representative number of different milk fats based on a wide range of feeding and lactation conditions as well as 123 German margarines, shortenings, cooking and dietetic fats were analyzed for a variety of trans-C18:2 isomers (exhibiting at least one trans double bond) by means of gas chromatography on a 100m Sil 88 capillary column. In milk fats contents of trans Δ9,trans Δ12,cis Δ9,trans Δ13 (+ trans Δ8,cis Δ12),trans Δ8,cis Δ13,cis Δ9,trans Δ12,trans Δ9,cis Δ12 and trans Δ11,cis Δ15 amounted to 0.09%, 0.11%, 0.11%, 0.10%, 0.07% and 0.33% on average and the content of total trans-C18:2 isomers (without cis Δ9, trans Δ11) was 0.99% in the mean. The content of conjugated linoleic acid cis Δ9,trans Δ11 amounted to 0.81% on average in 238 milk fats. In margarine among others the isomers trans Δ9,trans Δ12,cis Δ9,trans Δ13 (+ trans Δ8,cis Δ12), cis Δ9,trans Δ12 and trans Δ9,cis Δ12 were determined and quantified to 0.03%, 0.04%, 0.29% and 0.23% on average. The mean total content of trans-C18:2 isomers in margarines was 0.61%. Moreover, for all trans-C18:2 isomers the frequency distributions as well as the correlation coefficients towards the trans-C18:1 isomers trans Δ6 to trans Δ16 were derived.     

Polyunsaturated fatty acid supply with human milk

Polyunsaturated fatty acids in human milk may derive from diet, liberation from maternal body stores, or endogenous synthesis from precursor fatty acids. The contribution of each of these sources has not been studied in detail. Although maternal diet is a key factor affecting human milk composition, other factors such as gestational age, stage of lactation, nutritional status, and genetic background are known to influence the fat content and fatty acid composition in human milk. Both linoleic and α-linolenic acids, the essential fatty acids, are present in human milk, as are several other n−6 and n−3 longer chain polyunsaturated fatty acids that are required for optimal growth and development of infants. The fatty acid profile of human milk from lactating women of different countries is remarkably stable, but there is variability in some of the components, such as docosahexaenoic acid, which is mainly due to differences in dietary habits. Tracer techniques with stable isotopes have been valuable in assessing the kinetics of fatty acid metabolism during lactation and in determining the origin of fatty acids in human milk. Based on these studies, the major part of polyunsaturated fatty acids in human milk seems not to be provided directly from the diet but from maternal tissue stores.     

Fatty acid and conjugated linoleic acid isomer profiles in human milk fat

The fatty acid composition of 39 mature human milk samples from four Spanish women collected between 2 and 18 weeks during lactation was studied by gas chromatography. The conjugated linoleic acid (CLA) isomer profile was also determined by silver‐ion HPLC (Ag⁺‐HPLC) with three columns in series. The major fatty acid fraction in milk lipids throughout lactation was represented by the monounsaturated fatty acids, with oleic acid being the predominant compound (36–49% of total fatty acids). The saturated fatty acid fraction represented more than 35% of the total fatty acids, and polyunsaturated fatty acids ranged on average between 10 and 13%. Mean values of total CLA varied from 0.12 to 0.15% of total fatty acids. The complex mixture of CLA isomers was separated by Ag⁺‐HPLC. Rumenic acid (RA, cis‐9 trans‐11 C18:2) was the major isomer, representing more than 60% of total CLA. Trans‐9 trans‐11 and 7‐9 (cis‐trans + trans‐cis) C18:2 were the main CLA isomers after RA. Very small amounts of 8‐10 and 10‐12 C18:2 (cis‐trans + trans‐cis) isomers were detected, as were different proportions of cis‐11 trans‐13 and trans‐11 cis‐13 C18:2. Although most of the isomers were present in all samples, their concentrations varied considerably.     

