Ce travail a été mené afin de développer des systèmes polymériques particulaires chargés en facteur de croissance (TGF-β1) en vue d’une application à l’ingénierie tissulaire du cartilage. Tout d’abord, des nanoparticules d’acide poly(lactique-co-glycolique) (PLGA) ont été générées par un procédé basé sur la séparation de phase. De plus, des microparticules de PLGA ont été formées à l’aide d’un procédé d’émulsification/extraction du solvant en milieu CO2 pressurisé. L’un des avantages de ces procédés de formulation est l’utilisation de solvants injectables, non toxiques et non volatils. Les systèmes polymériques préparés ont été caractérisés et des particules sphériques à libération contrôlée ont été obtenues avec un rendement d’encapsulation satisfaisant tout en préservant l’activité biologique du facteur de croissance. Les particules chargées en TGF-β1 ont ensuite été incorporées dans un hydrogel injectable à base d’un dérivé cellulosique silanisé (Si-HPMC) contenant des cellules souches. Le biomatériau obtenu a été caractérisé en termes de morphologie, de propriétés rhéologiques et pour sa capacité à libérer le facteur de croissance. La libération contrôlée et localisée du TGF-β1 pourrait induire la survie, la prolifération ainsi que la différenciation chondrocytaire des cellules souches associées ce qui contribuerait à la régénération du cartilage. En conclusion, le biomatériau hybride élaboré au cours de ce travail possède un potentiel prometteur pour l’ingénierie tissulaire du cartilage. / The aim of this work is to develop polymeric particulate systems loaded with transforming growth factor (TGF-β1) for cartilage tissue engineering application. First, nanoparticles of PLGA (poly(lactic-co-glycolic) acid) were generated using a phase separation method while PLGA microparticles were prepared by an emulsification/extraction process in CO2 medium. Interestingly, non-toxic, non-volatile injectable solvents were used in the formulation processes. The prepared polymeric systems were characterized; spherical particles with sustained release were obtained and satisfactory encapsulation efficiency was achieved with preservation of the growth factor bioactivity. TGF-β1-loaded particles were then incorporated within injectable silanized cellulose-based hydrogel (Si-HPMC) containing stem cells. The obtained biomaterial was characterized in terms of morphology, rheological properties and release study. The local and sustained release of TGF-β1 could induce survival, proliferation and differentiation of stem cells into chondrocytes which may promote cartilage regeneration. To conclude, the elaborated hybrid biomaterial has a promising potential for cartilage tissue engineering.

L'objectif de cette thèse est de mettre en place des outils permettant l'éco-purification de produits naturels par chromatographie de partage centrifuge (CPC), une technique préparative permettant la séparation de composés grâce à un système solvant constitué de deux liquides non miscibles. Ce manuscrit expose dans un premier temps une brève présentation des différentes techniques préparatives fréquemment utilisées pour la purification de produits naturels, puis détaille la technique de chromatographie de partage centrifuge. Le concept de chimie verte est alors introduit avant d'exposer la manière dont il sera pris en compte par la suite. Ainsi, un développement de méthode CPC à petite échelle, dans le but de limiter la consommation de solvants et la production de déchets, tout en limitant la consommation d'énergie, est illustré par la purification du carnosol issu du romarin. Par la suite, une nouvelle méthodologie de changement d'échelle en CPC est introduite, afin de pouvoir augmenter de manière efficace la productivité d'une méthode de purification, sans avoir à la redévelopper à grande échelle. L'utilisation de cette méthodologie peut notamment permettre à un utilisateur CPC de réduire considérablement sa consommation de solvants et d'énergie, tout comme sa génération de déchets chimiques. Cette méthodologie de changement d'échelle est appliquée dans le cadre de la purification du carnosol développée en amont. Enfin, la dernière partie de ces travaux est consacrée à la substitution de solvants d'origine pétrolière, par des solvants plus respectueux de l'environnement, principalement d'origine agro-sourcée / The main objective of this thesis is to propose some clues for natural products eco-purification by centrifugal partition chromatography (CPC), a preparative separation technique that uses a biphasic solvent system. First, the manuscript describes the differents preparative techniques commonly used for natural products purification. Then, the centrifugal partition chromatography technique is detailed. The green chemistry concept is subsequently introduced and discussed in regard to its application in purification process. A method development is realized on a small scale CPC, illustrating the reduction of solvent consumption and waste production, through the carnosol purification from rosemary. Later, a new scale-up methodology for CPC is introduced to efficiently transfer the method on larger instrument and increase production. This new methodology will allow for a CPC user to reduce solvent and energy consumption and chemical wastes discharges. This scale-up methodology is then applied to carnosol purification from a rosemary solid extract. Finally, the last part of this manuscript is devoted to petroleum solvents substitution by more environmentally friendly solvents like bio-sourced solvents

