No / The skin produces bioactive lipids that participate in physiological and pathological states, including homeostasis, induction, propagation, and resolution of inflammation. However, comprehension of the cutaneous lipid complement, and contribution to differing roles of the epidermal and dermal compartments, remains incomplete. We assessed the profiles of eicosanoids, endocannabinoids, N-acyl ethanolamides, and sphingolipids, in human dermis, epidermis, and suction blister fluid. We identified 18 prostanoids, 12 hydroxy-fatty acids, 9 endocannabinoids and N-acyl ethanolamides, and 21 non-hydroxylated ceramides and sphingoid bases, several demonstrating significantly different expression in the tissues assayed. The array of dermal and epidermal fatty acids was reflected in the lipid mediators produced, whereas similarities between lipid profiles in blister fluid and epidermis indicated a primarily epidermal origin of suction blister fluid. Supplementation with omega-3 fatty acids ex vivo showed that their action is mediated through perturbation of existing species and formation of other anti-inflammatory lipids. These findings demonstrate the diversity of lipid mediators involved in maintaining tissue homeostasis in resting skin and hint at their contribution to signaling, cross-support, and functions of different skin compartments. Profiling lipid mediators in biopsies and suction blister fluid can support studies investigating cutaneous inflammatory responses, dietary manipulation, and skin diseases lacking biomarkers and therapeutic targets.

Les peptides Albumines 1 b sont des membres de la famille structurale des knottines et présentent un potentiel intéressant en tant que composés insecticides. À ce jour, leur diversité parmi les Fabacées a été essentiellement étudiée en utilisant des approches biochimiques et moléculaires. Les ressources bioinformatiques (le séquençage complet du génome, les bases de données transcriptomiques (EST…), les atlas d’expression...) de l’espèce modèle des légumineuses, Medicago truncatula Gaertn. (Mtr), nous a permis de développer une approche génomique de cette biodiversité. Deux objectifs principaux nous ont guidé à : 1) déchiffrer l'histoire évolutive de la famille A1 dans cette espèce et 2) explorer la biodiversité naturelle afin de découvrir de nouvelles molécules bioactives. L'exploration du génome de Mtr a révélé une remarquable expansion, à travers des duplications en tandem, des loci A1 qui retiennent presque toutes la même structure génique canonique (2 exons et 1 intron). L'analyse phylogénétique nous a permis de comprendre l’évolution des gènes A1 intraspécifique et l’analyse de leur expression (EST, puces à ADN) a révélé la distribution de la famille des gènes A1 dans les organes de la plante (tissus) : Cette dernière s’est révélée bien plus diverse que celle connue chez les autres espèces examinées de légumineuses, où la famille était jusque-là principalement graine-spécifique. Selon plusieurs critères, certains peptides ont été sélectionnés puis chimiquement synthétisés et repliés in vitro et testés pour leur activité biologique. Parmi eux, un peptide, nommé AG41, isoforme MtrA1013 et issu de l’EST orpheline TA24778_3880, a révélé un pouvoir insecticide élevé et inattendu. L’analyse à grande échelle en présence d’homologues d’A1 des légumineuses a montré l’ancestralité de la fonction insecticide et l’âge de la famille est estimé à plus de 58 million d’années. Notre étude s’est aussi orientée vers l’analyse de cette famille peptidique chez des légumineuses originaires du Liban. Cette approche par biologie moléculaire nous a permis de caractériser 9 nouveaux gènes chez 6 espèces de Papilionoideae. L’étude plus approfondie de ces gènes au niveau structural et fonctionnel est envisagée. Afin de relier les variations de structure et d'activité, un système d'expression hétérologue (baculovirus/cellules d’insectes Sf9) a été mis au point. Le peptide recombinant de référence PA1b (Pea Albumin 1 sub-unit b), même exprimé en faible quantité, présente une activité biologique et une masse bien conforme ainsi qu’une structure bien repliée. Ce système a permis, de même, de produire la proprotéine PA1, forme intermédiaire entre la préproprotéine et le peptide PA1b mature. Cette proprotéine, identifiée pour la première fois, ne présente aucune toxicité envers les cellules Sf9. / Albumin 1 b peptides are members of the knottin structural family and display an interesting potential as insecticidal compounds. To date their diversity among Fabaceae was essentially investigated using biochemical and molecular approaches. The bioinformatic resources (full-genome sequencing, EST database, gene expression atlas…) of the Legume model, Medicago truncatula Gaertn. (Mtr), prompted us to develop a large-scale approach in two ways: 1) to decipher the evolutionary history of A1 family in this species and 2) to explore the natural biodiversity to uncover new bioactive molecules. Exploring Mtr genome revealed a remarkable expansion, through tandem duplications, of A1 