A study was performed to investigate the effect of diets rich in polyunsaturated fatty acids and other related nutrients, in the effort to improve skin and hair coat conditions in canines. The study included 24 dogs fed a baseline diet (Ol’Roy®), with an acclimation period of 12 weeks (Phase I). Nine female beagles and 15 male hound mixed-bred dogs were used. For the next 12 weeks (Phase II) the dogs were randomly divided into three groups and fed one of three specially formulated complete and balanced diets. Differences among the three diets were as follows: Diet A contained lower but adequate amounts of dietary zinc and linoleic acid than Diet B. Diet C was similar to Diet B with respect to zinc but contained less linoleic acid and more alpha-linolenic acid. In the beginning, a preliminary study assessing skin lipids showed that hair is a suitable material to analyze. This study lead to the subsequent investigation, using plucked and shed hair samples obtained on weeks 3, 7, and 11 (Phase I) and again on weeks 1, 3, 7, and 11 (Phase II). One objective of this research was to investigate the dietary effects of the combination of linoleic acid in combination with zinc on canine sebum lipids during a 12 week diet phase. Another objective was to investigate if any correlation existed between the hair cycle anagen and telogen phase of the hair follicle and the lipid constituents present in the sebum during the 12 week acclimation phase and the 12 week feeding trial. Two hypotheses were tested. First, the lipid content of hair can be altered by fat and fatty acid composition of the diet. Second, the modification of dietary fat and polyunsaturated fatty acids lead to changes in hair lipids that may be related to improved skin and hair coat scores. Over the 12 week feeding period of Phase II, test results revealed statistically significant increases in both CE and CE/WD, and a prolonged growth of hair follicles in the anagen phase of diet B dogs, which are both consistent with improved skin and hair coat scores.

To examine how dietary n-3 polyunsaturated fatty acids affect Th2 cell development, female C57BL/6 mice were fed a washout corn oil (CO) diet for 1 wk followed by 2 wk of either the same CO diet or a fish oil (FO) diet. CD4+ T cells were isolated from spleens and cultured under both neutral (anti-CD3 and phorbol myristate acetate (PMA)) and Th2 polarizing conditions (anti-CD3 and PMA, in presence of rIL-4, rIL-2, and anti-IFN-γ) in the presence of homologous mouse serum (HMS) or fetal bovine serum (FBS) for 2 d. Dietary n-3 PUFA significantly enhanced Th2 cell development and suppressed Th1 development under neutral conditions as assessed by intracellular cytokine staining for IL-4 and IFN-γ as the two prototypic Th2 and Th1 cytokines, respectively. However, under Th2 polarizing conditions, while the suppression of Th1 cells was maintained in FO-fed mice, no dietary effect was observed in Th2 cells. Dietary FO increased the Th2/Th1 ratio under both neutral and Th2 polarizing conditions with HMS in the cultures. To examine the effect of dietary n-3 PUFA on Th1 development, DO11.10 Rag2-/- mice expressing transgenic T cell receptor specific for ovalbumin (OVA) peptide were used. CD4+ T cells were isolated from spleens and lymph nodes and stimulated with ovalbumin (OVA) peptide and irradiated BALB/c splenocytes in the presence of rIL-12, anti-IL-4, and rIL-2 in HMS for 2d. Cells were expanded for another 3 d in the presence of rIL-2 and rIL-12. Dietary n-3 PUFA did not affect Th1 differentiation as assessed by the proportion of IFN-γ+, IL-4- T cells in the cultures, but suppressed rIL-2 induced expansion. The suppressed expansion was due to suppressed proliferation (p<0.05). In vivo expansion of antigen-specific T cells was visualized by flow cytometric analysis of CFSE-positive transgenic T cells. Dietary n-3 PUFA did not appear to affect antigen-induced CD4+ T cell cycle progression in vivo. Overall, these results suggest dietary n-3 PUFA have no direct effect on Th2 cell development but do directly suppress Th1 cell development following both mitogenic and antigenic stimulation in vitro.