Effect of abomasal infusions of geometric isomers of 10,12 conjugated linoleic acid on milk fat synthesis in dairy cows

The trans-10, cis-12 isomer of conjugated linoleic acid (CLA) decreases TAG accumulation in 3T3-L1 adipocytes, reduces lipid accretion in growing animals, and inhibits milk fat synthesis in lactating mammals. However, there is evidence to suggest that other FA may also exert antilipogenic effects. In the current experiment, the effects of geometric isomers of 10,12 CLA on milk fat synthesis were examined using four Holstein-British Friesian cows in a 4×4 Latin Square experiment with 14-d periods. Treatments consisted of abomasal infusions of skim milk, or skim milk containing trans-10, cis-12 CLA (T1), trans-10, trans-12 CLA (T2), or a mixture of predominantly 10,12 isomers containing (g/100 g) trans-10, cis-12 (35.0), cis-10, trans-12 (23.2), trans-10, trans-12 (14.9), and cis-10, cis-12 (5.1). CLA supplements were prepared from purified ethyl linoleate and infused as nonesterified FA. Infusions were conducted over a 4-d period with a 10-d interval between treatments and targeted to deliver 4.5 g/d of 10,12 CLA isomers. Compared with the control, trans-10, trans-12 CLA had no effect (P>0.05) on milk fat yield, whereas treatments T1 and T3 depressed (P<0.05) milk fat content (19.8 and 22.9%, respectively) and decreased milk fat output (20.8 and 21.3%, respectively). Comparable reductions in milk fat synthesis to 4.14 and 1.80 g trans-10, cis-12/d supplied by treatments T1 and T3 indicate that other 10,12 geometric isomers of CLA have the potential to exert antilipogenic effects. The relative abundance of cis-10, trans-12 CLA in treatment T3 and the low transfer efficiency of this isomer into milk suggest that cis-10, trans-12 CLA was the active component.     

Lactational changes in the fatty acid composition of human milk gangliosides

The objectives of this work were to study the FA composition of milk gangliosides, as well as to gain further insight into the characterization of human milk gangliosides. The potential capacity of human milk gangliosides to adhere to human enterotoxigenic Escherichia coli (ETEC-strains) was also studied. Human milk gangliosides were isolated and identified by high-performance TLC or immunoassay. The latter also was used to assay bacterial adhesion. The FA composition of gangliosides was studied by GC. The presence of O-acetyl GD3 (Neu5,9Ac₂α2–8 NeuAcα2–3Galβ1–4GlcCer) and trace amounts of GM1 [Galβ1–3GalNAcβ1,−3(NeuAcα2–3)Galβ1–4GlcCer] in human milk was confirmed. Medium-chain FA were almost absent in colostrum, whereas in the subsequent stages they rose to 20%. The levels of long-chain FA decreased after colostrum. With respect to the degree of saturation, gangliosides from colostrum were richer in monounsaturated FA than gangliosides synthesized during the rest of the lactation period, opposite to the pattern for PUFA. A human-ETEC colonization factor antigen II-expressing strain showed binding capacity to human milk GM3 (NeuAcα2–3Galβ1–4GlcCer). New data on human milk gangliosides have been gathered. A thorough knowledge of their composition is needed since they may have important biological implications in regard to newborns' defense against infection. The ganglioside nomenclature of Svennerholm (34) is followed.     

Dual-labeled technique for human lipid metabolism studies using deuterated fatty acid isomers

Two deuterated fatty acids, elaidate-d ₂ and oleate-d ₄, were fed simultaneously to a human subject as a mixture of trielaidin-d ₆ and triolein-d ₁₂. Periodically, blood samples were drawn, and red blood cells were separated from the plasma. Red blood cells and plasma lipids were fractionated and analyzed by combined gas chromatography—multiple ion mass spectroscopy. Dual deuterium-labeling allows rate and extent of fatty acid incorporation to be followed in various plasma and red cell neutral and phospholipid fractions. Maximum amount of deuterated fat varied from 4% in cholesterol ester to 64% in phosphatidyl ethanolamine. The highest levels of deuterated fat occurred in either 6-, 8-, or 12-hr samples; generally, <1% labeled fatty acids could be detected in 72-hr samples. Because the method is based on dual-labeling, differences in the relative incorporation of both fatty acid isomers can be compared directly. Differences in rates of incorporation, rates of removal, and extent of incorporation of labeled fatty acids into blood plasma can also be determined reliably. Our experimental labeling of fats with deuterium permits for the first time the metabolism of two fatty acid isomers to be compared simultaneously in human subjects. This new method should be applicable to a variety of other lipid metabolic studies. Presented at the AOCS meeting Dallas, Texas, April 27–30, 1975.     

