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Method development for the analysis of trans-fatty acids in hydrogenated oils by capillary electrophoresis.

A novel capillary electrophoresis methodology using UV indirect detection (224 nm) for the analysis of trans-fatty acids in hydrogenated oils was proposed. The electrolyte consisted of a pH 7 phosphate buffer at 15 mmol x L(-1) concentration containing 4 mmol.L(-1) sodium dodecylbenzenesulfonate, 10 mmol x L(-1) polyoxyethylene 23 lauryl ether (Brij 35), 2% 1-octanol and 45% acetonitrile. Under the optimized conditions, ten fatty acids, C12:0, C13:0 (internal standard), C14:0, C16:0, C18:0, C18:1c, C18:1t, C18:2cc, C18:2tt and C18:3ccc were baseline-separated in less than 12 min. The proposed methodology was applied to monitor the formation of trans-fatty acids during hydrogenation of Brazilnut oil. A crude oil sample (42.1% linoleic acid, 37.3% oleic acid, 13.4% palmitic acid, and 7.0% stearic acid) was mixed with 0.25% of a nickel-based catalyst and submitted to two independent hydrogenation conditions: 175 degrees C, 3 atm, 545 rpm for 60 min (GH(1) sample), and 150 degrees C, 1 atm, 545 rpm for 30 min (GH(2) sample). For the most severe hydrogenation condition (higher temperature and pressure, under longer reactional period), a more complete conversion of linoleic and oleic acids into stearic acid occurred with concomitant formation of the trans-species, elaidic acid (C18:1t). For the milder hydrogenation procedure that generated sample GH(2), larger amounts of linoleic and oleic acids remained, in addition to the transformations already observed in the GH(1) sample.

Electrophoresis, Capillary↗

Saturated, unsaturated, and trans-fatty acids modulate oxidative burst induced by high dextrose in human umbilical vein endothelial cells.

OBJECTIVE: We determined whether hyperglycemia-induced oxidative burst is augmented in the presence of saturated, unsaturated, and trans-fatty acids (FAs). METHODS: The time course of superoxide (O2-) production in human umbilical vein endothelial cells treated with 5.5 mM (D100) or 27.5 mM (D500) of dextrose and select FAs were measured using the hydroethidene fluorescence method. RESULTS: The rate of O2- production in cells treated with D500 was significantly higher (55% to 125% increase) than the rate observed in control cells treated with D100 (P < 0.001). The rate of O2- production (mean +/- standard deviation, in arbitrary units) was increased when cells were treated with 250, 500, and 1000 muM of myristic acid (C14; 0.189 +/- 0.04, 0.240 +/- 0.04, 0.234 +/- 0.02, respectively; P > 0.1, P < 0.05, P < 0.05, respectively), palmitate (C16; 0.151 +/- 0.03, 0.218 +/- 0.01, 0.289 +/- 0.07; P > 0.1, P < 0.05, P < 0.01), and stearic acid (C18; 0.321 +/- 0.03, 0.259 +/- 0.02, 0.341 +/- 0.03; P < 0.01 for all comparisons) compared with control cells treated with D100 only (0.184 +/- 0.01). In the presence of D500, myristic acid did not significantly augment O2- production, whereas stearic acid caused a significant further increase in O2- production. The cis unsaturated FA (oleic, C18:1; linoleic, C18:2; or linolenic acid, C18:3) stimulated O2- production significantly more than did saturated FA or trans-unsaturated FA such as elaidic, linolelaidic, or linolenelaidic acid. CONCLUSIONS: Fatty acids have the ability to modulate oxidative load in hyperglycemia and this effect may be related to the length, saturation, and possibly cis/trans orientation of the FA.

Cells, Cultured↗

Trans fatty acids. 1. Growth, fertility, organ weights and nerve histology and conduction velocity in sows and offspring.

