Le stress oxydatif et la formation des radicaux libres sont impliqués dans plusieurs mécanismes de la mort neuronale qui caractérisent la maladie d'Alzheimer. Les antioxydants sont reconnus comme une source de protection contre le stress oxydatif et peuvent avoir un effet protecteur sur le développement de la maladie d’Alzheimer. Cette étude visait à évaluer le potentiel antioxydant, par le biais du score « oxygen radical absorbance capacity » (ORAC), de l’alimentation habituelle de personnes âgées atteintes de la maladie d’Alzheimer en comparaison avec des témoins appariés pour l’âge sans problèmes cognitifs. L’hypothèse stipulait que les patients atteints de la maladie d’Alzheimer ont une alimentation dont le potentiel antioxydant est inférieur à celui des témoins sans problèmes cognitifs. L’étude a consisté en des analyses secondaires de données provenant de l’étude « Nutrition-Mémoire » (NMS), durant laquelle quarante-deux patients avec une démence du type Alzheimer (DTA) probable et leurs aidants étaient suivis pendant une période de dix-huit mois. Pour la présente étude, les données provenaient de trois jours de collecte alimentaire, ont été colligées au début (T0) de l’étude NMS, selon la méthode « Multiple-Pass ». Le potentiel antioxydant de l’alimentation a été déterminé à l’aide de la description des aliments énumérés dans la base de données des valeurs ORAC de l’USDA. Les résultats de l’étude ont montré que les patients avaient une alimentation dont le potentiel antioxydant était inférieur à celui des témoins (13784,07 ± 7372,70 μmol TE/100g contre 23220,54 ± 10862,55 μmol TE/100g, patients et témoins, respectivement; p<0,0001). Les analyses de régression hiérarchique pas à pas montraient que l’IMC, l’éducation, et le groupe (patients, témoins) étaient des facteurs influençant le score ORAC total. La consommation des aliments riches en antioxydants est un comportement préventif à faible risque qui pourrait bénéficier des individus susceptibles de développer la maladie d'Alzheimer. / Oxidative stress and the formation of free radicals are involved in several mechanisms of neuronal death that are characteristic of Alzheimer's disease. Antioxidants are known to help defend against oxidative stress and may protect against the development of Alzheimer's disease. This study aims to evaluate the antioxidant potential, using the “oxygen radical absorbance capacity” (ORAC) score of the diet of older adults people with Alzheimer's disease compared with cognitively-intact age-matched controls. It was hypothesized that the antioxidant potential of the diet of elderly people with Alzheimer's disease will be lower than that of controls without cognitive problems. The current study is a secondary analysis of data obtained from the "Nutrition-Memory study" (NMS). The NMS study recruited forty-two patients with probable Alzheimer’s disease, and their caregivers, and followed them over a period of eighteen months. The current study focuses on three days of dietary data collection, compiled at the beginning (T0) of the NMS study. The antioxidant potential of the diet was determined using the list of ORAC scores highlighted in the USDA database for the oxygen radical absorbance capacity of selected foods. Our results showed that the diet of patients (13784.07 ± 7372.70 μmol TE/100g) had a lower antioxidant potential than that of controls (± 23220.54 10862.55 μmol TE/100g). Moreover, BMI, education and group-status were factors that influenced the total ORAC score. Eating foods rich in antioxidants is a low risk preventative behaviour that could benefit individuals susceptible to developing Alzheimer’s disease.

This thesis proposes a privacy protection framework for the controlled distribution and use of personal private data. The framework is based on the idea that privacy policies can be set directly by the data owner and can be automatically enforced against the data user. Data privacy continues to be a very important topic, as our dependency on electronic communication maintains its current growth, and private data is shared between multiple devices, users and locations. The growing amount and the ubiquitous availability of personal private data increases the likelihood of data misuse. Early privacy protection techniques, such as anonymous email and payment systems have focused on data avoidance and anonymous use of services. They did not take into account that data sharing cannot be avoided when people participate in electronic communication scenarios that involve social interactions. This leads to a situation where data is shared widely and uncontrollably and in most cases the data owner has no control over further distribution and use of personal private data. Previous efforts to integrate privacy awareness into data processing workflows have focused on the extension of existing access control frameworks with privacy aware functions or have analysed specific individual problems such as the expressiveness of policy languages. So far, very few implementations of integrated privacy protection mechanisms exist and can be studied to prove their effectiveness for privacy protection. Second level issues that stem from practical application of the implemented mechanisms, such as usability, life-time data management and changes in trustworthiness have received very little attention so far, mainly because they require actual implementations to be studied. Most existing privacy protection schemes silently assume that it is the privilege of the data user to define the contract under which personal private data is released. Such an approach simplifies policy management and policy enforcement for the data user, but leaves the data owner with a binary decision to submit or withhold his or her personal data based on the provided policy. We wanted to empower the data owner to express his or her privacy preferences through privacy policies that follow the so-called Owner-Retained Access Control (ORAC) model. ORAC has been proposed by McCollum, et al. as an alternate access control mechanism that leaves the authority over access decisions by the originator of the data. The data owner is given control over the release policy for his or her personal data, and he or she can set permissions or restrictions according to individually perceived trust values. Such a policy needs to be expressed in a coherent way and must allow the deterministic policy evaluation by different entities. The privacy policy also needs to be communicated from the data owner to the data