Tesi sul tema "Linoleate"

Kyoto University (京都大学)
0048
新制・課程博士
博士(農学)
甲第18313号
農博第2038号
新制||農||1020(附属図書館)
学位論文||H26||N4820(農学部図書室)
31171
京都大学大学院農学研究科食品生物科学専攻
(主査)教授 安達 修二, 教授 河田 照雄, 教授 保川 清
学位規則第4条第1項該当

Les interactions entre nutriments dans les produits alimentaires peuvent affecter leur bioaccessibilité. La β-lactoglobuline (βlg), protéine majeure du lactosérum, est connue pour lier des ligands hydrophobes tels que des acides gras (AG). Très sensible aux conditions industrielles, la βlg est souvent sous formes non natives dans les aliments transformés. Notre hypothèse est que les changements structuraux au niveau de la protéine modifient l'affinité pour les AG et par conséquent les propriétés biologiques des complexes AG/protéine. L'objectif était d'étudier l'interaction de la βlg bovine sous différentes structures (native, dimère covalent et nanoparticules) avec l’acide linoléique (LA) et l’acide linoléique conjugué (CLA), et l'impact de ces complexes sur leurs activités biologiques in vitro. Par fluorescence intrinsèque et calorimétrie de titration isotherme, nous avons montré que quel que soit l’état d'agrégation de la protéine, LA interagit avec la βlg au niveau de deux classes de sites. En augmentant le niveau d’agrégation de la βlg, la stœchiométrie des complexes LA/βlg augmente mais sans changement des constantes d'association. En présence de LA, la βlg native est plus sensible à la digestion gastrique in vitro en raison de l'augmentation du niveau de dénaturation/agrégation de la βlg. La quantification de l’absorption du LA dans une monocouche cellulaire imitant la barrière intestinale indique que son transport est ralenti en présence de βlg native ce que confirme la microscopie confocale. La cytotoxicité du LA pour les cellules Caco-2 a été réduite lorsque l’AG était lié à la βlg. Le CLA, qui est moins soluble dans l'eau que le LA, est plus cytotoxique lorsqu'il est complexé à la βlg que sous sa forme libre. Par conséquent, la βlg peut moduler le transport et la bioaccessibilité de l’AG en fonction de la solubilité de ce dernier
Food structure can have a profound influence on delivering health benefits. Bioaccessibility of nutrients can be affected by their interaction with food components. The dairy protein β-lactoglobulin (βlg) is known to bind hydrophobic ligands such as fatty acids (FA). However, this protein is highly sensitive to the process conditions used in the dairy industry. Therefore βlg is often present in non-native or aggregated form in processed food. This structural change may modify the protein affinity for FA and the biological properties of the FA/protein complexes. The aim of this thesis was to investigate the interaction of bovine βlg in different structural forms (native, covalent dimer and nanoparticles) with linoleate (C18:2, cis,cis-9,12-octadecadienoic acid) and conjugated linoleic acids (CLA, C18:2), and the impact of those complexes on their biological activity in vitro. Two different sets of binding sites were determined for the interaction between linoleate and βlg, regardless of its state of aggregation, using intrinsic fluorescence spectroscopy and isothermal titration calorimetry. By increasing the level of βlg aggregation, the linoleate/βlg stoichiometry increased but the association constants remained similar for both sets of binding sites. In the presence of linoleate, the native protein was more sensitive to gastric in vitro digestion, due to the increased level of denaturation/aggregation of βlg. Transport of linoleate in Caco-2 cells was decreased in presence of the native βlg as observed by confocal microscopy and a monolayer that mimics the intestinal barrier. Cytotoxicity of linoleate on Caco-2 cells was reduced when the FA was bound to βlg compared to free FA. CLA, which is less water soluble than linoleate, is more cytotoxic when complexed by βlg than in its free form. Therefore, it is proposed that βlg can act as a molecular carrier and alter the bioaccessibility of FA depending on their solubility

Emerging evidence suggests that an important mechanism for the negative feedback control of insulin signalling involves the inhibition of tyrosine phosphorylation of IRS-1 by its prior serine/threonine (ser/thr) phosphorylation. IRS-1 ser/thr phosphorylation has been linked to the dissociation of IRS-1 from the insulin receptor and PI3K, and its degradation via a proteasome-dependent pathway. Studies in animal models have shown that increases in plasma free fatty acids (FFAs) are associated with reduced IRS-1-signalling, and so it has been postulated that elevated FFA cause insulin resistance by activating pathways that negatively regulate insulin action, including hyper-phosphorylation of ser/thr residues in IRS-1. We have shown that in the case of linoleate-induced insulin resistance in L6 rat skeletal muscle cells, the inhibition of IRS-1-dependent signalling arises via effects on both the phosphorylation status and degradation of IRS-1, which are mediated, in part, by IKKb. In addition, the reduction of IRS-1 mRNA levels allude to transcriptional effects of linoleate treatment that also contribute to the observed reduction in the total levels of this protein. PtdOH, particularly dilinoleoyl PtdOH, was found to be significantly increased in linoleate treated L6 cells, and sufficient to induce at least some of the effects on insulin-signalling that are observed upon linoleate treatment. It is unlikely, however, that IKKb and PtdOH are components of the same inhibitory pathway, since inhibiting IKKb activity did not alleviate the effects of PtdOH on IRS-1 tyrosine (tyr) phosphorylation. Moreover, although an integral component of the mechanism by which linoleate induces insulin-resistance in L6 cells, it appears that restoring IRS-1 function in linoleate treated cells is not sufficient to reverse insulin resistance. Hence, we hypothesise that linoleate induces multiple inhibitory pathways in L6 cells, with at last two of these involving IKKb- and PtdOH-dependent inhibition of IRS-1 signalling, which act in parallel to reduce glucose disposal and cause insulin resistance in this model.

