Aumento nos níveis circulantes de alguns ácidos graxos (AGs) está relacionado com o quadro de resistência à insulina em músculo esquelético. Os mecanismos pelos quais os AGs diminuem a ação da insulina não estão elucidados, entretanto a participação destas biomoléculas no controle do NF-kB, SREBP-1c, HIF-1<font face=\"Symbol\">&#945;, LXR<font face=\"Symbol\">&#945; e PPARg considerados reguladores do Slc2a4, já foi sugerida. O objetivo deste estudo foi investigar a ação dos ácidos graxos, oléico (OFA) e linoléico (LFA), em células musculares L6, na regulação do Slc2a4. Redução no conteúdo protéico e de mRNA GLUT4 foi verificada na presença de ambos AGs. Esta redução foi relacionada com aumento na expressão e na atividade de ligação do NF-kB e diminuição na expressão e na atividade de ligação do SREBP-1 ao gene Slc2a4, na presença de OFA e LFA. Ambos AGs aumentaram a expressão do mRNA de LXR<font face=\"Symbol\">&#945;, PPARg and HIF-1<font face=\"Symbol\">&#945;, todavia apenas na presença de LFA foi detectada uma diminuição na ligação de PPARg ao Slc2a4. Ambos AGs reduzem a expressão do GLUT4 devido modulação da ligação do NF-kB e SREBP-1 ao gene Slc2a4. / High elevated levels of some free fatty acids (FFAs) are associated with insulin resistance in skeletal muscle. The mechanisms by which FFAs impair this hormone sensitivity need to be clarified; nevertheless, its effects in the modulation of NF-kB, SREBP-1c, HIF-1<font face=\"Symbol\">&#945;, LXR<font face=\"Symbol\">&#945; and PPARg which are related with Slc2a4 gene regulation have been suggested. The goal of this study was to investigate the action of oleic (OFA) and linoleic (LFA) fatty acids, in L6 muscle cells, in Slc2a4 regulation. The GLUT4 protein and mRNA expression decreased in the presence of OFA and LFA. The reduced GLUT4 expression was related to a significative enhancement of the NFkappaB mRNA expression and binding activity in presence of both FFAs and a decrease of SREBP-1 mRNA and binding activity in the Slc2a4. OFA and LFA increase LXR<font face=\"Symbol\">&#945;, PPARg and HIF-1<font face=\"Symbol\">&#945; mRNA expression, but only a reduction in PPARg binding activity was verified in presence of the LFA. A reduction in GLUT4 expression in the presence of OFA and LFA was detected and related with NF-kB and SREBP-1 binding activity in the Slc2a4 gene.

Please read the abstract in the section 00front of this document / Thesis (PhD (Food Science))--University of Pretoria, 2006. / Food Science / unrestricted

Fatty acid desaturase 2 (FAD2) enzymes are phosphatidylcholine desaturases occurring as integral membrane proteins in the endoplasmic reticulum membrane and convert monounsaturated oleic acid into polyunsaturated linoleic acid. The major objective of this research was to study the expression and function of two cotton FAD2 genes (the FAD2-3 and FAD2-4 genes) and their possible role in plant sensitivity to environmental stress, since plants may increase the polyunsaturated phospholipids in membranes under environmental stress events, such as low temperature and osmotic stress. Two FAD2 cDNA clones corresponding to the two FAD2 genes have been isolated from a cotton cDNA library, indicating both genes are truly expressed in cotton. Model yeast cells transformed with two cotton FAD2 genes were used to study the chilling sensitivity, ethanol tolerance, and growth rate of yeast cells. The expression patterns of the two FAD2 genes were analyzed by reverse transcription polymerase chain reactions (RT-PCR) and Western blot analyses in cotton plants under different treatment conditions. The coding regions of both FAD2 genes were inserted downstream from the CaMV 35S promoter in the pMDC gateway binary vector system. Five different FAD2/pMDC constructs were transformed into the Arabidopsis fad2 knockout mutant background, and multiple potential transgenic Arabidopsis plant lines harboring the cotton FAD2 genes were generated. The cotton FAD2 genes were amplified by the polymerase chain reaction (PCR) from the genomic DNAs isolated from the transgenic Arabidopsis T1 plant lines. Complementation of the putative transgenic Arabidopsis plants with the two cotton FAD2 genes was demonstrated by gas chromatography analyses of the fatty acid profiles of leaf tissues. The cellular localization of cotton FAD2-4 polypeptides with N-terminal green fluorescence protein (GFP) was visualized by confocal fluorescence microscopy. The phenotype of transgenic Arabidopsis plants transformed with the cotton FAD2-4 gene was compared to Arabidopsis knockout fad2 mutant plants and wild type Arabidopsis plants regarding their sensitivity to low temperature, and the size and height of the plants.