Diffusion is a fundamental process which plays a crucial role in many processes occurring in nature. It is governed by the Fickian laws of diffusion. The laws of diffusion explain how diffusive flux is related to the concentration gradient. However, diffusion occurs even when there is no concentration gradient. Chapter 1 introduces diffusion and related concepts such as random walk, Brownian motion, etc. Present understanding with relation to ionic conduction and diffusion in polar solvents and the anomalies observed in the variation of ionic conductivity with ionic radii has also been discussed. Walden’s rule states that the product of limiting ionic conductivity and viscosity is constant for a given ion in different solvents and it is inversely proportional to ionic radius in a given solvent. However, experimental observations indicate that in a given solvent limiting ionic conductivities show an increase followed by a decrease with increase in ionic radii. This is often referred to as the breakdown of Walden’s rule. Several theories have been proposed in the past to explain the breakdown in Waldens rule. Solvent-berg model, continuum based theories and microscopic theories are some of theories that have been proposed. These theories are discussed briefly. The limitations in these theories are also outlined. There are several computer simulation investigations of ions in water and these are discussed. Also described is diffusion of hydrocarbons in zeolites. Various interesting observations such as window effect, nest effect, single file diffusion and the levitation effect are discussed. In Chapter 2, we have analysed the experimental ionic conductivity data as a function of the ionic radius for monovalent cations and anions in aqueous solution. Molecular dynamics simulations on LiCl and CsCl dissolved in water are also reported. The results suggest that the activation energy is responsible for the anomalous dependence of ionic conductivity on ionic radii. It is seen that ions with high conductivity posses low activation energy. The reason for the variation of activation energy with ionic radii are explained in terms of Derouane’s mutual cancellation of forces or levitation effect. This provides an alternative to the existing theories. Experimental limiting ionic conductivity, λ0 of different alkali ions in water shows markedly different dependences on pressure. Existing theories such as that of Hubbard-Onsager are unable to explain this dependence on pressure of the ionic conductivity for all ions. Experimental ionic conductivity data shows that smaller ions such as Li+ exhibit a monotonic increase in λ0 with pressure. Intermediate sized ions such as K+ exhibit an increase in λ0 followed by a decrease at still higher pressures. Larger ions such as Cs+ exhibit a monotonic decrease in λ0 with increase in pressure. In the present thesis, we have explored this intriguing behaviour shown by alkali ions in water in the next few chapters. In Chapter 3, we report molecular dynamics investigation of potassium chloride solution (KCl) at low dilution in water at several pressures between 1 bar and 2 kbar. Two different potential models have been employed. One of the models successfully reproduces the experimentally observed trend in ionic conductivity of K+ ion in water over 0.001-2 kbar range at 298K. We also propose a theoretical explanation, albeit at a qualitative level, to account for the dependence of ionic conductivity on pressure in terms of the previously studied Levitation Effect. A number of properties of the solvent in the hydration shell are also reported. In Chapter 4, residence times of water in the solute and water hydration shell are reported for KCl in water as a function of pressure. Two different approaches – Impey, McDonald and Madden’s approach as well as the recently proposed stable state picture (SSP) of Laage and Hynes yield somewhat different values for the residence times. The latter suggests that the hydration shell is more labile. As pressure is varied, the analysis suggests drastic changes in the hydration shell around water and little or no change in the hydration shell of the ions at higher pressures. The residence times τIMM as well as τSSP show a decrease with increase in pressure upto 1.5 kbar and a small increase beyond this pressure. This correlates with the dependence of the ionic conductivity of potassium ion on pressure. Similar correlation is also seen for chloride ion between ionic conductivity and residence time in hydration shell. However, no such correlation is seen in the case of water. We also report variation of residence time as a function of t∗, the minimum time that a water has to leave the hydration shell to be excluded from it. In Chapter 5, a molecular dynamics study of LiCl dissolved in water is reported at several pressures between 1 bar and 4 kbars at 240K. Structural properties such as radial distribution function, distribution of the angle between ion-oxygen and dipole vector of water in the hydration shell, angle between ion-oxygen and OH vector, oxygen-ion oxygen angle for water in the hydration shell, mean residence times by two different approaches are reported. Self-diffusivity of both Li+ and Cl− exhibit an increase with pressure in agreement with the experimentally observed trend. We also report the velocity autocorrelation function as a function of pressure. We show that the changes in these can be understood in terms of the levitation effect. For the first time we report the self part of the intermediate scattering function, Fs(k, t), at different pressures. These show for Li+ at small wavenumber k, a bi-exponential decay with time at low pressures. At higher pressures when the ionic conductivity is high, Fs(k, t) exhibits a single exponential decay. We also report wavenumber dependence of the ratio of the full width at half