This Thesis investigates the effect of membrane-active agents, such as synthetic ether lipids (SEL), local anaesthetics and polyunsaturated fatty acids (PUFAs) on human leukaemia cells. The two cell lines used were human acute myeloblastic leukaemia (HL60) cells and human myelogenous leukaemia (K562) cells. SEL, local anaesthetics and PUFAs were found to be cytotoxic to both cell lines at certain concentrations. The SEL ET-18-OCH(_3) was found to be cytotoxic to both cell lines but the HL60 cells were found to be the more sensitive cell line. HL60 cells were found to be so sensitive to the action of the local anaesthetic dibucaine that a subtoxic concentration that killed ≤10% was not determined. However, in K562 cells the combination of a subtoxic dibucaine concentration together with a range of ET-I8-OCH(_3) concentrations increased the cytotoxicity over that of ether lipid alone. PUFAs were shown to incorporate into plasma membrane phospholipids at concentrations as low as 1 μM after an incubation of 48 hours. PUFAs were shown to be cytotoxic, but the addition of vitamin E reduced the cytotoxicity of arachidonic acid, eicosapentaenoic acid and docosahexaenoic acid in HL60 cells, and of docosahexaenoic acid in K562 cells. This implied that lipid peroxidation was involved in PUFA cytotoxicity. This was, however, not confirmed. PUFA in combination with ET-I8-OCH3 resulted in a slight decrease in cytotoxicity. PUFA combined with dibucaine did not alter cytotoxicity. Cells were also treated with a combination of PUFA and 1-β-D- arabinofliranosylcytosine (ara-C), which is an agent known to induce cell differentiation. Onset of differentiation was determined by following haemoglobin accumulation in K562 cells. PUFA on their own were found to promote accumulation of haemoglobin. The greatest accumulation of haemoglobin was observed with K562 cells treated with PUFA and ara-C.

Young and ovariectomized animals eating diets rich in omega-3 polyunsaturated fatty acids (n-3 PUFAs) exhibit enhanced bone formation and decrease bone loss, respectively. Eicosapentaenoic acid, an n-3 PUFA found in fish oil, competes with arachidonic acid, an n-6 PUFA, for the cyclooxygenase enzyme, modulating prostaglandin E2, a mediator of bone mechanotransduction. Whether this diet affects bone gains during exercise is not well defined. We hypothesized rats consuming a high n-3 PUFA diet would gain more bone mass with increased bone formation compared to the rats consuming a high n-6 PUFA diet in response to exercise. Virgin Sprague-Dawley rats (5-mo-old, n=18) were assigned to one of two groups: diet rich in corn oil with a n-6:n-3 dietary ratio of 23:1 (O6) or a diet rich in fish oil with an n-6:n-3 dietary ratio of 2:1 (O3). After acclimation, rats completed 9 sessions on alternate days of stimulated muscle contractions at 75% peak isometric strength. Structural and densitometric properties of proximal tibia were measured using in vivo peripheral quantitative CT. Bone formation rate was quantified on the periosteal the surface by standard bone histomorphometry after intraperitoneal injections of calcein. There was a significant main effect due to diet on total volumetric bone mineral density. The diet rich in n-3 PUFAs also allowed for increases in cancellous volumetric bone mineral density at the proximal tibia independent from exercise, as high as 28%. However, proximal tibia metaphysis bone size and shape was not modified due to changes in diet. The training protocol resulted in a robust increase in bone formation, mass, and area at the midshaft tibia. Mineral apposition rate and bone formation rate were significantly greater in the O3 group compared to the O6 group with exercise at the midshaft tibia, ~36% and ~38% respectively. However, the greater bone formation seen in the O3 groups did not translate over to significantly greater bone mass and size as noted by the pQCT results at the same bone site, because there were no detectable differences between groups. In summary, our data demonstrate that a diet high in n-3 PUFAs independently increases bone density at the proximal tibia. In addition, there was enhanced BFR due to a diet high in n-3 PUFAs with exercise, but those increases did not translate over to increased cortical bone mass or size. These data provide evidence that fish oil consumption with and without simulated resistance training exercise can be beneficial to bone outcomes.

