Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of: Pharmaceutical Technology Consultants: PharmDr. Ondřej Holas, Ph.D. Student: Simona Žemličková Title of thesis: Hybrid polymeric-lipid nanoparticles as drug carriers The work is focused on hybrid polymer-lipid nanoparticles, their advantages obtained from polymer and lipid part, purpose of surface modification, basic properties of nanoparticles, methods of preparation, modification of preparation conditions and use of nanoparticles in medicine. The aim of the experimental part was to prepare nanoparticles composed of polyester and lipid by emulsion evaporation method and nanoprecipitation. Two types of linear polymer poly (lactic-co-glycolic acid) and phosphatidylcholine were used in various ratios. The surfactant used for stabilization was poloxamer Pluronic® F127 and the organic solvents were ethyl acetate and acetone. Curcumin served as a model active substance. The effect of lipid and surfactant on the size and zeta potential of nanoparticles was evaluated. Modification of preparation conditions, which included many process parameters, also influenced the monitored parameters. Encapsulation effectivity and drug loading were also tested. Dissolution tests were performed. It was found that size of nanoparticles...

Les Poly(HydroxyAlcanoates) (PHA) sont des polymères naturels, biodégradables et biocompatibles, synthétisés par de nombreux organismes, et plus particulièrement des procaryotes. Il existe à ce jour plus de 150 types de monomères de PHA différents, accumulés chez différents genres bactériens, en tant que source d’énergie et de carbone. En effet, les granules de PHA intracellulaires sont produites en réponse à un apport en excès de sources de carbone dans l’environnement (glucides, acides gras…), couplé à une carence en éléments azotés nécessaires à la division cellulaire. De par leur caractère biodégradable et biocompatible, les PHA sont employés depuis plus de 20 ans comme biomatériaux dans les domaines pharmaceutiques et biomédicaux, notamment comme micro/nanovecteurs à visée thérapeutique. Ce doctorat met en évidence des méthodes de criblage moléculaire par PCR pour la sélection de bactéries productrices de PHA, isolées de sites hydrothermaux des océans Atlantique et Pacifique au cours de campagnes océanographiques Ifremer. Selon des protocoles de fermentation standardisés et optimisés, des polymères de poly(3-hydroxybutyrate-4-hydroxybutyrate) P(3HB4HB) d’intérêt biomédical ont été produits, puis des études taxonomiques et phylogénétiques ont été menées pour explorer la biodiversité microbienne associée aux environnements marins profonds. Ensuite, des PHA ont été modifiés par réaction thiol-ène photoactivée afin d’obtenir des copolymères hydrosolubles, adaptés pour l’enrobage de nanoparticules poreuses de type Metal-Organic Frameworks (MOF). La caractérisation physicochimique a été réalisée par différentes techniques, et notamment par SEM et STEM-EDX. Les systèmes hybrides poreux MOF-PHA ont ensuite été évalués quant à leur biocompatibilité vis-à-vis de cellules immunitaires (macrophages), par des tests de cytotoxicité et de prolifération cellulaire. Cette étude met en lumière les potentialités de cette nouvelle génération de nanovecteurs, synthétisés pour augmenter le bénéfice thérapeutique tout en minimisant les effets secondaires sur l’organisme humain. / Poly(HydroxyAlkanoates) (PHA) are natural polymers, biodegradable and biocompatible, synthesized by many organisms, especially prokaryotes. There are over 150 kinds of these polyesters, accumulated in a wide variety of bacteria as carbon and energy storage material. PHA granules are deposited intracellularly when microorganisms are cultivated in the presence of an excess of carbon source (glucids, fatty acids...) together with a nitrogenous nutrient deficiency. Due to their biodegradability and biocompatibility, PHA can be used as biomaterials in medical or pharmaceutical fields, and numerous therapeutic micro/nanovectors have already been developed over the past two decades.The present PhD research project highlighted molecular screening methods by PCR for the PHA producing Bacteria selection, isolated during Ifremer cruises from hydrothermal vents in Atlantic and Pacific oceans.According to standardized and optimized fermentation protocols, poly(3-hydroxybutyrate-4-hydroxybutyrate) P(3HB4HB) polymers of biomedical interest were produced, then taxonomic and phylogenetic studies were performed to explore microbial biodiversity associated with deep-sea environments. Next, PHA were modified by ‘click chemistry’ to obtain hydrosoluble copolymers, suitable for coating high porous Metal-Organic Frameworks (MOF) therapeutic nanoparticles. Physico-chemical characterization was performed using different techniques, and more particularly by SEM and STEM-EDX. MOF-PHA hybrid porous systems were then evaluated for their biocompatibility against immune cells (macrophages), by cytotoxicity and cellular proliferation tests. This study highlights potentials of these new generations of nanovectors, synthesized to increase the therapeutic benefit while minimizing side effects on the human body.