maximum to 2Dk2. These changes in these properties can be accounted for in terms of the levitation effect. The changes in the void structure of water with pressure plays a crucial role in the changes in ionic conductivity of both the ions. In Chapter 6, a detailed molecular dynamics study of self-diffusivity of model ions in water is presented as a function of pressure. First, we have obtained the dependence of self-diffusivity on ionic radius for both cations and anions by varying the radius of the ion, rion. Self-diffusivity exhibits an increase with ionic radius when rion is small and reaches a maximum at some intermediate value, before decreasing with increase in rion for rion > . The velocity autocorrelation function for different sizes of cations as well as anions suggest that the ion with maximum self-diffusivity has facile motion with little back scattering. These trends can be understood in terms of the levitation effect which relates the dependence of self-diffusivity on ionic radius to the bottleneck radius of the pore network provided by the solvent or water. The ratio ζ, defined as the full width at half maximum of the self part of the dynamic structure factor at wavenumber k to its value (2Dk2) at k = 0 is seen to increase with k for ions far away from the diffusivity maximum while a decrease with k is observed for ions closer to the diffusivity maximum. Calculations have also been carried out at pressures of 0.001, 2 and 4 kbars to obtain the variation of ionic conductivity with pressure for model ions of several different sizes. It is shown that for small ions (rion < ), self-diffusivity increases with pressure or exhibits an increase followed by a decrease. In contrast, we show that whenever ionic radius is large, (rion > ), a decrease in self-diffusivity with increase in pressure is seen. We suggest that there is a relation between the dependence of self-diffusivity on ionic radius and its dependence on pressure. The nature of this relationship arises through the levitation effect. Increase in pressure leads to decrease in the bottleneck radius, thus increasing the levitation parameter. For small ions (rion < ), this will lead to increase in diffusivity whereas for large ions (rion > ) this will lead to decrease in diffusivity. For small ions (rion < ), the increase in pressure leads to lowered back scattering in the velocity autocorrelation function. In contrast to this, for large ions (rion ≥ ), any increase in pressure leads to increase in back scattering in the velocity autocorrelation function. For the 1.7 °A anion, the ratio ζ is seen to exhibit a minimum at intermediate k and increase with k at large k for 0.001 kbar pressure. This changes to a less pronounced minimum at 2 kbars and by 4 kbars to a nearly monotonically decreasing function of k. These changes suggest, in agreement with the predictions of the levitation effect, the approach of the bottleneck radius to values similar to that of the ionic radius of 1.7 °A on increasing pressure to 4 kbars. Thus, this work offers an unification in our understanding of the dependence of ionic conductivity on ionic radius and pressure. It is seen that when the ionic radius is varied the numerator of the expression for levitation parameter is varied whereas by varying the pressure, the denominator is varied. The variation of diffusivity with density of the host medium and degree of disorder of the host medium is explored in Chapter 7. The system consists of a binary mixture of a relatively smaller sized solute (whose size is varied) and a larger sized solvent interacting via Lennard-Jones potential. Calculations have been performed at three different reduced densities of 0.7, 0.8 and 0.933. These simulations show that diffusivity exhibits a maximum for some intermediate size of the solute when the solute diameter is varied. The maximum is found at the same size of the solute at all densities which is at variance with the prediction of the levitation effect. In order to understand this anomaly, we have carried out additional simulations in which we have varied the degree of disorder at constant density and find that the diffusivity maximum gradually disappears with increase in disorder. We have also carried out simulations in which we have kept the degree of disorder constant but changed only the density. We find that the maximum in diffusivity is now seen to shift to larger distances with decrease in density. In these simulations we have characterized the disorder by constructing the minimal spanning tree. These results are in excellent agreement with the predictions of the levitation effect. They suggest that the effect of disorder is to shift the maximum in diffusivity towards smaller solute radius while that of the decrease in density is to shift it towards larger solute radius. Thus, in real systems where the degree of disorder is lower at higher density and vice versa, the effect due to density and disorder have opposing influences. These are confirmed by the changes seen in the velocity autocorrelation function, self part of the intermediate scattering function and activation energy. In Chapter 8 we report a molecular dynamics study of the dependence of diffusivity of the cation on cation radii in molten superionic salt containing iodine ion. In this study, we have employed modified Parinello-Rahman-Vashistha interionic pair potential proposed by Shimojo et al (F. Shimojo and M. Kobayashi, J. Phys. Soc. Jpn 60, 3725 (1991)). Our results suggest that the diffusivity of the cation exhibits an increase followed by a decrease as the ionic radius is increased. Several other properties like velocity auto correlation function, intermediate scattering function, activation energy are reported. The next two chapters deal with diffusion of hydrocarbon isomers containing aromatic moiety. Chapter 9 reports structure, energetics and dynamic properties of the three isomers of trimethyl benzene in β-zeolite. Monte Carlo and molecular dynamics simulations have been performed at 300K. Of the three isomers, it is observed that 1,2,4-trimethyl benzene(124 TMB) shows fast dynamics inside the channels of β-zeolite. It is seen that both translational and rotational diffusivities are in the order D (124 TMB) > D (123 TMB) > D (135 TMB). 