Thesis (M.S.) in Biological Engineering--University of Maine, 2008. / Includes vita. Includes bibliographical references (leaves 39-42).

In this work the synergy between a polymer containing chelate sites and gold ions was explored by the fabrication of a polymeric membrane with embedded gold nanoparticles inside its matrix and by developing a process to recover gold from acidic solutions. After realizing that the thiosemicarbazide groups present in the monomeric unit of poly-thiosemicarbazide (PTSC) formed strong complexes with Au ions, membrane technology was used to exploit this property to its maximum. The incorporation of metal nanoparticles into polymeric matrices with current technologies involves either expensive and complicated procedures or leads to poor results in terms of agglomeration, loading, dispersion, stability or efficient use of raw materials. The fabrication procedure described in this thesis solves these problems by fabricating a PTSC membrane containing 33.5 wt% in the form of 2.9 nm gold nanoparticles (AuNPs) by a three step simple and scalable procedure. It showed outstanding results in all of the areas mentioned above and demonstrated catalytic activity for the reduction of 4-Nitrophenol (4−NP) to 4-Aminophenol (4−AP). The current exponential demand of gold for electronics has encouraged the development of efficient processes to recycle it. Several adsorbents used to recover gold from acidic solutions can be found in the literature with outstanding maximum uptakes,yet, poor kinetics leading to an overall inefficient process. The method developed in this dissertation consisted in permeating the gold-containing solution through a PTSC membrane that will capture all the Au ions by forming a metal complex with them. Forcing the ions through the pores of the membrane eliminates the diffusion limitations and the adsorption will only depended on the fast complexation kinetics, resulting in a very efficient process. A flux as high as 1868 L/h m2 was enough to capture >90% of the precious metal present in a solution of 100 ppm Au. The maximum uptake achieved without sacrificing the mechanical stability was 5.4 mmol/g. The selectivity between gold and copper (the most common unwanted metal present along with gold) was 6.7 for 100 ppm initial concentration of both metals and 14.6 for 500 ppm.

Multicomponent ligand interactions are involved in essentially all nanoparticle (NP) applications. However, the ligand conformation and ligand binding mechanisms on NPs are highly controversial. The research reported here is focused on deepening the fundamental understanding of multicomponent ligand interactions with gold and silver nanoparticles (AuNPs and AgNPs) in water. We demonstrated that AuNPs passivated by saturated layer of poly(ethylene glycol) (PEG-SH) have large fractions of AuNP surface area available for ligand adsorption and exchange. The fraction of AuNP surface area passivated by PEG-SH with molecular weights of 2000, 5000, and 30000 g/mol was calculated to be ~ 25%, ~20%, and ~9% using 2-mercaptobenzimidazole and adenine as model ligands. The effect of both reduced and oxidized protein cysteine residues on protein interactions with AgNPs was investigated. The model proteins included wild-type and mutated GB3 variants with 0, 1, or 2 reduced cysteine residues. Bovine serum albumin containing 34 oxidized (disulfide-linked) and 1 reduced cysteine residues was also included. Protein cysteine content that were found to have no detectable effect on kinetics of protein/AgNP binding. However, only proteins that contain reduced cysteine induced significant AgNP dissolution. We further demonstrated that organothiols can induce both AgNP disintegration and formation under ambient conditions by simply mixing organothiols with AgNPs or AgNO3, respectively. Surface plasmon- and fluorescence-active AgNPs formed by changing the concentration ratio between Ag+ and organothiol. Organothiols also induced AuNP formation by mixing HAuCl4 with organothiols, but no AuNP disintegration occured. Finally, we proposed that multicomponent ligand binding to AuNPs can be highly dependent on the sequence of ligand mixing with AuNPs. Quantitative studies revealed that competitive adenine and glutathione adsorption onto both as-synthesized and PEG-SH functionalized AuNPs is predominantly a kinetically controlled process. Besides providing new insights on multicomponent ligand interactions with colloidal AuNPs and AgNPs, this study opens a new avenue for fabrication of novel nanomaterials in biological/biomedical applications.