Durant ce travail, la synthèse de nanoparticules hybrides composées d'un coeur organique cristallin fluorescent enrobé d'une coquille silicatée a été optimisée dans l'optique du développement d'un nouveau type de traceurs très brillants pour l'imagerie médicale (microscopie de fluorescence excitée à deux photons). La préparation des nanoparticules a pu être adaptée à différents types de fluorophores organiques. La composition de la coquille organosilicatée peut elle aussi être modulée afin de modifier les propriétés de dispersion en solution aqueuse, la charge de surface des particules ou les fonctions chimiques de surface. Le coeur organique de ces particules se dissolvant dans les solvants organiques, différentes stratégies de fonctionnalisation en milieu aqueux ont été réalisées afin d'augmenter la furtivité de ces objets in vivo. Ces différentes méthodes ont permis de faire circuler les nanoparticules dans le flux sanguins de souris. La durée de vie de circulation de ces particules restant cependant courte, de nouvelles stratégies de fonctionnalisation (chimie click) devraient permettre à l'avenir d'allonger ce temps de circulation. / This work has allowed the synthesis of hybrid nanoparticles composed by organic crystal cores embedded in organosilicate shells for the development of in vivo imaging applications(two-photon fluorescence microscopy). The synthesis of these nanoparticles has been adapted with different types of organic dyes. The composition of the silicate shell can also be modulated in order to modify the properties of dispersion in aqueous solution, the surface charge of particles or chemical functions present at the surface. Due to the dissolution of organic cores in organic solvents, different strategies of functionalization were developed in aqueous media to enhance the furtivity of these nanoparticles in vivo. These methods have led to the circulation of the nanoparticles into the bloodstream of mice. However, the circulation lifetime of these nanoparticles is short. New strategies of functionalization (click chemistry) should help in the future to extend this circulation time for the possible development of these new fluorescent tracers.

Cette thèse décrit l’élaboration de nouvelles nanosondes disposant de propriétés permettant leur utilisation pour la microscopie multiphotonique ainsi que la thérapie ciblée du cancer. Dans un premier temps, ce travail s’est concentré sur la synthèse de nanoparticules actives en optique non linéaire et pour la photothérapie. Différents types de nanoparticules ont ainsi été élaborées et caractérisées comme des nanobâtonnets d’or, des nanoparticules de carbure de silicium ou de niobate de potassium, et des nanohybrides couplant ces différentes briques de base. Les nanoparticules ont ensuite été fonctionnalisées par des biomolécules comme l’acide folique afin de leur conférer des propriétés de ciblage spécifique vis-à-vis des cellules cancéreuses. La fonctionnalisation de surface des nanoparticules a été caractérisée de manière approfondie par des techniques avancées telles que la spectroscopie infrarouge, XPS et ToF-SIMS. Dans un second temps, les propriétés optiques non linéaires et thérapeutiques de ces nanoparticules ont été étudiées. Ainsi, ces nanosondes ont été utilisées avec succès pour réaliser le marquage de cellules saines et le ciblage spécifique de cellules cancéreuses pour la microscopie multiphotonique. Enfin, les propriétés photothérapeutiques de ces nanoparticules ont également été étudiées pour réaliser la destruction photoinduite de cellules cancéreuses. / This thesis describes the synthesis of new nanoprobes with properties allowing their use for cancer-targeted multiphotonic microscopy and cancer phototherapy. On the one hand, this work was focused on the synthesis of nanoparticles with non-linear optical and phototherapeutic properties. Different nanoparticles were synthesized and used like gold nanorods, silicon carbide or potassium niobate nanoparticles, and nanohybrids coupling these previous nano-building blocks. These nanoparticles were functionalized with biomolecules like folic acid to provide specific cancer-targeting properties. The surface chemistry of these nanoparticles was carefully evaluated through advanced characterization techniques such as infrared spectroscopy, XPS and ToF-SIMS. On the other hand, optical and therapeutic properties of these nanoparticles were studied. These nanoprobes were successfully used to perform healthy cells labelling and cancer cells targeting for multiphotonic microscopy. Phototherapeutic properties of our nanoparticles were also used to induce light-triggered cancer therapy.