124 TMB seems to perform jumps between perpendicular channels more frequently whereas 123 and 135 isomers experience more hindrance to these jumps. It is also shown that there is a lower energetic barrier for 124 TMB across the window that separates two perpendicular channels in β-zeolite. Reorientational correlation functions suggest that reorientation of C6 axis (axis perpendicular to the plane of the phenyl ring) is highly restricted in case of 135 TMB. Reorientation of C2 axis (axis on the plane of the phenyl ring) seems to be more facile than that of C6 axis in case of both 123 TMB and 135 TMB. And interestingly, C6 and C2 axis reorientations are equally facile in case of 124 TMB. Chapter 10 presents molecular dynamics simulation results carried out on an equimolar binary mixture of cumene (isopropyl benzene) and pseudo-cumene (1,2,4-trimethyl benzene) in zeolite-NaY at four different temperatures. We compare different structural, energetic and dynamic properties of cumene and pseudo-cumene in zeolite-NaY. Our results suggest that both translational and rotational diffusivities are higher for cumene as compared to pseudo-cumene. Potential energy landscapes show that there is an energetic barrier for diffusion past the 12 MR window plane that separates two neighboring super cages. Such an energetic barrier is large for pseudo-cumene (3 kJ/mol) as compared to that of cumene (1.5 kJ/mol). Activation energies corresponding to both translational and rotational diffusion suggest that pseudo-cumene encounters larger energetic barriers for both translation and rotation as compared to cumene. Reorientational correlation functions suggest that reorientation of C2 axis is more facile than that of C6 axis in case of both cumene and pseudo-cumene. Activation energies corresponding to reorientational relaxations suggest that C6 axis encounters larger energetic barriers as compared to C2 axis in case of both cumene and pseudo-cumene. Chapter 11 discusses the main conclusions of the thesis and directions for future work.

Les huiles essentielles et les arômes des plantes constituent un réel potentiel pour l’industrie dans le but de substituer aux composés synthétiques ayant des effets néfastes sur la santé et l’environnement. Afin de contribuer aux principes de la chimie verte, cette étude porte sur l’éco-extraction et la valorisation des extraits naturels des plantes et le développement d’une nouvelle technologie «verte» pour l’extraction des composés aromatiques naturels. La première partie de ce manuscrit mets en évidence les propriétés biologiques des huiles essentielles et des arômes et l’importance de leurs applications dans différents domaines surtout le domaine agroalimentaire. Dans la deuxième partie, les études sur les propriétés biologiques des huiles essentielles des plantes des Alliacées montrent leurs bonnes activités antioxydantes et antimicrobiennes. Ces résultats encourageants ont permis dans la troisième partie de valoriser les sous-produits d’oignon issus de la turbo hydrodistillation et qui sont considérés normalement comme déchets. La technique offre une bonne extraction des composés phénoliques et des flavonoïdes utilisant l’eau comme solvant naturel. La quatrième et la dernière partie de ce travail s’est orientée vers l’optimisation et le développement d’une technologie « verte» utilisant les ultrasons et l’huile de tournesol comme solvant naturel pour l’extraction des composés aromatiques du thym. Cette nouvelle approche écologique permet l’extraction des absolues dépourvues de cire et des résidus de solvants pétroliers, contenant la teneur la plus élevée en thymol et exerçant la plus forte activité antioxydante. / Plant essential oils and aromas are a real potential for the industry to substitute the synthetic compounds that might have harmful effects on the human health and the environment. In order to contribute to the green chemistry principles, this study focuses on the “eco-extraction” and valorization of natural plant extracts and the development of a new «green» technology for the extraction of aromatic compounds. The first part of this manuscript highlights the biological properties of the essential oils and aromas and the importance of their applications in various sectors especially in the food industry. In the second part, studies on the biological properties of the essential oils from plants of the Alliaceae family show their good antioxidant and antimicrobial activities. These encouraging results have allowed in the third part to evaluate the onion by-products resulting from the turbo hydrodistillation, which are normally considered as waste. The technique offers a good extraction of flavonoids and phenols using water as natural solvent. The fourth and final part of this work deals with the optimization and the development of a «green» technology using ultrasound and sunflower oil as a natural solvent for the extraction of aromatic compounds from thyme. This new ecological approach allows the extraction of absolutes free from waxes and petroleum solvent residues, having the highest content in thymol and exerting the strongest antioxidant activity.