The lipoxygenase (LOX) pathway plays an important role in the oxidative metabolism of polyunsaturated N-acylethanolamines (PU-NAEs). The LOX pathway functions in conjugation with hydrolysis by fatty acid amide hydrolase (FAAH) and to produce oxidized NAEs during seed germination and early seedling development. When Arabidopsis seedlings were grown in low micromolar concentrations of lauroylethanolamide (NAE 12:0), growth retardation and elevated endogenous PU-NAE levels were observed due to the competitive inhibition of LOX by NAE 12:0. The elevated levels of endogenous PU-NAEs were more pronounced in genotypes with reduced NAE hydrolase capacity (faah knockouts), and less evident with overexpression of FAAH. Alterations in PU-NAE metabolism were studied in seedlings of various lox and FAAH mutants. The partitioning of PU-NAEs into oxylipin metabolites was exaggerated in the presence of exogenous linolenoylethanolamide (NAE18:3) and resulted in bleaching of cotyledons. The bleaching phenotype was restricted to a narrow developmental window (3-to-5 days after sowing), and was attributed to a reversible disruption of thylakoid membranes in chloroplasts. Biochemical and genetic evidence suggested that 9-hydro(pero)xy and 13-hydro(pero)xy octadecatrienoylethanolamides (9- and 13-NAE-H(P)OT), but not their corresponding hydro(pero)xy free fatty acids, induced cotyledon bleaching. The LOX-mediated metabolites of NAE18:3 shared some overlapping effects on seedling development with those of linoleoylethanolamide (NAE18:2) such as a reduction in seedling root growth. On the other hand, NAE18:3 oxylipin metabolites also exhibited distinct effects during seedling development such as the inhibition of photomorphogenesis. Biochemical and genetic evidence indicated that a LOX-mediated metabolite of NAE18:2, 9-hydro(pero)xy octadecadienoylethanolamide (9-NAE-H(P)OD), acted as a potent negative regulator of seedling root development, and this depended on an intact abscisic acid (ABA) signaling pathway. Synergistic inhibition of root elongation between 9-NAE-H(P)OD and ABA was restricted to a narrow developmental window (3-to-5 d after sowing) of seedling development. Genetic evidence with Arabidopsis mutants in ABA synthesis (aba1, aba2), perception (pyr1, pyl2, pyl4, pyl5, pyl8) and transcriptional regulation (abi3-1) suggested that negative regulation of growth by 9-NAE-H(P)OD likely was mediated through an increase in ABA synthesis, and this was confirmed biochemically. Induction of a secondary dormancy program in Arabidopsis seedlings by environmental stresses also requires an intact ABA signaling cascade, and our study has shown that this regulatory seedling program is dependent, in large part, on NAE oxylipin formation. Together, results presented here indicated that LOX-mediated metabolites of NAE18:3 and NAE18:2 in Arabidopsis represent a newly-discovered group of bioactive metabolites, and their accumulation during the embryo-to-seedling transition of plant development may act to synchronize seedling establishment with environmental cues.