Various innovative separation methods in chromatography have been proposed. The interaction of the 4-t-butylphenyl group with b-cyclodextrin is well-known; compounds tagged with the 4-t-butylphenyl group are separated from untagged compounds using b-cyclodextrin column. In this study, increasing the chain length of tagged molecules does not increase the retention time but depends on other functional groups present, while increasing the number of tags in a molecule increases retention time. The t-butyl group was also compared to adamantyl and lithocholic acid tags. In-house b-cyclodextrin columns were synthesized to observe the effect of the linkage of the cyclodextrin molecule to the support. Furthermore, tagged products could be separated from the starting materials using in-house columns employing flash b-cyclodextrin. Supported room temperature ionic liquids (SILs/MSILs) were explored for the extraction of polyunsaturated triacylglycerols (PUTAGs).Various room temperature ionic liquids (RTILs) with silver salts were tested for extraction effectiveness.The most hydrophobic RTIL [Hmim][PF6] in this study, with dissolved AgBF4 proved to be the best combination for effective extraction. RTILs supported on silica gel and mesoporous SBA-15 reacted with silver salts (psorbents) were synthesized and characterized. The later support revealed a conserved mesopore structure by SEM, TEM, SAXS and N2 isotherms. Trilinolenin (tri-18:3) quantification was explored by conversion to polyunsaturated fatty acid alcohols and methyl esters, Proton NMR spectroscopy and by non-aqueous reverse phase (NARP)-HPLC with evaporative light scattering detector (ELSD). Tri-18:3 and its free fatty acid (FFA-18:3) were extracted selectively and quantified using a two-step methodology. The extraction of a-tocopherol with RTILS modified with organic anions was also successfully demonstrated. Finally, studies of chiral stationary phases with on-resin cyclic oligoprolines were demonstrated. Cross linking of linear oligoprolines was shown to disrupt the poly-proline helix (II) thus leading to lower number of analytes resolved. On-resin cyclic oligoproline synthesis was successful for penta- and tetraproline but failed for the triproline. PyBOP/HOBt/DIPEA was employed for effective cyclization. The on-resin cyclic oligoproline CSP showed a lower number of resolved analytes, implying the importance of the poly-proline helix for chiral selectivity. The innovative separations in this study serve as starting points for developing mature separation methods.

The objective of two studies was to manipulate the essential fatty acid content of turkey semen by enhancing the dietary levels of either n-3 polyunsaturated fatty acids (PUFA) or n-6 PUFA and determine the effect on fertility. In 1999 (Trial 1), and again in 2000 (Trial 2), Large White tom turkeys, 37 weeks of age, were fed one of three diets substituted with chicken fat, soybean oil, or menhaden fish oil. Chicken fat provided the industry's standard ratio of n-6 to n-3, soybean oil provided a greater ratio of n-6 to n-3, and fish oil provided a lower ratio of n-6 to n-3. Contemporary hens were inseminated weekly with semen collected from each group of toms. The effects of dietary lipids on tom body weights, fertility, motility, perivitelline layer sperm penetration percent, and live vs. dead sperm were analyzed. Whereas body weight increased linearly from 31 to 56 weeks of age (WOA), there was no effect of dietary treatment. As measured by the Accudenz® procedure, there were differences in sperm motility due to dietary treatment during 48 and 51 WOA during Trial 1. During Trial 2, sperm motility differences were observed at 53 WOA with the soybean oil-treated toms having the largest absorbance reading and the chicken fat-treated toms having the largest absorbance reading during 56 WOA. The live vs. dead sperm cells during Trial 1 revealed differences among the toms prior to treatment and post treatment. No dietary effects on percent live vs. dead sperm cells were observed during Trial 2. Once per mo, eggs were collected for a one-week period to analyze for sperm penetration of the perivitelline layer. In Trial 1, sperm from toms fed chicken fat produced more penetrations (holes) during 