Les cellules photoacoustiques sont des capteurs optiques qui utilisent l'absorption des photons par des molécules de gaz pour générer une onde de pression proportionnelle à leur concentration. Le signal photoacoustique est également inversement proportionnel au volume de la cellule. La miniaturisation de la cuve permet donc l’amélioration des performances du capteur. Le travail de cette thèse consiste en la conception, la réalisation et la caractérisation de cellules photoacoustiques résonantes différentielles d’Helmholtz (DHR) miniaturisées. Dans un premier temps, des simulations par la méthode des éléments finis de cellules à l'échelle millimétrique ont permis de montrer que la miniaturisation de ce type de résonateur est une voie prometteuse. Aussi, la réalisation ambitieuse d’une cellule DHR sur silicium a été engagée en utilisant les techniques de la microélectronique. Cependant, cette voie de miniaturisation extrême s'est heurtée à des difficultés de réalisation, qui n'ont pas permis d'obtenir des dispositifs fonctionnels. Une alternative de miniaturisation, à l'échelle centimétrique, utilisant des microphones MEMS du commerce, a donc été engagée. Trois cellules fabriquées par différentes méthodes ont été réalisées et testées pour la détection de méthane. La dernière génération a permis la détection d'environ 100 ppb de méthane avec un laser à cascade interbande commercial à 3,357 µm de longueur d’onde. Pour préparer la prochaine génération de cellules, l'optimisation de la géométrie a été effectuée par simulation. Cette optimisation permet d'envisager une augmentation de 43 % du signal par rapport à la cellule la plus performante. / Photoacoustic cells are optical sensors based on the absorption of photons by gas molecules. The pressure wave created by gas relaxation is proportional to the trace gas concentration. Furthermore the photoacoustic signal is inversely proportional to the cell volume. Thus cell miniaturization enables performances improvements. This work consists in designing, realizing and characterizing miniaturized photoacoustic cells, based on the differential Helmholtz resonator (DHR) principle. In a first phase, modeling by the finite element method of millimeter scale cells has shown that the miniaturization of this type of resonator should effectively improve the detection limit. Thus, the ambitious realization of a DHR cell on silicon by the use of microelectronic techniques has been attempted. However, this extreme miniaturization direction encountered design and fabrication difficulties which made the produced devices unusable. To overcome these difficulties, a miniaturization alternative, at the centimeter scale, using commercial MEMS microphones, has been carried out. Three cells have been built by different methods and have been tested for methane detection. The last cell generation can detect around 100 ppb of methane with a commercial interband cascade laser at 3.357 µm of wavelength. Finally, to anticipate the next cell generation fabrication, a geometry optimization has been performed by simulation. This optimization shows that a 43 % signal improvement, compared to the most performant cell already built.

Optical coherence tomography (OCT) and Photoacoustic microscopy (PAM) are two noninvasive, high-resolution, three-dimensional, biomedical imaging modalities based on different contrast mechanisms. OCT detects the light backscattered from a biological sample either in the time or spectral domain using an interferometer to form an image. PAM is sensitive to optical absorption by detecting the light-induced acoustic waves to form an image. Due to their complementary contrast mechanisms, OCT and PAM are suitable for being combined to achieve multimodal imaging. In this dissertation, an optical coherence photoacoustic microscopy (OC-PAM) system was developed for in vivo multimodal retinal imaging with a pulsed broadband NIR light source. To test the capabilities of the system on multimodal ophthalmic imaging, the retina of pigmented rats was imaged. The OCT images showed the retinal structures with quality similar to conventional OCT, while the PAM images revealed the distribution of melanin in the retina since the NIR PAM signals are generated mainly from melanin in the posterior segment of the eye. By using the pulsed broadband light source, the OCT image quality highly depends on the pulse-to-pulse stability of the light source without averaging. In addition, laser safety is always a concern for in vivo applications, especially for eye imaging with a pulsed light source. Therefore, a continuous wave (CW) light source is desired for OC-PAM applications. An OC-PAM system using an intensity-modulated CW superluminescent diode was then developed. The system was tested for multimodal imaging the vasculature of a mouse ear in vivo by using Gold Nanorods (GNRs) as contrast agent for PAM, as well as excised porcine eyes ex vivo. Since the quantitative information of the optical properties extracted from the proposed NIR OC-PAM system is potentially able to provide a unique technique to evaluate the existence of melanin and lipofuscin specifically, a phantom study has been conducted and the relationship between image intensity of OCT and PAM was interpreted to represent the relationship between the optical scattering property and optical absorption property. It will be strong evidence for practical application of the proposed NIR OC-PAM system.

