La microspectroscopie Raman confocale apparait comme un outil à fort potentiel pour l'analyse in vivo de la peau, avec des applications innovantes en dermatologie et cosmétologie. Cette technique biophotonique permet d'accéder à des informations moléculaires très spécifiques d'un échantillon ; et ceci de façon totalement non destructive et sans aucun marquage ni préparation particulière. Les travaux référencés ont porté sur la caractérisation des constituants cutanés, l'évaluation du taux d'hydratation, le suivi de la perméation d'actif ou encore le diagnostic des lésions tumorales de la peau. Ces études ont été réalisées avec des configurations instrumentales très diverses. En vue de la conception d'une nouvelle micro-sonde Raman confocale, cette thèse a permis d'établir un cahier des charges précis ; les paramètres instrumentaux ont été définis et leurs effets sur la qualité des enregistrements Raman (fluorescence parasite, rapport signal sur bruit, atténuation du signal en profondeur) ont été évalués. En particulier, l'effet de la longueur d'onde d'excitation, paramètre clé en spectroscopie Raman, a été analysé. Dans nos expérimentations, la répétabilité et la variabilité du signal Raman ont également été prises en compte. Dans le but d'évaluer l'approche Raman pour l'analyse de la perméation cutanée d'actif, la diffusion cutanée de deux molécules (caféine et resvératrol) appliquées en faibles concentrations a été suivie sur un microspectromètre Raman confocal avec des paramètres d'acquisition similaires à ceux définis pour la micro-sonde. Nous avons montré la possibilité d'enregistrer des cinétiques de diffusion, et également mis en évidence des points limitatifs spécifiques de ces études expérimentales. / Confocal Raman microspectroscopy seems to be a tool with strong potential for in vivo skin analyses, and for innovative applications in dermatology and cosmetology fields. This biophotonic technique allows access to very specific molecular information non-destructively and without any particular skin labeling or sample preparation. The referenced work focused on the characterization of the cutaneous constituents, the evaluation of the rate of skin hydration, the monitoring of active ingredient permeation or diagnosis of tumoral skin lesions. These studies were performed with a variety of instrumental configurations. In view of the design of a new confocal Raman micro-probe, this thesis has established a registry of precise specifications; the instrumental parameters were defined, and their effects on the quality of the Raman records (parasite fluorescence, signal to noise ratio, signal attenuation in depth) were evaluated. In particular, the impact of the excitation wavelength, a key parameter in Raman spectroscopy, was analyzed. In our experiments, the reproducibility and the variability of the Raman signal were also considered. In order to evaluate the Raman approach for the analysis of cutaneous permeation of active ingredients, the cutaneous diffusion of two molecules (caffeine and resveratrol) applied in low concentrations were monitored by confocal Raman microspectroscopy using the acquisition parameters defined for the micro-probe. We have demonstrated the possibility of recording diffusion kinetics and at the same time unraveled specific limitations of these experimental studies.