36, 48, and 52 WOA. In Trial 2, sperm penetration values were lower for toms fed fish oil during 42, 47, and 51 WOA. Whereas there were significant differences in fertility, hatch of total eggs, and hatch of fertile eggs among treatments in Trial 1, a bacterial contamination on the farm during weeks seven through fourteen may have contributed to these findings. No significant differences due to treatment were found in these parameters during the second study. The fatty acid analysis of spermatozoa collected at the conclusion of Trial 2 revealed significant differences in total n-3 and total n-6 content, leading to significant differences in the ratio of total n-6 to total n-3. The mixed results indicated the fertilizing ability of domesticated turkey spermatozoa may not be affected by the n-6 to n-3 ratio in the diet of the tom. / Master of Science

The present research examined the hypothesis that omega-3 polyunsaturated fatty acids would increase seizure threshold in rats in vivo, and reduce neuronal excitability in mouse hippocampal slices. Seizure thresholds were measured in rats using the maximal pentylenetetrazol and electrical stimulation seizure tests following α-linolenic acid (ALA) or docosahexaenoic acid administration. ALA raised seizure threshold in the maximal PTZ seizure test, but this effect probably occurred because ALA displaced DHA from liver to the brain. DHA itself was therefore tested in the PTZ and electrical stimulation seizure tests. Direct administration of DHA by subcutaneous injection raised seizure thresholds in the PTZ seizure test, which models tonic-clonic attacks in humans. Dietary enrichment with DHA raised afterdischarge seizure thresholds in the cortex and amygdala, which model simplex and complex partial seizures in humans, although this effect took some time to occur. In vitro, the application of DHA also reduced the incidence of excitatory sharp waves in mouse hippocampal slices. This effect did not appear to be due to either an increase in GABAergic inhibitory tone, nor to a decrease in glutamatergic drive. The fatty acid composition of phospholipids and unesterified fatty acids were measured in the brain following microwave fixation in order to determine whether the effects of DHA on seizure thresholds were due to its de-esterification from the phospholipid membrane. The assay surprisingly revealed that subcutaneous administration of DHA at a dose that raised seizure threshold, increased unesterified arachidonic acid, but not unesterified DHA concentrations during seizures. The results of these studies support the hypothesis that DHA raises seizure threshold in rats, and reduces neuronal excitability in vitro. The effects of DHA on seizure threshold are possibly mediated by the de-esterification of arachidonic acid, which is known to have effects on the voltage-dependent sodium channel.

Conselho Nacional de Desenvolvimento CientÃfico e TecnolÃgico / A reparaÃÃo tecidual consiste em uma importante caracterÃstica de prevenÃÃo do processo inflamatÃrio; alguns fatores podem reduzir, retardar ou impedir este processo. Existe um interesse constante em relaÃÃo ao uso de correntes de baixa intensidade aliado da suplementaÃÃo com nutracÃuticos (alta relaÃÃo &#969;9:&#969;6 e baixa relaÃÃo &#969;6:&#969;3). O objetivo desse trabalho foi avaliar os efeitos do prÃ-condicionamento com misturas de Ãleos contendo &#969;3/&#969;6 e &#969;9/&#969;6 e correntes de baixa intensidade MENS( Microcurrent ElÃtric Neuromuscular Stimulation) sobre a reparaÃÃo tecidual apÃs incisÃes em pele de ratos. Foram utilizados 108 ratos Wistar, distribuidos em 3 grupos: G-1 controle (soro fisiolÃgico) n= 36 subdivididos em dois subgrupos: nÃo estimulado e estimulado; G-2 controle neutro (Ãleo de milho e soja) n=36, subdivididos em dois subgrupos: nÃo estimulado e estimulado e G-3 ALA (Ãleo de oliva+linhaÃa+cÃnola), n=36 subdivididos em dois subgrupos: nÃo estimulado e estimulado. As misturas de Ãleos foram administradas por via orogÃstrica, e a aplicaÃÃo da corrente elÃtrica, ocorreram 1 hora antes da incisÃo cirÃrgica. Os animais foram sacrificados no sÃtimo, dÃcimo quarto e vigÃsimo primeiro dia. Amostra de pele de 1cm/1cm foram coletadas entre o segundo e terceiro ponto da sutura. As