Le dichlorométhane (DCM ; CH2Cl2) est un polluant chloré toxique émis dans l’environnement principalement par les activités industrielles. Ce polluant peut être dégradé par des bactéries méthylotrophes qui utilisent des composés en C1 réduits comme seule source de carbone et d’énergie. La protéobactérie Methylorubrum extorquens DM4 porte quatre gènes dcm au sein du transposon catabolique dcm très conservé chez les bactéries dégradant le DCM. Le gène dcmA code la DCM déshalogénase de la famille des glutathion-S transférases essentielle à la croissance avec le DCM. Son activité est modulée par DcmR, un facteur de transcription qui régule l’expression de dcmA ainsi que son propre gène, orienté de manière divergente par rapport à dcmA. DcmR porte un domaine hélice-tour-hélice de fixation à l'ADN et un second domaine appelé methanogen / methylotroph, DcmR sensory (MEDS) potentiellement impliqué dans la fixation d’un composé hydrocarboné ligand. Les objectifs de ma thèse étaient de répondre à plusieurs questions : i) quel est le niveau d’expression des transcrits dcm et des protéines correspondantes par rapport à d’autres gènes et protéines dont l’abondance est modifiée en réponse à la croissance sur le DCM ? ii) Comment le facteur de transcription DcmR intervient-il dans la régulation des gènes dcm ? iii) Quelle est la variabilité du gène dcmR et de son environnement génétique in situ ? Des méthodes globales « -omiques » de transcriptomique et de protéomique ont permis d’inventorier les ARN et les protéines dont l’abondance varie chez la souche sauvage DM4 cultivée soit avec le DCM ou le méthanol, le substrat de référence de la méthylotrophie. Les gènes dcm sont parmi les plus exprimés en présence de DCM, ce qui confirme leur régulation en présence de DCM. Deux approches complémentaires ciblant la détermination des sites d’initiation de la transcription (TSS-Seq) et de la traduction (N-terminome) ont permis la recherche de motifs de régulation dans les régions 5’UTR (5’-untranslated terminal region) et les promoteurs de gènes régulés en réponse à la croissance avec le DCM. Le rôle régulateur de dcmR a été étudié en comparant les phénotypes de croissance, l’activité promotrice par fusion transcriptionnelle, la quantification des ARN et des protéines des gènes dcm chez la souche sauvage par rapport à des mutants du gène dcmR seul ou combiné à d’autres gènes dcm mutés. Ces travaux ont permis de confirmer qu’en absence de DCM, DcmR inhibe la transcription de son propre gène ainsi que celle de dcmA. Outre DcmR, la répression nécessite aussi l’expression d’au moins un des autres gènes dcm et ceci par un mécanisme indépendant des boîtes de 12 pb conservées dans les promoteurs des gènes dcmR et dcmA. Lors de la croissance en condition DCM, l’absence du gène dcmR confère une vitesse de croissance ralentie, qui ne résulte pas d’une différence de production des ARN et des protéines codées par le transposon dcm. Pour que l’expression du gène dcmA soit activée, l’ensemble de la région intergénique entre dcmR et dcmA doit être présente, ce qui suggère la présence de sites de régulation pour la fixation d’un facteur de transcription indépendant de DcmR. L’ensemble de ces résultats a permis de proposer un nouveau modèle de régulation des gènes dcm. Alors que le gène dcmR a été détecté en quantité similaire à celle du gène dcmA par qPCR dans des échantillons de sites contaminés par le DCM, une analyse bioinformatique à partir des données de séquences indique que des gènes dcmR-like sont trouvés dans d’autres contextes génétiques que celui du transposon catabolique dcm. Ainsi, DcmR pourrait exercer un rôle de régulateur dans d’autres contextes ouvrant de nouvelles pistes pour l’identification des ligands du domaine MEDS. / Dichloromethane (DCM; CH2Cl2) is a toxic chlorinated pollutant mainly emitted in the environment through industrial activities. Some methylotrophic bacteria that utilize reduced one carbon compounds as sole source of carbon and energy can degrade DCM. The four dcm genes found in the catabolic dcm transposon are highly conserved among DCM-degrading bacteria, including in the Proteobacterium Methylorubrum extorquens DM4. The dcmA gene encodes a DCM dehalogenase of the glutathione-S transferase family which is essential for growth with DCM. The transcriptional factor DcmR regulates the expression of dcmA and its own gene, two genes which are divergently oriented. DcmR consists of a helix-turn-helix domain for DNA fixation and of a domain, called MEDS for ‘methanogen / methylotroph, DcmR sensory’, proposed to bind a hydrocarbon ligand. The aim of my PhD was to address the following questions: i) What is the expression level of dcm transcripts and their corresponding proteins compared to other genes and proteins whose abundance is modulated in response to growth with DCM? ii) How does DcmR regulate the expression of the dcm genes? iii) How variable is dcmR and its genetic environment in situ? Global transcriptomic and proteomic approaches were followed to inventory transcripts and proteins whose abundance varied in the wild-type DM4 strain grown with DCM compared to with methanol, the reference substrate for methylotrophy. The dcm genes were among the most expressed in cultures grown with DCM, confirming their regulation in response to DCM. The identification of transcription start sites (TSS-Seq) and translation starts (N-terminome) allowed the subsequent search for regulation motifs in