The methyl ester of an unsaturated fatty acid, methyllinoleate was reacted with oxygen in a pressurized system at a controlled temperature. A natural autoxidation of methyl linoleate was observed without the addition of an initiating reagent. This autoxidation process could be used to mimic the course of lipid peroxidation, which is the major cause ofradical damage to living cells. The technology of GC-MS was employed to monitor the autoxidation of methyllinoleate. Eight of the autoxidation products separated by GC column were identified by interpreting the corresponding EI ion mass spectra. The products from 9-alkoxy methyl linoleate radical were methyl octanate, 2,4-decadienal, nonanoic acid, 9-oxo-methyl ester, and its further oxidation product, nonanedioic acid, monomethyl ester. All of them formed through a pathway of beta-cleavage. The products from 13-alkoxy methyl linoleate radical were tridecanoic acid, 9, 11-diene-13-oxo-methyl ester, hex anal, and its further oxidation product, hexanoic acid. They were also formed through a mechanism of beta-cleavage. The fourth product from 13-alkoxy methyllinoleate radical was 13-keto-9, 11-octadecadienoic acid, methyl ester, which was obtained through a pathway of keto formation. Observation of their concentrations in the samples at different autoxidation periods revealed the time-course of formation of these products.

The methyl ester of an unsaturated fatty acid, methyllinoleate was reacted with oxygen in a pressurized system at a controlled temperature. A natural autoxidation of methyl linoleate was observed without the addition of an initiating reagent. This autoxidation process could be used to mimic the course of lipid peroxidation, which is the major cause ofradical damage to living cells. The technology of GC-MS was employed to monitor the autoxidation of methyllinoleate. Eight of the autoxidation products separated by GC column were identified by interpreting the corresponding EI ion mass spectra. The products from 9-alkoxy methyl linoleate radical were methyl octanate, 2,4-decadienal, nonanoic acid, 9-oxo-methyl ester, and its further oxidation product, nonanedioic acid, monomethyl ester. All of them formed through a pathway of beta-cleavage. The products from 13-alkoxy methyl linoleate radical were tridecanoic acid, 9, 11-diene-13-oxo-methyl ester, hex anal, and its further oxidation product, hexanoic acid. They were also formed through a mechanism of beta-cleavage. The fourth product from 13-alkoxy methyllinoleate radical was 13-keto-9, 11-octadecadienoic acid, methyl ester, which was obtained through a pathway of keto formation. Observation of their concentrations in the samples at different autoxidation periods revealed the time-course of formation of these products.

Vegetable oils are a cheap and abundant chemical feedstock which can be readily broken down into fatty acid methyl esters (FAMEs) by transesterification using methanol and a base catalyst. These FAMEs contain reactive unsaturated double bonds which can be targeted for modification. In this study, enzymatic and chemical modifications of the unsaturated double bonds of FAMEs are explored with the goal of producing higher value products. Specifically the enzymatic modification of methyl linoleate using soybean lipoxygenase and the chemical modification of methyl oleate using hypobromantion are studied.

Dioxygenase-cytochrome P450 (DOX-CYP) fusion enzymes are present in diverse human and plant pathogenic fungi. They oxygenate fatty acids to lipid mediators which have regula­tory functions in fungal development and toxin production. These enzymes catalyze the for­mation of fatty acid hy­droperoxides which are subsequently converted by the P450 activities or reduced to the corresponding alcohols. The N-terminal DOX domains show catalytic and structural homology to mammalian cyclooxygenases, which belong to the most thoroughly studied human enzymes. 7,8-Linoleate diol synthase (LDS) of the plant pathogenic fungus Gaeumannomyces graminis was the first characterized member of the DOX-CYP fusion enzyme family. It catalyzes the conversion of linoleic acid to 8R-hydroperoxylinoleic acid (HPODE) and subse­quently to 7S,8S-dihy­droxylinoleic acid by its DOX and P450 domains, respectively. By now, several enzymes with homology to 7,8-LDS have been identified in im­portant fungi, e.g., psi fac­tor-producing oxygenase (ppo)A, ppoB, and ppoC, of Aspergillus nidulans and A. fumigatus. By cloning and recombinant expression, ppoA of A. fumigatus was identi­fied as 5,8-LDS. Partial expression of the 8R-DOX domains of 5,8-LDS of A. fumigatus and 7,8-LDS of G. graminis yielded active protein which demonstrates that the DOX activities of LDS are independent of their P450 domains. The latter domains were shown to contain a conserved motif with catalytically important amide residues. As judged by site-directed mutagene­sis studies, 5,8- and 7,8-LDS seem to facilitate heterolytic cleavage of the oxygen-oxygen bond of 8R-HPODE by aid of a glutamine and an asparagine residue, respectively. Cloning and expression of putative DOX-CYP fusion proteins of A. terreus and Fusarium oxysporum led to the discovery of novel enzyme activities, e.g., linoleate 9S-DOX and two allene oxide synthases (AOS), specific for 9R- and 9S-HPODE, respectively. The fungal AOS are present in the P450 domains of two DOX-CYP fusion enzymes and show higher se­quence homology to LDS than to plant AOS and constitute therefore a novel class of AOS. In summary, this thesis describes the discovery of novel fatty acid oxy­genases of human and plant pathogenic fungi and the characterization of their reaction mechanisms.