user, so that it can be enforced. Data and policy are stored together as a Protected Data Object that follows the Sticky Policy paradigm as defined by Mont, et al. and others. We developed a unique policy combination approach that takes usability aspects for the creation and maintenance of policies into consideration. Our privacy policy consists of three parts: A Default Policy provides basic privacy protection if no specific rules have been entered by the data owner. An Owner Policy part allows the customisation of the default policy by the data owner. And a so-called Safety Policy guarantees that the data owner cannot specify disadvantageous policies, which, for example, exclude him or her from further access to the private data. The combined evaluation of these three policy-parts yields the necessary access decision. The automatic enforcement of privacy policies in our protection framework is supported by a reference monitor implementation. We started our work with the development of a client-side protection mechanism that allows the enforcement of data-use restrictions after private data has been released to the data user. The client-side enforcement component for data-use policies is based on a modified Java Security Framework. Privacy policies are translated into corresponding Java permissions that can be automatically enforced by the Java Security Manager. When we later extended our work to implement server-side protection mechanisms, we found several drawbacks for the privacy enforcement through the Java Security Framework. We solved this problem by extending our reference monitor design to use Aspect-Oriented Programming (AOP) and the Java Reflection API to intercept data accesses in existing applications and provide a way to enforce data owner-defined privacy policies for business applications. / Im Rahmen der Dissertation wurde ein Framework für die Durchsetzung von Richtlinien zum Schutz privater Daten geschaffen, welches darauf setzt, dass diese Richtlinien oder Policies direkt von den Eigentümern der Daten erstellt werden und automatisiert durchsetzbar sind. Der Schutz privater Daten ist ein sehr wichtiges Thema im Bereich der elektronischen Kommunikation, welches durch die fortschreitende Gerätevernetzung und die Verfügbarkeit und Nutzung privater Daten in Onlinediensten noch an Bedeutung gewinnt. In der Vergangenheit wurden verschiedene Techniken für den Schutz privater Daten entwickelt: so genannte Privacy Enhancing Technologies. Viele dieser Technologien arbeiten nach dem Prinzip der Datensparsamkeit und der Anonymisierung und stehen damit der modernen Netznutzung in Sozialen Medien entgegen. Das führt zu der Situation, dass private Daten umfassend verteilt und genutzt werden, ohne dass der Datenbesitzer gezielte Kontrolle über die Verteilung und Nutzung seiner privaten Daten ausüben kann. Existierende richtlinienbasiert Datenschutztechniken gehen in der Regel davon aus, dass der Nutzer und nicht der Eigentümer der Daten die Richtlinien für den Umgang mit privaten Daten vorgibt. Dieser Ansatz vereinfacht das Management und die Durchsetzung der Zugriffsbeschränkungen für den Datennutzer, lässt dem Datenbesitzer aber nur die Alternative den Richtlinien des Datennutzers zuzustimmen, oder keine Daten weiterzugeben. Es war daher unser Ansatz die Interessen des Datenbesitzers durch die Möglichkeit der Formulierung eigener Richtlinien zu stärken. Das dabei verwendete Modell zur Zugriffskontrolle wird auch als Owner-Retained Access Control (ORAC) bezeichnet und wurde 1990 von McCollum u.a. formuliert. Das Grundprinzip dieses Modells besteht darin, dass die Autorität über Zugriffsentscheidungen stets beim Urheber der Daten verbleibt. Aus diesem Ansatz ergeben sich zwei Herausforderungen. Zum einen muss der Besitzer der Daten, der Data Owner, in die Lage versetzt werden, aussagekräftige und korrekte Richtlinien für den Umgang mit seinen Daten formulieren zu können. Da es sich dabei um normale Computernutzer handelt, muss davon ausgegangen werden, dass diese Personen auch Fehler bei der Richtlinienerstellung machen. Wir haben dieses Problem dadurch gelöst, dass wir die Datenschutzrichtlinien in drei separate Bereiche mit unterschiedlicher Priorität aufteilen. Der Bereich mit der niedrigsten Priorität definiert grundlegende Schutzeigenschaften. Der Dateneigentümer kann diese Eigenschaften durch eigene Regeln mittlerer Priorität überschrieben. Darüber hinaus sorgt ein Bereich mit Sicherheitsrichtlinien hoher Priorität dafür, dass bestimmte Zugriffsrechte immer gewahrt bleiben. Die zweite Herausforderung besteht in der gezielten Kommunikation der Richtlinien und deren Durchsetzung gegenüber dem Datennutzer (auch als Data User bezeichnet). Um die Richtlinien dem Datennutzer bekannt zu machen, verwenden wir so genannte Sticky Policies. Das bedeutet, dass wir die Richtlinien über eine geeignete Kodierung an die zu schützenden Daten anhängen, so dass jederzeit darauf Bezug genommen werden kann und auch bei der Verteilung der Daten die Datenschutzanforderungen der Besitzer erhalten bleiben. Für die Durchsetzung der Richtlinien auf dem System des Datennutzers haben wir zwei verschiedene Ansätze entwickelt. Wir haben einen so genannten Reference Monitor entwickelt, welcher jeglichen Zugriff auf die privaten Daten kontrolliert und anhand der in der Sticky Policy gespeicherten Regeln entscheidet, ob der Datennutzer den Zugriff auf diese Daten erhält oder nicht. Dieser Reference Monitor wurde zum einen als Client-seitigen Lösung implementiert, die auf dem Sicherheitskonzept der Programmiersprache Java aufsetzt. Zum anderen wurde auch eine Lösung für Server entwickelt, welche mit Hilfe der Aspekt-orientierten Programmierung den Zugriff auf bestimmte Methoden eines Programms kontrollieren kann. In dem Client-seitigen Referenzmonitor werden Privacy Policies in Java Permissions übersetzt und automatisiert durch den Java Security Manager gegenüber beliebigen Applikationen durchgesetzt. Da dieser Ansatz beim Zugriff auf Daten mit anderer Privacy Policy den Neustart der Applikation erfordert, wurde für den Server-seitigen Referenzmonitor ein anderer Ansatz gewählt. Mit Hilfe der Java Reflection API und Methoden der Aspektorientierten Programmierung gelang es Datenzugriffe in existierenden Applikationen abzufangen und erst nach Prüfung der Datenschutzrichtlinie den Zugriff zuzulassen oder zu verbieten. Beide Lösungen wurden auf ihre Leistungsfähigkeit getestet und stellen eine Erweiterung der bisher bekannten Techniken zum Schutz privater Daten dar.