the promoter and 5’-untranslated terminal regions (5’UTR) of regulated genes in response to DCM utilization. The role of dcmR was studied using growth phenotypes, promoter activities in transcription fusion assays, dcm transcript and corresponding protein product quantification, comparing the wild-type strain with strains mutated only in dcmR, or also in other dcm genes. In absence of DCM, DcmR inhibited the transcription of its own gene and of dcmA. In addition to DcmR, repression requires the expression of one of the other dcm genes by a mechanism that does not involve previously predicted DcmR-binding sites (a 12 bp conserved box shared by dcmR and dcmA promoters). Mutants with impaired dcmR show slower growth with DCM, although no difference in transcript or protein abundance encoded by the dcm transposon was observed. The complete dcmR-dcmA intergenic region was shown to be required to activate dcmA expression. This suggests that regulatory sites present in the intergenic region may be involved in DcmR-independent transcription activation. Taken together, these data allowed to propose a new model of dcm gene regulation. The dcmR gene was detected and quantified in similar amounts as dcmA in DCM-contaminated environmental samples, despite the fact that bioinformatics analysis of sequence databases suggests that dcmR-like genes are not necessary associated with the catabolic dcm transposon. Thus, DcmR may play a role in other contexts. This may provide new leads for future investigations of potential ligands of the MEDS domain.

No description available.

Contexte : Le rôle de l’exposition professionnelle aux solvants dans la survenue de cancers des voies aéro-digestives supérieures (VADS), suggéré dans quelques études, n’a été que peu examiné, malgré leur utilisation très répandue en milieu de travail. Objectif : L’objectif de cette thèse état d’évaluer les associations entre les expositions professionnelles aux solvants et le risque de cancer des VADS. Méthodes : Ce travail est basé sur les données de l’étude Icare, une large étude cas-témoins en population générale conduite en France entre 2001-2007. L’analyse a été restreinte aux hommes et a porté sur 1857 cas de carcinome épidermoïde de la cavité buccale, du pharynx et du larynx, et 2780 témoins. L’histoire professionnelle détaillée ainsi que les consommations de tabac et d’alcool ont été recueillies par questionnaire. Les expositions aux solvants ont été évaluées à l’aide de matrices emplois-expositions et incluaient cinq solvants chlorés (perchloroéthylène, trichloroéthylène, chlorure de méthylène, chloroforme, tétrachlorure de carbone), cinq solvants pétroliers (benzène ; essences carburant ; gazole, fiouls et kérosène ; essences spéciales ; white spirits) et cinq solvants oxygénés (cétones et esters ; alcools ; éther éthylique ; éthylène glycol ; tétrahydrofurane). Les odds-ratios ajustés sur les consommations de tabac et d’alcool et d’autres facteurs de confusion potentiels, et les intervalles de confiances à 95% ont été estimés par régression logistique. Résultats : Aucune association significative n’a été mise en évidence entre exposition professionnelle aux solvants chlorés, pétroliers et oxygénés étudiés et le risque de cancer de l’ensemble des VADS. Dans l’analyse par localisation, le risque de cancer du larynx augmentait significativement avec l’exposition cumulée au perchloroéthylène. Des risques élevés de cancer de l’hypopharynx, bien que non significatifs, étaient observés pour les hommes exposés à des niveaux élevés de chlorure de méthylène, de white spirits et de tétrahydrofurane. Une association entre exposition au tétrahydrofurane et cancer de la cavité buccale était également suggérée. Aucune association claire n’était observée pour les autres solvants, quelle que soit la localisation de cancer. Conclusion : Des associations positives ont été observées avec plusieurs solvants spécifiques. Toutefois, dans l’ensemble, nos résultats ne sont pas en faveur d’un rôle majeur de l’exposition aux solvants dans la survenue de cancer des VADS. / Background : The role of occupational exposure to solvents in the risk of head and neck cancer has been suggested in some studies but has been few investigated, despite their widespread use in the workplace. Objective: The objective of this thesis was to examine the associations between occupational exposure to solvents and the risk of head and neck cancer. Methods: This work is based on data from the ICARE study, a large population-based case-control study conducted in France between 2001 and 2007. The analysis was restricted to men and included 1,857 cases of squamous cell carcinoma of the oral cavity, pharynx and larynx and 2,780 controls. Detailed occupational lifetime as well as alcohol and tobacco consumptions were collected by questionnaires. Exposure to solvents was assessed by job exposure matrices and included five chlorinated solvents (perchloroethylene, trichloroethylene, methylene chloride, chloroform, carbon tetrachloride), five petroleum-based solvents (benzene; gasoline; diesel, fuels and kerosene; special petroleum products; white spirits) and five oxygenated solvents (ketones and esters; alcohols; diethyl ether; ethylene glycol; tetrahydrofuran). Odds-ratios adjusted for smoking, alcohol drinking and other potential confounders and 95% confidence intervals were estimated with logistic models. Results: No significant association was found between occupational exposure to chlorinated, petroleum-based and oxygenated solvents and the risk of head and neck cancer overall. In subsite analysis, the risk of laryngeal cancer increased with cumulative exposure to perchloroethylene. Non-significantly elevated risks of hypopharyngeal cancer were found in men exposed to high cumulative levels of methylene chloride, white spirits and tetrahydrofuran. An association between exposure to tetrahydrofuran and oral cavity cancer was also suggested. No other clear association was found for the other solvents under study, for any cancer site. Conclusion: Although positive associations were observed for several solvents, overall the results do not suggest a substantial role of exposure to solvents in head and neck cancer risk.