7,8-Linoleate diol synthase (7,8-LDS) of the take-all pathogen of wheat, Gaeumannomyces graminis, converts linoleic acid to 8R-hydroperoxyoctadecadienoic acid (8-HPODE) by 8-dioxygenase activity (8-DOX), and further isomerizes the hydroperoxide to 7S,8S-dihydroxyoctadecadienoic acid (7,8-DiHODE) by hydroperoxide isomerase activity. Sequence alignment showed homology to prostaglandin H synthase (PGHS), and both enzymes share structural and catalytic properties. The 8-DOX of 7,8-LDS was successfully expressed in Pichia pastoris and in insect cells (Sf21). Site-directed mutagenesis confirmed His379 as the proximal heme ligand and Tyr376 as a residue, which forms a tyrosyl radical and initiates catalysis. Furthermore, mutagenesis suggested His203 could be the proposed distal histidine, and Tyr329 of catalytic relevance for substrate positioning at the active site. Aspergilli are ubiquitous environmental fungi. Some species, in particular Aspergillus fumigatus, are responsible for invasive aspergillosis, which is a life-threatening disease for immunocompromised patients. A. fumigatus and A. nidulans metabolized linoleic acid to 8R-HPODE, 10R-hydroperoxyoctadecadienoic acid (10R-HPODE), 5S,8R-dihydroxyoctadecadienoic acid, and 8R,11S-dihydroxyoctadecadienoic acid. When the genomes of certain Aspergilli strains were published, several species showed at least three homologous genes (ppoA, ppoB, ppoC- psi producing oxygenases) to 7,8-LDS and PGHS. Gene deletion identified PpoA as an enzyme with 8-DOX and 5,8-hydroperoxide isomerase activities, designated 5,8-LDS in homology to 7,8-LDS. In the same way, PpoC was identified as a 10-dioxygenase (10-DOX), which converts linoleic acid to 10R-HPODE. 10-DOX differs from LDS, since it dioxygenates linoleic acid at C-10, after hydrogen abstraction at C-8 and double bond migration. 10-DOX was cloned and expressed in insect cells. Leu384 and Val388 were found to be critical for dioxygenation at C-10. Mutation to the homologous residues of 5,8- and 7,8-LDS (Leu384Val, Val388Leu) increased oxygen insertion at C-8. LDS and 10-DOX are fusion proteins with a dioxygenase and a hydroperoxide isomerase (cytochrome P450) domain with a cysteine heme ligand. The P450 domain of 10-DOX lacked the crucial cysteine heme ligand and was without hydroperoxide isomerase activity. LDSs and 10-DOX are newly characterized heme containing fungal dioxygenases, with homology to PGHS of vertebrates. Their metabolites regulate reproduction, development, and act as signal molecules with the host after pathogen attack.

The dioxygenase-cytochrome P450 fusion proteins (DOX-CYP) comprise a heme-containing enzyme family that shares structural and catalytic properties with mammalian prostaglandin H (PGH) synthases. 7,8-Linoleate diol synthase (7,8-LDS) of Gaeumannomyces graminis was first characterized, and DOX-CYP enzymes are of mechanistic and biological interest. The growing number of fungal genome sequences has revealed DOX-CYP homologues in medically and economically important species. The aim of this thesis was to identify novel members of the DOX-CYP fusion protein family. The devastating rice pathogen Magnaporthe oryzae contains two DOX-CYP genes. The fungus synthesizes 7S,8S-dihydroxyoctadecadienoic acid (7,8-DiHODE) by dioxygenation of linoleic acid to 8R-hydroperoxyoctadecadienoic acid (8R-HPODE), and subsequent isomerisation to the diol. 7,8-LDS of M. oryzae was identified by gene deletion, but the infection and reproduction processes of the Δ7,8-LDS strain were not altered. A mutant with constitutive protein kinase A activity profoundly changed the oxygenation profile, possibly due to post-translational modification. The human pathogens Aspergillus fumigatus and A. clavatus contain three DOX-CYP, designated psi producing oxygenase A (ppoA), ppoB, and ppoC, and form three oxylipins: 5S,8R-DiHODE, 8R,11S-DiHODE, and 10R-hydroxyoctadecadienoic acid.  PpoA was identified as 5,8-LDS, and ppoC as 10R-DOX. The 8,11-linoleate hydroperoxide isomerase activity was reduced by two imidazole-containing P450 inhibitors, miconazole and 1-benzylimidazole. PpoB could not be linked to the biosynthesis of 8,11-DiHODE for the following reasons: First, the 8,11-hydroperoxide isomerase activity was retained in A. fumigatus ΔppoB strains. Second, the P450 domain of the deduced ppoB of A. clavatus lacks a heme-thiolate cysteine ligand, presumably essential for hydroperoxide isomerase activity. Linoleate 9R-DOX activities of Aspergillus terreus and Lasiodiplodia theobromae were discovered. 9R-HPODE was further converted into unstable allene oxides, as judged by the accumulation of their hydrolysis products, α- and γ-ketols. These allene oxide synthase activities were specific for 9R-hydroperoxides. The 9R-DOX and AOS were found to have unique characteristics. In conclusion, novel DOX-CYP enzymes were identified in human and plant pathogenic fungi. These enzymes might be involved in biological processes, and show interesting catalytic similarities to human PGH synthase and thromboxane synthase (CYP5A).