La peau assure la protection de l’organisme contre les agressions extérieures. Pourtant, la peau n’est pas à l’abri des effets inéluctables du vieillissement chronologique. En effet, ce processus a pour conséquence d’altérer la structure des différentes couches cutanées au point d’en affecter leurs caractéristiques fonctionnelles. Ce travail de thèse a pour but de caractériser les modifications moléculaires et structurales de la peau au cours du vieillissement chronologique à l’aide de méthodes non-invasives que sont les spectroscopies optiques vibrationnelles. Ainsi avons-nous pu mettre en évidence des altérations structurales du réseau de collagène dans des échantillons de peaux d’âges différents à l’aide d’une approche basée sur l’imagerie IR-TF (Infrarouge à Transformée de Fourier) en mode polarisé. Par la suite, nous avons démontré l’influence des molécules d’eau sur les changements d’orientation des fibres de collagène avec l’âge en adoptant une méthodologie basée sur la substitution, thermodynamiquement favorable, des molécules d’eau liée au collagène (H2O) par des molécules d’eau deutérée (D2O). Lors d’études réalisées in vivo, nous avons établi des corrélations entre les propriétés physiques (mécanique et statut d’hydratation) et les informations moléculaires du stratum corneum (SC) en traitant les données Raman et biométriques par la méthode des moindres carrés partiels (PLS). En s’appuyant sur la complémentarité des techniques biophysiques employées, ces différentes études ont permis d’illustrer le potentiel des spectroscopies vibrationnelles pour évaluer le vieillissement cutané et en déterminer certaines bases moléculaires. / The skin protects the body against external aggressions. However, the skin is not immune to the inevitable effects of the chronological aging. Indeed, this process leads to several structural alterations of the different cutaneous layers to the point of affecting their functional characteristics. This thesis work aims to assess the molecular and structural changes of the skin during chronological aging by using non-invasive methods such as optical vibrational spectroscopies. To do this, we highlighted the structural modifications of the collagen network in different-aged skin samples by using an approach based on FT-IR imaging (Fourier Transform Infrared) in polarized mode. Subsequently, we demonstrated the influence of water molecules on the changes of collagen fibers with age by adopting a methodology based on the substitution, thermodynamically favorable, of the collagen-bound water molecules (H2O) by deuterated water molecules (D2O). In in vivo studies, we have established correlations between physical and molecular properties of the stratum corneum (SC) by analyzing the Raman and biometric measurements with the Partial Least Squares (PLS) processing. Based on the complementarity of the biophysical techniques employed, these studies permitted to evaluate the impact of the chronological aging on the skin and could open some interesting prospects in both cosmetology and dermatology.

Esta tese tem como objetivo contribuir para o conhecimento e desenvolvimento das células solares sensibilizadas por corantes, através da elaboração de novas espécies supramoleculares e de estudos fundamentais de caracterização do filme de TiO2-P25 e da interface TiO2-Corante pela técnica de microscopia Raman confocal. Os estudos de microcopia Raman confocal da distribuição das fases cristalinas rutilo e anatase em filmes de TiO2-P25 (Degussa) mostraram que a fase rutilo está presente em agregados de diferentes tamanhos (de 250 nm a 3 µm) distribuídos na fase anatase e que estes agregados permaneceram mesmo após os processos de preparação da suspensão coloidal. Na presença do corante N3, verificou-se que a irradiação com laser em 532 nm, usando potência acima de 25 mW cm-2, provoca a absorção e posterior combustão do filme interfacial, com drásticas mudanças na morfologia visualizadas por microscopia Raman confocal, revelando a ocorrência da transformação de anatase em rutilo na região de incidência do feixe de laser. Outro ponto investigado nesta tese, foi a síntese e aplicação de novos sensibilizadores supramoleculares em células solares sensibilizadas por corante (DSC), visando diminuir os processos