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Progress has been made in using human serum albumin nanoparticles (HSAPs) as carrier systems for targeted treatment of cancer. Human serum albumin (HSA), the most abundant human blood protein, can form HSAPs via a desolvation and crosslinking method, with the size of the HSAPs having crucial importance for drug loading and in vivo performance. Gold nanoparticles have also gained medicinal attention due to their ability to absorb near-infrared (NIR) light. These relatively non-toxic particles offer combinational therapy via imaging and photothermal therapy (PPTT) capabilities. A desolvation and crosslinking approach was employed to encapsulate gold nanoparticles (AuNPs), hollow gold nanoshells (AuNSs), and gold nanorods (AuNRs), into efficiently sized HSAPs for future tumor heat ablation via PPTT. The AuNR-HSAPs, AuNP-HSAPs and AuNS-HSAPs had average particle diameters of 222 ± 5, 195 ± 9 and 156 ± 15, respectively. We simultaneously encapsulated AuNRs and the anticancer drug paclitaxel (PAC), forming PAC-AuNR-HSAPs with overall average particle size of 299 ± 6 nm. Loading of paclitaxel into PAC-AuNR-HSAPs reached 3μg PAC/mg HSA. PAC-AuNR-HSAPs experienced photothermal heating of 46 ˚C after 15 minutes of NIR laser exposure; the temperature necessary to cause severe cellular hyperthermia. There was a burst release of paclitaxel up to 188 ng caused by the irradiation session, followed by a temporal drug release. AuNR-HSAPs were tested for ablation of renal cell carcinoma using NIR irradiation in vitro. Particles created with the same amount of AuNRs, but varying HSA (1, 5 or 20 mg) showed overall particle size diameters 409 ± 224, 294 ± 83 and 167 ± 4 nm, respectively. Increasing HSAPs causes more toxicity under non-irradiated treatment conditions: AuNR-HSAPs with 20 mg versus 5 mg HSA caused cell viability of 64.5% versus 87%, respectively. All AuNR-HSAPs batches experienced photothermal heating above 42 ˚C. Coumarin-6, was used to visualize the cellular uptake of AuNR-HSAPs via fluorescence microscopy. Finally, camptothecin (CPT) an antineoplastic agent and BACPT (7-butyl-10-aminocamptothecin) were loaded into HSAPs to combat their aqueous insolubility. BACPT-HSAPs loaded up to 5.25 micrograms BACPT/ mg of HSA. CPT encapsulation could not be determined. BACPT-HSAPs and CPT-HSAPs showed cytotoxicity to human sarcoma cells in vitro.