La valorisation des minéraux calciques est un problème mondial. La flottation est une technique utilisée pour séparer ces minéraux puisqu’elle joue sur leurs propriétés superficielles. Cependant, c’est un défi scientifique important puisque les propriétés de surface des minéraux calciques sont très proches. Les difficultés de séparation de ces derniers sont donc dues, aux similitudes entre leurs propriétés chimiques de surface, mais aussi leurs propriétés électrocinétiques et leur stabilité en milieu aqueux. L’application des résultats de recherche fondamentale porte sur les minéraux purs dont 4 calcites et une apatite d’origines différentes. L’objectif principal est d’étudier les propriétés et les paramètres impliqués dans les mécanismes de séparation des minéraux calciques tels que l’importance de la solubilité, la spéciation de surface du minéral, la charge globale de la surface du minéral et la densité d’adsorption des tensio-actifs. Les propriétés électrocinétique permettent de déterminer le type de collecteur à employer pour faire flotter sélectivement les minéraux et pour choisir le pH optimal de séparation. Deux collecteurs, l’oléate et le linoléate de sodium (structure de la chaîne hydrocarbonée différente) ont été utilisés pour étudier la flottation des minéraux calciques. Une étude des mélanges de ces deux collecteurs avec différents ratios molaires a montré un contraste de séparation à pH 5 avec le ratio molaire 2 :1 à pH 5 et le ratio 1 :1 à pH 9. De plus, Les effets synergiques entre collecteur ionique (oléate ou linoléate de sodium) et non-ionique (PX type alcool) en présence ou non des déprimants tels que l’amidon et le silicate de sodium ont été étudiés pour améliorer la la sélectivité de la séparation calcite apatite. Les isothermes d’adsorption de l’oléate et du linoléate de sodium, en présence ou non d’un collecteur non-ionique, obtenus par la méthode de Gregory mettent en évidence leurs co-adsorption sur la surface de la calcite et de l’apatite. Ceci a aussi été confirmé par les déplacements des bandes de vibration symétriques et asymétriques des groupements CH2-CH3 sur les spectres infrarouges en réflexion diffuse. L’adsorption du linoléate de sodium sur la calcite orange présente différentes régions, ce qui peut être expliqué par une adsorption en multicouche résultant de la condensation bidimensionnelle du collecteur sur une surface hétérogène. Cependant, l’adsorption de l’oléate de sodium sur l’échantillon de calcite avec les impuretés de Mg est linéaire. L’apatite de Madagascar et la calcite optique présentent une saturation des sites d’adsorption à partir d’une concentration 3.10-5 M en oléate de sodium (CMC)
Valorization of calcium minerals is a global problem. Flotation is a technique used for the separation of these minerals since it plays on their surface properties. However, this is an important scientific challenge because the surface properties of calcium minerals are very similar. The difficulties of separation of these minerals are then due to the similarities between their chemical surface properties, but also their electrokinetic and their stability in aqueous solutions. The application of the basic research results will focus on 4 calcites and one apatite of different origins. The main objective is to study the properties and parameters involved in the separation mechanisms of calcium minerals such as the importance of the solubility, the surface speciation of the mineral, the overall loading of the mineral surface and the density of adsorption of surfactants. The electrokinetic properties therefore make it possible to determine the type of collector to be used to selectively float the minerals and to choose the optimum separation pH. Two collectors, sodium oleate and linoleate (different semi-developed formula), were used to study the flotation of calcium minerals. The mixture of these two collectors with different molar ratios was also studied showing a separation contrast at pH 5 with the molar ratio 2: 1 and the ratio 1: 1 at pH 9. In addition, the synergistic effects between ionic collector (oleate or sodium linoleate) and non-ionic (PX alcohol type) in the presence or absence of depressants such as starch and sodium silicate have been studied to improve recovery and selectivity. The adsorption isotherms of sodium oleate and linoleate, in the presence or absence of a non-ionic collector, obtained by the Gregory method demonstrate their co-adsorption on the surface of calcite and apatite. This was also confirmed by the displacements of the symmetric and antisymmetric vibration bands of the CH2-CH3 groups on the infrared spectra in diffuse reflectance. The adsorption of sodium linoleate on one of the calcite minerals (calcite orange) shows different regions which can be explained by multi-layer adsorption resulting from the two-dimensional condensation of the collector on a heterogeneous surface. However, the adsorption of sodium oleate on calcite orange is linear. The apatite from Madagascar and calcite optical have a saturation of the adsorption sites from a concentration of 3.10-5 M in sodium oleate (CMC)