Effects of dietary trans fatty acids on the pre- and postnatal growth and development in pigs were studied with special emphasis on nervous tissue. In experiment 1, female pigs were fed partially hydrogenated fish oil (PHFO) (28% trans) or soybean oil (PHSBO) (36% trans), in comparison with lard (0% trans) from weaning (3 wk) through the first reproduction cycle (up to 2 yr). In experiment 2, female pigs were fed two fish oils (33 and 19% trans) in comparison with coconut oil (0% trans) in diets with low and high levels of linoleic acid (18:2n-6 cis, cis) from gestation until their offspring were three wk old. Compared with the trans-free fats, the trans-containing fats had no effect on growth and development, feed consumption and utilization or on the weight of the brain, heart, kidneys, liver, lungs or spleen in the adult sows and their offspring. No effects from the experimental fats were found on histology and conduction velocity of the peroneal nerve. An increased number of the sows fed PHFO had fertility problems compared with those fed lard and PHSBO in Expt. 1, but no similar effects were seen in Expt. 2. It is concluded that consumption of trans fatty acids with 18-22 carbon atoms from PHFO and with 18 carbon atoms from PHSBO at levels that were 5 to 12 times higher than those normally consumed by humans had no detrimental effects on female pigs or their offspring during pregnancy and lactation.

Animals↗

Trans-fatty acids and radical stress: what are the real culprits?

Free radical-catalysed cis-trans isomerization of unsaturated lipids in biomimetic models and their significance in eukaryotic cells have been explored in the last few years, an integrating hypothesis being that trans-fatty acids have their origins in both dietary sources and from isomerization of natural isomers by an endogenous radical stress. In this perspective, a summary of the achievements and a discussion of the possible biological sources of isomerizing radical species are given, indicating a need for further research on thiyl radical generation in biological systems. In this context, crucial questions remain to be answered by free radical research involving membrane lipids, thus contributing to lipidomics and embracing biology and medicine.

Fatty Acids↗

Trans fatty acids and conjugated linoleic acids in the milk of urban women and nomadic Fulani of northern Nigeria.

BACKGROUND: Trans fatty acids (TFAs) and conjugated linoleic acids (CLAs) are present in dairy products and human milk and can have detrimental and beneficial effects in humans. The content of TFAs and CLAs in milk is determined largely by the diet of the mother. METHODS: We compared the proportions of TFAs and CLAs in the milk of rural Fulani in northern Nigeria who consume dairy products to that of women living in an urban center who consume little in the way of dairy products. Lactating Fulani women (n=41) and women residing in the city of Jos, Nigeria (n=41) were recruited into the study. We predicted that the milk of the Fulani pastoralists would contain higher amounts of TFAs and CLAs compared to their urban counterparts. RESULTS: The mean total TFA proportions for the Fulani and urban women were 0.22% and 0.34%, respectively, and were not significantly different. The percentages of CLAs in milk fat were not different between rural and urban women (0.16% vs 0.14%). These TFA and CLA values were 4- to 10-fold lower than for milk of women elsewhere in the world. CONCLUSIONS: The percentages of TFAs and CLAs in milk were not different between rural and urban dwellers in northern Nigeria whose diets differ greatly in the amounts of dairy products they contain. However, the fact that the percentages of TFAs and CLAs in the milk of Nigerian women were much lower than the percentages reported from other parts of the world may have implications for the long-term growth and development of infants in the northern Nigeria and elsewhere in the Western Sahel.

Adult↗

Rationale and design of the TRANSFACT project phase I: a study to assess the effect of the two different dietary sources of trans fatty acids on cardiovascular risk factors in humans.

BACKGROUND: Detrimental effects of consumption of industrial trans fatty acids (TFA) from partially hydrogenated vegetable oils (PHVO) on cardiovascular disease (CVD) risk factors are well documented. However, very little information is available on the effect of natural sources of TFA coming from milk fat, dairy products and ruminant meat. In fact, due to the naturally low level of TFA in milk fat, it is almost impossible to conduct a clinical trial with a limited number of subjects (<200). METHODOLOGY: To compare the effects of industrial and natural dietary sources of TFA, two specific test fats have been designed and produced. A substantial amount of milk fat (130 kg) enriched in TFA has been produced by modification of the cow's diet and selection of cows with the highest TFA content. The level obtained was approximately 4- to 7-fold higher than typically present in milk fat (approximately 20 instead of 3-6 g/100 g of total fatty acids). The control fat is composed of PHVO balanced in saturated fatty acids (lauric, myristic and palmitic). Both experimental fats contain about 20-22% of monounsaturated TFA and the volunteers' daily experimental fat intake (54 g), will represent about 12.0 g/day of TFA or 5.4% of the daily energy (based on 2000 kcal/day). These two test fats have been incorporated into food items and will be provided to 46 healthy subjects under a randomised, double blind, controlled, cross-over design. The primary outcome is high-density lipoprotein cholesterol (HDL-C), which is an independent risk factor for CVD. Other parameters such as low-density lipoprotein cholesterol (LDL-C), very low-density lipoprotein cholesterol (VLDL-C), and HDL-C level and subclasses will be also to be evaluated. CONCLUSION: We have shown that it is technically feasible to perform a clinical trial on the comparative effects of natural and industrial sources of TFA isomers on CVD risk factors. Results are expected by mid-2006.