amostras foram analisadas a partir de lÃminas histopatolÃgicas coradas com hematoxilina-eosina (HE) e forneceram a anÃlise geral dos cortes de tecido com espessura de 5Âm. Os dados obtidos pela tÃcnica de (HE) foram classificados de acordo com a intensidade inflamatÃria e transformados em variÃveis quantitativas mediante atribuiÃÃo de Ãndice para os achado histolÃgicos segundo (VIDINSK, 2006). Os dados estatÃsticos foram analisados utilizando-se o programa graphpad prisma for Windows, versÃo 6,0 para avaliaÃÃo conjunta dos efeitos dos grupos G-1,.G-2 e G-3, e dos subgrupos nÃo estimulado e estimulado, e dos dias (7Â, 14Â e 21Â). As variÃveis da avaliaÃÃo histolÃgica foram analisadas pelo teste nÃo paramÃtrico de Mann-Whitney e pÃs-teste de Dunn. O nÃvel de significÃncia foi de 5% (p<0,05). Os resultados obtidos mostram que a eletroestimulaÃÃo prÃ-condicionante promoveu elevaÃÃo do exsudato e reduÃÃo da reepitelizaÃÃo no 7Â dia. O uso dos prÃ-condicionantes Ãleo 1(milho e soja) e 2 (oliva+canÃla+linhaÃa) reduziram a vascularizaÃÃo no 7Â e 14Â dias respectivamente.Ambos os grupos dos Ãleos 1 e 2 induziram a reduÃÃo da fibrose no 14Â dia. O prÃcondicionamento com Ãleo1 levou a elevaÃÃo da expressÃo do NFkB no 7Â dia. Conclui-se que a eletroestimulaÃÃo promoveu elevaÃÃo de marcadores da fase inflamatÃria com consequente aceleraÃÃo desse processo e o uso dos Ãleos 1 e 2 reduziram a fibrose e a vascularizaÃÃo na fase proliferativa. O prÃcondicionamento com o Ãleo 1(milho e soja) promoveu mais inflamaÃÃo no 7Âdia. / The tissue repair consists in an important feature of the inflammatory process prevention, some factors may reduce, delay or prevent this process. There is a continued interest in the use of currents with low intensity combined by supplementation with nutraceuticals (high ratio w9:w6 and low ratio w6:w3). The objective of this work was to evaluate the effects of preconditioning with oil blends containing (w3/w6) and (w9/w6), and currents of low intensity MENS on the tissue repair after incisions in mice skin. One hundred and eight (108) Wistar rats were used , divided into three groups: G-1 control (saline) n = 36 subdivided into two subgroups: unstimulated and stimulated; G-2 neutral control (corn and soybean oil) n= 36, subdivided into two subgroups: unstimulated and stimulated and G-3 ALA (olive oil + flaxseed + canola), n= 36 subdivided into two subgroups unstimulated and stimulated. The oil mixtures were administered by orogastric, and the application of electric current directly into mouse skin occurred 1 hour before surgical incision. The animals were sacrificed on the seventh, fourteenth and twenty-first day. Skin sample of 1cm/1 cm were collected between the second and third suture point. The samples were analyzed from surgical specimens stained with hematoxylin-eosin (HE) and provided a general analysis of tissue sections with a thickness of 5 um. The data obtained by the (HE) technique were classified according to the inflammatory intensity and transformed into quantitative variables by assigning index for the hestological finding according to (VIDINSK, 2006). Statistical data were analysed using the GraphPad Prism for Windows, version 6.0, for joint assessment of the effects of the groups G-1, G-2 e G-3, and the subgroups unstimulated and stimulated, and days (7, 14, 21). The variables of histological evaluation were analysed by nonparametric Mann-Whitney. The significance level was 5 % (p<0,05). The results show that the preconditioning electrical stimulation caused the elevation of exudate and reduced the reepitheliazation on the 7th day. The use of preconditons oil 1 (corn and soybeans) and oil 2 (olive+ canola + flaxseed) reduced the vascularization on the 7th and 14th days respectively. Both groups, 1 and 2 of oils, induced the reduction of fibrosis on the 14th day. The preconditioning with oil 1 led to elevated expression of NFKB on the 7th day. It is concluded that the electrical stimulation caused the elevation of markers of inflammatory phase with consequent acceleration of this process and the use of oils 1 and 2 reduced the fibrosis and vascularization on proliferative phase. The preconditioning with the oil 1 (corn and soybeans) caused more inflammation on the 7th day.