Comme de nombreux colloïdes, des nanoparticules métalliques recouvertes de ligands en suspension s’organisent au-delà d’une fraction volumique seuil et forment ce que l’on appelle un « supracristal ». Ce sont ainsi des systèmes modèles, déjà largement étudiés à partir de suspensions dans des solvants volatils, qui permettent de mieux comprendre l’auto-assemblage de sphères déformables. Les interactions qui conduisent à l’auto-assemblage sont couramment décrites par une compétition entre une attraction de van der Waals entre les cœurs métalliques et une répulsion entre les ligands qui va dépendre de l’affinité entre les ligands et le solvant. Un effet du solvant a déjà été observé sur l’auto-organisation de nano-objets. En mesurant par diffusion de rayons X aux petits angles le facteur de structure de suspensions de nanoparticules d’or greffées, j’ai pu sonder de façon systématique les interactions entre des nanoparticules en suspension avec plusieurs tailles de cœur, des ligands alcane-thiols de longueur différente et dans différents solvants à la fois volatils et non volatils. J’ai ainsi pu mettre en évidence une interaction attractive inattendue dans des alcanes linéaires flexibles et dont l’intensité augmente avec la longueur de l’alcane. Pour corréler les interactions entre particules à leur diagramme de phase, j’ai suivi le processus de cristallisation dans des suspensions en solvant volatil ou partiellement volatil ainsi qu’en émulsion, techniques qui permettent d’augmenter lentement la concentration en nanoparticules. Les interactions attractives induites par le solvant contribuent ainsi à la formation de supracristaux à de très faibles fractions volumiques. A de fortes concentrations, la structure des supracristaux ne dépend pas du solvant utilisé mais, à forte densité de greffage, du rapport R entre la longueur des ligands et le diamètre du cœur d’or. Pour un rapport R voisin de 0.7, la structure finale observée est cubique centrée, la structure à concentration intermédiaire étant cubique à faces centrées. Pour un rapport R deux fois plus petit, une structure originale a été mise en évidence. Il s’agit d’une structure hexagonale de grand paramètre de maille, analysée comme une phase de Frank et Kasper de type MgZn2 ou C14. C’est la première fois qu’une telle phase à empilement local tétraédrique est observée dans un système de sphères monodisperses molles. L’existence de cette phase ainsi que le rôle du rapport R a pu être interprétée en estimant quantitativement la compétition entre l’attraction de van der Waals forte et l’entropie des ligands. / As many colloids, metallic nanoparticles grafted with hydrophobic ligands self-assemble above a volume fraction threshold and thus build superlattices. These model systems, which are widely studied when suspended in volatile oils, enable a better understanding of soft spheres self-assembly.Interactions which lead to self-assembly are commonly described by the combination of van der Waals attraction with interaction between the ligand shells. The shell behavior is controlled by the ligand affinity with the solvent. An effect of the solvent on the self-assembly of nanoparticles has already been observed. Using a small angle X-ray scattering, I measured, through the structure factor, the interactions between gold nanoparticles grafted with alkanethiols in different oils, at various concentrations, for different lengths of ligands and core diameters. I noticed an attractive interaction when using flexible linear alkanes as solvent. It has also been shown that the attraction intensity increases with the solvent length.In order to correlate the interactions between particles to their phase diagram, I studied the crystallization process by concentrating nanoparticles using evaporation in capillaries or Ostwald ripening in emulsions. I showed that attractive interactions induced by the solvent lead to superlattices formation at very low volume fractions.At high concentrations, the superlattice structure depends on the ratio of the ligand length over the gold core diameter. For a ratio around 0.7, the final structure observed is body centered cubic, whereas at lower concentration, it is face centered cubic. When this ratio is halved, an unexpected structure is observed. It is a hexagonal structure with a large lattice parameter. It has been analyzed as a Frank and Kasper’s phase named MgZn2 or C14. It is the first time that this topologically close-packed structure is observed for monodisperse soft spheres. The existence of this phase and the role of the ratio R have been interpreted by considering quantitatively the competition between ligands entropy and the strong van der Waals attraction.

