abstract: This work aims to characterize protein-nanoparticle interactions through the application of experimental techniques to aid in controlled nanoparticle production for various applications from manufacturing through medical to defense. It includes multiple steps to obtain purified and characterized protein and then the production of nanoparticles using the protein. This application of protein requires extremely pure homogenous solution of the protein that was achieved using numerous protein separation techniques which were experimented with. Crystallization conditions, protein separation methods and protein characterization methods were all investigated along with the protein-nanoparticle interaction studies. The main protein of study here is GroEL and the inorganic nanoparticle used is platinum. Some studies on MBP producing gold nanoparticles from an ionic gold precursor were also conducted to get a better perspective on nanoparticle formation. Protein purification methods, crystallization conditions, Car-9 tag testing and protein characterization methods were all investigated along with the focus of this work. It was concluded that more Car9 studies need to be carried out before being used as in the form of a loop in the protein. The nanoparticle experiments were successful and platinum nanoparticles were successfully synthesized using GroEL. The direction of further research in protein-nanoparticle studies are outlined towards the end of the thesis. / Dissertation/Thesis / Masters Thesis Chemical Engineering 2019

Introduction: Over the past few decades, nanoparticles (NPs) have gained immeasurable interest in the field of drug delivery. Various NP formulations have been disseminated in drug development in an attempt to increase efficacy, safety and tolerability of incorporated drugs. In this context, NP formulations that increase solubility, control release, and/or affect the in vivo disposition of drugs, were developed to improve the pharmacokinetic and pharmacodynamic properties of encapsulated drugs.Areas covered: In this article, important properties related to NP function such as particle size, surface charge and shape are disseminated. Also, the current understanding of how NP characteristics affect particle uptake and targeted delivery is elucidated. Selected NP systems currently used in delivery of drugs in biological systems and their production methods are discussed as well. Emphasis is placed on current NP formulations that are shown to reduce drug-induced adverse renal complications.Expert opinion: Formulation designs utilizing NP-encapsulated drugs offer alternative pharmacotherapy options with improved safety profiles for current and emerging drugs. NPs have been shown to increase the therapeutic index of several entrapped drugs mostly by decreasing drug localization and side effects on organs. Recent studies on NP-encapsulated chemotherapeutic and antibiotic medications show enhanced therapeutic outcomes by altering drug degradation, increasing systemic circulation and/or enhancing cell specific targeting. They may also reduce the distribution of encapsulated drugs into the kidneys and attenuate drug-associated adverse renal complications. The usefulness of NP formulation in reducing the nephrotoxicity of nonsteroidal anti-inflammatory drugs is an underexplored territory that deserves more attention.

We suggest a computational approach for estimating the ring-like deposition of nanoparticles contained in a drying liquid droplet. The proposed method involves a Monte Carlo scheme, based on three independent probabilistic processes: (a) evaporation at the liquid surface, (b) convective motion of nanoparticles to the contact line, and (c) treatment of the nanoparticles floating in the air. According to the computational results, while the liquid is evaporating in nanoparticle-suspended liquid droplet (NSLD), the nanoparticles are moved to the contact line as the mass of droplet decreases linearly with time. Since the resulting ring-like deposition can be accounted for in terms of nanoparticle mobility and liquid evaporation from the droplet, our computational approach achieves a morphological and kinematical description of NSLD drying. Some other important features, such as self-pinning of the contact line, reduction of the droplet radius, and pattern formation, are also obtained from this simulation.

Cerium oxide nanoparticles (nanoceria) have recently been shown to protect cells against oxidative stress in both cell culture and animal models. Nanoceria has been shown to exhibit superoxide dismutase (SOD) activity using a ferricytochrome C assay, and this mimetic activity that has been postulated to be responsible for cellular protection by nanoceria. The nature of nanoceria's antioxidant properties, specifically what physical characteristics make nanoceria effective at scavenging superoxide anion, is poorly understood. In this study electron paramagnetic resonance (EPR) analysis confirms the reactivity of nanoceria as an SOD mimetic. X-ray photoelectron spectroscopy (XPS) and UV-visible analyses of nanoceria treated with hydrogen peroxide demonstrate that a decrease in the Ce 3+/4+ ratio correlates directly with a loss of SOD mimetic activity. These results strongly suggest that the surface oxidation state of nanoceria plays an integral role in the SOD mimetic activity of nanoceria and that ability of nanoceria to scavenge superoxide is directly related to cerium(III) concentrations at the surface of the particle.