Hybride Nanostrukturen verbinden die Vorzüge von individuellen Materialien, die neue Eigenschaften hervorrufen können. In dieser Arbeit wurden verschiedene Metal Nanostrukturen synthetisiert und deren optische Eigenschaften analysiert. Die Herstellung eines Spasers oder Lichteinfang in Solarzellen wurde untersucht. Der Einfluß von Größe, Form und Brechungsindex auf die Metal-Plasmonen wurde erforscht. Die gewonnen Erkenntnisse genutzt um Metal Nanopartikel mit gezielten Eigenschaften herzustellen. Hybride Gold Nanostrukturen (funktionalisiert mit Farbstoffen oder Quantenpunkten) wurden hergestellt und Energie-Transfereffekte untersucht. Diese hybriden Nanostrukturen wurden optisch gepumpt um Spasing zu erreichen. Allerdings wurde festgestellt, dass eine unrealistisch hohe Verstärkung benötigen wird, um die charakteristischen Verluste im Metal zu überwinden. Silber und Gold Nanopartikel wurden synthetisiert um diese in Dünnschichtsolarzellen einzusetzen. Es konnte gezeigt werden, dass Silber chemisch instabil ist und, wenn oxidiert, hohe Absorption auftritt. Durch hohe Temperaturen konnte die Oxidschicht auf den Silberpartikeln reduziert werden und damit auch die Verluste. Stabilere Gold Partikel wurden in Perovskit-Solarzellen eingebaut, wodurch die Effizienz einer solch modifizierten Solarzelle um ~40% gesteigert werden konnte. Dies wurde durch eine erhöhte Anzahl an generierten Ladungsträgern mittels metallischen Lichtfallen erreicht. Zusätzlich wurden anisotrope Janus Trägerpartikel synthetisiert und mit Metal Nano-partikeln funktionalisiert. Gold Nanopartikel wurden abgeschieden und zu einer Gold Hülle gewachsen. Dies erfolgte entweder gleichförmig über das gesamte Hantel-Trägerpartikel oder einseitig unter Ausnutzung der chemischen Anisotropie. Desweiteren wurden Platin Nano-partikel einseitig abgeschieden und in Wasserstoffperoxid Lösung gegeben. Die Partikel wurden daraufhin mittels dynamischer Lichtstreuung auf Selbstvortrieb untersucht. / Hybrid nanostructures combine the assets of the individual materials with a vast amount of new properties. In this work various metal nanoparticles have been synthesized and investigated on their optical properties. The synthesized metal nanoparticles have been implemented for potential applications, e.g. fabrication of a spaser or in solar cells. At first, the size, shape and refractive index effects of gold and silver nanoparticles have been investigated. The insight gained helps to optimize the synthesis of metal nanoparticles with specific optical properties needed for further applications. Optimized hybrid gold nanostructures have been synthesized and functionalized with dye molecules or quantum dots to investigate energy transfer effects. These hybrid structures have been optically pumped to achieve spasing. However, comparison with a theory showed that such metal nanostructures need unrealistic high gain to overcome the inherent losses and achieve spasing. Silver and gold nanoparticles have been synthesized for applications in thin film solar cells. It has been shown that silver lacks chemical stability and thus, if oxidized, the nanoparticles exhibit weak scattering and strong Ohmic losses. The oxide layer of silver nano-spheres could be via annealing. By contrast, gold nanoparticles, known for their higher stability, have been implemented in a perovskite solar cell. Such a modified solar cell showed an increase in efficiency by ~40% through increased generation of carriers. Anisotropic Janus carrier systems have been synthesized and functionalized with metal nanoparticles. Gold nanoparticles have been deposited either uniformly or on one lobe only of the dumbbell-shaped carrier system by using its chemical anisotropy. These gold nano¬particles have been grown to a gold shell. Platinum nanoparticles have been deposited on a single lobe and its self-propelling ability in a chemical fuel was investigated by means of dynamic light scattering.