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Данная работа предполагает разработку методики анализа остаточных органических растворителей (ООР) в субстанциях натрия 3,8-диэтоксикарбонил-4-оксо-4Н-пиразоло[5,1-c][1,2,4]триазинид моногидрата (AB-19) и натрия 7-метилтио-4-оксо-3-циано-4Н-[1,2,4]триазоло[5,1-c][1,2,4]триазинида тригидрата (MNR-857). Анализ литературных данных показал, что наиболее предпочтительным методом определения ООР является метод газожидкостной хроматографии с парофазным вводом пробы. Для определения целевых аналитов в исследуемых субстанциях рассмотрены схемы синтеза и строение соединений, и определено, что образцы могут содержать органические растворители II (гексан, пиридин, метанол) и III (этанол, этилацетат, 2-пропанол) классов опасности. Однако опытные данные показали наличие в образцах субстанций только этанола и пропан-2-ола. В связи с этим проводили количественное определение только указанных растворителей. На основе литературных данных были определены исходные условия проведения хроматографического анализа. На практике исходная методика была скорректирована: изменение массы навески, разведения образца, времени выдерживания виалы, установление диапазона концентраций применения методики и метода количественного определения. Разработанную методику анализа подвергали испытаниям на пригодность по таким показателям как специфичность, линейность, предел количественного определения, сходимость, промежуточная прецизионность и правильность. Согласно проведенным исследованиям была доказана достоверность и надежность получаемых с использованием данной методики результатов. Количественное определение ООР с помощью разработанной методики в образцах субстанций AB-19 (3 партии синтеза) и MNR-857 (5 партий синтеза) подтвердило их соответствие требованиям ОФС.1.1.0008.15 по показателю остаточные органические растворители (не более 0,5 % для растворителей III класса опасности). / This paper discusses the development of a method for assay of residual organic solvents in substances in substances such as sodium 3,8-diethoxycarbonyl-4-oxo-4H-pyrazolo[5,1-c][1,2,4]triazinide monohydrate (AB-19) and sodium 7-methylthio-4-oxo-3-cyano-4H-[1,2,4]triazolo[5,1-c][1,2,4]triazinide trihydrate (MNR-857). An analysis of the literature data showed that the most preferred method for determining residual organic solvents is headspace gas-liquid chromatography. To determine the target analytes in the studied substances, synthesis schemes and the structure of compounds are considered. Thanks to this, it was decided that the samples might contain organic solvents of hazard classes II and III. Class II includes hexane, pyridine, methanol, and сlass III includes ethanol, ethyl acetate, 2-propanol. However, experimental data showed the presence of only ethanol and propan-2-ol in the samples of substances. In this regard, an assay was carried out only for these two solvents. Based on literature data, the initial conditions for the chromatographic analysis were determined. The original method was adjusted by changing the test portion of sample, diluting the samples, the holdup time of the vial, as well as by establishing ranges of concentrations and the method of quantitative measurement. The developed analysis method was tested for suitability by such indicators as specificity, linearity, limit of quantification, repeatability, intermediate precision, and accuracy. According to the research, it was demonstrated that the analytical technique is acceptable for solving the task. The residual organic solvents were quantified using the developed procedure for three batches of synthesis of substance AB-19 and for five batches of synthesis of substance MNR-857. The percentage of residual organic solvents in all samples was less than 0.5%. Thus, the results obtained satisfy the requirements of the article OFS.1.1.0008.15 of the State Pharmacopoeia of the Russian Federation in terms of residual organic solvents.