Tesis presentada para optar al Título de Ingeniero en Alimentos y Grado de Magíster en Alimentos, mención en Gestión, Calidad e Inocuidad de Alimentos
Las materias grasas, en general los aceites que contienen ácidos grasos poliinsaturados son susceptibles a sufrir deterioro oxidativo cuando se someten a procesos térmicos, alterando las propiedades organolépticas y nutricionales de los aceites. Para retardar el deterioro oxidativo se ha recurrido a la utilización de antioxidantes sintéticos, sin embargo, estos han sido cuestionados por sus posibles efectos adversos en la salud a altas concentraciones. En este contexto, investigaciones en la utilización de extractos vegetales como fuente de antioxidantes, se están llevando a cabo. El objetivo de esta tesis fue estudiar el efecto de la adición de extractos hidroalcohólicos y acuosos de hojas y cáscaras de palto en polvo, sobre la estabilidad oxidativa de linoleato de metilo sometido a 80 °C, en comparación con la adición de un antioxidante sintético (BHA). Los extractos de cáscaras y hojas de palto (variedad Hass) se prepararon utilizando como solvente agua (EAC, EAH) o etanol/agua (EHC, EHH), posteriormente se secaron a 140 °C, usando Capsul® como carrier. Los extractos de palto (EAC, EAH, EHC y EHH) antes y después del secado, se caracterizaron de acuerdo al contenido de polifenoles totales (Folin-Ciocalteau) y capacidad antioxidante (FRAP y DPPH), mientras que la identificación de los principales polifenoles (HPLC-detector UV y espectómetro de masas) se realizó en los extractos en polvo. Los resultados obtenidos mostraron, un aumento en el contenido de polifenoles totales y capacidad antioxidante en los cuatros extractos después del secado por atomización, siendo mayor para el EHC. Los principales polifenoles identificados en los extractos en polvo fueron ácido clorogénico, procianidina dímero y epicatequina. La estabilidad térmica-oxidativa de muestras de linoleato de metilo (LM) con la adición de extractos de cáscaras y hojas de palto en polvo (EAC, EAH, EHC y EHH) (equivalente a 400 ppm de polifenoles) se sometieron a un almacenamiento a 80 °C por 240 horas. Los resultados mostraron que los extractos de palto no tuvieron un efecto significativo (p≤0,05) en disminuir la formación de compuestos polares en LM, sólo el BHA mejoró la estabilidad termo-oxidativa de LM. La extracción de los polifenoles ácido clorogénico (AC) y procianidina dímero (PD) desde los sistemas de LM con los extractos de cáscaras y hojas de palto en polvo, después del almacenamiento a 80 °C, se realizó por extracción en fase sólida (cartuchos diol). Los resultados mostraron que los extractos de hojas (hidroalcohólico y acuoso) presentaron mayores concentraciones de AC, mientras que los extractos de cáscaras (hidroalcohólico y acuoso) presentaron mayores concentraciones de PD. No se observó un efecto del tipo de extracto (solvente de extracción y tipo de material vegetal (hoja o cáscara)) sobre la constante de velocidad de degradación de AC y PD en LM con extractos de palto almacenados a 80 °C hasta 216 horas
In general oils that contain polyunsaturated fatty acids are susceptible to oxidative deterioration under thermal processes. Plant extracts can be considered as a source of natural antioxidants for inhibition of lipid oxidation. The aim of this work was to study the effect of avocado extracts (leaves and peels) on the oxidative stability of methyl linoleate at 80 °C. The avocado extracts of leaves and peels (var. Hass) were elaborated using water (PAE and LAE, respectively) or ethanol:water (50:50 v/v) (PHE and LHE, respectively) as extracting solvents. The aqueous and hydroalcoholic extracts were dried by spray-drying at 140 ºC using Capsul® as carrier. The extracts (PAE, LAE, PHE and LHE) before and after spray-drying were characterized according to total content of polyphenols (Folin-Ciocalteau) and antioxidant activity (FRAP and DPPH) while the identification of the main polyphenols (HPLC-UV detector and mass spectrometer) was performed on powder extracts. The results showed an increase of total polyphenol content and antioxidant capacity in the four extracts after spray- drying, being the highest PHE. The main polyphenols identified in powder extracts were: chlorogenic acid, procyanidin dimer and epicatechin. The thermal-oxidative stability of metyl linoleate (ML) samples with the addition of avocado extracts powder (LAE, PAE, LHE y PHE) (400 ppm polyphenols) were storaged at 80 ºC until 240 h. The results showed that the avocado extracts did not have a significant effect (p≤0.05) in decreasing the formation of polar compounds in ML, only BHA improved the thermo-oxidative stability of ML. The extraction of chlorogenic acid (CA) and procyanidin dimer (PD) from ML with avocado extracts during storage at 80 °C, were performed by solid phase extraction (cartridges Diol). The avocado leaves extracts (hydroalcoholic and aqueous) had a higher CA content wereas the avocado peels extracts (hydroalcoholic and aqueous) had a higher PD content. The extract type (extracting solvent and plant material (leaf or peel)) did not have influence on the degradation rate constants of CA and PD in ML with avocado extracts at 80 °C until 216 hours