loci that retain nearly all the primary structure (2 exons and 1 intron) . Phylogenetic analysis has allowed us to understand the evolution of intraspecific A1 genes and the analysis of their expression (EST, microarrays), and revealed the distribution of the A1 gene family in plant organs (tissue): the latter proved to be much more diverse than that seen in other examined legumes species, where the family until then was mainly seed-specific. Selected upon several criteria some peptides were chemically synthezised, folded in vitro and assayed for their biological activity. Among them one peptide, named AG41: isoform MtrA1013 (orphan EST : TA24778_3880), revealed a high and unexpected insecticidal power. The large-scale analysis in the presence of legumes A1 homologous showed the ancestry of the isecticidal function and the age of this family is estimated to be more than 58 million years. Our study is also directed towards the analysis of this family of peptide in legumes from Lebanon. This approach based on molecular biology has allowed us to characterize nine new genes in six species of Papilionoideae. The further study of these genes at the structural and functional level is considered. To link changes in structure and activity, a heterologous expression system (baculovirus / insect Sf9 cells) was developed. The reference recombinant peptide PA1b (Pea Albumin 1 sub-unit b), even expressed in small quantities, was biologicaly active and harbouring the expected mass as well as a well-folded structure. This system has enabled also to produce the proprotein PA1, intermediate form between the preproprotein and the mature peptide PA1b. This proprotein, identified for the first time, has no toxicity towards Sf9 cells.

[Truncated abstract] Antimicrobial growth promoters are added to feed to prevent lactic acidosis in ruminant animals by selectively inhibiting rumen bacteria that produce lactic acid. However, recently imposed or impending bans on the use of antimicrobial growth promoters in animal production have lead to a critical need to find practical alternatives that are safe for the animal and consumer and that obtain similar production benefits. I investigated bioactive plants of Australia for their potential to prevent lactic acidosis in ruminants. The unifying hypothesis tested was that plants would be identified that selectively inhibit lactic acid-producing bacteria and consequently protect against lactic acidosis. This hypothesis was tested in a three phase process: phase 1, plant selection and collection; phase 2, a three stage protocol for screening plants and essential oils; phase 3, in vivo experiments and chemical fractionation of the most promising plant. I developed an in vitro bioassay that simulated acidosis by adding glucose to rumen fluid in Bellco tubes and incubating for 5 h (Chapter 4). The pH and gas production were used as indicators of acidosis and fermentation activity. I used this bioassay to screen ninety-five plants (dried and ground material from 79 species) and ten essential oils and included a negative control (oaten chaff) and a positive control (virginiamycin). One plant, Eremophila glabra, produced a similar pH (5.63) to the positive control (5.43) although it inhibited gas production to a moderate extent (P < 0.05). ... Seven serrulatane diterpenes were identified to be the major secondary metabolites in E. glabra. The metabolites were screened using a broth dilution and microtitre spectrophotometry method and were selective against S. bovis at between 320 and 1077 [mu]g/ mL. The serrulatanes from E. glabra were probably responsible for the activity against acidosis that I observed in vitro, because they selectively inhibited lactateproducing bacteria. It is also possible that a synergy between serrulatanes and possibly other metabolites are responsible for the activity observed in vitro. The results from my experiments support the role that bioactive plants may have to replace the antibiotics that are added to livestock feed. Australian plants were identified containing compounds that were active against the bacterial processes responsible for ruminant acidosis. To my knowledge this is the first work undertaken to identify bioactive plants of Australia for their potential to prevent acidosis. I developed in vitro screening bioassays that targeted key indicators of acidosis. These bioassays enabled me to identify 5 plants from the 104 screened that could potentially control acidosis. One of these plants in particular, E. glabra, showed a level of activity in vitro that was comparable to antibiotic protection against acidosis. The exciting in vitro results were not demonstrated in vivo but only one dose level of E. glabra was used, which was based on the in vitro work. In contrast to the in vitro system the rumen is a continuous flow system with greater complexity and it is possible that the concentration of E. glabra that I used in vivo was not optimum. This places importance on future dose response experiments to confirm the efficacy of E. glabra in vivo.