Le stress oxydatif et la formation des radicaux libres sont impliqués dans plusieurs mécanismes de la mort neuronale qui caractérisent la maladie d'Alzheimer. Les antioxydants sont reconnus comme une source de protection contre le stress oxydatif et peuvent avoir un effet protecteur sur le développement de la maladie d’Alzheimer. Cette étude visait à évaluer le potentiel antioxydant, par le biais du score « oxygen radical absorbance capacity » (ORAC), de l’alimentation habituelle de personnes âgées atteintes de la maladie d’Alzheimer en comparaison avec des témoins appariés pour l’âge sans problèmes cognitifs. L’hypothèse stipulait que les patients atteints de la maladie d’Alzheimer ont une alimentation dont le potentiel antioxydant est inférieur à celui des témoins sans problèmes cognitifs. L’étude a consisté en des analyses secondaires de données provenant de l’étude « Nutrition-Mémoire » (NMS), durant laquelle quarante-deux patients avec une démence du type Alzheimer (DTA) probable et leurs aidants étaient suivis pendant une période de dix-huit mois. Pour la présente étude, les données provenaient de trois jours de collecte alimentaire, ont été colligées au début (T0) de l’étude NMS, selon la méthode « Multiple-Pass ». Le potentiel antioxydant de l’alimentation a été déterminé à l’aide de la description des aliments énumérés dans la base de données des valeurs ORAC de l’USDA. Les résultats de l’étude ont montré que les patients avaient une alimentation dont le potentiel antioxydant était inférieur à celui des témoins (13784,07 ± 7372,70 μmol TE/100g contre 23220,54 ± 10862,55 μmol TE/100g, patients et témoins, respectivement; p<0,0001). Les analyses de régression hiérarchique pas à pas montraient que l’IMC, l’éducation, et le groupe (patients, témoins) étaient des facteurs influençant le score ORAC total. La consommation des aliments riches en antioxydants est un comportement préventif à faible risque qui pourrait bénéficier des individus susceptibles de développer la maladie d'Alzheimer. / Oxidative stress and the formation of free radicals are involved in several mechanisms of neuronal death that are characteristic of Alzheimer's disease. Antioxidants are known to help defend against oxidative stress and may protect against the development of Alzheimer's disease. This study aims to evaluate the antioxidant potential, using the “oxygen radical absorbance capacity” (ORAC) score of the diet of older adults people with Alzheimer's disease compared with cognitively-intact age-matched controls. It was hypothesized that the antioxidant potential of the diet of elderly people with Alzheimer's disease will be lower than that of controls without cognitive problems. The current study is a secondary analysis of data obtained from the "Nutrition-Memory study" (NMS). The NMS study recruited forty-two patients with probable Alzheimer’s disease, and their caregivers, and followed them over a period of eighteen months. The current study focuses on three days of dietary data collection, compiled at the beginning (T0) of the NMS study. The antioxidant potential of the diet was determined using the list of ORAC scores highlighted in the USDA database for the oxygen radical absorbance capacity of selected foods. Our results showed that the diet of patients (13784.07 ± 7372.70 μmol TE/100g) had a lower antioxidant potential than that of controls (± 23220.54 10862.55 μmol TE/100g). Moreover, BMI, education and group-status were factors that influenced the total ORAC score. Eating foods rich in antioxidants is a low risk preventative behaviour that could benefit individuals susceptible to developing Alzheimer’s disease.

