Achievement of dietary fatty acid intakes in long-term controlled intervention studies: approach and methodology

Objective: To describe the calculations and approaches used to design experimental diets of differing saturated fatty acid (SFA) and monounsaturated fatty acid (MUFA) compositions for use in a long-term dietary intervention study, and to evaluate the degree to which the dietary targets were met. Design, setting and subjects: Fifty-one students living in a university hall of residence consumed a reference (SFA) diet for 8 weeks followed by either a moderate MUFA (MM) diet or a high MUFA (HM) diet for 16 weeks. The three diets were designed to differ only in their proportions of SFA and MUFA, while keeping total fat, polyunsaturated fatty acids (PUFA), trans-fatty acids, and the ratio of palmitic to stearic acid, and n-6 to n-3 PUFA, unchanged. Results: Using habitual diet records and a standardised database for food fatty acid compositions, a sequential process of theoretical fat substitutions enabled suitable fat sources for use in the three diets to be identified, and experimental margarines for baking, spreading and the manufacture of snack foods to be designed. The dietary intervention was largely successful in achieving the fatty acid targets of the three diets, although unintended differences between the original target and the analysed fatty acid composition of the experimental margarines resulted in a lower than anticipated MUFA intake on the HM diet, and a lower ratio of palmitic to stearic acid compared with the reference or MM diet. Conclusions: This study has revealed important theoretical considerations that should be taken into account when designing diets of specific fatty acid composition, as well as practical issues of implementation.

The influence of dietary fatty acid composition on N-ethyl-Nnitrosourea-induced mammary tumour incidence in the rat and on the composition of inositol- and ethanolamine-phospholipids of normal and tumour mammary tissue

This study has investigated the influence of dietary fatty acid composition on mammary tumour incidence in N-ethyl-N-nitrosourea (ENU)-treated rats and has compared the susceptibility to dietary fatty acid modification of the membrane phospholipids phosphatidyliuositol (PI) and phosphatidylethanolamine (PE) from normal and tumour tissue of rat mammary gland. The incidence of mammary tumours was significantly lower in fish oil- (29%), compared with olive oil- (75%; P < 0.04) but not maize oil- (63%; P < 0.1) fed animals. No differences in PI fatty acid composition were found in normal or tumour tissue between rats fed on maize oil, olive oil or fish oil in diets from weaning. When normal and tumour tissue PI fatty acids were compared, significantly higher amounts of stearic acid (18:O) were found in tumour than normal tissue in rats given olive oil (P < 0.05). A similar trend was found in animals fed on maize oil, although differences between normal and tumour tissue did not reach a level of statistical significance (P < 0.1). In mammary PE, maize oil-fed control animals had significantly higher levels of linoleic acid (18:2n-6) than either olive oil- or fish oil-fed animals (P < 0.05, both cases) and levels of arachidonic acid were also higher in maize oil- compared with fish oil-fed animals (P < 0.05). In tumourbearing animals no differences in PE fatty acid composition were found between the three dietary groups. When normal and tumour tissue PE fatty acids were compared, significantly lower amounts of liuoleic acid (18:2n-6; P < 0.01) and significantly greater amounts of arachidonic acid (20:4n-6; P < 0.05) were found in tumour than normal tissue of rats fed on maize oil. The present study shows that the fatty acid composition of PI from both normal and tumour tissue of the mammary gland is resistant to dietary fatty acid modification. The PE fraction is more susceptible to dietary modification and in this fraction there is evidence of increased conversion of linoleic acid to arachidonic acid in tumour compared with normal tissue. Lower tumour incidence rates in rats given fish oils may in part be due to alteration in prostanoid metabolism secondary to displacement of arachidonic acid by eicosapentaenoic acid, but PE rather than PI would appear to be the most likely locus for diet-induced alteration in prostanoid synthesis in this tissue. Effects of dietary fatty acids other than on the balance of n-6 and n-3 fatty acids, and on prostanoid metabolism, should also be considered. The significance of increased stearic acid content of PI in tumours of olive oil-fed animals and the possible influence of dietary fatty acids on the capacity for stearic acid accumulation requires further study.