CoordenaÃÃo de AperfeiÃoamento de NÃvel Superior / O presente trabalho teve como objetivo avaliar as caracterÃsticas fÃsicas, fÃsico-quÃmicas e quÃmicas e atividade antioxidante total de frutos de viuvinha (Myrcia splendens (Sw.) DC.), mapirunga (Eugenia tinctoria Gagnep.) e araticum-do-brejo (Annona glabra L.) oriundos da vegetaÃÃo litorÃnea do CearÃ, em diferentes estÃdios de maturaÃÃo. Os frutos foram caracterizados fisicamente quanto à massa fresca, dimensÃes, coloraÃÃo, rendimento e firmeza. Para a avaliaÃÃo das caracterÃsticas fÃsico-quÃmicas e quÃmicas foram feitas as seguintes anÃlises: sÃlidos solÃveis (SS), acidez total titulÃvel (ATT), pH, relaÃÃo SS/ATT, aÃÃcares solÃveis totais e redutores, pectina solÃvel, vitamina C, clorofila e carotenÃides totais, antocianinas e flavonÃides amarelos, polifenÃis extraÃveis totais (PET) e atividade antioxidante total pelo mÃtodo ABTS. Os resultados indicaram que os frutos de viuvinha e mapirunga apresentaram um aumento na massa fresca, comprimento, diÃmetro e rendimento em polpa no decorrer do amadurecimento. Nos frutos de araticum-do-brejo observou-se um declÃnio nestes parÃmetros no final da maturaÃÃo. Observou-se tambÃm um aumento, no teor de sÃlidos solÃveis totais, na relaÃÃo SS/ATT, aÃÃcares solÃveis totais e aÃÃcares redutores nos frutos analisados à medida em que ocorria a hidrÃlise do amido. Apenas nos frutos de viuvinha ocorreu diminuiÃÃo da acidez ao longo do amadurecimento. Houve solubilizaÃÃo da pectina ao longo do amadurecimento, exceto nos frutos de mapirunga. Dentre os compostos bioativos observou-se aumento na vitamina C no decorrer do amadurecimento das frutas analisadas. Constatou-se alto teor de antocianinas nos Ãltimos estÃdios de maturaÃÃo na polpa de mapirunga. Apenas nos frutos de viuvinha houve uma diminuiÃÃo de flavonÃides amarelos no decorrer do amadurecimento. Foram encontrados altos valores de polifÃnois extraÃveis totais e elevada atividade antioxidante nos frutos de viuvinha e mapirunga. Houve correlaÃÃo positiva e significativa entre a atividade antioxidante total e o teor de polifenÃis extraÃveis totais de todas as espÃcies analisadas. As polpas de viuvinha e mapirunga sÃo promissoras tendo em vista considerÃveis teores de compostos bioativos como, carotenÃides, antocianinas, flavonÃides amarelos e polifenÃis totais extraÃveis que as fazem produtos diferenciais no mercado de frutas nativas. / O presente trabalho teve como objetivo avaliar as caracterÃsticas fÃsicas, fÃsico-quÃmicas e quÃmicas e atividade antioxidante total de frutos de viuvinha (Myrcia splendens (Sw.) DC.), mapirunga (Eugenia tinctoria Gagnep.) e araticum-do-brejo (Annona glabra L.) oriundos da vegetaÃÃo litorÃnea do CearÃ, em diferentes estÃdios de maturaÃÃo. Os frutos foram caracterizados fisicamente quanto à massa fresca, dimensÃes, coloraÃÃo, rendimento e firmeza. Para a avaliaÃÃo das caracterÃsticas fÃsico-quÃmicas e quÃmicas foram feitas as seguintes anÃlises: sÃlidos solÃveis (SS), acidez total titulÃvel (ATT), pH, relaÃÃo SS/ATT, aÃÃcares solÃveis totais e redutores, pectina solÃvel, vitamina C, clorofila e carotenÃides totais, antocianinas e flavonÃides amarelos, polifenÃis extraÃveis totais (PET) e atividade antioxidante total pelo mÃtodo ABTS. Os resultados indicaram que os frutos de viuvinha e mapirunga apresentaram um aumento na massa fresca, comprimento, diÃmetro e rendimento em polpa no decorrer do amadurecimento. Nos frutos de araticum-do-brejo observou-se um declÃnio nestes parÃmetros no final da maturaÃÃo. Observou-se tambÃm um aumento, no teor de sÃlidos solÃveis totais, na relaÃÃo SS/ATT, aÃÃcares solÃveis totais e aÃÃcares redutores nos frutos analisados à medida em que ocorria a hidrÃlise do amido. Apenas nos frutos de viuvinha ocorreu diminuiÃÃo da acidez ao longo do amadurecimento. Houve solubilizaÃÃo da pectina ao longo do amadurecimento, exceto nos frutos de mapirunga. Dentre os compostos bioativos observou-se aumento na vitamina C no decorrer do amadurecimento das frutas analisadas. Constatou-se alto teor de antocianinas nos Ãltimos estÃdios de maturaÃÃo na polpa de mapirunga. Apenas nos frutos de viuvinha houve uma diminuiÃÃo de flavonÃides amarelos no decorrer do amadurecimento. Foram encontrados altos valores de