Plasma membranes are essential to both the structure and function of mammalian cells. The first unifying paradigm of membrane structure, the Fluid Mosaic Model, is no longer considered adequate to describe the many non-homogeneous lipid structures that have been observed in both natural and model membranes over the past approximately thirty years. The field of membrane biophysics now appreciates that the complex mixture of different lipid species found in natural membranes produces a range of dynamic, laterally segregated, non-homogeneous structures which exist on time scales ranging from microseconds to minutes. When sphingomyelin (SM), POPC and cholesterol are all present in a bilayer there is wide range of compositional ratios where the bilayer consists of a coexistence between two fluid phases designated liquid ordered (lo) and liquid disordered (ld). The lo phase is cholesterol-rich phase characterized by relatively high molecular order and slow rotational and translational motion, while the ld phase generally has low molecular order and relatively rapid rotational and translational motion. The driving force for the formation of these two phases is the ability of cholesterol to form favorable van der Waals contacts with the two saturated acyl chains on PSM and the one saturated acyl chain on POPC. The ternary system is an important model system for examining the physical properties and functional implications of co-existing lo and ld phases. However, it does not include one of the most significant compositional variables found in many important mammalian membranes. Membranes in the nervous system contain high concentrations of the highly polyunsaturated fatty acid docosahexaenoic acid (DHA), which contains 22 carbons and 6 double bonds. A wide range of experimental evidence shows that DHA-containing phospholipids are important for optimal performance of a number of membrane signaling systems and membrane protein functions. The goal of this study is to determine how addition of a DHA-containing phospholipid, PDPC alters the biologically important lo and ld co-existence region.

Les bifidobactéries sont exposées à de nombreux stress, liés aux conditions environnementales rencontrées lors de la production, du stockage au froid, et pendant la digestion des laits fermentés. Afin d'améliorer leur survie, cette étude vise la compréhension des mécanismes de dégradation de l'état physiologique de différentes souches de Bifidobacterium soumises aux stress froid et acide et au stress gastro-intestinal simulé in vitro. Elle ambitionne également d'établir des relations entre la résistance aux différents stress et la teneur en acides gras membranaires et des laits biologiques et conventionnels. Les résultats montrent que l'activité acidifiante des bifidobactéries est souche-dépendante et qu'elle augmente lorsque les bactéries sont associées aux bactéries lactiques du yaourt, avec du lait biologique et lorsque la température d'incubation est fixée à 42°C au lieu de 37°C. La cultivabilité et la survie des souches ont été déterminées après fermentation, après stockage à 4°C pendant 7 à 28 jours, et pendant un processus de digestion simulé in-vitro dans un digesteur dynamique reproduisant le tractus gastro-intestinal. Ces caractéristiques sont améliorées dans les laits fermentés biologiques par rapport aux produits conventionnels, lorsque la fermentation est effectuée à 42°C jusqu'à pH 4,4, et lorsque les laits fermentés sont maintenus à 28°C pendant 12 heures avant d'être refroidi à 4°C. Ces procédures de fabrication spécifiques génèrent ainsi une adaptation physiologique des bifidobactéries aux stress. Pendant la digestion in-vitro, la cultivabilité des bifidobactéries se dégrade moins lorsque la fermentation se déroule en lait biologique plutôt qu'en lait conventionnel et, dans une moindre mesure, lorsque les procédures d'adaptation sont appliquées pendant la fabrication du lait fermenté. Ces résultats sont liés aux teneurs plus élevées en acides gras insaturés, en particulier en acides trans-vaccénique, linoléique conjugué et α-linolénique, qui caractérisent les produits biologiques. Ces profils d'acides gras particuliers aux laits biologiques permettent aux bifidobactéries de modifier leur composition en acides gras membranaires, en augmentant leur teneur en acides gras insaturés et en raccourcissant la longueur moyenne des chaînes d'acides gras saturés, adaptant ainsi leur fluidité membranaire. Lorsque les procédures de fabrication spécifiques sont mises en oeuvre pour induire une adaptation physiologique des bifidobactéries, la composition en acides gras des membranes se modifie différemment de ce qui est observé en lait biologique. Cette différence indique ainsi que d'autres mécanismes biologiques d'adaptation sont probablement impliqués, en particulier au niveau protéomique. Finalement, cette étude démontre que les modifications au niveau de la membrane contribuent à moduler la résistance aux stress technologique et gastro-intestinal de souches de Bifidobacterium. / Bifidobacteria are exposed to various stress, as a result of environmental conditions encountered during fermented milk production, cold storage and during digestion of the products inside gastrointestinal tract. In order to improve their survival, this study aimed at understanding the degradation mechanisms of the physiological state of various Bifidobacterium strains when exposed to cold, acid and in vitro simulated gastrointestinal stress. It also intended to establish relationships between stress resistance and milk and membrane fatty acids contents, in organic and conventional milks. The results showed that acidification activity of bifidobacteria was strain-dependent and increased when bifidobacteria were associated to yogurt cultures, when organic milk was used and when incubation temperature was set at 42°C instead of 37°C. Cultivability and survival of the Bifidobacterium strains were determined after fermentation, after storage at 4°C for 7 to 28 days, and during in-vitro digestion that was simulated in a dynamic gastrointestinal tract model. These characteristics were improved in organic fermented milks as compared to conventional products, when fermentation was performed at 42°C until pH 4.4, and when the fermented milks were kept at 28°C for 12 hours before being cooled to 4°C. These specific manufacture procedures thus generated physiological adaptation of the bifidobacteria to the stress. During in-vitro digestion, cultivability of bifidobacteria was less deteriorated when they were grown in organic instead of conventional milk, and to a less extent, when the adaptation procedures were applied during fermented milk manufacture. These results were related to the higher unsaturated fatty acids content, including trans-vaccenic, conjugated linoleic and α-linoleic acids that characterize organic products. These particular fatty acids profiles of organic milks allowed bifidobacteria to modify their membrane fatty acids composition, by increasing their unsaturated fatty acids contents and by shortening the length of medium chain saturated fatty acids, thus adapting their membrane fluidity. When specific manufacture procedures were carried out to trigger physiological adaptation of the bifidobacteria, membrane fatty acid composition changed différently from what is observed in organic milk. This difference indicates that other biological adaptation mechanisms are probably involved, especially at the proteomic level. Finally, this study demonstrated that modifications at membrane level contribute to modulate resistance against technology and gastro-intestinal stress of Bifidobacterium strains to better withstand technological and gastro-intestinal stress.