Doctor en Nutrición y Alimentos
La oxidación lipídica es una de las principales reacciones que afectan la calidad de los aceites, transformándose en uno de los principales problemas para la industria alimentaria por la aparición de olores y sabores indeseables. Para la protección de la oxidación lipídica, se pueden utilizar micropartículas de flavonoides de liberación controlada para extender la vida útil de las materias grasas y disminuir la utilización inefectiva del antioxidante natural. De acuerdo a estos antecedentes el objetivo de este estudio fue estudiar la cinética y mecanismo de liberación de flavonoides desde micropartículas con y sin la adición de un agente canalizante en linoleato de metilo (LM) y su efecto en la formación de compuestos polares. Se encapsularon los flavonoides quercetina (Q), naringenina (N) o epicatequina (E), mediante secado por atomización, utilizando inulina (IN) o hidroxipropilcelulosa (HPC) como agentes encapsulantes y Capsul® (C) como agente canalizante, de acuerdo a un diseño Box-Behnken, con un total de 15 experimentos para cada sistema de micropartículas estudiado (Q-(IN-C), N-(IN-C), E-(IN-C), Q-(HPC-C), N-(HPC-C) y E-(HPC-C)). Las variables independientes fueron la relación flavonoide/agente encapsulante, temperatura del aire entrada y contenido de Capsul. Las variables dependientes fueron la eficiencia de encapsulación y liberación de flavonoides a los 14 días (t14) y a los 28 días (t28) de almacenamiento en hexano (H) para los sistemas con IN y HPC, respectivamente. Se utilizó la metodología de Superficie Respuesta para la obtención de micropartículas bajo condiciones óptimas. Los sistemas de micropartículas obtenidos bajo condiciones óptimas (Q-IN, Q-(IN-C), N-IN, N-(IN-C), E-IN, E-(IN-C), Q-(HPC-C), N-(HPC-C) y E-(HPC-C)) se utilizaron para determinar la EE y el perfil de liberación de flavonoides en H y LM. La determinación de flavonoides se realizó mediante HPLC. Las micropartículas obtenidas mostraron EE superiores al 60% para los sistemas de Q y E y menores para los sistemas con N. Un incremento en el número de grupos OH en el flavonoide aumentó la EE debido a la interacción entre los grupos OH del flavonoide con los sitios OH de IN y HPC mediante puentes de hidrógeno. El perfil de liberación de flavonoides desde las micropartículas en H y en LM durante el almacenamiento en estufa a 30 ºC, mostró que para las micropartículas con IN sin y con C, la constante de velocidad de liberación de N fue significativamente mayor (p≤0,05) respecto a las de Q y E en LM y en H. Mientras que la constante de velocidad de liberación de E fue significativamente menor (p≤0,05) respecto a Q y N en LM y en H. El parámetro “n” en el modelo de Peppas para las micropartículas con IN sugiere que el mecanismo de liberación de los flavonoides se atribuye a la presencia de un agente canalizante, independiente de la naturaleza del flavonoide. Para las micropartículas con HPC solamente se observó la liberación de N en LM y correspondió a liberación de los flavonoides superficiales. Mientras que en H se observó liberación de N y Q sin diferencias significativas (p>0,05) entre las constantes de liberación. El parámetro “n” en el modelo de Peppas sugiere que el mecanismo de liberación de los flavonoides se atribuye a la interacción agente encapsulante-medio de disolución y flavonoide-agente encapsulante. Se realizaron ensayos de estabilidad oxidativa en linoleato de metilo a 60 ºC con la adición de flavonoides libres (LM-N, LM-Q y LM-E) y encapsulados (LM-Q-IN, LM-Q-(IN-C), LM-Q-(HPC-C), LM-E-IN, LM-E-(IN-C) y LM-E-(HPC-C)) (200 mg/Kg). Las micropartículas de flavonoides se elaboraron utilizando la misma relación flavonoide-agente encapsulante, mientras que el contenido de C y temperatura de entrada al secador correspondieron a las utilizadas en la elaboración de micropartículas obtenidas bajo condiciones óptimas. La determinación de compuestos de oxidación se realizó por HPSEC y la retención de flavonoides por HPLC. La adición de flavonoides libres mostró un factor de protección y tiempo de inducción significativamente mayor para E, en relación a Q y N, mientras que N no ejerció ningún efecto sobre la estabilidad oxidativa. La adición de flavonoides encapsulados al LM mostró un aumento significativo en el factor de protección y tiempo de inducción, respecto al LM y fue mayor para el sistema LM-Q-(IN-C) mostrando el efecto del agente canalizante sobre el control de la liberación. En todos los sistemas el final de la fase lag e inicio de la propagación fue claramente marcado por la desaparición de Q y E y la iniciación de la formación de polímeros coincide con el tiempo de inducción Se puede concluir que conocer el perfil de liberación y capacidad antioxidante determina la aplicabilidad de las micropartículas en sistemas lipídicos