de recombinação e melhor aproveitamento do espectro solar através da estabilização do corante foto-oxidado e da transferência vetorial de elétron/energia. Além dos trabalhos com novas porfirinas supramoleculares, reproduzidas no apêndice, esta tese apresenta um estudo detalhado de uma série, formada por três espécies diméricas, constituídas pelas unidades [Ru(dcbH2)Cl]+ e [Ru(dmb)2Cl]+, unidas por ligantes ponte com dimensões lineares crescentes, isto é: bpy, bpe e bpeb (dcbH2= ácido-2,2\'-bipiridina-4,4\'-dicarboxílico, dmb= 4,4\'-dimetil-2,2\'- bipiridina, bpy= 4,4\'-bipiridina, bpe= trans-1,2-bis(4-piridil)-etileno e bpeb= trans-1,4- bis[2-(piridil)etenil]-benzeno). O rendimento global das DSCs correspondentes cresceram com o aumento do tamanho da ponte, passando de 2,78 % para o corante dim-Ru-bpy, para 2,89 e 3,25 % nos corantes dim-Ru-bpe e dim-Rbpeb, respectivamente. Foi observado que os valores de Jsc e η têm correlação linear com a distância de separação do buraco e da superfície do eletrodo de TiO2, ratificando que o processo de retardação da recombinação de carga, causado pelo aumento da ponte, é um fator muito importante para esta classe de compostos. Os resultados de IPCE mostraram que a unidade [Ru(dmb)2Cl(P)]+ também é responsável pela fotoinjeção de elétrons no TiO2. Assim, a excitação de qualquer uma das unidades, sempre acaba formando a espécie TiO2(e-)-Ru-Ru(h+) no filme interfacial. Os resultados confirmam as expectativas esperadas no planejamento energético vetorial, no qual o HOMO foi centrado na unidade distante da superfície e o LUMO ficou sobre a dcbH2 diretamente ancorada no superfície do TiO2. / This thesis encompasses our efforts to improve the knowledge and contribute to the development of dye-sensitized solar cells, by focusing on supramolecular design of new dyes and on critical aspects of the morphology of the TiO2-P25 and TiO2-Dye interfaces, as probed by confocal Raman microscopy. According to Raman imaging of the crystalline phase distribution of TiO2-P25 (Degussa) rutile was present as aggregates of different sizes (from 250 nm to 3 µm) dispersed in the 25 nm anatase powder, persisting even after applying criterious procedures for generating uniform colloidal suspensions. In addition, the irradiation of TiO2-P25 films containing adsorbed N3 dye, using a 532 nm laser (power over to 25 mW cm-2) led to the absorption and instantaneous combustion of the dye, promoting drastic local changes associated with the anatase to rutile conversion. The design and application of new supramolecular sensitizers in DSC has also been pursued, aiming an improvement of efficiency by slowing down the electron-hole recombination process, while enhancing the light harvesting effects in the visible range by means of vectorial electron/energy transfer. Studies concerning another efficient supramolecular porphyrin dye, have also been performed and published (Appendix). As the main subject, a series of dimeric species have been reported. They were based on the [Ru(dcbH2)Cl]+ and [Ru(dmb)2Cl]+ units connected by linear bridging ligands of increasing lengths, such as: bpy, bpe and bpeb (dcbH2= 2,2\'-bipyridine-4,4\'-dicarboxylic acid, dmb= 4,4\'-dimethyl-2,2\'-bipyridine, bpy= 4,4\'-bipyridine, bpe= trans-1,2-bis(4- pyridyl)-ethylene e bpeb= trans-1,4-bis[2-(4-pyridyl)ethenyl]-benzene). The overall yield of the corresponding DSCs increased with the bridging ligand length, from 2.78 % for dim-Ru-bpy, to 2.89 % and 3.25 % for dim-Ru-bpe and dim-Ru-bpeb, respectively. The electrochemical parameters associated with the short circuit current exhibited a linear correlation with the hole separation distance between the TiO2 electrode and the binuclear dye terminal, confirming that the retardation of charge recombination through the increasing distance is indeed a relevant factor for this series of compounds. The IPCE results indicated that the [Ru(dmb)2Cl(P)]+ unit is also involved in electron transfer, such that the dye excitation always leads the TiO2(e-)-Ru-Ru(h+) species. This conclusion confirmed the success of the supramolecular design and vectorial transfer strategy, in which the HOMO center was placed far away from the surface, but in communication with the LUMO center located at the dcbH2 group anchored on TiO2.