Omega-3 fatty acids are increasingly recognised as playing an important role in human brain development and mental health. The polyunsaturated fatty acids (PUFAs) include omega-3 and omega-6 fats which are essential fatty acids (EFAs), consumed via the diet. Omega-3 fatty acids are particularly abundant in fish oils. The omega-3 fatty acids are being focused on for their role in depression, the main types being docosahexaenoic acid (DHA), which is abundant in neural tissue, and also eicosapentaenoic acid (EPA) which is biologically very active. There is an emerging literature in relation to omega-3 fatty acid blood levels in depression and the effects of treatment with omega-3. Strong epidemiological evidence has also been published indicating an association between a population's fish intake and depression rates. A specific research focus on omega-3 as a treatment for depression in the perinatal period is also starting to emerge. The importance of this particular area is enhanced by the knowledge that omega-3 depletion occurs during the perinatal period due to fetal diversion for neurodevelopment. In view of the lay public promotion of omega-3 and its appeal to women as a 'natural therapy', there is a need to scientifically evaluate its effectiveness to treat depression in the perinatal period. It is also important to investigate omega-3 as an alternative to antidepressants given the ongoing uncertainties regarding their safety in pregnancy. In this thesis a literature review presents current research relating to this field. This is followed by a description of the methodology and results for the two trials conducted. The results of the double-blind randomised placebo controlled trial of omega-3 as a treatment for depression in the perinatal period were essentially negative. However this result is limited by the small sample size in the study and therefore it may be unwise to interpret the result as conclusive. The case-control study confirmed the hypothesis that omega-3 levels were more depleted in depressed women compared to non-depressed women. A discussion of the results and trial limitations then follows in the thesis. It is concluded that further larger studies are warranted in this area.

<p>Several abnormal conditions, including some cancers, have been associated with changes in the membrane lipid and FA composition. Dietary fat serves as a major source of lipids and FA, particularly the polyunsaturated fatty acids (PUFA), n-6 and n-3. High intakes of n-6 PUFA have been linked to the development of colon cancer in association with low n-3 PUFA intake. Therefore understanding the differences in the lipid and FA profiles between cancer and normal cells in the colon, and the role diet plays in these factors may be invaluable in understanding their role in carcinogenesis. This study compares the lipid profile of azoxymethane (AOM) induced colon polyps to that of the surrounding mucosa tissue in rats fed a diet high in n-6 PUFA. Male Fischer rats were fed the AIN-76A diet containing sunflower oil that has high n-6 PUFA content for a period of nine months. Results indicate that the lipid and FA content of the colon polyps differs significantly from the surrounding mucosa. Colon polyps had an increase in membrane phopholipids phosphatidylcholine (PC) and phosphatidylethanolamine (PE). Changes in membrane fluidity were indicated by the decrease (p&lt / 0.05) in the PC/PE and cholesterol/phospholipids (chol/PL) ratios, and increase (p&lt / 0.05) in the polyunsaturated FA/saturated FA (P/S) ratio. Metabolism of FA was significantly altered in the polyps favouring n-6 FA metabolism and the production of prostaglandin E2. No clear indication of impaired &Delta / 6-desauturase enzyme activity was noticed. Increases in the n-6 PUFA content could be a reflection of the dietary FA intake that increases FA incorporation in the polyps. Changes in the FA parameters of the polyps, particularly an increase in C20:4n-6 and the n6/n3 ratio have been shown to contribute to the rapid growth of cancer tissue. These lipid changes associated with the development of colon polyps could provide unique targets for developing strategies in chemoprevention by dietary manipulation.</p>