Adult↗

Labeling of trans fatty acid content in food, regulations and limits-the FDA view.

With the scientific evidence associating trans fatty acid (TFA) intake with an increased risk of coronary heart disease (CHD), the U.S. Food and Drug Administration (FDA) issued a final rule that requires the declaration of the amount of TFA present in foods, including dietary supplements, on the nutrition label by January 1, 2006. The addition of TFA to the nutrition label will lead to the prevention of 600 to 1200 cases of CHD and 240-480 deaths each year saving Dollars 900 million to Dollars 1.8 billion per year in medical costs, lost productivity, and pain and suffering. For the purpose of nutrition labeling, TFA are defined as the sum of all unsaturated fatty acids that contain one or more isolated (i.e. non-conjugated) double bonds in a trans configuration. There are many issues that FDA has yet to resolve: (1) defining nutrient content claims for "free" and "reduced" levels of trans fat, (2) placing limits on the amount of TFA in conjunction with saturated fat limits for nutrient content claims, health claims, and disclosure and disqualifying levels, (3) a daily value, and (4) a possible footnote or disclosure statement to enhance consumer understanding of cholesterol raising lipids. FDA issued an Advanced Notice of Proposed Rulemaking (ANPR) requesting comments on the unresolved issues. FDA will also be conducting consumer research to determine consumer understanding of various TFA labeling possibilities. Comments to the ANPR, results of consumer research and current science will be used by FDA to resolve these issues and to determine future rulemaking for TFA labeling.

Cardiovascular Diseases↗

Differential effects of dietary saturated and trans-fatty acids on expression of genes associated with insulin sensitivity in rat adipose tissue.

OBJECTIVE: Trans-fatty acids (TFAs) are formed during partial hydrogenation of vegetable oils and are shown to be more atherogenic than saturated fatty acids (SFAs). Our previous study showed that dietary TFAs decrease adipose tissue insulin sensitivity to a greater extent than SFAs in rats. We hypothesized that the effects of these fatty acids on insulin sensitivity could be mediated through an alteration in gene expression. In the current study we have investigated the effects of dietary TFAs or SFAs on expression of genes associated with insulin sensitivity in rat adipose tissue. DESIGN AND METHODS: Male weanling Wistar/NIN rats were divided into four groups and fed one of the following diets containing 10% fat (g/100 g diet) differing only in the fatty acid composition for 3 months: control diet (3.7% linoleic acid (LA)), SFA diet (5% SFA), TFA diet 1 (1.5% TFA + 1% LA) and TFA diet 2 (1.5% TFA + 2% LA). The mRNA expression of peroxisome proliferator-activated receptor gamma (PPARgamma), lipoprotein lipase (LPL), glucose transporter-4 (GLUT4), resistin and adiponectin was analyzed in epididymal fat using RT-PCR. The effects of TFA were studied at two levels of LA to understand the beneficial effects of LA over the effects of TFA. RESULTS: Both dietary SFA and TFA upregulated the mRNA levels of resistin. Dietary SFA downregulated adiponectin and GLUT4 and upregulated LPL, while TFA downregulated PPARgamma and LPL. The effects of dietary TFA on PPARgamma and resistin were not counteracted by increased LA (TFA diet 2). CONCLUSION: The effects of SFAs on the aforementioned genes except PPARgamma could be extrapolated towards decreased insulin sensitivity, while only the alteration in the mRNA levels of PPARgamma and resistin could be associated with insulin resistance in TFA-fed rats. These findings suggest that dietary SFAs and TFAs alter the expression of different genes associated with insulin sensitivity in adipose tissue.

Adiponectin↗

Cell growth with trans fatty acids is affected by adenosine 3',5'-monophosphate and membrane fluidity.