Identification of positional isomers of lysophosphatides in butter serum

The separation of individual classes of phospholipids by two-dimensional thin layer chromatography revealed the presence of lysophosphatides (monoacyl glycerophosphoryl compounds) in minor quantities in butter serum. The naturally occurring lysophosphatidyl ethanolamine (LPE) and lysophosphatidyl choline (LPC) were prepared by the combined use of a diethylaminoethyl (DEAE) cellulose column and thin layer chromatography. Gas chromatographic analysis of methyl esters of LPE and LPC showed that both lysophosphatides contained, in addition to saturated fattya cids, considerable quantities of unsaturated fatty acids. A comparison of the fatty acid pattern of naturally occurring LPE and LPC with that of LPE and LPC prepared by phospholipase A hydrolysis of phosphatidyl ethanolamine (PE) and phosphatidyl choline (PC) suggested that both the 1-and 2-forms of positional isomers were present. Hydrolysis of naturally occurring LPE and LPC with phospholipase C, and separation of the monoglycerides formed, confirmed the occurrence of two positional isomers of lysophosphatides.     

Biological utilization of fatty acid isomers

Summary A review of the literature has shown that in the hydrogenation of vegetable oils, positional and stereoisomers of the unsaturated fatty acids are formed in appreciable quantities and that some isomers are intermediates in the development of others as hydrogenation proceeds. Using a microbiological assay technique, it was demonstrated that the iso-oleic acids formed during hydrogenation are not antimetabolites for natural oleic acid but are utilized as nutrients. Fatty acids with conjugated double bonds are not antimetabolites for the essential fatty acids but are readily metabolized to carbon dioxide and water. Hydrogenated fats compare favorably with a natural fat of comparable firmness in serving as a source of essential fatty acids. In hydrogenating vegetable oils, isomers of linoleic acid can be formed which resist spectrophotometric detection but exhibit essential fatty acid activity. Paper presented at the 1953 Conference on Food and Nutrition, Gordon Research Conferences, AAAS, Colby Junior College, New London, N. H. Contribution 351 of the Department of Biochemistry and Nutrition, University of Southern California.     

Biochemistry of unsaturated fatty acid isomers

Recognition that catalytic hydrogenation changes the configuration and position of double bonds and alters the physical properties of unsaturated fats prompted numerous early investigations on the bio-chemical effects of ldtrans isomers.” Recent research has provided data on positional isomer metabolism. Some aspects of fatty acid isomer metabolism are now reasonably well understood, but other issues are not resolved. Human and animal data have provided good evidence that isomers in partially hydrogenated oils are well adsorbed and incorporated into all organs and tissues. Analyses of human tissues also indicate that hydrogenated oils are the major source of fatty acid isomers in the US diet. Tissue composition data combined with isolated enzyme studies and isotope tracer experiments with whole organisms show unquestionably that structural differences between various fatty acid isomers influence specific biochemical transformations. Examples are differences in the reaction rates and/ or specificities of acyl transferase, lipase, desaturase and cholesteryl esterase/hydrolase for various positional fatty acid isomers. Isolated microsomes and mitochondria also have been used to identify dif-ferences in acyl CoA activation, oxidation, and elongation of posi-tional isomers. In addition, isotope tracer experiments show that preferential metabolism of individual positional isomers occurs in vivo. In vivo studies with hydrogenated vegetable oil diets containing adequate levels of linoleic acid produced no obvious physiological changes. Experiments with specific polyunsaturated isomers have produced changes in blood cell properties, pulmonary weight, lino-leic acid requirements and tissue lipid composition. These changes may be related to a number of factors such as membrane fluidity and permeability, cell function, synthesis of arachidonic acid, homo-gamma-linoleic acid or prostaglandins. Whether differences in the biochemistry of fatty acid isomers are desirable or undesirable and whether these differences contribute to long-term or subtle effects important to the etiology of atherosclerosis and cancer are not resolved. Presented at the 73rd AOCS annual meeting, Toronto, 1982.     

The use of SP2340 glass capillary columns for the estimation of thetrans fatty acid content of foods

Glass capillary gas chromatography (GCGC) on 100-m and 60-m SP2340 columns was used for quantitation of thetrans unsaturated fatty acids in shortenings and fast foods. The separation of thecis andtrans octadecenoates on GCGC was evaluated by preparatory argentation thin layer chromatography. In addition, thetrans content of shortening samples obtained by GCGC was compared totrans content determined by infrared analysis. This research conducted by the Science and Education Administration, USDA, on the commercial foods as reported in this paper was limited to analysis of their lipid compositions. The data are reported solely as factual information and are limited to the samples analyzed. No warranty or guarantee is made or implied that other samples of these products will have the same or similar composition. It is the policy of the USDA not to endorse those commercial products used in research over those that were not included in the research.