Orientador: Flavia Maria Netto / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-19T21:19:52Z (GMT). No. of bitstreams: 1 Silva_FernandaGuimaraesDrummonde_M.pdf: 1363127 bytes, checksum: 68b1a97c95798f4425019ee900814160 (MD5) Previous issue date: 2012 / Resumo: Existem evidências numerosas sobre o papel dos radicais livres em uma série de condições patológicas, incluindo envelhecimento, câncer, esclerose múltipla, doenças cardiovasculares. Hidrolisados protéicos de diferentes fontes têm sido estudados por seu potencial antioxidante. A atuação antioxidante da proteína, na maioria das vezes, encontra-se limitada devido à conformação espacial, que concentra resíduos capazes de neutralizar radicais livres no interior da molécula, dificultando o acesso das espécies reativas aos centros nucleofílicos. A hidrólise da proteína contribui para aumentar a exposição desses resíduos de aminoácidos, aumentando sua atuação como antioxidante. Compostos fenólicos podem estar presentes em hidrolisados proteicos de origem vegetal, devido a sua associação com as proteínas. Métodos in vitro que simulam as condições do trato gastrointestinal permitem estudar como a digestão pode interferir na atividade antioxidante de peptídeos e compostos fenólicos. O presente trabalho tem como objetivos obter hidrolisados proteicos com capacidade antioxidante a partir da farinha de linhaça e avaliar o efeito da digestão in vitro pode interferir nessa atividade. A farinha de linhaça marrom foi desengordurada, obtendo-se a farinha de linhaça marrom desengordurada (FLMD). O concentrado proteico de linhaça (CPL) foi obtido a partir da FLMD por extração alcalina e precipitação no ponto isoelétrico seguida de neutralização. Para obtenção dos hidrolisados proteicos (HPL), a partir do CPL, com Alcalase, foi realizado um delineamento composto central rotacional (DCCR) 2². As variáveis independentes foram pH que variou entre 7,5 a 9,5 e relação enzima: substrato (E:S) que variou de 1:150 a 1:30. As variáveis dependentes foram grau de hidrólise (GH), teor de substâncias redutoras do reagente de Folin-Ciocalteau e atividade antioxidante, determinada por FRAP e ORAC. Teor de substâncias redutoras e atividade antioxidante foram avaliados a partir dos extratos aquosos e metanólico (metanol 70%). Os hidrolisados de maior atividade antioxidante, a FLMD e o CPL foram submetidos à digestão in vitro, simulando as condições da digestão gastrintestinal. As amostras antes e após a digestão in vitro foram caracterizadas por eletroforese em sistema SDS-PAGE Tricina e por cromatografia liquida de alta eficiência de fase reversa (HPLC- RP). O teor de substâncias redutoras e da atividade antioxidante das amostras FLMD, CPL e HPL foram avaliados antes e após a digestão in vitro. As condições ótimas para obtenção de HPL de maior GH (21,0%) são pH entre 7,5 e 8,0 e E:S entre 1:60 e 1:30, indicando que a faixa de pH ótimo da enzima e a alta E:S favorecem maior hidrólise do CPL. Para obtenção de HPL com maior teor de substâncias redutoras para os extratos aquoso (24 mg EAG/ g HPL) e metanólico (20 mg EAG/ g HPL) as condições ótimas são pH ~ 8,5 /E:S 1:30. Este resultado parece estar relacionado à liberação de compostos fenólicos ligados a proteína e também de peptídeos durante a hidrólise. Açúcares e aminoácidos aromáticos presentes no hidrolisado podem interferir na reação e superestimar o teor de fenóis dos HPL. A maior atividade antioxidante determinada pelo método de FRAP para o extrato aquoso (42 mg SF/ g HPL) se dá nas condições de pH ~ 9,5/E:S ~1:150 e para o extrato metanólico (40 mg SF/ g HPL) pH entre 8,5 e 9,0/E:S entre 1:90 a 1:150. Para o método de ORAC, as condições ótimas para maior atividade antioxidante no extrato aquoso (300 µmol TE/ g HPL) são pH entre 7,5 a 9,5/E:S ~ 1:30 ou ~1:150 e para o extrato metanólico (330 µmol TE/ g HPL) são pH ~ 8,5/E:S entre 1:150 e 1:30. Os hidrolisados de maior atividade antioxidante foram os obtidos em pH 8,5/E:S 1:90, e em pH 9,2/E:S 1:133 denominados HPL 0 e HPL 3, respectivamente. Para a FLMD, CPL e os hidrolisados, após a digestão in vitro, observou-se que o teor de substâncias redutoras totais aumentou (9 a 20 vezes) para todas as amostras. O teor de substâncias redutoras do CPL (~24 mg EAG/ g amostra), em ambos os extratos, após a digestão in vitro se igualou ao teor dos hidrolisados (~23 mg EAG/ g amostra). Este resultado sugere que tanto a hidrólise com Alcalase quanto o processo digestório liberam compostos redutores, dentre eles fenólicos da proteína de linhaça. A atividade antioxidante dos extratos de FLMD e CPL, determinada por FRAP, também aumentou (de 3 a 10 vezes) após a digestão, mas não se igualou à atividade antioxidante dos hidrolisados (48 mg SF/g amostra). No entanto, o CPL apresentou atividade antioxidante determinada por ORAC semelhante à dos hidrolisados no extrato aquoso (~420,24 µmol TE/ g amostra) e 10 % maior que o encontrado para os hidrolisados (~365 µmol TE/ g amostra) no extrato metanólico. Após a digestão in vitro, os hidrolisados apresentaram a maior atividade antioxidante medida por FRAP (50 mg SF/ g amostra), e o CPL, a maior atividade determinada pelo método de ORAC (~430 µmol TE/ g amostra). Estes resultados sugerem o processo digestório é tão ou mais eficiente que a Alcalase em liberar os compostos com atividade redutora no CPL. Uma vez que a metodologia de determinação da atividade antioxidante por ORAC tem maior proximidade com o mecanismo de oxirredução que ocorre in vivo, esses resultados sugerem o uso do CPL como melhor produto protéico da linhaça com maior potencial antioxidante para a formulação de nutracêuticos e alimentos funcionais / Abstract: There are several evidences which indicate the role of free radicals on a series of pathological conditions, including aging, cancer, multiple sclerosis and cardiovascular disease. Hydrolysates from different sources have been studied because of their antioxidant potential. The antioxidant activity of the protein, in most cases, is limited due to their conformation, which concentrates residues capable of neutralize free radicals in the molecule¿s core, hampering the access of the reactive species to nucleophilic sites. The protein hydrolysis contributes to increasing the exposure of these amino acid residues, increasing their role as antioxidants. Phenolic compounds may also be present in vegetable protein hydrolysates because of their association with proteins. In vitro methods that simulate the