We demonstrate a versatile phase-change sub-micron contrast agent providing three modes of contrast enhancement: 1) photoacoustic imaging contrast, 2) ultrasound contrast with optical activation, and 3) ultrasound contrast with acoustic activation. This agent, which we name 'Cy-droplet', has the following novel features. It comprises a highly volatile perfluorocarbon for easy versatile activation, and a near-infrared optically absorbing dye chosen to absorb light at a wavelength with good tissue penetration. It is manufactured via a 'microbubble condensation' method. The phase-transition of Cy-droplets can be optically triggered by pulsed-laser illumination, inducing photoacoustic signal and forming stable gas bubbles that are visible with echo-ultrasound in situ. Alternatively, Cy-droplets can be converted to microbubble contrast agents upon acoustic activation with clinical ultrasound. Potentially all modes offer extravascular contrast enhancement because of the sub-micron initial size. Such versatility of acoustic and optical 'triggerability' can potentially improve multi-modality imaging, molecularly targeted imaging and controlled drug release. (C) 2017 The Authors. Published by Elsevier GmbH.

Ce travail de thèse s'inscrit dans le domaine de l'imagerie photoacoustique en tant que modalité d'imagerie prometteuse pour la médecine. Il a consisté en l'expérimentation du guidage photoacoustique de la thérapie par ultrasons focalisés de haute intensité (HIFU), et en l'étude de la génération par des agents de contraste spécifiques que sont les nanoparticules d'or. Dans un premier temps lors d'expériences in vitro, une sonde conçue à la fois pour l'imagerie photoacoustique et la thérapie ultrasonore a été utilisée pour démontrer la faisabilité du guidage photoacoustique pour traiter par HIFU une zone tissulaire cible. L'usage de la photoacoustique pour le contrôle des HIFU révèle ainsi son caractère prometteur. Dans un second temps la modélisation physique de la génération photoacoustique par des nanoparticules d'or permet de quantifier les mécanismes non-linéaires thermoélastiques impactant le signal émis. Dans le cas d'une nanosphère d'or unique, nous décrivons en fonction des paramètres du problème l'importance de la contribution de ces phénomènes non-linéaires, qui sont causés par la dépendance en température de la dilatation thermique de l'eau. Nous en tirons un ensemble de prédictions quantitatives quant à l'influence et le poids de ces non-linéarités sur le signal photoacoustique. Puis nous généralisons ces résultats théoriques à une collection de nanosphères d'or, et les confrontons à nos résultats expérimentaux. Nous mettons ainsi en évidence une nouvelle forme prometteuse de contraste en imagerie photoacoustique: le contraste de non-linéarité thermoélastique. / This work falls within the field of photoacoustic imaging as a promising modality for biomedical applications. It consists in experimenting photoacoustic guidance of high intensity focused ultrasound (HIFU) for therapy, and in studying photoacoustic generation by gold nanoparticles as contrast agents. First of all, a probe designed for both photoacoustic imaging and ultrasound therapy was employed for \textit{in vitro} experiments. We demonstrate the feasability of photoacoustic guidance to treat a target embedded in biological tissue, as a promising tool for HIFU control. In a second part we model the physical mechanisms of photoacoustic generation by gold nanoparticles. This allows to quantify thermoelastic nonlinearities impacting the emitted signal, which origins derive from the temperature dependence of the thermal expansion coefficient of water. In the case of a single gold nanosphere, we describe the nonlinear contribution to the signal according to the different parameters of the problem. We infer a set of quantitative predictions concerning the weight of nonlinearities on photoacoustic signals. Then we generalize these theoretical results to a collection of gold nanospheres, and confront them to our experimental results. Thermoelastic nonlinearity thereby offers a promising new type of contrast for photoacoustic imaging.