Zero-dimensional graphene (carbon) quantum dots have been drawing attention in bio-related applications since their discovery, especially for their optical properties, chemical stability, and easily modifiable surface.  This thesis focuses on the green synthesis of nitrogen-doped graphene quantum dots (GQDs) for dual-mode bioimaging with X-ray fluorescence (XRF) and optical fluorescence. Both conventional and microwave- (MW-)assisted solvothermal methods were followed to investigate the precursors’ effect on the synthesized GQDs. The MW-assisted method permitted the synthesis of uniform GQDs with an excitation-independent behavior, due to highly controllable reaction conditions. It was demonstrated that the molecular structure of the precursors influenced the optical fluorescence properties of the GQDs. Thus, both blue- (BQDs) and red-emitting (RQDs) GQDs were obtained by selecting specific precursors, leading to emission maxima at 438 and 605 nm under the excitation wavelengths of 390 and 585 nm, respectively.  Amine-functionalized Rh nanoparticles (NPs) were chosen as the X-ray fluorescence (XRF) active core, synthesized via MW-assisted hydrothermal method with a custom designed sugar ligand as the reducing agent. These NPs were conjugated with BQDs using EDC-NHS treatment. The hybrid Rh-GQDs NPs exhibited green emission (520 nm) under 490 nm excitation and led to a reduced cytotoxicity with respect to bare Rh NPs, highlighting the passivation role of the GQDs via the real-time cell analysis (RTCA) assay. The hybrid complex constituted a multimodal bioimaging contrastagent, tested with confocal microscopy (in vitro) and XRF phantom experiments. / Sedan deras upptäckt har nolldimensionella kvantprickar av grafen (kol) uppmärksammats inom biorelaterade applikationer, särskilt för deras optiska egenskaper, kemiska stabilitet och enkelt modifierbara yta. Denna avhandling fokuserar på en grön syntesmetod av kvävedopade grafen-kvantprickar för bimodal bioavbildning med röntgenfluorescens och optisk fluorescens. Både konventionella och mikrovågs-assisterade solvotermiska syntesmetoder användes för att undersöka metodernas effekt på de syntetiserade kvantprickarna. Den mikrovågs-assisterade metoden möjliggjorde syntes av uniforma kvantprickar med exciteringsoberoende egenskaper på grund av mycket kontrollerbara reaktionsförhållanden. Det demonstrerades att den molekylära strukturen hos prekursorerna påverkade de optiska fluorescensegenskaperna hos grafen-kvantprickarna. Genom att välja specifika prekursorer erhölls kvantprickar som emitterar i både blått och rött ljus, motsvarande emissionsmaxima vid 438 respektive 605 nm under excitering vid 390 respektive 585 nm. Amin-funktionaliserade Rh-nanopartiklar valdes som en aktiv kärna för röntgenfluorescens, syntetiserad genom en mikrovågs-assisterad hydrotermisk metod med en specialdesignad sockerligand som reduktionsmedel. Dessa nanopartiklar konjugerades med blåemitterande kvantprickar genom EDC-NHS-behandling. De hybrida nanopartiklarna uppvisade grön emission (520 nm) under 490 nm excitation och ledde till en minskad cytotoxicitet uppmätt genom cellanalys i realtid (RTCA) jämfört med endast Rh-nanopartiklar, vilket framhävde passiveringsrollen som kvantprickarna spelar. Hybridkomplexet utgjorde ett multimodalt kontrastmedel för bioavbildning, vilket demonstrerades med konfokalmikroskopi (in vitro) och fantomexperiment med röntgenfluorescens.