Lipid oxidation is one of the main reactions affecting the quality of oils; it is also responsible for the appearance of unwanted smells and tastes, negatively impacting the food industry. However, the use of controlled-release flavonoid microparticles can reduce the effects of lipid oxidation, thus extending the shelf life of fats, and reducing the ineffective use of natural antioxidants. Based on the above, the objective of this research was to study the kinetics and mechanism of flavonoid release from microparticles with and without the addition of a channeling agent in methyl linoleate (ML), and its effect on the formation of polar compounds. The flavonoids quercetin (Q), naringenin (N) or epicatechin (E) were encapsulated by spray-drying, using Inulin (IN) or Hydroxypropylcellulose (HPC) as encapsulating agents, and Capsul (C) as channeling agent, using a Box-Behnken design. A total of 15 experiments were conducted for each microparticles system (Q-(IN-C), N-(IN-C), E-(IN-C), Q-(HPC-C), N-(HPC-C) and E-(HPC-C)). The independent variables were the flavonoid/encapsulating agent ratio, air inlet temperature and Capsul content, whereas the dependent variables were encapsulation efficiency and flavonoid release at 14 (t14) and 28 (t28) days of storage in hexane (H) for the IN and HPC systems, respectively. The Response Surface methodology was used to obtain microparticles under optimal conditions. The microparticles systems obtained under optimal conditions (Q-(IN-C), N-(IN-C), E-(IN-C), Q-(HPC-C), N-(HPC-C) and E-(HPC-C)) were used to determine the EE, and the flavonoid release profile in H and ML. The determination of flavonoids was performed using HPLC. The obtained microparticles indicated that the encapsulation efficiency (EE) to be greater than 60% for the Q and E systems, and less for the N systems. An increase in the number of OH groups in the flavonoid increased the EE due to the interaction between the OH groups of the flavonoid with the OH sites of IN and HPC through hydrogen bonds. The release profile of flavonoids from the microparticles in H and in ML during oven storage at 30° showed that, for the microparticles with IN and with and without C, the constant release rate of N was significantly greater than (p≤0.05) with respect to the one of Q and E in ML and in H. While the release rate constant of E was significantly lower (p≤0.05) with respect to Q and N in ML and in H, the “n” parameter for Peppas model for the microparticles with IN suggests that the release mechanism of flavonoids is attributed to the presence of a channeling agent, irrespective of the nature of the flavonoid. For the microparticles with HPC, only the release of N in ML was observed, and it corresponded to the release of superficial flavonoids, whereas for H the release of N and Q was observed without significant differences (p>0,05) amongst the release constants. The “n” parameter in Peppas model suggests that the release mechanism of flavonoids is attributed to the interaction of both the encapsulating agent-dissolution medium and flavonoid-encapsulating agent medium. Oxidative stability trials were performed in methyl linoleate at 60° with the addition of free flavonoids (ML-N, ML-Q and ML-E) and encapsulated flavonoids (ML-Q-IN, ML-Q-(IN-C), ML-Q-(HPC-C), ML-E-IN, ML-E-(IN-C) and MLE-E-(HPC-C)) (200 mg/kg). The flavonoid microparticles were elaborated using the same flavonoid-encapsulating agent ratio, while the content of C and inlet temperature of the dryer corresponded to those used in the elaboration of microparticles obtained under optimal conditions. The determination of oxidative compounds was performed using HPSEC, and the retention of flavonoids using HPLC. The addition of free flavonoids presented a protection factor and induction time significantly higher for E in relation to Q and N, whereas N did not have an effect on the oxidative stability. The addition of encapsulated flavonoids to ML showed a significant increase in the protection factor and induction period with respect to ML, and it was greater for the ML-Q-(IN-C) system, displaying the effect of the channeling agent over the release control. In all systems, the end of lag phase and the initiation of propagation were clearly marked by the vanishing of Q and E. The appearance of the formation of polymers coincides with the induction period. It can be concluded that knowing the release profile and antioxidant capacity determine the applicability of microparticles in a lipid systems
Fondecyt