Quantification of ruminal Clostridium proteoclasticum by real-time PCR using a molecular beacon approach

Aims: All members of the ruminal Butyrivibrio group convert linoleic acid (cis-9,cis-12-18 : 2) via conjugated 18 : 2 metabolites (mainly cis-9,trans-11-18 : 2, conjugated linoleic acid) to vaccenic acid (trans-11-18 : 1), but only members of a small branch, which includes Clostridium proteoclasticum, of this heterogeneous group further reduce vaccenic acid to stearic acid (18 : 0, SA). The aims of this study were to develop a real-time polymerase chain reaction (PCR) assay that would detect and quantify these key SA producers and to use this method to detect diet-associated changes in their populations in ruminal digesta of lactating cows. Materials and Results: The use of primers targeting the 16S rRNA gene of Cl. proteoclasticum was not sufficiently specific when only binding dyes were used for detection in real-time PCR. Their sequences were too similar to some nonproducing strains. A molecular beacon probe was designed specifically to detect and quantify the 16S rRNA genes of the Cl. proteoclasticum subgroup. The probe was characterized by its melting curve and validated using five SA-producing and ten nonproducing Butyrivibrio-like strains and 13 other common ruminal bacteria. Analysis of ruminal digesta collected from dairy cows fed different proportions of starch and fibre indicated a Cl. proteoclasticum population of 2-9% of the eubacterial community. The influence of diet on numbers of these bacteria was less than variations between individual cows. Conclusion: A molecular beacon approach in qPCR enables the detection of Cl. proteoclasticum in ruminal digesta. Their numbers are highly variable between individual animals. Signifance and Impact of the Study: SA producers are fundamental to the flow of polyunsaturated fatty acid and vaccenic acid from the rumen. The method described here enabled preliminary information to be obtained about the size of this population. Further application of the method to digesta samples from cows fed diets of more variable composition should enable us to understand how to control these bacteria in order to enhance the nutritional characteristics of ruminant-derived foods, including milk and beef.

Protein-lipid interactions at the air/water interface

Surface pressure measurements and external reflection FTIR spectroscopy have been used to probe protein-lipid interactions at the air/water interface. Spread monomolecular layers of stearic acid and phosphocholine were prepared and held at different compressed phase states prior to the introduction of protein to the buffered subphase. Contrasting interfacial behaviour of the proteins, albumin and lysozyme, was observed and revealed the role of both electrostatic and hydrophobic interactions in protein adsorption. The rate of adsorption of lysozyme to the air/water interface increased dramatically in the presence of stearic acid, due to strong electrostatic interactions between the negatively charged stearic acid head group and lysozyme, whose net charge at pH 7 is positive. Introduction of albumin to the subphase resulted in solubilisation of the stearic acid via the formation of an albumin-stearic acid complex and subsequent adsorption of albumin. This observation held for both human and bovine serum albumin. Protein adsorption to a PC layer held at low surface pressure revealed adsorption rates similar to adsorption to the bare air/water interface and suggested very little interaction between the protein and the lipid. For PC layers in their compressed phase state some adsorption of protein occurred after long adsorption times. Structural changes of both lysozyme and albumin were observed during adsorption, but these were dramatically reduced in the presence of a lipid layer compared to that of adsorption to the pure air/water interface.

Differences in cell morphology, lipid and apo B secretory capacity in caco-2 cells following long term treatment with saturated and monounsaturated fatty acids

The suitability of the caco-2 cell line as a model for studying the long term impact of dietary fatty acids on intestinal lipid handling and chylomicron production was examined. Chronic supplementation of caco-2 cells with palmitic acid (PA) resulted in a lower triacylglycerol secretion than oleic acid (OA). This was coupled with a detrimental effect of PA, but not OA, on transepithelial electrical resistance (TER) measurements, suggesting a loss of structural integrity across the cell monolayer. Addition of OA reversed the adverse effects of PA and stearic acid on TER and increased the ability of cells to synthesise and accumulate lipid, but did not normalise the secretion of lipids by caco-2 cells. Increasing amounts of OA and decreasing amounts of PA in the incubation media markedly improved the ability of cells to synthesise apolipoprotein B and secrete lipids. Real time RT-PCR revealed a down regulation of genes involved in lipoprotein synthesis following PA than OA. Electron microscopy showed adverse effects of PA on cellular morphology consistent with immature enterocytes such as stunted microvilli and poor tight junction formation. In conclusion, previously reported differences in lipoprotein secretion by caco-2 cells supplemented with saturated fatty acids (SFA) and OA may partly reflect early cytotoxic effects of SFA on cellular integrity and function. (C) 2007 Elsevier B.V. All rights reserved.

