Thesis (M.S.)--Wisconsin. / Includes bibliographical references (leaves 62-64).

Call number: LD2668 .T4 1965 C55 / Master of Science

Le travail réalisé au cours de cette thèse est le fruit d’une collaboration étroite entre une équipe de l’Université du MAINE et la société ASHLAND qui est spécialisée dans la synthèse de polymères.Le sujet de cette thèse concerne la synthèse d’un polymère biomimétique de la gélatine afin de l’engager dans un procédé de coacervation complexe permettant l’encapsulation de principes actif ou de colorants.La micro-encapsulation est une méthode permettant d’envelopper de petites particules individuelles ou de gouttelettes dans une couche protectrice de polymères. Largement utilisée en pharmacie, agroalimentaire, cosmétique, les biotechnologies, le phénomène d’encapsulation nécessite la protection des matières et de contrôler la libération de l’actif.Parmi les nombreux procédés d’encapsulation existant, nous nous sommes particulièrement orientés vers une méthodologie de coacervation complexe.Cette dernière est basée sur la complexation entre deux polyélectrolytes de charges opposées. Ainsi, cette technique requiert un polymère anionique et un polymère cationique qui interagissent ensemble (interactions électrostatiques, liaisons hydrogène…) pour former des coacervats.Un des systèmes les plus utilisés de la coacervation complexe est le système gomme d’acacia avec la gélatine en raison de leur abondance et de leur biodégradabilité. Cependant, à cause de son origine animale, l’objectif du travail a consisté à remplacer la gélatine dans le processus d’encapsulation. Pour ce faire, nous avons synthétisé plusieurs polymères spécifiques à partir de différents monomères méthacryliques / méthacrylamides permettant la coacervation et ensuite la réticulation des coacervats formées. Dans un premier temps, nous avons créé une approche permettant de déterminer les conditions optimales de formation des coacervats (le pH d’encapsulation, le ratio de polymère, la force ionique) par potentiel zêta, mesure de la turbidité et par spectroscopie infra-rouge. Par la suite, des essais d’encapsulation avec plusieurs polymères anioniques ont permis de confirmer que la méthodologie établie précédemment était adéquate. Ensuite, un test de réticulation a été réalisé à l’aide d’agents de réticulation afin de rigidifier les parois des coacervats. Enfin, des analyses de force de rupture et de stabilité dans des détergents standards ont été réalisées afin de valider l’application de ces capsules en cosmétique. / The work realized during the last three years CIFRE PhD program consist in a collaborative research between a polymer team of the University of LE MANS and ASHLAND company which the first goal is the synthesis of polymers.The aim of this thesis is the synthesis of biomimetic polymers of the gelatin which were further engaged in the process of encapsulation by complex coacervation.Micro-encapsulation is an effective method of wrapping small individual particles or droplets in polymers protective coating widely used in the fields of food, pharmaceutics, cosmetics, pesticides, biotechnologies. The encapsulation phenomenon target is to protect functional materials and control its release. Among many existing processes, we particularly focused our attention on complex coacervation to produce microcapsules. Complex coacervation is a methodology based on the complexation between two oppositely charged polymers as polyelectrolyte. Thus, this technique requires one anionic and one cationic polymer which are capable to interact together (through electrostatic attractions, hydrogen bond) to form coacervats. One of the most used systems for complex coacervation is the gelatin / acacia gum system. These two natural polymers are widely used as wall material for the capsules due to their abundance and biodegradability. However, gelatin polymer needs to be replaced in the encapsulation process due to its animal origin. In order to substitute gelatin by synthetic polymers in the complex coacervation process, we synthesized several polymers from various monomers authorizing coacervation and further crosslinking. Firstly, we stetted up an approach allowing the evaluation of the optimal conditions of coacervation (pH, ratio, ionic strength) by zetâ potential, and infra-red analysis.Furthermore, encapsulation tests with several anionic polymers were performed which tended to confirm that the methodology was appropriate. Moreover, a test of crosslinking was successfully realized using crosslinkers in order to rigidify coacervats walls. Finally, break strength analysis and the stability in surfactants were conducted to validate the process for cosmetic applications.