conditions of the gastrointestinal digestion are an important way to evaluate how the digestion affects the antioxidant activity of phenolic compounds and peptides. This study aims at obtaining hydrolysates with antioxidant capacity from defatted flaxseed flour and evaluate the effect of the in vitro digestion on this activity. The brown flaxseed flour was defatted, resulting in the brown defatted flaxseed meal (BDFM). The flaxseed protein concentrate (FPC) was obtained from the BDFM by alkaline extraction and precipitation at the isoelectric point followed by neutralization. To obtain the flaxseed protein hydrolysates (FPL), using FPC and Alcalase, a central composite rotational design (DCCR) was performed. The independent variables were pH ranging from 7.5 to 9.5 and enzyme: substrate ratio (E: S) that ranged from 1:150 to 1:30. The dependent variables were the degree of hydrolysis (DH), total phenolic content and antioxidant activity, determined by FRAP and ORAC. Phenolic and antioxidant activity were evaluated from the aqueous and methanol (70% methanol). The hydrolysates with the highest antioxidant activity, the CPL FLMD were submitted to the in vitro digestion. The samples obtained before and after the in vitro digestion were characterized by electrophoresis SDS-PAGE- tricine and HPLC. The total phenolic content and antioxidant activity of FLMD, CPL and HPL were evaluated before and after in vitro digestion. The optimum conditions to obtain HPL with the highest GDH (21.0%) are pH (7.5-8) and E:S ratio (1:60-1:30), which indicates that the Alcalase optimum pH and highest E:S ratio collaborates to highest hydrolysis of CPL. To obtain HPL with higher content of Folin-Ciocalteau reducing compounds content in aqueous (EAG 24 mg / g HPL) and methanol (20 mg EAG / g HPL) extracts, the optimum conditions were pH ~ 8.5 / E: S 1:30. This result seems to be related to the release of phenolic compounds bound to protein and also of peptides during hydrolysis. The highest antioxidant activity determined by the FRAP method in the aqueous extract (42 mg SF / g HPL) occurs under pH ~ 9.5 / E: S ~ 1:150 and the methanol extract (40 mg SF / g HPL) pH 8.5-9.0 / E: S 1:90-1:150. For the ORAC method, optimum conditions for increased antioxidant activity in aqueous extract (300 µmol TE / g HPL) are pH 7.5-9.5 / E: S ~ 1:30 or 1:150 and the methanol extract (330 µmol TE / g HPL) are pH ~ 8.5 / E: S 1:30-1:150. The hydrolysates with the highest antioxidant activities were obtained at pH 8.5 / E: S 1:90, and at pH 9.2 / E: S 1:133 denominated HPL ) and HPL 3, respectively. For FLMD, CPL and hydrolysates, after in vitro digestion, the content increased (9-20 times) for all samples. The Folin-Ciocalteau reducing capacity of the CPL (EAG ~ 24 mg / g sample) in both extracts after in vitro digestion equaled the content of hydrolysates (EAG ~ 23 mg / g sample). This result suggests that both hydrolysis with Alcalase and the digestion process are able to release phenolic compounds from the flaxseed products. The antioxidant activity of extracts of FLMD, CPL determined by FRAP, also increased (from 3 to 10 times) after digestion, but did not reached the antioxidant activity of hydrolysates (48 mg SF / g sample). However, when the activity was determined by ORAC, the FPC showed value similar to the hydrolysates, measured on the aqueous extract (~ 420.24 µmol TE / g sample) and 10% higher than on the methanol extract (~ 365 µmol TE / g sample). After in vitro digestion, hydrolysates showed the highest antioxidant activity measured by FRAP (SF 50 mg / g sample), and the FPC, the highest activity determined by ORAC method (~ 430 micromol TE / g sample). These results suggest that digestive process are equally or more effective than Alcalase in releasing peptides and phenolic compounds present in the FPC. Since the methodology for determining the antioxidant activity by ORAC utilizes a biologically relevant radical source, these results suggest the use of FPC as the best protein product of flaxseed with potential antioxidant in the formulation of nutraceuticals and functional foods / Mestrado / Nutrição Experimental e Aplicada à Tecnologia de Alimentos / Mestre em Alimentos e Nutrição

La prevalencia de los trastornos metabólicos está creciendo a nivel mundial. Botánicos, como el romero (Rosmarinus officinalis L.) y las semillas de fresno (Fraxinus excelsior L.), pueden ser una alternativa para mejorar estos trastornos. El objetivo de esta tesis doctoral fue generar extractos industriales de las partes comestibles de estas plantas, identificar sus compuestos por HPLC, HPLC-MS y RMN, y determinar su eficacia y seguridad. Se desarrollaron tres extractos de romero - estandarizados al 20% de ácido carnósico, 20% de ácido rosmarínico, y 40% de ácido ursólico. El extracto rico en ácido ursólico mostró la menor capacidad antioxidante en modelos ORAC y FRAP in vitro, mientras que los otros dos extractos tuvieron capacidades más elevadas. Sin embargo, el extracto rico en ácido carnósico (RE) fue superior en la inhibición de la oxidación de LDL ex vivo, y fue el único seleccionado para estudios posteriores. Se desarrolló un extracto de semilla de fresno (FE) de acuerdo a su uso tradicional en Marruecos, siendo identificados salidrosido y 9 glucósidos secoiridoides, dos de ellos fueron descubiertos por primera vez: Excelside A y Excelside B. En estudios in vitro, RE y FE activaron receptores nucleares que regulan la homeostasis de la glucosa y la energía, y actuaron sobre mecanismos contra la obesidad y la dislipidemia - RE activa PPAR? e inhibe la lipasa pancreática, y FE estimula PPAR? y evita la diferenciación de preadipocitos. La eficacia de ambos extractos para el control de trastornos metabólicos fue confirmada en ratones C57BL/6J administrados con una dieta baja en grasa, una dieta alta en grasa, o una dieta alta en grasa más 0.5% RE (HFD.RE) o 0.5% FE (FED) durante 16 semanas. HFD.RE limitó el aumento del peso corporal y grasa en el epidídimo en un 69% (P<0.01) y 79% (P<0.001), respectivamente. HFD.RE también redujo la glucemia (72%, P<0.01) y los niveles de colesterol (68%, P<0.001). FED redujo la glucemia (76.52%, P<0.001), insulinemia (53.43%, P< / Ibarra, SA. (2011). OBTENTION AND CHARACTERIZATION OF ROSEMARY AND ASH TREE SEED EXTRACTS AND STUDY OF THEIR PREVENTIVE EFFECTS ON METABOLIC DISORDERS [Tesis doctoral no publicada]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/10795 / Palancia