京都大学 / 新制・課程博士 / 博士(理学) / 甲第22990号 / 理博第4667号 / 新制||理||1670(附属図書館) / 京都大学大学院理学研究科物理学・宇宙物理学専攻 / (主査)教授 山本 潤, 教授 石田 憲二, 教授 田中 耕一郎 / 学位規則第4条第1項該当 / Doctor of Science / Kyoto University / DFAM

Indiana University-Purdue University Indianapolis (IUPUI) / The reaction between elemental sulfur and sulfide is a lynchpin in the biotic and abiotic cycling of sulfur. This dissertation is focused on the reactivity of elemental sulfur nanoparticles (S8weimarn, S8raffo) among other forms of elemental sulfur (S8aq, S8aq-surfactant, α-S8), and how the variation of their surface area, character and coatings reflect on the analytical, physical-chemical and geochemical processes involving sulfur cycling. A comprehensive electrochemical investigation utilizing mercury-surface electrodes showed that elemental sulfur compounds are represented by three main voltammetric signals, corresponding to potentials at -1.2V, -0.8V, and -0.6V in the absence of organics at circumneutral pH. Dissolved S8aq-surfactant signals can be found from -0.3V up to -1.0V, depending on the surfactant in the system. Variations in current response resulted from differences in electron transfer efficiency among the forms of S8, due to their molecular structural variability. Based on this observation a new reaction pathway between S8 and Hg-surface electrodes is proposed, involving an amalgam-forming intermediate step. The kinetics of the nucleophilic dissolution of S8nano by sulfide, forming polysulfides, were investigated under varying surface area, surface character and presence or absence of surfactant coatings on S8nano. Hydrophobic S8weimarn and hydrophilic S8raffo show kinetic rate laws of 𝑟𝑆8𝑤𝑒𝑖𝑚𝑎𝑟𝑛 = 10−11.33 (𝑒 −700.65 𝑅𝑇 ) (Molar(S8)/second/dm-1) and𝑟𝑆8𝑟𝑎𝑓𝑓𝑜 = 10−4.11 𝑖−0.35 (𝑒 −615.77 𝑅𝑇 ) (Molar(S8)/second), respectively. The presence of surfactant molecules can influence the reaction pathways by dissolving S8nano and releasing S8aqsurfactant, evolving the rate-limiting step as a function of the degree of the solubilization of S8nano. The reaction rate of S8biological can be compared with those of S8raffo and S8weimarn in circumneutral pH values and T=50oC, making the forms of S8nano successful abiotic analogue models of microbially produced S8biological. Field observations and geochemical kinetic modeling in the geothermal features of Yellowstone indicate that the nucleophilic dissolution reaction appears to be a key abiotic pathway for the cycling of sulfur species and the enhancement of elemental sulfur bioavailability. Furthermore, in situ and ex situ voltammetry in the same geothermal waters disclosed chaotic variability in chemical gradients of sulfide (observed over small temporal and spatial scales) which can be considered as an ecological stressor capable of influencing single cell physiology and microbial community adaptation.