polifÃnois extraÃveis totais e elevada atividade antioxidante nos frutos de viuvinha e mapirunga. Houve correlaÃÃo positiva e significativa entre a atividade antioxidante total e o teor de polifenÃis extraÃveis totais de todas as espÃcies analisadas. As polpas de viuvinha e mapirunga sÃo promissoras tendo em vista considerÃveis teores de compostos bioativos como, carotenÃides, antocianinas, flavonÃides amarelos e polifenÃis totais extraÃveis que as fazem produtos diferenciais no mercado de frutas nativas. / The objective of this work was to characterize physical, chemical and physico-chemical and evaluate the total antioxidant activity of fruits of viuvinha (Myrcia splendens (Sw.) DC.), mapirunga (Eugenia tinctoria Gagnep.) and araticum-do-brejo (Annona glabra L.) from the coastal vegetation of CearÃ, at different stages of maturation. Fruits were physically characterized by size, color, total weight, yield, and firmness. For the evaluation of the physico-chemical and chemical analyzes were made as follows: soluble solids (SS), acidity, pH, SS / TA ratio, total soluble sugars and reducing sugars, soluble pectin, vitamin C, chlorophyll and total carotenois, anthocyanins and yellow flavonoids , total extractable polyphenols (TEP) and total antioxidant activity (TAA) by ABTS method. The study found that the fruits of viuvinha and mapirunga showed an increase in total weight, length, diameter and yield of the pulp during ripening. In fruits of araticum observed a decrease in these parameters at the end of maturation. Were also observed an increase in soluble solids, SS / TA ratio, total soluble sugars, reducing sugars in the fruits analyzed as starch hydrolysis occurred. Only the fruits of viuvinha there was a decrease in acidity during the ripening. There solubilization of pectin during ripening, except in the fruits of mapirunga. Among the bioactive compounds reported an increase in vitamin C during ripening of fruits analyzed. There was a high content of anthocyanins in late-stage of viuvinha and mapirunga. Only in the fruits of viuvinha there were yellow flavonoids during ripening. Emphasis was placed high values of extractable polyphenols and total antioxidant activity in fruits of viuvinha and mapirunga. There was significant positive correlation between total antioxidant activity and total extractable polyphenol content of all species analyzed. The pulp of viuvinha and mapirunga are promising in view of considerable amounts of bioactive compounds such as carotenoids, anthocyanins, yellow flavonoids and total polyphenols total extractable differentials that make products on the market of native fruits. / The objective of this work was to characterize physical, chemical and physico-chemical and evaluate the total antioxidant activity of fruits of viuvinha (Myrcia splendens (Sw.) DC.), mapirunga (Eugenia tinctoria Gagnep.) and araticum-do-brejo (Annona glabra L.) from the coastal vegetation of CearÃ, at different stages of maturation. Fruits were physically characterized by size, color, total weight, yield, and firmness. For the evaluation of the physico-chemical and chemical analyzes were made as follows: soluble solids (SS), acidity, pH, SS / TA ratio, total soluble sugars and reducing sugars, soluble pectin, vitamin C, chlorophyll and total carotenois, anthocyanins and yellow flavonoids , total extractable polyphenols (TEP) and total antioxidant activity (TAA) by ABTS method. The study found that the fruits of viuvinha and mapirunga showed an increase in total weight, length, diameter and yield of the pulp during ripening. In fruits of araticum observed a decrease in these parameters at the end of maturation. Were also observed an increase in soluble solids, SS / TA ratio, total soluble sugars, reducing sugars in the fruits analyzed as starch hydrolysis occurred. Only the fruits of viuvinha there was a decrease in acidity during the ripening. There solubilization of pectin during ripening, except in the fruits of mapirunga. Among the bioactive compounds reported an increase in vitamin C during ripening of fruits analyzed. There was a high content of anthocyanins in late-stage of viuvinha and mapirunga. Only in the fruits of viuvinha there were yellow flavonoids during ripening. Emphasis was placed high values of extractable polyphenols and total antioxidant activity in fruits of viuvinha and mapirunga. There was significant positive correlation between total antioxidant activity and total extractable polyphenol content of all species analyzed. The pulp of viuvinha and mapirunga are promising in view of considerable amounts of bioactive compounds such as carotenoids, anthocyanins, yellow flavonoids and total polyphenols total extractable differentials that make products on the market of native fruits.