Elastin is a polymeric protein of the extracellular matrix that imparts the characteristics of extensibility and elastic recoil to tissues. Recombinant polypeptides based on the domain structures and sequences of human elastin self-assemble into organized fibrous structures, with physical properties similar to those of native polymeric elastin. Elastin self-assembly is initiated by a temperature-induced phase separation, called coacervation. Previous to this work, coacervation temperature had been the only parameter available to measure propensity for self-assembly. A variety of techniques were developed using spectrophotometry, microscopy, and rheometry to differentiate the stages of self-assembly, thereby enabling independent observation and quantitation of each stage, and allowing investigations into properties of polypeptides and solution conditions affecting these stages. Kinetic analysis of self-assembly yielded two additional parameters: coacervation velocity and maturation velocity. Examining the effects of agitation, salt concentration, temperature, polypeptide concentration, size of a polypeptide, hydrophobic domain sequence, and cross-linking domain structure on the kinetics demonstrated that coacervation and maturation are independent stages of self-assembly involving distinct mechanisms. Microscopic observations showed that protein-rich droplets of coacervate grew by coalescence to a stable droplet size, which correlated to differences in maturation velocities between polypeptides. Coacervate droplet growth appeared limited by the formation of organized polypeptide at the surface of the droplets, decreasing surface fluidity. Many of the general principles of the physical chemistry of colloids and emulsions appeared to apply to the formation, growth and stabilization of coacervates of the elastin-like polypeptides. Self-assembly in the presence of non-elastin, matrix-associated proteins showed that these proteins maintained the coacervate as small droplets, which sometimes flocculated into fibre-like structures. Rheometry demonstrated a second temperature-induced transition above the coacervation temperature, which resulted in gelation and viscoelastic characteristics similar to microgels. Together, these observations have resulted in a greater level of understanding of the entire self-assembly process, and provided a comprehensive model of elastin-like polypeptide self-assembly that relates to in vivo assembly of elastic fibres.

Elastin is a polymeric protein of the extracellular matrix that imparts the characteristics of extensibility and elastic recoil to tissues. Recombinant polypeptides based on the domain structures and sequences of human elastin self-assemble into organized fibrous structures, with physical properties similar to those of native polymeric elastin. Elastin self-assembly is initiated by a temperature-induced phase separation, called coacervation. Previous to this work, coacervation temperature had been the only parameter available to measure propensity for self-assembly. A variety of techniques were developed using spectrophotometry, microscopy, and rheometry to differentiate the stages of self-assembly, thereby enabling independent observation and quantitation of each stage, and allowing investigations into properties of polypeptides and solution conditions affecting these stages. Kinetic analysis of self-assembly yielded two additional parameters: coacervation velocity and maturation velocity. Examining the effects of agitation, salt concentration, temperature, polypeptide concentration, size of a polypeptide, hydrophobic domain sequence, and cross-linking domain structure on the kinetics demonstrated that coacervation and maturation are independent stages of self-assembly involving distinct mechanisms. Microscopic observations showed that protein-rich droplets of coacervate grew by coalescence to a stable droplet size, which correlated to differences in maturation velocities between polypeptides. Coacervate droplet growth appeared limited by the formation of organized polypeptide at the surface of the droplets, decreasing surface fluidity. Many of the general principles of the physical chemistry of colloids and emulsions appeared to apply to the formation, growth and stabilization of coacervates of the elastin-like polypeptides. Self-assembly in the presence of non-elastin, matrix-associated proteins showed that these proteins maintained the coacervate as small droplets, which sometimes flocculated into fibre-like structures. Rheometry demonstrated a second temperature-induced transition above the coacervation temperature, which resulted in gelation and viscoelastic characteristics similar to microgels. Together, these observations have resulted in a greater level of understanding of the entire self-assembly process, and provided a comprehensive model of elastin-like polypeptide self-assembly that relates to in vivo assembly of elastic fibres.