Esta tese tem como objetivo contribuir para o conhecimento e desenvolvimento das células solares sensibilizadas por corantes, através da elaboração de novas espécies supramoleculares e de estudos fundamentais de caracterização do filme de TiO2-P25 e da interface TiO2-Corante pela técnica de microscopia Raman confocal. Os estudos de microcopia Raman confocal da distribuição das fases cristalinas rutilo e anatase em filmes de TiO2-P25 (Degussa) mostraram que a fase rutilo está presente em agregados de diferentes tamanhos (de 250 nm a 3 µm) distribuídos na fase anatase e que estes agregados permaneceram mesmo após os processos de preparação da suspensão coloidal. Na presença do corante N3, verificou-se que a irradiação com laser em 532 nm, usando potência acima de 25 mW cm-2, provoca a absorção e posterior combustão do filme interfacial, com drásticas mudanças na morfologia visualizadas por microscopia Raman confocal, revelando a ocorrência da transformação de anatase em rutilo na região de incidência do feixe de laser. Outro ponto investigado nesta tese, foi a síntese e aplicação de novos sensibilizadores supramoleculares em células solares sensibilizadas por corante (DSC), visando diminuir os processos de recombinação e melhor aproveitamento do espectro solar através da estabilização do corante foto-oxidado e da transferência vetorial de elétron/energia. Além dos trabalhos com novas porfirinas supramoleculares, reproduzidas no apêndice, esta tese apresenta um estudo detalhado de uma série, formada por três espécies diméricas, constituídas pelas unidades [Ru(dcbH2)Cl]+ e [Ru(dmb)2Cl]+, unidas por ligantes ponte com dimensões lineares crescentes, isto é: bpy, bpe e bpeb (dcbH2= ácido-2,2\'-bipiridina-4,4\'-dicarboxílico, dmb= 4,4\'-dimetil-2,2\'- bipiridina, bpy= 4,4\'-bipiridina, bpe= trans-1,2-bis(4-piridil)-etileno e bpeb= trans-1,4- bis[2-(piridil)etenil]-benzeno). O rendimento global das DSCs correspondentes cresceram com o aumento do tamanho da ponte, passando de 2,78 % para o corante dim-Ru-bpy, para 2,89 e 3,25 % nos corantes dim-Ru-bpe e dim-Rbpeb, respectivamente. Foi observado que os valores de Jsc e η têm correlação linear com a distância de separação do buraco e da superfície do eletrodo de TiO2, ratificando que o processo de retardação da recombinação de carga, causado pelo aumento da ponte, é um fator muito importante para esta classe de compostos. Os resultados de IPCE mostraram que a unidade [Ru(dmb)2Cl(P)]+ também é responsável pela fotoinjeção de elétrons no TiO2. Assim, a excitação de qualquer uma das unidades, sempre acaba formando a espécie TiO2(e-)-Ru-Ru(h+) no filme interfacial. Os resultados confirmam as expectativas esperadas no planejamento energético vetorial, no qual o HOMO foi centrado na unidade distante da superfície e o LUMO ficou sobre a dcbH2 diretamente ancorada no superfície do TiO2. / This thesis encompasses our efforts to improve the knowledge and contribute to the development of dye-sensitized solar cells, by focusing on supramolecular design of new dyes and on critical aspects of the morphology of the TiO2-P25 and TiO2-Dye interfaces, as probed by confocal Raman microscopy. According to Raman imaging of the crystalline phase distribution of TiO2-P25 (Degussa) rutile was present as aggregates of different sizes (from 250 nm to 3 µm) dispersed in the 25 nm anatase powder, persisting even after applying criterious procedures for generating uniform colloidal suspensions. In addition, the irradiation of TiO2-P25 films containing adsorbed N3 dye, using a 532 nm laser (power over to 25 mW cm-2) led to the absorption and instantaneous combustion of the dye, promoting drastic local changes associated with the anatase to rutile conversion. The design and application of new supramolecular sensitizers in DSC has also been pursued, aiming an improvement of efficiency by slowing down the electron-hole recombination process, while enhancing the light harvesting effects in the visible range by means of vectorial electron/energy transfer. Studies concerning another efficient supramolecular porphyrin dye, have also been performed and published (Appendix). As the main subject, a series of dimeric species have been reported. They were based on the [Ru(dcbH2)Cl]+ and [Ru(dmb)2Cl]+ units connected by linear bridging ligands of increasing lengths, such as: bpy, bpe and bpeb (dcbH2= 2,2\'-bipyridine-4,4\'-dicarboxylic acid, dmb= 4,4\'-dimethyl-2,2\'-bipyridine, bpy= 4,4\'-bipyridine, bpe= trans-1,2-bis(4- pyridyl)-ethylene e bpeb= trans-1,4-bis[2-(4-pyridyl)ethenyl]-benzene). The overall yield of the corresponding DSCs increased with the bridging ligand length, from 2.78 % for dim-Ru-bpy, to 2.89 % and 3.25 % for dim-Ru-bpe and dim-Ru-bpeb, respectively. The electrochemical parameters associated with the short circuit current exhibited a linear correlation with the hole separation distance between the TiO2 electrode and the binuclear dye terminal, confirming that the retardation of charge recombination through the increasing distance is indeed a relevant factor for this series of compounds. The IPCE results indicated that the [Ru(dmb)2Cl(P)]+ unit is also involved in electron transfer, such that the dye excitation always leads the TiO2(e-)-Ru-Ru(h+) species. This conclusion confirmed the success of the supramolecular design and vectorial transfer strategy, in which the HOMO center was placed far away from the surface, but in communication with the LUMO center located at the dcbH2 group anchored on TiO2.