The biggest hurdle in success of gene and drug therapy is designing and preparation of suitable bio-nanomaterials to carry the desired nucleic acid and drug to the targeted site. The work described in the present thesis encompasses two different approaches for the delivery of bioactive oligo/polynucleotides and drugs in vitro and in vivo using either cationic lipids or their nanocomposites with different carbon nanomaterials. The idea of using carriers for oligo/polynucleotides and drugs came into existence because of numerous physiological barriers in pathway of delivery of naked oligo/polynucleotides or drugs which reduces the overall activity of these bioactives in biological systems. These barriers trigger scientific research toward the preparation of appropriate biomaterials which can overcome the physiological barriers and improve the activity of bioactive oligo/polynucleotides and drugs in cellular systems. Toward this end, the design and synthesis of different cationic lipids and carbon nanomaterials were undertaken as described in seven chapters of the thesis. A series of novel cationic lipids with structural variability was prepared and used for gene delivery in vitro. They were further tuned chemically to sustain delivery efficiency in high serum percentage during in vitro transfection. These serum compatible lipids were used to perform transfection of reporter gene plasmid and found to be more efficient compared to the some well known commercial products for the same purpose. Another series of novel lipids were synthesized for the targeted gene delivery in vitro. These tryptophan based cholesteryl lipids were used to prepare mixed liposomes. These mixed liposomes were highly efficient in targeting sigma receptor rich HEK293T over sigma receptor negative HeLa cells. Mixed liposomes were also prepared for selective targeting of αvβ3 and αvβ5 integrins in gene transfection protocol using a palmitoyl-RAFT-RGD4 template. A mixed liposomal formulation was developed to carry out anti-sense siRNA mediated knockdown of Smad-2 protein with better efficiency compared to some of the best known commercial products for the same purpose. These mixed liposomes were also highly efficient for regression via induction of p53 mediated apoptosis in xenograft tumors developed in nude mice. Carbon nanomaterials have been extensively explored as nanoscale gene/drug carriers for potential applications. But the challenge is to solubilize these highly hydrophobic materials in aqueous medium for use in biological systems. Although there are reports for covalent modifications of such nanomaterials but it could be done only with the loss of some beneficial features of these materials. Herein a non-covalent technique has been efficiently used to suspend single walled carbon nanotubes in water using biocompatible cationic lipids. These nanosuspensions were used to complex plasmid DNA and transfect them in vitro. They proved to be highly serum compatible DNA carriers which did not drop the efficiency even in very high percentage of serum. Similarly exfoliated graphene was modified with cationic lipid and serum components to improve IC50 of Tamoxifen citrate and Methotrexate to a considerable extent in vitro. The improved Methotrexate formulations were highly efficient for regression in size of xenograft tumors developed in nude mice. Thus, the present thesis entails generation of cationic lipids and carbon nanomaterials based nanocomposites which were not only highly biocompatible themselves but their efficiency was found many fold better compare to some of the best commercial delivery agents. These were useful for the delivery of various bioactive oligo/polynucleotides and drugs in vitro and in vivo.