The growing incidences of orthopedic problems globally have created a huge demand for strong bioactive materials for bone tissue engineering. Over the years, studies have shown chemical, physical, and mechanical properties of biomaterials influence the cellular interactions at the material-tissue interface, which subsequently controls biological response to materials. Strong biomaterials with surface properties that actively direct cellular response hold the key for engineering the next generation orthopedic implants. With its unique properties graphene can be used to reinforce poly (ε-caprolactone) (PCL) to prepare strong and bioactive polymer nanocomposites for bone tissue regeneration. The thesis entitled ―Engineering bioactive and multifunctional graphene polymer composites for bone tissue regeneration” systematically studies the effect of different chemically functionalized and metal-graphene hybrid nanoparticles in PCL composites for bone tissue engineering. The thesis comprises of seven chapters. Chapter 1 is an outline review on the impact of graphene and graphene derived particles to prepare supporting substrates for tissue regeneration and the associated cell response to multifunctional graphene substrate. This chapter discusses how cells interact with different graphene based particles and the interplay between cells performance and multifunctional properties of graphene based substrates. Chapter 2 describes the role, if any, of the functionalization of graphene on mechanical properties, stem cell response and bacterial biofilm formation. PCL composites of graphene oxide (GO), reduced GO (RGO) and amine-functionalized GO (AGO) were prepared at different filler contents (1%, 3% and 5%). Although the addition of the nanoparticles to PCL markedly increased the storage modulus, this increase was higher for GO and AGO than with RGO. In vitro cell studies revealed that the AGO and GO particles significantly increased human mesenchymal stem cell (hMSC) proliferation. AGO was most effective in augmenting stem cell osteogenesis leading to mineralization. Bacterial studies revealed that interaction with functionalized GO induced bacterial cell death due to membrane damage which was further accentuated by amine groups in AGO. The synergistic effect of oxygen containing functional groups and amine groups on AGO-reinforced composites renders the optimal combination of improved modulus, favorable stem cell response and biofilm inhibition desired for orthopaedic applications. In Chapter 3, toward preparing strong multi-biofunctional materials, poly(ethylenimine) (PEI) conjugated graphene oxide (GO_PEI) was synthesized using poly(acrylic acid) (PAA) as spacer and incorporated in PCL at different fractions. GO_PEI significantly promoted proliferation and formation of focal adhesions in hMSCs on PCL. GO_PEI was highly potent in inducing stem cell osteogenesis leading to 90% increase in alkaline phosphatase activity and mineralization over neat PCL with 5% filler content and was 50% better than GO. Remarkably, 5% GO_PEI was as potent as soluble osteo-inductive factors. Increased adsorption of osteogenic factors due to the amine and oxygen containing functional groups on GO_PEI augment stem cell differentiation. GO_PEI was also highly efficient in imparting bactericidal activity with 85% reduction in counts of E. coli colonies compared to neat PCL at 5% filler content and was more than twice as efficient as GO. This may be attributed to the synergistic effect of the sharp edges of the particles along with the presence of the different chemical moieties. Thus, in contrast to using labile biomolecules, GO_PEI based polymer composites can be utilized to prepare bioactive resorbable biomaterials for fabricating orthopedic devices for fracture fixation and tissue engineering. Chapter 4 describes the preparation of hybrid nanoparticles of graphene sheets decorated with strontium metallic nanoparticles and its advantages in bone tissue engineering. Strontium-decorated reduced graphene oxide (RGO_Sr) nanoparticles were synthesized by facile reduction of graphene oxide and strontium nitrate. X-ray diffraction, transmission electron microscopy, and atomic force microscopy revealed that the hybrid particles were composed of RGO sheets decorated with 200 – 300 nm metallic strontium particles. Thermal gravimetric analysis further confirmed the composition of the hybrid particles as 22 wt% of strontium. Macroporous tissue scaffolds were prepared incorporating RGO_Sr particles in PCL. The PCL/RGO_Sr scaffolds were found to elute strontium ions in aqueous medium. Osteoblast proliferation and differentiation was significantly higher in the PCL scaffolds containing the RGO_Sr particles in contrast to neat PCL and PCL/RGO scaffolds. The increased biological activity can be attributed to the release of strontium ions from the hybrid nanoparticles. This study demonstrates that composites prepared using hybrid nanoparticles that