A espectroscopia fotoacústica (PA) vem sendo largamente utilizada em diversos ramos de pesquisa, principalmente nas investigações de materiais quanto às suas propriedades ópticas e térmicas, demonstrando a sua vasta versatilidade. No presente trabalho, desenvolveu-se uma câmara fotoacústica de uso geral e uma metodologia baseada na técnica fotoacústica, técnica não intrusiva e não destrutiva, para a determinação da porosidade aberta de membranas vítreas. A porosidade aberta de membranas vítreas está relacionada diretamente à permeabilidade e à eficiência dessas membranas em processos de filtração. Essas membranas foram processadas utilizando-se a técnica de preenchimento e garrafas de vidro, encontradas comercialmente, como matéria-prima. Através da utilização de NaCl como o material inerte, foram processadas membranas com diferentes percentagens de poros. A metodologia fotoacústica desenvolvida neste trabalho teve como base modelos teóricos fotoacústicos tradicionais de análise. Esta metodologia foi capaz de determinar a porosidade aberta das membranas e de distinguí-las quanto as suas superfícies. Os resultados obtidos através da técnica fotoacústica foram comparados com os resultados obtidos por porosimetria de mercúrio (técnica intrusiva e destrutiva) e por ensaios de permeabilidade (técnica intrusiva e não destrutiva) com as mesmas membranas. Estas duas técnicas de análise são as usualmente utilizadas em estudos de estruturas porosas. A comparação entre os dados experimentais comprovou a validade dos resultados obtidos com a nova metodologia. Além desta metodologia fornecer a porosidade aberta com relativa facilidade, ela demonstrou ser rápida, não-destrutiva e de baixo custo para cada ensaio nas análises / The photoacoustic (PA) spectroscopy has been used in several research lines, mainly in the investigation of thermal and optical properties related to different materials, showing in this way high versatility. In this work is described the development of a general-purpose photoacoustic cell, and of a methodology based on photoacoustic technique for the determination of the opened porosity of vitreous membranes. The technique is non-intrusive and non-destructive in relation to the membranes. The opened porosity of vitreous membranes is directly related to the permeability and also directly related to the efficiency of these membranes in the filtration processes. Membranes with different percentage of pores were processed by means of filler principle. For this purpose were used glass bottles found commercially as raw material and NaCl as inert material. The photoacoustic methodology developed in this work was based on classical theoretical models found in the literature. This methodology was capable to determine the open porosity of the membranes and to distinguish membranes with different surfaces. The results obtained with the photoacoustic technique were compared with that obtained by means of mercury porosimetry (intrusive and destructive technique) and permeability measurements (intrusive and non destructive technique) with the same membranes. These two techniques are usually used in studies of porous structures. The comparison between the experimental data showed the validity of the results obtained with this new methodology. Moreover, it supplies the opened porosity with relative easiness, in a faster and non-destructive way, and with low cost in the analyses

Small ligand diffusion in heme proteins is not fully understood. To help better understand CO diffusion, three systems were investigated: L29H/F43H site-directed sperm whale myoglobin, horse heart myoglobin in a heavy water buffer, and calix[4]resorcinarene. Binding of copper to calix[4]resorcinarene was photophysically characterized to unravel transient binding of small molecules in heme-copper proteins. Copper binding was found to have a low dissociation constant of approximately 8.6 micrometers.. Reaction profiles using photoacoustic calorimetry were constructed for the myoglobin systems. In deuterium oxide, ligand escape is not rate limited by water entry. Large enthalpy differences arise from the thermodynamic properties of deuterium oxide and the extensive hydrogen bonding network in myoglobin. In the mutant, CO rebinds primarily to the heme and is exothermic with a large volume contraction because of altered electrostatics within the binding pocket and higher water occupancy.

京都大学 / 0048 / 新制・課程博士 / 博士(薬科学) / 甲第21713号 / 薬科博第104号 / 新制||薬科||11(附属図書館) / 京都大学大学院薬学研究科薬科学専攻 / (主査)教授 小野 正博, 教授 山下 富義, 教授 髙倉 喜信 / 学位規則第4条第1項該当 / Doctor of Pharmaceutical Sciences / Kyoto University / DFAM

付記する学位プログラム名: 充実した健康長寿社会を築く総合医療開発リーダー育成プログラム / 京都大学 / 新制・課程博士 / 博士(人間健康科学) / 甲第23127号 / 人健博第89号 / 新制||人健||6(附属図書館) / 京都大学大学院医学研究科人間健康科学系専攻 / (主査)教授 杉本 直三, 教授 黒木 裕士, 教授 松田 秀一 / 学位規則第4条第1項該当 / Doctor of Human Health Sciences / Kyoto University / DFAM