Iminosugars are polyhydroxylated alkaloids, and can be generally defined as sugar mimetics in which the endocyclic oxygen atom has been replaced with a basic nitrogen. A common affect of this atomic substitution is to bestow these compounds with the ability to inhibit various sugarprocessing enzymes; most significantly the glycosidases (glycoside hydrolases) which areintimately involved in a huge array of biological functions. Compounds which inhibit these enzymes concordantly possess much potential as medicinal agents for the treatment of a variety of diseases. Several iminosugars have already achieved market approval as drugs, and many more are promising candidates in the late stages of clinical development. As such there remains considerable interest in this class of compound, both in terms of the exploration of novel iminosugar structures, as well as the continual development of more efficient general methodology for their synthesis. The densely-packed functionality and stereochemical information present in iminosugars makes them challenging targets for asymmetric chemical synthesis, whereas carbohydrates are clearly very attractive as chiral-pool starting materials for this purpose. Indeed, the majority of the most successful syntheses of iminosugars use the latter approach, and such is the focus of this thesis. Chapter 1 presents a relatively brief introduction to iminosugars, including their types of structure, natural occurrence and biological mode of action. The rationale behind their use as therapeutic agents for the treatment of some significant disease targets is also discussed. Chapter 2 is concerned with the preparation of a number of novel polyhydroxylated azetidines, and their evaluation as glycosidase inhibitors. Such compounds represent an almost entirely neglected class of iminosugars within the literature. An overview of natural and synthetic products incorporating an azetidine motif is given, as well as a brief review of preparative methods and known azetidine iminosugars. A highly efficient and flexible method for the key azetidine ring formation is demonstrated by the cyclisations of 3,5-di-O-triflates of pentoses and hexoses, and of a 2,4-di-O-triflate of glucose, with various primary amines. In this manner, many azetidine triols and tetrols were prepared in good yield. Furthermore, this process is readily adaptable to the installation of added functionality to the azetidine scaffold, as demonstrated by the preparation of 1-acetamido analogues. The initial biological screening of these compounds showed a promising array of glycosidase inhibition, including that of selective inhibition of fungal enzymes. Chapter 3 describes a strategy with which to prepare all sixteen stereoisomers of a known piperidine iminosugar, alpha-homonojirimycin (alpha-HNJ), in a highly divergent manner from just four of the possible thirty-two 6-azidoheptitols using traditional chemical synthesis in tandem with biotechnological transformations. One half of the execution of this strategy is described in this thesis. Two 6-azidoheptitols were prepared from D-mannose, thereby providing access to four 6-azidoketoheptoses through a combination of microbial oxidation and enzymatic epimerisation. Catalytic hydrogenation of these 6-azidoketoheptoses furnished four diastereomeric mixtures of 2,6-iminoheptitols, with varying degrees of stereoselectivity. Purification of these mixtures allowed six 2,6-iminoheptitols to be isolated, two of which have never previously been tested for glycosidase inhibition. Significantly, one of them was found to be a potent and highly selectiveinhibitor of alpha-galactosidases, and may therefore be of interest in the treatment of Fabry disease.

Bluthochdruck zählt zu einer der häufigsten Zivilisationskrankheiten und ist der Hauptfaktor für die Entstehung kardiovaskulärer Erkrankungen. Das Präventionspotenzial bei Hypertonie ist sehr hoch, da lebensstilassoziierte Faktoren wie Übergewicht, hoher Kochsalz- und Alkoholkonsum oder Stress die Entstehung eines erhöhten Blutdrucks wesentlich begünstigen. Daher wird eine antihypertensive Therapie meist mit nicht-medikamentösen Maßnahmen eingeleitet. Für die Regulation des Blutdrucks ist die nähere Betrachtung des Angiotensin-Converting Enzymes (ACE) wichtig, da es eines der Schlüsselenzyme des Renin-Angiotensin-Aldosteron-Systems und des Kallikrein-Kinin-Systems darstellt. Die Möglichkeit, dass ACE-inhibierende Peptide aus Lebensmittelproteinen über die Nahrungsmittelaufnahme einen positiven physiologischen Effekt auf den Blutdruck ausüben, ist ein vielversprechender Ansatz zur Unterstützung einer nicht-medikamentösen Therapie bei Hypertonie. In der Literatur sind zahlreiche Peptide beschrieben, welche eine inhibitorische Wirkung auf das ACE in vitro besitzen. Die vorliegende Arbeit beschäftigte sich mit der Klasse der tryptophanhaltigen Dipeptide, die in der Literatur als potente, natürliche ACE-Inhibitoren beschrieben sind. Die tryptophanhaltigen Peptide wurden hinsichtlich ihrer Gewinnung, ihrer Hemmwirkung auf das Zielenzym und bezüglich ihrer Bioverfügbarkeit in vitro und in vivo untersucht.