Two positional isomers (9 and 11) of trans octadecenoates did not support growth on glucose of an Escherichia coli mutant that requires unsaturated fatty acids. However, the trans fatty acids provided sufficient fluidity to produce much higher cell yields when the concentration of adenosine 3',5'-monophosphate was raised. The effectiveness of the trans acids rose from 0 to 1 cell per femtomole to 15 to 20 cells per femtomole as the concentration of adenosine 3',5'-monophosphate was increased. The corresponding cis positional isomers supported high yields (35 to 40 cells per femtomole) independent of supplementation. The enhanced growth with adenosine 3',5'-monophosphate supplementation is not due to an increased uptake and incorporation of the trans isomers relative to the cis isomers, since the 9-trans isomer was incorporated more rapidly than the 9-cis isomer into the membrane phospholipids under all growth conditions and represented 21 +/- 2 mole percent of the acids. The finding that cells growing with trans fatty acid isomers have a higher requirement for adenosine 3',5'-monophosphate may indicate that some fatty acids can alter the metabolic regulation normally exerted by the cyclic nucleotide.

Cell Division↗

The effects of trans fatty acids on fatty acyl delta 5 desaturation by human skin fibroblasts.

The effectiveness of different fatty acids as inhibitors of fatty acyl delta 5 desaturation activity in human skin fibroblasts has been investigated. When incubated with 2.25 microM [14C] eicosatrienoate (20:3 omega 6) in otherwise lipid-free medium, these cells rapidly incorporate the radiolabeled fatty acid into cellular glycerolipids and desaturate it to produce both [14C] arachidonate and [14C]docosatetraenoate. The delta 5 desaturation activity can be enhanced by prior growth of the cells without serum lipids. Elaidate (9t-18:1) is a potent inhibitor of delta 5 desaturation while trans-vaccenate (11t-18:1) is virtually without effect. Oleate and linoleate are only mildly inhibitory. Linoelaidate (9t,12t-18:2) is more inhibitory than linoleate but significantly less effective than elaidate. The effects of elaidate can be readily overcome by increasing the concentration of exogenous eicosatrienoate. Studies with a variety of trans monounsaturates of differing chain lengths indicate that the omega 9 trans fatty acids are potent inhibitors of delta 5 desaturation, while omega 7 trans fatty acids are relatively ineffective. Intact human fibroblasts could thus be important in characterizing novel fatty acids as selective inhibitors of arachidonate synthesis in vivo.

Arachidonic Acids↗

Intake of trans fatty acids and risk of coronary heart disease among women.

Trans isomers of fatty acids, formed by the partial hydrogenation of vegetable oils to produce margarine and vegetable shortening, increase the ratio of plasma low-density-lipoprotein to high-density-lipoprotein cholesterol, so it is possible that they adversely influence risk of coronary heart disease (CHD). To investigate this possibility, we studied dietary data from participants in the Nurses' Health Study. We calculated intake of trans fatty acids from dietary questionnaires completed by 85,095 women without diagnosed CHD, stroke, diabetes, or hypercholesterolaemia in 1980. During 8 years of follow-up, there were 431 cases of new CHD (non-fatal myocardial infarction or death from CHD). After adjustment for age and total energy intake, intake of trans isomers was directly related to risk of CHD (relative risk for highest vs lowest quintile 1.50 [95% Cl 1.12-2.00], p for trend = 0.001). Additional control for established CHD risk factors, multivitamin use, and intakes of saturated fat, monounsaturated fat, and linoleic acid, dietary cholesterol, vitamins E or C, carotene, or fibre did not change the relative risk substantially. The association was stronger for the 69,181 women whose margarine consumption over the previous 10 years had been stable (1.67 [1.05-2.66], p for trend = 0.002). Intakes of foods that are major sources of trans isomers (margarine, cookies [biscuits], cake, and white bread) were each significantly associated with higher risks of CHD. These findings support the hypothesis that consumption of partially hydrogenated vegetable oils may contribute to occurrence of CHD.

Age Factors↗

Isomers of trans fatty acids modify the activity of platelet 12-P lipoxygenase and cyclooxygenase/thromboxane synthase.