<p>Chromatography is a powerful technique, essential in chemical analyses and preparative separation in industry and research. Many different kinds of chromatographic material are needed, due to the large variety of applications. Detailed methods of characterisation are needed to design new chromatographic materials and understand their properties. In this thesis, confocal Raman spectroscopy and surface enhanced Raman spectroscopy (SERS) have been applied to micrometer-size chromatographic beads, for which these techniques have not been used earlier. New methodology, optimized for use with the chromatographic beads, has been developed and evaluated. </p><p>Confocal spectroscopy has been used to determine distributions of functional groups within single chromatographic beads. This distribution is of great importance in determining the chromatographic properties, since the material is porous and the solute molecules can diffuse inside the beads. Most of the confocal experiments have been performed with Raman spectroscopy; fluorescence spectroscopy, using Nd³⁺ ions or dye-labelled proteins as fluorescence probes, has been used for comparison. </p><p>The concentration of adsorbed analytes is very low within the beads. SERS was therefore used to enhance the Raman signal. SERS-active surfaces were prepared by incorporating gold nano-particles into the interior of the bead. TEM measurements showed that the gold nano-particles could be observed throughout, and it was possible to record analyte spectra from different positions within the bead. Enhanced spectra could be obtained both for small test molecules and for larger bio-molecules, although the spectra for the smaller analytes were much more intense.</p>

Chromatography is a powerful technique, essential in chemical analyses and preparative separation in industry and research. Many different kinds of chromatographic material are needed, due to the large variety of applications. Detailed methods of characterisation are needed to design new chromatographic materials and understand their properties. In this thesis, confocal Raman spectroscopy and surface enhanced Raman spectroscopy (SERS) have been applied to micrometer-size chromatographic beads, for which these techniques have not been used earlier. New methodology, optimized for use with the chromatographic beads, has been developed and evaluated. Confocal spectroscopy has been used to determine distributions of functional groups within single chromatographic beads. This distribution is of great importance in determining the chromatographic properties, since the material is porous and the solute molecules can diffuse inside the beads. Most of the confocal experiments have been performed with Raman spectroscopy; fluorescence spectroscopy, using Nd3+ ions or dye-labelled proteins as fluorescence probes, has been used for comparison. The concentration of adsorbed analytes is very low within the beads. SERS was therefore used to enhance the Raman signal. SERS-active surfaces were prepared by incorporating gold nano-particles into the interior of the bead. TEM measurements showed that the gold nano-particles could be observed throughout, and it was possible to record analyte spectra from different positions within the bead. Enhanced spectra could be obtained both for small test molecules and for larger bio-molecules, although the spectra for the smaller analytes were much more intense.