O objetivo deste estudo foi avaliar os efeitos do ômega-3 (&omega;-3) e do ômega-6 (&omega;-6) do leite e sua importância na nutrição e saúde humana, sobre as características reprodutivas e produtivas, sobre o hemograma, leucograma e perfil de ácidos graxos (AG) do sangue das matrizes e dos leitões, visando a relação ideal de 1:1 a 5:1 (&omega;-6/ &omega;-3). O trabalho foi realizado com 30 matrizes, Large White x Landrace, divididas nos tratamentos controle (TC), leite enriquecido naturalmente com &omega;-3 (TW3) e leite enriquecido naturalmente com &omega;-6 (TW6), que receberam esta suplementação desde 34 dias de idade até o desmame dos seus leitões. Foi avaliado o desempenho das matrizes e dos leitões, e colhidas amostras de sangue, colostro e leite, para análises laboratoriais. Os resultados mostraram uma melhora no desempenho das matrizes, associado à fase de desenvolvimento em que se encontravam. Dos 34 aos 190 dias, a concentração absoluta de leucócitos foi maior (P=0,049) para TW3 e TW6, mostrando uma alteração no sistema imune das matrizes. No 1º dia de lactação (D1L) houve uma redução (P&lt;0,05) de ureia e VLDL e aumento (P&lt;0,05) de proteína total e LDL para TC. Para as matrizes de 34 a 190 dias de idade e no D1L, o perfil de AG foi alterado (P&lt;0,05) e para os leitões aos 0 e 14 dias de idade, houve maior (P=0,01) concentração de EPA e menor (P=0,045) proporção de AA/EPA para TW3. Os leitões do TW3 e TW6 apresentaram maior peso médio (P=0,004) em relação a TC, pela possível alteração da razão &omega;-6/&omega;-3 no organismo, que ocorreu por meio da suplementação das mães. A conclusão do presente estudo é que o fornecimento do leite enriquecido com uma melhor razão &omega;-6/&omega;-3, pode induzir alterações metabólicas, com base nos parâmetros sanguíneos analisados, assim como na produtividade dos leitões, afetando seu peso médio. / The objective of this study was to evaluate the effects of omega-3 (&omega;-3) and omega-6 (&omega;-6) on milk and its importance in nutrition and human health. the reproductive and productive characteristics, on the hemogram, leukogram and fatty acid profile of the blood of the dams and piglets, aiming at the ideal ratio of 1: 1 to 5: 1 (&omega;-6 / &omega;-3). The work was performed with 30 matrices, Large White x Landrace, divided in control treatments (TC), milk enriched naturally with &omega;-3 (TW3) and milk enriched naturally with &omega;-6 (TW6), which received this supplementation from 34 days to the weaning of their piglets. The performance of the dams and piglets was evaluated, and samples of blood, colostrum and milk were collected for laboratory analysis. The results showed an improvement in the performance of the matrices, associated to the development phase in which they were. From 34 to 190 days, the absolute leukocyte concentration was higher (P = 0.049) for TW3 and TW6, showing a change in the immune system of the matrices. On the 1st day of lactation (D1L) there was a reduction (P &lt;0.05) of urea and VLDL and increase (P &lt;0.05) of total protein and LDL for CT. For the matings of 34 to 190 days of age and in D1L, the fatty acid profile was altered (P &lt;0.05) and for the piglets at 0 and 14 days of age, there was a higher (P = 0.01) concentration of EPA and lower (P = 0.045) ratio of AA / EPA to TW3. The TW3 and TW6 piglets had a higher mean weight (P = 0.004) compared to CT, due to the possible alteration of the &omega;-6 / &omega;-3 ratio in the organism, which occurred through the mothers\' supplementation. The conclusion of the present study is that the supply of enriched milk with a better &omega;-6 / &omega;-3 ratio may induce metabolic alterations, based on the blood parameters analyzed, as well as the piglet productivity, affecting its average weight.