OBJECTIVES: Trans isomers of fatty acids (TFA) have been implicated in the initiation and progression of atherosclerosis. Previous studies have shown that trans-unsaturated fatty acids like their cis-unsaturated counterparts exert a modifying effect on platelet aggregation. The aim of this work was to determine the influence of TFA on the aforementioned enzymes: lipoxygenase and cyclooxygenase/thromboxane synthase. METHODS: Two C18 cis-trans fatty acid pairs: oleic/elaidic and linoleic/linolelaidic were chosen. Fasting blood was sampled from 30 healthy volunteers without any lipid abnormalities and not on medication for at least 7 days prior to sampling. Platelet-rich plasma (PRP) was prepared and the platelet count was adjusted to 3 x 10(8) cells x ml(-1). Fatty acids in hexane were added to a final concentration of 7 microM and evaporated to dryness under nitrogen, 500 microl of PRP was pipetted into each tube, thoroughly mixed and incubated at 37 degrees C for 60 min. 12-P LOX activity was measured with a spectrophotometric method and expressed per mg protein. Cyclooxygenase/thromboxane synthase activity was determined by TXB(2) production with an ELISA-based assay. Statistics were done with the Kruskal-Wallis test and Statistica software package. RESULTS: We have found that the activity of 12-P LOX was suppressed by all cis-trans fatty acids used by us. Cyclooxygenase/thromboxane synthase activity was significantly inhibited by polyunsaturated fatty acids only. Linolelaidic acid was more potent in comparison to its monounsaturated (elaidic acid) counterpart. CONCLUSIONS: We believe that the effects of fatty acids are demonstrated at the membrane level where fatty acids may produce a perturbation in specific lipid domains. TFA are able to interact with the platelet membrane and transmembrane proteins just as the cis isomers. By interacting with proteins exposed on the cytoplasmic membrane, TFA may modify the activity of receptors and other membrane proteins. In this way, changes on the membrane surface are propagated into the cell affecting the activity of cytoplasmic enzymes like 12-P LOX and cyclooxygenase/thromboxane synthase.

Blood Platelets↗

Dietary fatty acids and insulin resistance.

High-fat diets have been associated with insulin resistance, a risk factor for both Type II diabetes and heart disease. The effect of dietary fat on insulin varies depending on the type of fatty acid consumed. Saturated fatty acids have been consistently associated with insulin resistance. On the other hand, medium and long-chain fatty acid intakes are associated with insulin sensitivity, as are high intakes of w3 fatty acids. Trans fatty acids appear to potentiate insulin secretion, at least in the short-term, to a greater degree than cis fatty acids. This may reflect chronic alterations in insulin sensitivity, although this remains to be tested. In summary, although it must be emphasized that all diets high in fat cause insulin resistance relative to high-carbohydrate diets, it appears that dietary saturated, short-chain, and w6 fatty acids have the most deleterious effects on insulin action.

Coronary Disease↗

Association of fatty acids in serum phospholipids with hay fever, specific and total immunoglobulin E.

The dietary intake of certain fatty acids might contribute to the development of allergic diseases such as hay fever and asthma. We investigated the association between the concentrations of fifteen fatty acids in serum phospholipids, as a marker of dietary intake and metabolism, and hay fever, allergic sensitisation and total IgE in adults. Data from a population-based cross-sectional study on respiratory health, including the measurement of fatty acids in the serum phospholipids of 740 adults between 20 and 64 years of age, were analysed. Positive associations were found between hay fever and arachidonic acid, and allergic sensitisation and oleic acid. No other fatty acids showed any association with hay fever or allergic sensitisation. Elevated levels of total IgE were not related to fatty acids. Concentrations of long-chain n-3 fatty acids, trans fatty acids or saturated fatty acids in serum phospholipids were not associated with allergic diseases in adults in this study. The present result on the association between hay fever and arachidonic acid is consistent with current hypotheses but warrants further research.

Adult↗

Saturated fats: what dietary intake?