Indiana University-Purdue University Indianapolis (IUPUI) / Non-invasive brain stimulation is valuable for studying neural circuits and treating various neurological disorders in human. However, current technologies of noninvasive brain stimulation usually have low spatial and temporal precision and poor brain penetration, which greatly limit their application. A new class of nanoparticles known as magneto-electric nanoparticles (MENs) is highly efficient in coupling an externally applied magnetics wave with generating local electric fields for neuronal activity modulation. Here, a new type of MENs was developed that consisted of CoFe2O4- BaTiO3 and had excellent magneto-electrical coupling properties. Calcium imaging technique was used to demonstrate their efficacy in evoking neuronal activity in organotyic and acute cortical slices that expressed GCaMP6 protein. For in vivo noninvasive delivery of MENs to brain, fluorescently labeled MENs were intravenously injected and attracted to pass through blood brain barrier to a targeted brain region by applying a focal magnet field. Magnetic wave (~450 G at 10 Hz) applied to mouse brain was able to activate cortical network activity, as revealed by in vivo two-photon and mesoscopic imaging of calcium signals at both cellular and global network levels. The effect was further confirmed by the increased number of c-Fos expressing cells after magnetic stimulation. Histological analysis indicated that neither brain delivery of MENs nor the subsequent magnetic stimulation caused any significant increases in the numbers of GFAP and IBA1 positive astrocytes and microglia in the brain. MENs stimulation also show high efficacy in short-term pain relieve when tested with a tibial nerve injury mouse model. The study demonstrates the feasibility of using MENs as a novel efficient and non-invasive technique of brain stimulation, which may have great potential for translation.

Indiana University-Purdue University Indianapolis (IUPUI) / Glioblastoma (GBM) is a deadly, malignant brain tumor with a poor long-term prognosis. The current median survival is approximately fifteen to seventeen months with the standard of care therapy which includes surgery, radiation, and chemotherapy. An important factor contributing to recurrence of GBM is high resistance of GBM cancer stem cells (CSCs), for which a systemically delivered single drug approach will be unlikely to produce a viable cure. Therefore, multi-drug therapies are needed. Currently, only temozolomide (TMZ), which is a DNA alkylator, affects overall survival in GBM patients. CSCs regenerate rapidly and over-express a methyl transferase which overrides the DNA-alkylating mechanism of TMZ, leading to drug resistance. Idasanutlin (RG7388, R05503781) is a potent, selective MDM2 antagonist that additively kills GBM CSCs when combined with TMZ. By harnessing the strengths of nanotechnology, therapy can be combined with diagnostics in a truly theranostic manner for enhancing personalized medicine against GBM. The goal of this thesis was to develop a multi-drug therapy using multi-functional nanoparticles (NPs) that preferentially target the GBM CSC subpopulation and provide in vivo preclinical imaging capability. Polymer-micellar NPs composed of poly(styrene-b-ethylene oxide) (PS-b-PEO) and poly(lactic-co-glycolic) acid (PLGA) were developed investigating both single and double emulsion fabrication techniques as well as combinations of TMZ and RG7388. The NPs were covalently bound to a 15 base-pair CD133 aptamer in order to target a specific epitope on the CD133 antigen expressed on the surface of GBM CSC subpopulation. For theranostic functionality, the NPs were also labelled with a positron emission tomography (PET) radiotracer, zirconium-89 (89Zr). The NPs maintained a small size of less than 100 nm, a relatively neutral charge and exhibited the ability to produce a cytotoxic effect on CSCs. There was a slight increase in killing with the aptamer-bound NPs compared to those without a targeting agent. This work has provided a potentially therapeutic option for GBM specific for CSC targeting and future in vivo biodistribution studies.

京都大学 / 0048 / 新制・課程博士 / 博士(エネルギー科学) / 甲第18382号 / エネ博第294号 / 新制||エネ||61(附属図書館) / 31240 / 京都大学大学院エネルギー科学研究科エネルギー基礎科学専攻 / (主査)教授 佐川 尚, 教授 八尾 健, 教授 萩原 理加 / 学位規則第4条第1項該当 / Doctor of Energy Science / Kyoto University / DGAM

京都大学 / 0048 / 新制・論文博士 / 博士(工学) / 乙第12923号 / 論工博第4116号 / 新制||工||1625(附属図書館) / 32133 / (主査)教授 木村 俊作, 教授 跡見 晴幸, 教授 岩田 博夫 / 学位規則第4条第2項該当 / Doctor of Philosophy (Engineering) / Kyoto University / DFAM