This Master’s thesis is dealing with theoretical analysis of vegetable, mineral oils and synthetic fluids, and with measuring of density and viscosity of selected samples of electric insulating fluids. The main part of the thesis is focused on vegetable oils and their elements called fatty acids. Those are more detail described in separate capitols. In those capitols is described their utilization in engineering practice with focus on energetics, where vegetable oils are used in larger scale. In experimental part of the work is measurement of density and dynamic viscosity. Viscosity was measured on two different machines, first the Hoppler viscometer and on vibrating viscometer. Results of these measurement are evaluated in tables and graphs.

Thesis (M.S.)--University of Wisconsin--Madison, 1985.
Typescript. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 74-81).

Thesis (M.S.)--University of Wisconsin--Madison, 1985.
Typescript. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 76-88).

碩士
輔仁大學
食品營養學系
87
Conjugated linoleic acid (CLA) belongs to a group of octadecadienoic acids that contain two conjugated double bonds. CLA is present in meats and dairy products of ruminants origin, and is stable under various processing and storage conditions. Several studies have shown that CLA possesses anticarcinogenic properties, however, the mechanism is still unclear. Also, the antioxidant activity of CLA remains controversial. Thus, it is necessary to study the antioxidant activity of CLA towards the various model lipids. In addition, the interaction between CLA and model lipids during heating and illumination remains unknown. The objectives of this study are to: (1) investigate the interaction between methyl conjugated linoleate and model lipids during heating and illumination; (2) compare the stability of methyl conjugated linoleate and model lipids during heating and illumination; and (3) evaluate the antioxidant activity of methyl conjugated linoleate towards the model lipids. Results showed that the optimum conditions for the separation of MCLA and model lipids consisted of a splitless injection system with helium as carrier gas at a flow rate of 1.1 mL/min. Oven temperature was programmed at 50 ℃, held for 1 min, then increased to 180 ℃ at a rate of 20 ℃/min, and held for 30 min. Injector temperature was 200 ℃. Detector temperature was 240 ℃. MS showed the highest stability, followed by MO, ML and MCLA. The stability of each sample could be associated with the degree of unsaturation. Model lipids with high degree of saturation or with low temperature treatment was able to retard the degradation of MCLA. In most cases, MCLA either facilitated degradation of model lipids or had no effect. In other cases, MCLA was able to retard the degradation of model lipids during heating. At 100 and 200 ℃ heating, the effect of MCLA on the degradation of model lipids was not pronounced. However, the degradation rate of model lipids increased with increasing MCLA levels at 150 ℃. The peroxide values of model lipids increased with increasing levels of MCLA at low temperature. At high temperature, the peroxide values of model lipids increased initially and then declined afterwards, and, meanwhile, the addition of MCLA did not show significant influence. The peroxide values of samples under light storage were higher than those under heating, and the addition of MCLA did not show significant effect on the peroxide values of model lipids during illumination. In this study MCLA did not show significant antioxidant ability against lipid oxidation.

While replacement of dietary saturated fat with unsaturated fat has been advocated to reduce cardiovascular disease risk, diets high in polyunsaturated fatty acids (PUFA) could increase low density lipoprotein (LDL) susceptibility to oxidation, potentially contributing to the pathology of atherosclerosis. To assess in vivo lipid peroxidation and susceptibility, of LDL surface and core lipids to ex vivo oxidation, in women consuming increased amounts of specific unsaturated fatty acids, 15 postmenopausal women took daily supplements of sunflower oil providing 12.3 g/day of oleate, safflower oil providing 10.5 g/day of linoleate, and fish oil providing 2.0 g/day of eicosapentaenoate (EPA) and 1.4 g/day of docosahexaenoate (DHA) during a crossover trial. Plasma F₂-isoprostanes (F₂-isoP), malondialdehyde (MDA), and thiobarbituric acid reacting substances (TEARS) were measured to assess lipid peroxidation in vivo. Ex vivo oxidation of LDL was monitored by measuring the formation of phosphatidylcholine hydroperoxides (PCOOH) and cholesteryl linoleate hydroperoxides (CE18:200H) during coppermediated oxidation. Plasma free F₂-isoP and MDA concentrations were lower after EPA/DHA supplementation than after oleate (P = 0.001, F₂-isoP and 0.02, MDA) and linoleate supplementation (P = 0.04 for both F₂-isoP and MDA). However, plasma TBARS concentrations were higher after EPA/DHA than after oleate (P = 0.001) and linoleate supplementation (P = 0.0004). During LDL oxidation, the lag phase for PCOOH formation was shorter in EPA/DHA- than oleate- (P = 0.0001) and linoleate-enriched LDL (P = 0.002), while the lag phase for CE18:200H was shorter in EPA/DHA- than oleate- (P = 0.01) but not linoleate-enriched LDL. The maximal rate of PCOOH formation was lower in EPA/ DHA- than linoleate- (P = 0.007) but not oleate-enriched LDL, while the maximal rate of CE18:200H formation was lower in EPA/DHA- than oleate- (P = 0.03) and linoleate-enriched LDL (P [less than or equal to] 0.0001). The maximal concentrations of PCOOH and CE18:200H were lower in EPA/DHA- than oleate- (P [less than or equal to] 0.05) and linoleate-enriched LDL (P [less than or equal to] 0.01). Oleate-enrichment generally decreased the oxidative susceptibility of LDL surface and core lipids, while EPA/DHA-enrichment did not increase LDL oxidative susceptibility compared to linoleate-enrichment. This study emphasizes the need for more than one relevant assay of in vivo lipid peroxidation.
Graduation date: 2000