Les taches blanches sont liées à l'hypominéralisation sous la surface de l'émail et sont la 1ère étape du développement de la carie dentaire. La détection précoce des caries dentaires naissantes avant qu'elles n'atteignent le stade de la cavitation offre une opportunité pour des soins dentaires efficaces. Pour réaliser les objectifs de cette étude qui sont de renforcer l'idiome des soins minimalement invasifs, nous avons identifié un modèle de cycle de pH modifié qui permet d’imiter les conditions intra-buccales qui conduisent à la formation de lésions de type tache blanche, dans un court laps de temps. Des techniques optiques non invasives, comme la microscopie Raman confocale et la microscopie multiphotonique, ont été utilisées dans cette étude pour détecter de petits changements dans la composition chimique de l'émail in vitro.De plus, la technique de nano-indentation a été appliquée pour étudier les changements dans les propriétés mécaniques de l'émail et les relier à ceux affectant sa composition chimique après l'induction des caries, afin d‘ajouter de la spécificité chimique-mécanique aux informations sur les lésions sous-surface de l'émail. Les résultats obtenus démontrent un grand potentiel pour les techniques examinées, fournissant une base pour des applications intéressantes dans le diagnostic clinique de différentes pathologies dentaires. Pour résoudre le problème du traitement des lésions primitives sans intervention chirurgicale, l'efficacité de la crème GC Tooth Mousse et du dentifrice contenant de la nano-hydroxyapatite «KAREX» dans l'amélioration de la reminéralisation de l'émail déminéralisé par la localisation du phosphate de calcium amorphe à la surface de la dent a été inspectée. L'étude a indiqué le manque de preuves fiables soutenant l'efficacité des agents reminéralisants dans le traitement des taches blanches. Ce travail doit être poursuivi par d'autres études in-vitro et par des études cliniques. / White spot lesion is the subsurface hypomineralization of enamel indicating the 1st stage of dental caries development. Early detection of incipient dental caries before it reaches the stage of cavitation offers an opportunity for effective dental care. The objectives of this study were to strengthen the idiom of minimally invasive treatment. In order to achieve the specified goals; we have identified a modified pH cycling model that can mimic the intraoral conditions leading to white spot lesions formation in a short time. In addition, non-invasive optical techniques, such as confocal Raman microscopy and multiphoton microscopy were used in this study to detect small changes in the enamel chemical composition in vitro.Furthermore, nano-indentation technique was used to detect the changes in the mechanical properties of enamel and relate them to those affecting its chemical composition after caries induction in order to add chemico-mechanical specificity in providing important information about subsurface lesions in enamel. The obtained results demonstrate a great potential for the examined techniques, providing a basis for interesting applications in the clinical diagnosis of various pathological conditions in dentistry. To treatment the incipient carious lesions non-invasively, the effectiveness of GC Tooth Mousse cream and nHA containing-dentifrice "KAREX" in the remineralization of demineralized enamel through localizing amorphous calcium phosphate at tooth surface has been inspected. The study indicated a lack of reliable evidence supporting the efficacy of remineralizing agents in the treatment of white spot lesions. Within the limitations of this study, further laboratory studies together with clinical research are therefore required to increase the available knowledge on this prevalent subject.

Nous avons étudié l'influence de la fabrication de guides d'ondes optiques par échange protonique doux(SPE) sur les cristaux de niobate de lithium (LN) polarisé périodiquement et nous avons montré que,dans certains cas, ce processus conduit à la création de nanodomaines en surface. Ces nanodomaines enforme d'aiguille peuvent être responsables de la réduction de l'efficacité de conversion non linéaireobservée dans les guides qui sont affectés. Nous avons également étudié l'influence de différents typesd'échange protonique sur la formation, par application d'un champ électrique, de domaines dans le LNcongruent. Cette étude montre que le seuil de nucléation peut être fortement réduit par la présence duguide d'onde et que l'apparition et le développement des domaines en forme de traits est fortementmodifiée. Elle montre également que la fusion des nanodomaines existants au voisinage des parois dedomaine aboutit à la formation de parois élargies et de domaines en forme de dendrites. En irradiantavec un faisceau d'électrons la surface Z- d'un échantillon de LN préalablement soumis à un échangeprotonique doux et recouvert d'une couche de résine électronique, nous avons réussi à former desdomaines avec des formes arbitraires. Par cette technique, nous avons fabriqué des domainespériodiques d'excellente qualité dans des cristaux présentant des guides canaux SPE. Des expériences degénération de deuxième harmonique dans ces guides nous ont permis d'obtenir des efficacités deconversion de 48%/W.cm2 ce qui est conforme aux prédictions ainsi que la forme des spectres d'accordde phase que nous avons observés. Ceci