Masters of Science / The aim of this study was therefore to evaluate the antioxidant and anti-diabesity potential of a hot water extract of C. maculata in non-cell based assays and correlate the activities with phenolic composition. Total antioxidant capacity (TAC) was assessed in terms of free radical scavenging and iron reducing ability. The DPPH, ABTS, ORAC and FRAP assays were employed. Anti-diabesity potential was assessed in terms of the inhibition of the digestive enzymes, α-glucosidase and pancreatic lipase

The evolution of aerobic metabolic processes unavoidably led to the production of reactive oxygen species (ROS). ROS have the ability to cause harmful oxidative damage to biomolecules. Increased ROS generation and subsequent oxidative stress have been associated with aging and neurodegenerative disorders such as Parkinson’s and Alzheimer’s diseases as a result of the extreme sensitivity of the central nervous system to damage from ROS. Antioxidant defence systems have co–evolved with aerobic metabolic processes to counteract oxidative damage inflicted by ROS. The impact of neurodegenerative disorders on society is increasing rapidly as the life expectancy of the global population increases. In this day and age, a much younger group of the population is also experiencing neurodegenerative symptoms as a result of the harmful effect of the human immunodeficiency virus (HIV) on the central nervous system. Plants are an invaluable source of medicinal compounds. The use of plants for their healing properties is rooted in ancient times. The aim of this study was to select from twenty one plants, the plant with the most promising antioxidant activity and to determine whether extracts of this plant could act as free radical scavengers, comparing the results to Trolox, a known free radical scavenger. The next step was to isolate and characterize a compound from an extract exhibiting promising antioxidant activity. Bioassay–guided fractionation was followed to achieve this. During screening trials, twenty one plants, namely Berula erecta, Heteromorpha arborescens, Tarchonanthus camphoratus, Vernonia oligocephala, Gymnosporia buxifolia, Acacia karroo, Elephantorrhiza elephantina, Erythrina zeyheri, Leonotis leonurus, Plectranthus ecklonii, P. rehmanii, P. venteri, Salvia auretia, S. runcinata, Solenostemon latifolius, S. rotundifolius, Plumbago auriculata, Clematis brachiata, Vangueria infausta, Physalis peruviana and Lippia javanica were selected from literature, based on reported antioxidant activity within the plant families, for screening of their antioxidant activity. One hundred and ten extracts were prepared from the leaves, using Soxhlet extraction and the solvents petroleum ether (PE), dichloromethane (DCM), ethyl acetate (EtOAc) and ethanol (EtOH), consecutively. The focus during initial screening trials was on chemistry–based assays. The oxygen radical absorbance capacity (ORAC) and ferric reducing antioxidant power (FRAP) assays were employed for the primary screening of the one hundred and ten leaf extracts. The ORAC assay was used to determine whether the plant extracts were able to scavenge peroxyl radicals and the FRAP assay was used to determine the reducing abilities of the extracts. Quantification of the peroxyl radical scavenging activity by the ORAC assay revealed that activity was observed for most of the extracts, with the ethyl acetate and ethanol extracts of L. javanica exhibiting the most promising activity. This pattern of activity was also found with the reducing capacity evaluated by the FRAP assay in which the EtOAc and EtOH extracts of L. javanica also exhibited the most promising activity. L. javanica was selected for further study by screening for biological activity, employing the nitro–blue tetrazolium (NBT) assay and thiobarbituric acid reactive substances (TBARS) assay. Using a cyanide model to induce neurotoxic effects in rat brain homogenate, the neuroprotective properties of the extracts of L. javanica leaves were examined using the NBT assay and compared to that of Trolox. The NBT assay determines the level of superoxide anions. All the extracts of L. javanica significantly reduced superoxide anion generation at all concentrations used. The petroleum ether and ethyl acetate extracts, at all concentrations, reduced superoxide anion generation to values lower than that of the control, suggesting that these extracts may be able to attenuate normal free radical processes in the brain. The petroleum ether extract exhibited the most promising activity at a concentration of 1.25 and 2.5 mg/ml and also exhibited similar results as the ethyl acetate extract at a lower concentration than the ethyl acetate extract (2.5 mg/ml compared to 5 mg/ml). A toxin–solution consisting of hydrogen peroxide (H2O2), iron(III)chloride (FeCl3) and ascorbic acid was used to induce lipid peroxidation and the ability of the extracts of the leaves of L. javanica to attenuate lipid peroxidation was investigated in rat brain homogenate and compared to that of Trolox. All of the extracts of L. javanica significantly attenuated toxininduced lipid peroxidation at all concentrations used. All of the extracts were also able to significantly attenuate toxin–induced lipid peroxidation to values lower than that of the control. These results suggest that all of the extracts of L. javanica possess the ability to attenuate not only toxin–induced lipid peroxidation, but also lipid peroxidation that occurs during normal processes in the brain. The petroleum ether extract was subjected to bioassay–guided fractionation using column and thin–layer chromatography and the NBT and TBARS assays. Fraction DD1 was investigated by means of nuclear magnetic resonance, infrared and mass spectrometry. The exact structure of fraction DD1 was not elucidated. Considering all the results, it is clear that L. javanica shows great potential as a medicinal plant with antioxidant activity and may therefore be beneficial in diminishing the destructive oxidative effects inflicted by free radicals. There are however still many compounds to be isolated from L. javanica. Key words: Verbenaceae, Lippia javanica, antioxidant, neurodegeneration, oxygen radical absorbance capacity (ORAC), ferric reducing antioxidant power (FRAP), nitro–blue tetrazolium assay (NBT), thiobarbituric acid reactive substances assay (TBARS). / Thesis (M.Sc. (Pharmaceutical Chemistry))--North-West University, Potchefstroom Campus, 2011.