The focus of this research was to develop a plant-based microcapsule for flax oil by complex coacervation. Complex coacervation involves the electrostatic attraction between two polymers of opposing charges. Specifically, the research aimed to: a) identify the ideal biopolymer and solvent conditions required for complex coacervation involving pea protein isolate (PPI) and gum Arabic (GA); b) understand the functional behaviour of PPI-GA complexes as food and biomaterial ingredients; and c) develop methodologies for encapsulating flax oil within PPI-polysaccharide capsules. Complex coacervation between PPI-GA was found to be optimized at a biopolymer weight mixing ratio of 2:1 in the absence of salt. The functional behaviours of the optimized biopolymer mixture were then investigated as a function of pH (4.30-2.40) within a region dominated by complex coacervation. Emulsion stability was found to be greater for PPI-GA mixture systems relative to PPI alone at pH values between 3.10 and 4.00; emulsions produced under one-step emulsification exhibited higher stability compared to those of two-step emulsification at all pH values. Foam expansion was independent of both biopolymer content and pH, whereas foam stability improved for the mixed system between pH 3.10 and 4.00. The solubility minimum was broadened relative to PPI at more acidic pH values. These findings suggested that the admixture of PPI and GA under complexing conditions could represent a new food and/or biomaterial ingredient, and has potential as an encapsulating agent. Two encapsulation processes were employed in this research: high speed mixing (two-step emulsification) and low speed mixing (one-step emulsification). Flax oil capsules formed using the gelatin-GA mixture (as control) under high speed mixing exhibited low moisture content, water activity and surface oil, and afforded adequate protection against oxidation relative to free oil over a 25 d storage period. The PPI-GA mixture failed to produce acceptable capsules using either high or low speed mixing. In contrast, PPI-alginate capsules were produced and exhibited similar chemical properties as gelatin-GA capsules, except with lower encapsulated flax oil content (30% vs. 50% w/w). However, oxidative stability may adversely affected by the low speed mixing condition during encapsulation.

The focus of this research was to develop a plant-based microcapsule for flax oil by complex coacervation. Complex coacervation involves the electrostatic attraction between two polymers of opposing charges. Specifically, the research aimed to: a) identify the ideal biopolymer and solvent conditions required for complex coacervation involving pea protein isolate (PPI) and gum Arabic (GA); b) understand the functional behaviour of PPI-GA complexes as food and biomaterial ingredients; and c) develop methodologies for encapsulating flax oil within PPI-polysaccharide capsules. Complex coacervation between PPI-GA was found to be optimized at a biopolymer weight mixing ratio of 2:1 in the absence of salt. The functional behaviours of the optimized biopolymer mixture were then investigated as a function of pH (4.30-2.40) within a region dominated by complex coacervation. Emulsion stability was found to be greater for PPI-GA mixture systems relative to PPI alone at pH values between 3.10 and 4.00; emulsions produced under one-step emulsification exhibited higher stability compared to those of two-step emulsification at all pH values. Foam expansion was independent of both biopolymer content and pH, whereas foam stability improved for the mixed system between pH 3.10 and 4.00. The solubility minimum was broadened relative to PPI at more acidic pH values. These findings suggested that the admixture of PPI and GA under complexing conditions could represent a new food and/or biomaterial ingredient, and has potential as an encapsulating agent. Two encapsulation processes were employed in this research: high speed mixing (two-step emulsification) and low speed mixing (one-step emulsification). Flax oil capsules formed using the gelatin-GA mixture (as control) under high speed mixing exhibited low moisture content, water activity and surface oil, and afforded adequate protection against oxidation relative to free oil over a 25 d storage period. The PPI-GA mixture failed to produce acceptable capsules using either high or low speed mixing. In contrast, PPI-alginate capsules were produced and exhibited similar chemical properties as gelatin-GA capsules, except with lower encapsulated flax oil content (30% vs. 50% w/w). However, oxidative stability may adversely affected by the low speed mixing condition during encapsulation.

No description available.