elute strontium ions can be used to prepare scaffolds with osteoinductive property. These findings have important implications for designing the next generation of biomaterials for use in tissue regeneration. Chapter 5 discusses the use of hybrid graphene-silver particles (RGO_Ag) to reinforce PCL and compared with PCL/RGO and PCL/Ag composites containing RGO and silver nanoparticles (AgNPs), respectively. RGO_Ag hybrid particles were well dispersed in the PCL matrix unlike the RGO and AgNPs due to enhanced exfoliation. RGO_Ag led to 77 % increase in the modulus of PCL and provided a conductive network for electron transfer. Electrical conductivity increased four orders of magnitude from 10-11 S/cm to 10-7 S/cm at 5 wt % filler that greatly exceeded the improvements with the use of RGO and AgNP in PCL. RGO_Ag particles reinforced in PCL showed sustained release of silver ions from the PCL matrix unlike the burst release from PCL/Ag. PCL/RGO_Ag and PCL/RGO composites were non-toxic to hMSCs and supported osteogenic differentiation unlike the PCL/Ag composites which were highly toxic at ≥3% filler content. The PCL/RGO_Ag composites exhibited good antibacterial effect due to a combination of silver ion release from the AgNPs and the mechanical rupture induced by the RGO in the hybrid nanoparticles. Thus, the synergistic effect of Ag and RGO in the PCL matrix uniquely yielded a multifunctional material for use in implantable biomedical devices and tissue engineering. Chapter 6 presents investigation of potential differences in the biological response to graphene in polymer composites in the form of 2D substrates and 3D scaffolds. Results showed that osteoblast response to graphene in polymer nanocomposites is markedly altered between 2D substrates and 3D scaffold due to the roughness induced by the sharp edges of graphene at the surface in 3D but not in 2D. Osteoblast organized into aggregates in 3D scaffolds in contrast to more well spread and randomly distributed cells on 2D discs due to the macro-porous architecture of the scaffolds. Increased cell-cell contact and altered cellular morphology led to significantly higher mineralization in 3D scaffolds compared to 2D. This study demonstrates that the cellular response to nanoparticles in composites can change markedly by varying the processing route. Chapter 7 summarizes the important results and future directions of the work. This chapter provides general conclusions arising from this study, and makes suggestions for future work designed to provide a greater understanding of the in vivo response in terms of bio-distribution of the released functionalized graphene from the scaffold or substrate must be assessed with special attention on their accumulation or excretion.

Optical properties of nanoscopic materials have been intensively perused over last couple of decades due to their tunable optical properties. Recent interests in this field have been mainly focused on the preparation of ordered arrays of nanoscopic materials and study of their optical properties. These interests have been motivated by the usability of such systems for nano photonic devices. Theoretical predictions from such systems reveal complex absorption and emission properties, different from individual ones mainly because of energy transfer between them. These properties can be controlled further by preparing hybrid arrays of nanostructures, including nano crystals of different types. Hybrid arrays with semiconducting quantum dots and metallic nanoparticles are an example of such system. Optical properties of such a system can be tuned by controlling the interaction between excitons and plasmons. This the-sis presents the experimental studies on optical properties of polymer capped polymer nanoparticles, quantum dot arrays and hybrid arrays with semiconducting quantum dot and metal nanoparticles. A brief summary of the experi-mental methods and results have been highlighted below. First chapter deals with the theoretical aspects of confined nanoscopic materials, especially describing the physics of zero dimensional systems and its optical properties. The discussions are mostly focused on two types of nano materials cadmium selenide (CdSe) quantum dot (QDs) and gold nano particles (Au NPs), used for the experimental study. Variation of energy levels of CdSe QDs and its absorption and emission properties under strong confinement regime has been discussed with respect to effective mass approximation (EMA) model. This is followed by the discussion on optical properties of Au NPs, describing absorption properties, based on Mie theory. Size dependent variation of absorption spectra of Au NPs and the modifications based on different models has been discussed. Second part of the chapter describes the physics of QD arrays and theory of exciton plasmon interactions based on the recent literatures. Energy transfer mechanism between semiconducting QDs and metal nanoparticles has been discussed