Chronic disease risk associated with different dietary saturated fatty acids

This chapter compares the risks of chronic disease, and cardiovascular disease in particular, associated with consumption of different saturated fatty acids. Emphasis is placed on the effects of stearic acid as this has potential to replace trans fatty acids in certain manufactured food products. The chapter first reviews the effects of individual saturated fatty acids on blood lipids, including cholesterol, as these are commonly used as markers of disease risk. It then looks directly at evidence in relation to health outcomes. Finally, recent evidence specifically on the effect of stearic acid relative to other fatty acids, including trans fatty acids, is summarised.

Effect of TAG composition on performance of low saturate shortenings in puff pastry

Four blends formulated with low saturated fatty acid content, with the saturated component rich in stearic acid, were prepared from shea stearin, interesterified shea stearin, fully hardened soybean oil and high oleic sunflower oil in order to study their performance as shortenings in puff pastry products. The blends had a low saturated fatty acid content (30.1 ± 1.1%) compared to butter (65.9%). Saturates in the four blends examined came mainly from SSS, SOS, SSO and SOO. Puff pastry prepared from the blend that contained SOS as the main source of saturates had better properties than the other blends. It was similar to butter in compressibility of the baked product. The β-polymorphic form was present in all blends, although blends containing the highest levels of SSS also showed some β′ crystals.

Dairy fat: perceptions and realities.

Milk provides many key nutrients but the saturated and trans fatty acids in milk fat are associated with perceived negative effects on human health, especially cardiovascular disease. Recent epidemiological studies and dietary intervention trials challenge this perception, however; available evidence does not support the concept that consumption of saturated fats or dairy products adversely affects the risk of coronary heart disease (although replacing some saturated fats with mono or polyunsaturated fats is likely to provide benefit). Furthermore, the trans fats found in dairy products are consumed in very low amounts and do not appear to have the negative health effects associated with the consumption of industrial sources of trans fat. Milk fat is an excellent source of oleic acid that originates mainly by endogenous synthesis from stearic acid, but increasing the milk fat content of unsaturated fatty acids requires dietary formulations that bypass rumen biohydrogenation. Recent research indicates that long-chain omega-3 fatty acids and conjugated linoleic acids have potential beneficial effects in health maintenance and the prevention of chronic diseases. Enhancing the milk fat content of these fatty acids offers exciting possibilities, but educating consumers about inaccurate and inappropriate generalisations about fat remains the primary challenge. Finally, individuals do not simply consume milk-fat-derived fatty acids on their own, but rather as components in dairy foods which are highly complex and may contain many beneficial ingredients. Overall, dairy products are critical in providing many of the essential nutrients in the human diet. Nevertheless, dairy products vary in their nutrient composition, including fat, and this needs to be considered in the context of dietary recommendations and our need to consume a balanced diet.

Influence of harvest date and crop yield on the fatty acid composition of virgin olive oils from cv. Picual

In this study was analyzed the effect of crop year and harvesting time on the fatty acid composition of cv. Picual virgin olive oil. The study was carried out during the fruit ripening period for three crop seasons. The mean fatty acid composition of Picual oils was determined. The oils contained palmitic acid (11.9%), oleic acid (79.3%), and linoleic acid (2.95%). The content of palmitic acid and saturated fatty acids decreased during fruit ripening while oleic and linoleic acids increased. The amount of stearic and linolenic acids decreased. The amount of saturated acids, palmitic and stearic, and the polyunsaturated acids linoleic and linolenic was dependent on the time of harvest, whereas the amount of oleic acid varied with the crop year. The differences observed between crop years for both palmitic and linoleic acid may be explained by the differences in the temperature during oil biosynthesis and by the amount of summer rainfall for oleic acid content. A significant relationship was observed between the MUFA/PUFA ratio and the oxidative stability measured by the Rancimat method.