Cette thèse porte sur la mise en œuvre de procédés innovants d'extraction de composés naturels bioactifs de plantes. Nous avons considéré ici l'extraction d'un composé anti-oxydant, l'acide rosmarinique, à partir du végétal mélisse (Melissa officinalis L.), qui contient également d'autres composés d'intérêt (citral et caryophyllène). Différentes techniques d'extraction-purification, soit à haute pression (extraction au CO2 supercritique) ou à pression atmosphérique (extraction Soxhlet, extraction batch, nanofiltration etc.) ont été envisagées. L'objectif de ce travail a été d'étudier expérimentalement l'influence des paramètres opératoires (débit, composition et concentration du solvant, taille des particules, pression et température) sur la cinétique des processus afin de sélectionner les meilleures conditions pour chaque opération. Les résultats expérimentaux ont été ensuite comparés avec plusieurs modèles mathématiques décrivant les phénomènes de transfert de masse et l'écoulement au travers du milieu poreux constitué par la matière végétale broyée. Cette démarche, à partir de la détermination des paramètres physiques du modèle, a fourni les éléments pour une extrapolation potentielle à l'échelle industrielle. Du point de vue du procédé complet d'extraction-purification, l'originalité du travail a été de proposer plusieurs scénarii d'enchainement d'opérations, couplant en synergie des opérations conventionnelles à pression atmosphériques (macération, nano-filtration etc.) et des opérations de traitement au CO2 supercritique avec co-solvant. / This thesis deals with the extraction of natural bioactive compounds from plants (case study with Lemon Balm (Melissa officinalis L.)) by using different separation techniques at high (supercritical extraction) and atmospheric pressure (Soxhlet extraction, batch extraction, nanofiltration etc.). The influence of main operational parameters (solvent composition and flow rate, particle size of the raw material, pressure, temperature) on the process kinetics is studied experimentally with the aim to determine appropriate operational conditions for better extraction. The experimental results are confronted to a number of mathematical models in order to estimate the applicability of different theoretical concepts to the particular process and to select and apply appropriate models for determination of important parameters, characterizing the mass transfer process and necessary for scale-up and design purposes. Coupling between different separation methods is also considered and a number of integrated process schemes are proposed resulting in better yield of the targeted compounds.

Streptomycetes are soil filamentous Gram-positive bacteria that produce wide variety of pigments and biologically active substances including macrolides. Some of them are used as very efficient antibiotics and strong antifungal agents in medicine, others have became useful tools for staining biomembranes and detecting cholesterol via their internal fluorescence. Actinomycete Streptomyces durmitorensis (wild type strain MS405T ) is a bacteria isolated from Durmitor National Park in Montenegro soil samples. It produces secondary metabolite that has been identified as 32,33-didehydroroflamycoin (DDHR) closely related to the macrolides roflamycoin and generaly used filipin. DDHR exhibits cytototoxic activity against mammalian cells and yeast Saccharomyces cerevisiae strain EGY48. In addition it has interesting fluorescence properties allowing visualization of some membrane components. DDHR interacts with biomembranes, causes their disintegration leading to changes of the actin and tubulin cytoskeleton organization and in higher concentrations it causes cells necrosis. DDHR-sterol interaction in cell membranes decreases fluorescence intensity of DDHR. The compound is able to fluorescently stain aberrant lysosomes and could be therefore potentially used in diagnostics of some lysosomal storage disease.