<p><a>Molecular imaging revolutionized the way researchers and clinicians visualize and investigate complex biochemical phenomena, and it is beneficial </a><a>for disease diagnosis, drug design and therapy assessment</a>. Among a variety of different imaging techniques, the non-ionizing and non-invasive photoacoustic (PA) imaging is attracting increased attentions, owing to its high spatial and temporal resolutions with reasonable penetration depth in tissue. Parallel efforts have been the preparation of PA imaging agents which has high PA efficacy and can specifically label the targets at cellular or molecular level. Particularly, there is exponentially growing interest in imaging in the second near-infrared (NIR-II) window (1000–1700 nm), where offers reduced tissue background and improved penetration depth. However, study of PA imaging in the NIR-II window is incomplete, partly due to the lack of suitable materials. Therefore, in my dissertation work I studied NIR-II PA imaging through semiconducting polymer. </p> <p>Firstly, the performance of PA imaging in the NIR-II window is explored by using a semiconducting polymer nanoparticle (SPN) which has strong absorption in the NIR-II window. Compared with lipid, blood and water, such SPN shows outstanding PA contrast in the NIR-II window <i>in situ</i> and <i>in vivo</i>, and an imaging depth of more than 5 cm at 1064 nm excitation is achieved in chicken-breast tissue. These results suggest that SPN as a PA contrast in the NIR-II window opens new opportunities for biomedical imaging with improved imaging contrast and centimeter-deep imaging depth.</p> <p>Next, targeted PA imaging of prostate cancer is achieved by functionalizing a NIR-II absorbing SPN with prostate-specific membrane antigen (PSMA)-targeted ligands. Insights into the interaction of the imaging probes with the biological targets are obtained from single-cell to whole-organ by transient absorption (TA) microscopy and PA imaging. TA microscopy reveals the targeting efficiency, kinetics, and specificity of the functionalized SPN to PSMA-positive prostate cancer at cellular level. Meanwhile, the functionalized SPN demonstrates selective accumulation and retention in the PSMA-positive tumor after intravenous administration <i>in vivo</i>. Taken together, it is demonstrated that BTII-DUPA SPN is a promising targeted probe for prostate cancer diagnosis by PA imaging. </p> <p>Lastly, PA imaging in the NIR-II window is also achieved water-soluble semiconducting polymer, which is prepared by oxygen-doping. After doping, it shows broadband absorption in the entire NIR-II window, with great chemical stability, photostability and biocompatibility. Owing to its merit of broadband absorption, the imaging depth comparison among different NIR-II wavelengths is also achieved. Moreover, this doped semiconducting polymer is readily soluble in normal physiological pH by virtue of carboxyl groups on side chains and tends to aggregate at an acidic environment which results in a 7.6-fold PA enhancement at pH 5.0. Importantly, a 3.4±1.0-fold greater signal in tumor tissue than that in muscle is revealed <i>in vivo</i>. This study provides a more attainable yet effective platform to the field for achieving water-soluble NIR-II absorbing contrast agents for activatable PA imaging. </p>

The aim of the thesis is to study mechanical properties of nanomaterials (multi-layer graphene, silicon, mica) suitable to be used as novel pressure sensors in laser photoacoustic spectroscopy. Membranes (diameter ~ 4 mm, thickness ~ 100 nm) were prepared by mechanical exfoliation method and then attached to a glass window in several slightly different designs. Movement of these membranes was detected using HeNe laser beam reflected from the membrane's surface onto a position sensitive detector. Methanol was used as a model gas and the signal was collected from studied element and microphone simultaneously. Acoustic wave, induced inside a measuring cell by periodic thermal variations, causes the membranes to move. The movement of a membrane is influenced by its mechanical properties, which is possible to determine by fitting the measured data into a mathematical model. Comparison of the output data of all membranes' measurements shows, that the signal intensity is influenced by the method of attaching membrane to a glass window and by volume of free space on a side of a membrane. Metallization of the membrane's surface (~ 70 nm) decreases its springiness thus decreases the sensitivity. Several membranes reached sensitivity comparable with top class microphone.