The evolution of aerobic metabolic processes unavoidably led to the production of reactive oxygen species (ROS). ROS have the ability to cause harmful oxidative damage to biomolecules. Increased ROS generation and subsequent oxidative stress have been associated with aging and neurodegenerative disorders such as Parkinson’s and Alzheimer’s diseases as a result of the extreme sensitivity of the central nervous system to damage from ROS. Antioxidant defence systems have co–evolved with aerobic metabolic processes to counteract oxidative damage inflicted by ROS. The impact of neurodegenerative disorders on society is increasing rapidly as the life expectancy of the global population increases. In this day and age, a much younger group of the population is also experiencing neurodegenerative symptoms as a result of the harmful effect of the human immunodeficiency virus (HIV) on the central nervous system. Plants are an invaluable source of medicinal compounds. The use of plants for their healing properties is rooted in ancient times. The aim of this study was to select from twenty one plants, the plant with the most promising antioxidant activity and to determine whether extracts of this plant could act as free radical scavengers, comparing the results to Trolox, a known free radical scavenger. The next step was to isolate and characterize a compound from an extract exhibiting promising antioxidant activity. Bioassay–guided fractionation was followed to achieve this. During screening trials, twenty one plants, namely Berula erecta, Heteromorpha arborescens, Tarchonanthus camphoratus, Vernonia oligocephala, Gymnosporia buxifolia, Acacia karroo, Elephantorrhiza elephantina, Erythrina zeyheri, Leonotis leonurus, Plectranthus ecklonii, P. rehmanii, P. venteri, Salvia auretia, S. runcinata, Solenostemon latifolius, S. rotundifolius, Plumbago auriculata, Clematis brachiata, Vangueria infausta, Physalis peruviana and Lippia javanica were selected from literature, based on reported antioxidant activity within the plant families, for screening of their antioxidant activity. One hundred and ten extracts were prepared from the leaves, using Soxhlet extraction and the solvents petroleum ether (PE), dichloromethane (DCM), ethyl acetate (EtOAc) and ethanol (EtOH), consecutively. The focus during initial screening trials was on chemistry–based assays. The oxygen radical absorbance capacity (ORAC) and ferric reducing antioxidant power (FRAP) assays were employed for the primary screening of the one hundred and ten leaf extracts. The ORAC assay was used to determine whether the plant extracts were able to scavenge peroxyl radicals and the FRAP assay was used to determine the reducing abilities of the extracts. Quantification of the peroxyl radical scavenging activity by the ORAC assay revealed that activity was observed for most of the extracts, with the ethyl acetate and ethanol extracts of L. javanica exhibiting the most promising activity. This pattern of activity was also found with the reducing capacity evaluated by the FRAP assay in which the EtOAc and EtOH extracts of L. javanica also exhibited the most promising activity. L. javanica was selected for further study by screening for biological activity, employing the nitro–blue tetrazolium (NBT) assay and thiobarbituric acid reactive substances (TBARS) assay. Using a cyanide model to induce neurotoxic effects in rat brain homogenate, the neuroprotective properties of the extracts of L. javanica leaves were examined using the NBT assay and compared to that of Trolox. The NBT assay determines the level of superoxide anions. All the extracts of L. javanica significantly reduced superoxide anion generation at all concentrations used. The petroleum ether and ethyl acetate extracts, at all concentrations, reduced superoxide anion generation to values lower than that of the control, suggesting that these extracts may be able to attenuate normal free radical processes in the brain. The petroleum ether extract exhibited the most promising activity at a concentration of 1.25 and 2.5 mg/ml and also exhibited similar results as the ethyl acetate extract at a lower concentration than the ethyl acetate extract (2.5 mg/ml compared to 5 mg/ml). A toxin–solution consisting of hydrogen peroxide (H2O2), iron(III)chloride (FeCl3) and ascorbic acid was used to induce lipid peroxidation and the ability of the extracts of the leaves of L. javanica to attenuate lipid peroxidation was investigated in rat brain homogenate and compared to that of Trolox. All of the extracts of L. javanica significantly attenuated toxininduced lipid peroxidation at all concentrations used. All of the extracts were also able to significantly attenuate toxin–induced lipid peroxidation to values lower than that of the control. These results suggest that all of the extracts of L. javanica possess the ability to attenuate not only toxin–induced lipid peroxidation, but also lipid peroxidation that occurs during normal processes in the brain. The petroleum ether extract was subjected to bioassay–guided fractionation using column and thin–layer chromatography and the NBT and TBARS assays. Fraction DD1 was investigated by means of nuclear magnetic resonance, infrared and mass spectrometry. The exact structure of fraction DD1 was not elucidated. Considering all the results, it is clear that L. javanica shows great potential as a medicinal plant with antioxidant activity and may therefore be beneficial in diminishing the destructive oxidative effects inflicted by free radicals. There are however still many compounds to be isolated from L. javanica. Key words: Verbenaceae, Lippia javanica, antioxidant, neurodegeneration, oxygen radical absorbance capacity (ORAC), ferric reducing antioxidant power (FRAP), nitro–blue tetrazolium assay (NBT), thiobarbituric acid reactive substances assay (TBARS). / Thesis (M.Sc. (Pharmaceutical Chemistry))--North-West University, Potchefstroom Campus, 2011.