Public health recommendations for the US population in 1977 were to reduce fat intake to as low as 30% of calories to lower the incidence of coronary artery disease. These recommendations resulted in a compositional shift in food materials throughout the agricultural industry, and the fractional content of fats was replaced principally with carbohydrates. Subsequently, high-carbohydrate diets were recognized as contributing to the lipoprotein pattern that characterizes atherogenic dyslipidemia and hypertriacylglycerolemia. The rising incidences of metabolic syndrome and obesity are becoming common themes in the literature. Current recommendations are to keep saturated fatty acid, trans fatty acid, and cholesterol intakes as low as possible while consuming a nutritionally adequate diet. In the face of such recommendations, the agricultural industry is shifting food composition toward lower proportions of all saturated fatty acids. To date, no lower safe limit of specific saturated fatty acid intakes has been identified. This review summarizes research findings and observations on the disparate functions of saturated fatty acids and seeks to bring a more quantitative balance to the debate on dietary saturated fat. Whether a finite quantity of specific dietary saturated fatty acids actually benefits health is not yet known. Because agricultural practices to reduce saturated fat will require a prolonged and concerted effort, and because the world is moving toward more individualized dietary recommendations, should the steps to decrease saturated fatty acids to as low as agriculturally possible not wait until evidence clearly indicates which amounts and types of saturated fatty acids are optimal?

Cholesterol, Dietary↗

[Spanish contribution to the creation of a European analytical database of trans-fatty acids].

Within the AAIR Program of the EU titled Evaluation of the Ingestion of Trans Fatty Acids (FA) and its association with cardiovascular risk factors in European countries (TRANSFAIR), which is being carried out in 16 countries with different alimentary habits, among which is our country, we have developed the following study. Based on the information derived from the last National Nutrition and Feeding Study (ENNA-3), we have made up a list of foods which make up 95% of the total ingestion of lipids, and it also includes those which although they are not included within this percentage, may have an especially high trans isomer content as a result of their processing. The foods selected for the analysis belong to different food groups: cereals, milk products, oils and fats, meats, various, and pre-cooked foods, until making up a total of 100 foods for each country. The central analysis laboratory is that of the Department of Human Nutrition, TNO Nutrition and Food Research, Zeist (The Netherlands). In each sample, in addition to the total lipids, one determines the saturated and unsaturated fatty acids, including the cis and trans isomers. The trans FA's measured were: C14:1 T9, C16:1 T9, C18:1 T, C18:2 T, C18:3 T + C20:1 T, C20:2 T11,14, and C22:1 T13. Of the samples analyzed, the highest percentages of trans FA with respect to the total FA corresponded to the following foods: French fries, pre-cooked and frozen croquettes sliced bread, margarine, cakes, and frozen mille feuilles dough of different industrial brands. The lowest percentages of trans FA's were found in refined vegetable oils (sunflower and olive), those used for deep frying, and those discarded in catering, as well as in some commercial brands of cookies and ice creams. Pure chocolate, different brands of sweetened powdered cocoa, and ready to make chocolate, did not contain and trans FA.

Databases as Topic↗

Platelet trans fatty acids in relation to angiographically assessed coronary artery disease.

Epidemiological and metabolic studies indicate that a higher intake of trans fatty acids (TFA) may be associated with increased risk of coronary heart disease (CHD). In a cross-sectional study of patients who underwent coronary angiography, the relationships between TFAs, measured in platelets, and the degree of coronary artery disease (CAD) were examined in 191 non-diabetic patients (134 men and 57 women). The degree of CAD was quantified by using an angiographic scoring system developed to provide an estimate of the extent of coronáry atherosclerosis: an "extent score'. The TFA composition of platelets, including palmitelaidic (16:1 omega 7t), elaidic (18:1 omega 9t), trans-10-octadecaenoic acid (18:1 omega 8t), trans vaccenic (18:1 omega 7t), trans-12-octadecaenoic acid (18:1 omega 6t) and linoelaidic (18:2 omega 6tt) acids, was measured by using gas chromatography and quantified as a percentage of total fatty acids. After adjustment for established CHD risk indicators, including age, gender, cigarette smoking, hypertension and serum total cholesterol concentration, elaidic acid (P = 0.0300) and trans-10-octadecaenoic acid (P = 0.0434) were positively associated with the extent score of CAD. The adjusted associations between other individual TFAs, including palmitelaidic acid (P = 0.1189), vaccenic acid (P = 0.7651), trans-12-octadecaenoic acid (P = 0.0582) and linoelaidic acid (P = 0.8793), and the extent score were not significant. The results of this study, therefore, provide evidence for an association between particular platelet TFAs and the degree of CAD in the patient population studied.

Adolescent↗