démontre tout l'intérêt de ce processus / The investigation of influence of the soft proton exchange (SPE) optical waveguide (WG) creation onperiodically poled lithium niobate (PPLN) has been done. It has been shown that the WG fabricationprocess can induce the formation of needle like nanodomains, which can be responsible for thedegradation of the nonlinear response of the WG created in PPLN crystals. The domain structure (DS)evolution has been studied in congruent lithium niobate (LN) crystals with surface layers modified bythree different proton exchange techniques. The significant decrease of the nucleation threshold fieldand qualitative change of domain rays nucleation and growth have been revealed. The formation of abroad domain boundary and dendrite domain structure as a result of nanodomains merging in front ofthe moving rays has been demonstrated. The formation of DS in LN with SPE by irradiation of coveredby electron resist polar surface of LN has been investigated. Formation of domains with arbitrary shapesas a result of discrete switching has been revealed. Finally, it has been demonstrated that electron beamirradiation of lithium niobate crystals with surface resist layer can produce high quality periodical domainpatterns after channel waveguide fabrication. Nonlinear characterizations show that the conversionefficiencies and the phase matching spectra conform to theoretical predictions, indicating that thiscombination presents a great interest for device fabrication. Second harmonic generation withnormalized nonlinear conversion efficiency up to 48%/(W cm2) has been achieved in such waveguides

Biologiska processer hos olika organismer påverkas av ultraviolett-B-strålning (UV-B). Kunskapen om förändringar i UV-B-flödet som nått planetens yta under det geologiska tidsspann sedan flercelligt liv uppstod är dock begränsad. Följaktligen har olika biologiska proxies använts som ett sätt att försöka rekonstruera UV-B-flödet som når jordens yta. Växter reagerar på fluktuationen av UV-B genom att variera produktionen av fenolföreningar såsom parakumarsyra (pCA) och ferulinsyra, och därför kan dessa beståndsdelar användas som UV-B-proxys. UV-B-absorberande föreningar (UAC) finns i växternas yttersta skikt, den så kallade kutikulan men även i pollen och sporer, vilka oftaär välbevarade i det geologiska arkivet. Ändå är användningen av kutikula som UV-B-proxy inte lika undersökt. Därför syftar denna magisteruppsats till att analysera kemiska förändringar i fossila växters från trias – jura-gränsen (c. 200 miljoner år gamla)  och deras potential som UV-B-proxys. Eftersom de fossila löven behövde rengöras från sediment och från förkolnat mesofyll, gjordes också en analys av de kemiska förändringarna som orsakades av de olika rengöringsstegen. Icke-destruktiva tekniker, som mikro-FTIR och konfokalt Raman-mikroskop, användes för analysen. Försöken visade att analysen endast kunde utföras med mikro-FTIR på grund av den höga auto-fluorescensen hos de fossila bladen  vilketförhindrade analys med Raman. Signifikanta förändringar orsakades då Schulzes reagens användes för borttagning av mesofyllet, vilket ledde till nitrering av kutikulan. Ett mindre aggressivt och mer hållbart alternativ till denna process har visat sig vara väteperoxid, vilket orsakade mindre förändringar av den fossila kutikulans kemiska sammansättning. Dessutom upptäcktes indikation på förekomst av UAC endast i Ginkgoites regnellii, vilket indikerar att eventuell nedbrytning av dessa föreningar inträffade under diagenesen av de andra analyserade fossilerna. / Biological processes of different organisms are impacted by ultraviolet-B radiation (UV-B). However, knowledge about alterations in the UV-B flux reaching the planet’s surface during the geological past is limited. Consequently, different biological proxies have been used as an approach to attempt reconstructing the UV-B flux reaching Earth’s surface. Plants respond to the fluctuation of UV-B by varying the production of phenolic compounds such as p-coumaric acid (pCA) and ferulic acid, and therefore these constituents can be used as UV-B proxies. UV-B-absorbing compounds (UACs) can be found in the cuticle, pollen, and spores, which are all well-preserved in the geological record. Nevertheless, the use of cuticles as UV-B proxy is not as explored. Hence, this Master’s thesis aimed to analyse chemical changes in the plant cuticles from the Triassic–Jurassic boundary and their potential as UV-B proxies. Moreover, as the cuticles needed to be cleaned from rocks and isolated from coalified mesophyll, an analysis of the chemical changes caused by the cleaning steps was also done. Non-destructive techniques, as micro-FTIR and confocal Raman microscope, were used for the analysis.  Yet, the analysis could only be conducted with micro-FTIR due to auto-fluorescence of the leaves preventing success with Raman. Main changes during the cleaning steps were caused using Schulze’s reagent, which led to nitration of the cuticles. A less aggressive and more sustainable alternative to this process is shown to be hydrogen peroxide, which caused minor alteration of the fossil cuticle’s chemical composition.  Moreover, indication of the presence of UACs was detected only in Ginkgoites regnellii, indicating that possible degradation of these compounds occurred during diagenesis of the other fossils analysed.