Le bénéfice de l’encapsulation des molécules bioactives a séduit les industries agroalimentaires depuis plusieurs décennies. Plus récemment des études ont montré la capacité de protéines alimentaires de charge opposée à s’assembler en microsphères par coacervation complexe. La compréhension des forces gouvernant le processus de coacervation entre protéines et l’influence exercée par la présence de bioactifs demeurent des prérequis pour l’utilisation des coacervats complexes comme agent d’encapsulation. Dans ce contexte, l’objectif de mon projet de thèse a été de comprendre le mécanisme de coacervation complexe entre la ¿-lactoglobuline (¿-LG) chargée négativement, et la lactoferrine (LF) chargée positivement, en absence et en présence de petits ligands. La LF a présenté une coacervation préférentielle avec le variant A de la¿¿-LG qui se distingue du variant B par la substitution de 2 acides aminés. Au niveau moléculaire, deux sites de fixation de la ¿-LG sur la LF ont été identifiés.En outre, par la mesure d’une part des coefficients de diffusion rotationnel et d’autre part de la cinétique de diffusion des entités moléculaires constituant les coacervats, il est suggéré que ces derniers sont formés à partir de -LG libre¿¿de pentamère, LF(-LG2)2, ainsi que des entités plus larges, (LF-LG2)n. Afin d’évaluer l’effet de la présence de petits ligands sur la coacervation complexe entre la -LG et la LF, des ligands modèles (ANS et acide folique) ont été utilisés. Dans les conditions expérimentales testées ces deux ligands n’ont pas d’affinité pour la -LG, mais après interact / Encapsulation of bioactives has been used by the food industries for decades and represents a great potential for the development of innovative products. Given their versatile functional properties, milk proteins in particular from whey have been used for encapsulation purposes using several encapsulation techniques. In parallel, recent studies showed the ability of oppositely charged food proteins to co-assemble into microspheres through complex coacervation. Understanding the driving forces governing heteroprotein coacervation process and how it is affected by the presence of ligands (bioactives) is a prerequisite to use heteroprotein coacervates as encapsulation device. In this context, the objective of my thesis work was to understand the mechanism of complex coacervation between -lactoglobulin (-LG) and lactoferrin (LF) in the absence and presence of small ligands. The conditions of optimal ¿-LG - LF coacervation were found at pH range 5.4-6 with a molar excess of ¿-LG. RemarkabAt molecular level, the presence of two binding sites on LF for -LG was evidenced. Moreover, the heterocomplexes such as pentamers LF(-LG2)2 and quite large complexes (LF-LG2)n were identified as the constituent molecular species of the coacervate phase. To evaluate the -LG - LF complex coacervation in the presence of small ligands, models of hydrophobic (ANS) and hydrophilic molecules (folic acid) were used. Although under the experimental conditions tested the small ligands did not interact with -LG, both interacted with LF inducing its self-association into nanoparticles. High relati

Ce travail porte sur le développement de systèmes d’encapsulation à base de polysaccharides et de molécules antimicrobiennes de nature protéique comme le lysozyme du blanc d’œuf et la nisine produite par la bactérie Lactococcus lactis. La coacervation complexe de l’alginate ou la pectine (deux polysaccharides anioniques) avec le lysozyme ou la nisine, chargés positivement sur une large gamme de pH, serait une solution pour protéger ces molécules et assurer une libération contrôlée lors de la conservation d’un aliment. Les polysaccharides utilisés ont été choisis en fonction de leur sensibilité aux enzymes produites par les bactéries cibles. L’ensemble des travaux menés à différentes échelles a mis en évidence (i) le rôle de certains facteurs physico-chimiques (ratio, pH, force ionique,…) sur les interactions mises en jeu entre le lysozyme ou la nisine et l’alginate ou la pectine, (ii) l’influence de ces facteurs sur les propriétés des complexes formés et (iii) le rôle des polysaccharides sélectionnés dans la stabilisation de la structure du lysozyme ou de la nisine lors du séchage par atomisation. Enfin, la structure et l’activité antimicrobienne des films obtenus par voie solvant (casting) et des microcapsules obtenues par atomisation sont étudiées en relation avec les propriétés des complexes formés. Ce travail a permis une meilleure compréhension des mécanismes impliqués dans la formation des complexes peptides/polysaccharides ou protéines/polysaccharides, leur résistance au séchage par atomisation ainsi que leur capacité à protéger et à libérer des molécules actives / This work deals with the development of encapsulation systems based on polysaccharides and antimicrobial molecules of protein nature such as hen egg-white lysozyme and nisin produced by Lactococcus lactis. Complex coacervation of alginate or pectin (two anionic polysaccharides) with lysozyme or nisin, positively charged over a wide range of pH, would be a solution to protect these molecules and ensure their controlled release during food preservation. The used polysaccharides were chosen according to their sensitivity to the enzymes produced by the targeted bacteria. The whole of this work performed at various levels showed (i) the impact of some physicochemical factors (ratio, pH, ionic strength,…) on the interactions between lysozyme or nisin and alginate or pectin, (ii) the influence of these factors on the properties of the formed complexes, and (iii) the role of the selected polysaccharides in stabilizing the structure of lysozyme or nisin during spray-drying. Finally, the structure and the antimicrobial activity of films produced by casting and microcapsules obtained by spray-drying were studied in relation to the properties of the formed complexes. This work makes it possible to contribute to a better understanding of the various mechanisms implied in the formation of peptide/polysaccharide or protein/polysaccharide complexes, their resistance to spray-drying and their ability to protect and release active molecules