based on numerical method and dipole approximation. Second chapter deals with the discussion on experimental techniques used for the study. Chapter 2 starts with the discussion on the synthesis method for CdSe QDs and Au NPs with different capping ligands. Preparation of QD ar-rays and hybrid arrays using self assembly technique has been discussed in this chapter. Preparation CdSe QD arrays and hybrid arrays with CdSe QDs and Au NPs using block copolymer (BCP) template and Langmuir Blodgett (LB) technique has been the main focus in the discussion. This is followed by the discussion on optical microscopy techniques, confocal, near field scanning microscopy (NSOM), Brewster angle microscopy and electron microscopy techniques, transmission electron microscopy and scanning electron microscopy. Studies on variation of band structure of small polymer capped Au NPs, with respect to the size and grafting density of the capping polymer is discussed in chapter 3. Polymer capped Au NPs with sizes 2-5 nm was used for the study. Dielectric constants of Au NPs were extracted from the absorption spectra by fitting the data using modified Mie theory. Dielectric constants of Au NPs were reproduced using an analytical expression, describing the contribution from different transitions in the optical regions. Results indicate systematic variations of the band structure with respect to the particle size and grafting density. The observations have been interpreted in terms of variation of co ordination number and chemical interaction of capping polymer with the surface atoms. Our new method analysis points to the importance of both quantum and surface effects in determining optical and electronic properties of polymer capped gold nanoparticles. Chapter 4 describes the study on morphology of the CdSe QD arrays prepared using different BCP templates and its correlation with optical properties. Spatially resolved spectra from the thin films of QD arrays were collected in near field and the compared with the spectra collected in far field. Spectra collected in near field mode shows sharp features in the emission spectra, possibly indicating the interaction of optical near field with QD excitation. It has been suggested that such fine structure could be induced by coupling between optical near filed and excitons and this coupling seems to be determined by local heterogeneity in QD density and disorder. Variation of exciton life time with respect to QD density and absorption spectra from the QD -BCP system is also described in chapter 4. Chapter 5 and 6 deals with the experimental studies on exciton -plasmon interaction in hybrid arrays of CdSe QDs and Au NPs. Emission properties hybrid arrays prepared using BCP templates has been the focus of chapter 5. Photoluminescence (PL) and lifetime measurements were performed on hybrid arrays and their variation with respect to the density and dispersion of Au NPs has been described. Optical measurements were performed on two sets of films using two different sizes of CdSe QDs, with the smaller QD emission overlapping with the plasmon resonance of Au NPs, while a red shifted emission peak for larger QDs. PL emission from hybrid arrays with smaller QDs shows en-hancement/quenching with respect to the dispersion of Au NPs, also showing systematic reduction of life time of CdSe QDs with Au NP density. Even though enhancement/quenching of emission properties of hybrid film with large QD shows similar behavior, PL decay measurements from such films shows non monotonic variation of exciton life time with respect to Au NP density. The enhancement/quenching behavior of the PL emission has been explained in terms of two competing mechanism, electromagnetic field enhancement and non radiative energy transfer. However to explain the energy transfer mechanism in hybrid arrays requires more systematic calculations. Chapter 6 describes the optical properties of highly compact hybrid arrays prepared using LB techniques. Hybrid arrays prepared at the air water inter-face were transferred to a glass substrates. The main focus on chapter 6 is to study the emission properties of highly compact hybrid arrays with respect to the spectral overlap between exciton energy of CdSe QDs and plasmon band of Au NPs with respect to their surface density (inter particle distance). Hybrid arrays were prepared with three types of QDs, with smaller QDs emission peak overlapping with plasmon band of Au NPs and clearly separated exciton and plasmon band for largest QDs. The PL emission from hybrid arrays with smaller QDs shows quenching, compared to strong enhancement in the emission from hybrid films with larger QDs. The disagreement of the observed results with respect to the theoretical calculations based on dipole approximation has been highlighted in the chapter. Chapter 7 includes the summary of the experimental results and the future works to be carried out as a continuation of the work presented in this thesis.