La peau est l’organe le plus étendu du corps humain. Doté d’une membrane biologique fine appelée la couche cornée, celle-ci le protège du desséchement et des agressions extérieures chimiques ou mécaniques auxquelles le corps humain doit faire face. Ce travail de thèse a consisté, dans un premier temps, à décrire via la littérature existante les effets de l’âge,dûs au vieillissement intrinsèque et extrinsèque,sur la physiologie cutanée du Stratum Corneum (SC).La partie expérimentale basée sur la microscopie vibrationnelle traitera des variations de la fonction barrière et de l’hydratation du SC lors du vieillissement chronologique et photo-vieillissement. D’autres méthodes ont également été utilisées comme la chromatographie liquide en phase normale couplée à la spectrométrie de masse haute résolution dotée d’une source APCI et d’un détecteur Orbitrap pour l’étude de la composition détaillée des lipides du SC ainsi que des méthodes plus globales comme la PIE ou la cornéométrie. Le caractère non invasif de toutes ces méthodes a permis de réaliser ces études in vivo. L’évolution de la fonction barrière a été étudiée aux niveaux tissulaire, moléculaire et supramoléculaire par micro-spectroscopie confocale Raman et spectroscopie Infrarouge. Puis le lien moléculaire a été fait entre le vieillissement intrinsèque et les céramides de la matrice lipidique intercornéocytaires par chromatographie en phase liquide couplée à la spectrométrie de masse. Les molécules discriminantes entre population jeune et âgée ont été déterminées par analyse chimiométrique. L’évolution de l’hydratation cutanée aux niveaux tissulaire, moléculaire et supramoléculaire a également été l’objet d’une investigation approfondie. Les variations de la composition des NMF et la teneur en eau dans le SC lors du vieillissement cutané ont été mises en lumière en utilisant des descripteurs spectraux Raman. Les variations structurelles des molécules d’eau impactant l’organisation supramoléculaire des édifices lipidiques ont également été évaluées. Au cours du vieillissement, la fonction barrière cutanée et hydratation sont conservées. / Skin is the external surface defining the human body in space. Its outer-most layer is a thin biological membrane, called Stratum Corneum(SC), that protects the internal organs from desiccation as well as chemical or mechanical external aggressions. The present thesis aimsin a first step, to summarize the current knowledge regarding the effects of intrinsic and extrinsic aging on SCphysiology,based on available literature. The experimental part addresses the gaps in our understanding of the effects of chronological aging and photoaging on the SC barrier function and hydration, using traditional methods (such as trans epidermal water loss and skin conductance) as well as more advanced ones (vibrational spectroscopies, liquid chromatography in normal phase tandem mass spectrometry high resolution with an APCI source and an Orbitrap detector. As these methods are non-invasive, all studies have been carried out in vivo. The evolution of the barrier function has been studied at the tissular, molecular and supramolecular levels using confocal Raman micro-spectroscopy and infrared spectroscopy. Then the link between the intrinsic aging and the ceramides of the intercorneocytary lipid matrix has been studied by liquid chromatography tandem mass spectrometry. The discriminant molecules between young and old population have been identified by a chemometric analysis. The evolution of cutaneous hydration at the tissular, molecular and supramolecular level has also been investigated. The variations in the NMF composition and the SC water content have been studied by Raman spectral descriptors. Moreover, the structural variations of water molecules impacting the supramolecular organization of the lipid structures have been evaluated. Chronological aging and chronic exposure to environmental factors mildly affect SC barrier function and hydration levels. However, the processes controlling these properties are affected by aging in a site-dependent fashion.