Ce travail de thèse s'attache à identifier des plantes et/ou molécules à activités cytotoxique, antioxydante, anti-inflammatoire et antivirale ciblant le virus du chikungunya (CHIKV) dans le but de trouver des alternatives thérapeutiques vis-à-vis du stress oxydatif et de l'inflammation, mécanismes impliqués dans les maladies chroniques non transmissibles (diabète, obésité…), et de la maladie du chikungunya, maladie vectorielle réémergente. La première partie de ces travaux présente les résultats obtenus lors d'un criblage d'activités biologiques réalisé sur une sélection de dix-huit plantes endémiques et indigènes de La Réunion. Les activités ciblées ont été les activités cytotoxiques sur une lignée cellulaire humaine (cellules THP-1), les activités antioxydantes évaluées par un test in cellulo d'hémolyse et par quatre tests chimiques (TEAC/DPPH/FRAP/ORAC) ainsi qu'une évaluation de la teneur en composés phénoliques (test FOLIN) et les activités anti-inflammatoires testées sur des macrophages murins (cellules RAW-BlueTM). Les résultats obtenus ont permis de mettre, plus particulièrement, en évidence les activités de différents extraits : cytotoxique pour Carissa spinarum, antioxydantes pour Agarista buxifolia et Dryopteris wallichiana et anti-inflammatoire pour Stillingia lineata et Indigofera ammoxylum. La deuxième partie du travail est consacrée à l'étude phytochimique d'une espèce indigène de La Réunion, Stillingia lineata, choisie en raison des résultats obtenus lors de ce criblage biologique préliminaire et de ceux du programme Phytochik. Un fractionnement bioguidé par un test antiviral, réalisé sur des cellules Vero (cellules rénales de singe vert Cercopithecus aethiops) contaminées par le CHIKV, a conduit à l'isolement de trois macrocycles diterpéniques rares de type tonantzitlolone dont l'un présente une structure non caractérisée jusque-là, et d'un pimarane de structure nouvelle. La 4'-acétoxytonantzitlolone a été identifiée comme molécule candidate contre le CHIKV (CE50 = 7 μM). Des relations structure-activité ont pu être définies ; la présence d'un groupement oxygéné sur la chaîne latérale des tonantzitlolones semble jouer un rôle important sur la réponse antivirale de ces squelettes diterpéniques. / The aims of this PhD work were to identify plants and/or molecules with cytotoxic, antioxidant, anti-inflammatory or antiviral (chikungunya virus , CHIKV) activities in order to find therapeutic alternatives towards oxidative stress and inflammation, mechanisms involved in chronic noncommunicable diseases (diabetes, obesity ...), and chikungunya disease, reemerging vector-borne disease. The first part of this work presents the results obtained from a biological screening carried out on a selection of eighteen endemic and indigenous plants of La Réunion. The targeted activities were cytotoxicity on a human cell line (THP-1), antioxidant activities evaluated using an in cellulo hemolysis assay and four chemical tests (TEAC / DPPH / FRAP / ORAC) together with an evaluation of the content of phenolic compounds (FOLIN test) and anti-inflammatory activity tested in murine macrophages (RAW cells-BlueTM). The results allowed to highlight activities of different extracts in particular : cytotoxic for Carissa spinarum, antioxidant for Dryopteris wallichiana and Agarista buxifolia and anti-inflammatory for Stillingia lineata and Indigofera ammoxylum.The second part of this work is devoted to the phytochemical study of Stillingia lineata, an indigenous species of La Réunion chosen because of the results obtained in this preliminary biological screening and those carried out in Phytochik programme. Bioassay-guided fractionation performed on Vero cells (green monkey kidney cells Cercopithecus aethiops) infected with CHIKV led to the isolation of three rare macrocycle-type diterpenes called tonantzitlolone and a new pimarane. The 4'-acetoxytonantzitlolone was identified as a candidate molecule against CHIKV (EC50 = 7 μM). Structure-activity relationships have been defined, the presence of an oxygenated group on the side chain of tonantzitlolones seems to play an important role in the antiviral response of the diterpene skeleton.