Os sucos e bebidas de frutas são altamente consumidos pela população brasileira, feitos de modo caseiro ou industrializados, porém pouco se sabe sobre o conteúdo de compostos bioativos e capacidade antioxidante destes diferentes tipos de produtos. O objetivo deste trabalho foi a determinação e comparação de diferentes compostos bioativos e da capacidade antioxidante de sucos naturais, integrais, néctares, preparados líquidos e sólidos para refresco nos sabores de tangerina, goiaba, maracujá, laranja e uva. Os sucos naturais foram preparados descartando as cascas e sementes e para cada tipo de bebida industrializada foram selecionadas três marcas distintas adquiridas no comércio local em Ribeirão Preto/SP. Foram determinados os teores dos seguintes compostos bioativos: fenólicos totais, flavonoides totais e amarelos, antocianinas totais, ácido ascórbico, licopeno e beta caroteno nas bebidas, assim como, a capacidade antioxidante foi determinada pelos métodos ABTS, DPPH e FRAP, sendo os dados analisados estatísticamente atráves da diferença entre as médias, também foram feitas correlações entre os diferentes métodos de capacidade antioxidante. Entre os achados observa-se que os preparados sólidos para refresco de diferentes marcas e sabores apresentaram os menores teores de compostos bioativos e capacidade antioxidante em relação às demais bebidas. O conteúdo de ácido ascórbico, fenólicos, flavonoides totais e amarelos apresentaram uma variabilidade grande entre as bebidas. As diferentes marcas de mesmo tipo de bebida e sabor apresentaram diferença significativa no teor de compostos bioativos e capacidade antioxidante. As antocianinas totais apresentaram-se em maior quantidade nos preparados líquidos e sucos integrais de uva. Os maiores teores de licopeno foram encontrados nos sucos natural e integrais de goiaba. O beta caroteno foi detectado apenas em algumas amostras sendo que as bebidas de tangerina apresentaram os maiores valores deste carotenoide, no suco natural, alguns sucos integrais, néctares e preparados líquidos para refresco. Os três diferentes métodos de capacidade antioxidante apresentaram resultados divergentes entre si e dados de correlação variáveis em sabores de frutas, porém, em todos os métodos, os preparados sólidos para refresco de diferentes marcas e sabores apresentaram medidas mais baixas ou nulas de capacidade antioxidante, mostrando que estes produtos são os mais inadequados do ponto de vista nutricional. Os resultados deste trabalho também mostram a dificuldade na caracterização destes produtos, comparando diferentes formulalções assim como diferentes produtos industrializados, mas apontam para valorização dos produtos naturais e menos processados / Juices and fruit drinks produced at home or industrialized are highly consumed by the Brazilian population, but the content of bioactive compounds and antioxidant capacity of these different types of products is unknown.The objective of this work was the determination and comparison of bioactive compounds and antioxidant capacity of natural and whole juices, nectars, liquid and solid prepared for soft drinks of tangerine, guava, passion fruit, orange and grape. The natural juices were prepared with the fruit pulp, scorning the barks and seeds and for each type of industrialized beverages, three different brands were collected in the local market of Ribeirão Preto -SP.Total phenolics, total and yellow flavonoids, total anthocyanins, ascorbic acid, lycopene and beta carotene were determined and analyzed by the difference between the averages, the antioxidant capacity was evaluated by ABTS, DPPH and FRAP assays, also their correlations were determined. Prepared solids for soft drinks of different brands and flavors had the lowest levels of bioactive compounds and antioxidant capacity compared to other beverages. The content of ascorbic acid, phenolics, total and yellow flavonoids presented a great variability between the drinks. The different brands of the same type of beverage and flavor presented significant difference in the content of bioactive compounds and antioxidant capacity.The total anthocyanins were higher in liquid prepared for soft drinks and whole grapes juices. The highest levels of lycopene were found in natural and whole juices of guava. Beta carotene was not found in all beverages studied, tangerine beverages presented the highest values of this carotenoid in natural juice, some whole juices, nectars and liquid prepared for soft drinks. Three different methods of antioxidant capacity presented divergent results and variable correlation data on fruit flavors, however, in all methods, solid prepared for soft drinks of different brands and flavors presented the smallest and zero antioxidant capacity, showing that these products are the least nutritionally suitable. The results of this work show the difficulty in characterizing these products, comparing different formulations as well as different industrialized products, but stand out for the valorization of natural and less processed products