University of Macau / Institute of Chinese Medical Sciences

No description available.

The study was designed to develop a nutraceutical product from by-products of fish processing. Fish oil was extracted from mackerel tissues (skin, viscera and muscle) using hexane-isopropanol (3:2 v/v) and chloroform-methanol (2:1 v/v) solvent systems. An oxidative study was carried out to determine the best processing and storage conditions to minimize autooxidation of mackerel oil. Urea complexation was used to concentrate polyunsaturated fatty acids (PUFA) from the extracted fish oil. The urea complexation process was optimized to determine the best reaction conditions for high yield of the omega-3 fatty acids in particular, and total PUFA. / Conjugated linoleic acid (CLA) was synthesized from four vegetable oils (sunflower, canola, soybean and corn) by alkaline isomerization. The CLA isomers and PUFA concentrated from mackerel tissues were used to synthesize triglycerides by lipase-catalyzed esterification. The effect of temperature, reaction medium, enzyme, moisture removal system and glycerol to fatty acid ratio on extent of synthesis were investigated. The synthesis process was also optimized using central composite design to determine the best conditions for high synthesis yield. The fatty acid composition and positional analyses were determined by GC-FID and electrospray ionization mass spectrometry (EI-MS) / The results showed that mackerel skins were most suitable for concentrating PUFA. The eicosapentaenoic acid (EPA) and -docosahexaenoic acid (DHA) content of fish oil from the tissues examined ranged between 6.3--9.7 (wt%) with an iodine value of 134 +/- 5.0. The baseline total PUFA content was increased from ca 21.0 to ca 83.0 (wt%) with an associated iodine value of 296 +/- 7.0 using urea complexation. Low (50 ppm and 100 ppm) levels of alpha-tocopherol in combination with low storage temperature (-40°C) showed lowest oxidation after 66 days of storage. High levels (250 and 500 ppm) of alpha-tocopherol were observed to be prooxidant based on TBARS, peroxide and conjugated diene measurements. Urea to fatty acid ratio and temperature were predominant effectors influencing the amounts of individual omega-3 fatty acids and total PUFA concentrated by urea complexation. The model developed for the optimized urea complexation process were capable of predicting the yields of EPA, DHA, total PUFA and Iodine values to a high degree of accuracy at R2 = 0.87, 0.96, 0.95, and 0.92 respectively. / Sunflower oil was most suitable for synthesizing conjugated linoleic acid by alkaline isomerization, compared with soybean, canola and corn oil. Two CLA isomers, c9,t11 and t10, c12 were most abundant and occurred in approximately equal proportions irrespective of vegetable oil used. Total CLA synthesized from sunflower oil was 93.5 +/- 3.5 (wt%) with the two major isomers making up 89 +/- 3.5 (wt%). Candida antartica lipase showed more synthesis activity than Mucor meihie in both organic and solvent-free systems. Analysis of isolated synthesized triglycerides by GC-FID and mass spectrometry showed that DHA, EPA, CLA and linolenic acid were the main fatty acids incorporated into the triglycerides. DHA and EPA were mostly esterified at the sn-2 position.