>Magister Scientiae - MSc / Different nanoparticles have the ability to improve plant tolerance to drought stress. In the study we report for the first time novel morpholin-4-ium 4-methoxyphenyl (morpholino) phosphinodithioate capped- gold nanoparticles (GYY4137-capped AuNPs). The GYY4137 is a slow releasing hydrogen sulfide (H2S) donor. The GYY4137 AuNPs compared to preliminary experiments of L-serine and L-threonine gold nanoparticles. The nanoparticles were prepared using a simple reflux reduction method in a rolling boil flask at 80 oC. The uncapped GYY4137-AuNPs were relatively stable and had surface plasmon resonance at 562 nm compared to 524 nm and 560 nm of serine-AuNPs and threonine-AuNPs. The nanoparticles were capped with different concentrations (0.1-5 %) of water-soluble poly (ethylene) glycol (PEG) (Mw300) and 0.2% chitosan. The PEG did not fully encapsulate the gold nanoparticles, while the chitosan successfully produced positively charged gold nanoparticles. The formation of chitosan capped GYY4137-AuNPs were verified with UV-Visible spectroscopy (UV-Vis), High Resolution Transmission electron microscopy (HRTEM), Dynamic Light scattering (DLS) and the Zetasizer. The UV-Vis, HRTEM and STEM verified chitosan capped nanoparticles had a surface plasmon resonance peak at 560 nm, with icosahedral, tetrahedron and spherical shaped nanoparticles as the serine-AuNPs that absorb at 560 nm. The agglomerated threonine-AuNPs had a maximum absorbance peak at 524 nm. The chitosan GYY4137-AuNPs had hydrodynamic size of 347.9 nm and zeta potential of + 47 mV, while serine-AuNPs and threonine-AuNPs had hydrodynamic size of 110 nm, zeta potential of -2.9 mV and -230 mV respectively. The polydispersity index (PDI) of the chitosan capped gold nanoparticles was 0.357 compared to 0.406 of both the amino acid gold nanoparticles. The polydispersity index (PDI) showed that the nanoparticles were polydispersed nanoparticles with broad size range as confirmed by the HRTEM and STEM results/ of chitosan capped GYY4137-AuNPs. The sizes of the nanoparticles were 100 nm and 60 nm for GYY4137-AuNPs while the size serine-AuNPs were 60 nm. The gold nanoparticles structural compositions were further confirmed by energy-dispersive X-ray spectrometry (EDX) and Attenuated total reflection infrared spectroscopy (ATR-IR). EDX results proved successful gold nanoparticles synthesis by presence of the element Au in all three nanoparticles and the chitosan GYY4137-AuNPs had 48. 56 wt. % of gold. The FTIR-ATR new bands formation shows that new chemical bonds are formed between the reducing agents, the precursor gold salt solution and capping agents. The shifts showed successful encapsulation with chitosan in GYY4137-AuNPs. The chitosan encapsulation improved surface charge and reactivity of the gold nanoparticles to improve delivery of the hydrogen sulfide donor GYY4137 for later applications to plants.

Over the past few decades, tremendous efforts have been made to develop nanomaterials for biotechnological applications such as therapeutics. Understanding and engineering interfaces between biomacromolecules and nanomaterials is a key to the creation of successful therapeutic systems. My research has been oriented toward developing therapeutic systems using gold nanoparticles (AuNPs) incorporating material science, organic synthesis, and biology. For this purpose, mixed monolayer protected AuNPs (~2 nm core size) with various functional groups have been employed for triggering therapeutic effects. Several strategies have been accomplished using anticancer drugs that non-covalently and covalently incorporate onto AuNPs as a drug delivery carrier. Alternatively, AuNPs were developed by regulating host-guest complexation processes inside the cell, allowing control of the therapeutic effect of the AuNP. In addition, by using host-guest chemical events on the AuNPs, exocytosis of the AuNPs was controlled, enabling their prolonged retention inside of the cells, providing new strategies for improving conventional drug delivery systems. Therefore, engineering of the AuNP surface can afford new pathways for designing and improving therapeutics.

Doxorubicin is one of the most widely used and effective anticancer agents to treat a wide spectrum of tumors. But its success in cancer therapy is greatly compromised by its cumulative dose-dependent side effects of cardiotoxicity and tumor cell resistance. For the purpose of addressing these side effects, a gold nanoparticles-based anticancer drug delivery system was designed. Five novel thiolated doxorubicin analogs were designed and synthesized and their biological activities have been evaluated. These doxorubicin analogs and the poly(ethylene glycol) (PEG) stabilizing ligands were conjugated to gold nanoparticles via formation of a gold-thiol bond. The systems were evaluated in vitro and in vivo, and the results show that controlled drug release can be achieved either by acidic conditions or by reducing agents in cancer cells, depending on the design of the thiolated drug construct. The overall drug delivery system should achieve enhanced drug accumulation and retention in cancer cells and favorable drug release kinetics, and should demonstrate therapeutic potential and the ability to address some of the current problems of doxorubicin in cancer therapy. / Master of Science

Yes / Peptides have demonstrated unique capabilities to fabricate inorganic nanomaterials of numerous compositions through noncovalent binding of the growing surface in solution. In this contribution, we demonstrate that these biomolecules can control all facets of Au nanoparticle fabrication, including Au3+ reduction, without the use of secondary reagents. In this regard using the AuBP1 peptide, the N-terminal tryptophan residue is responsible for driving Au3+ reduction to generate Au nanoparticles passivated by the oxidized peptide in solution, where localized residue context effects control the reducing strength of the biomolecule. The process was fully monitored by both time-resolved monitoring of the growth of the localized surface plasmon resonance and transmission electron microscopy. Nanoparticle growth occurs by a unique disaggregation of nanoparticle aggregates in solution. Computational modeling demonstrated that the oxidized residue of the peptide sequence does not impact the biomolecule’s ability to bind the inorganic surface, as compared to the parent peptide, confirming that the biomolecule can be exploited for all steps in the nanoparticle fabrication process. Overall, these results expand the utility of peptides for the fabrication of inorganic nanomaterials, more strongly mimicking their use in nature via biomineralization processes. Furthermore, these capabilities enhance the simplicity of nanoparticle production and could find rapid use in the generation of complex multicomponent materials or nanoparticle assembly. / Air Force Office of Scientific Research, grant FA9550-12-1-0226.

Yes / Peptide-mediated synthesis and assembly of nanostructures opens new routes to functional inorganic/organic hybrid materials. However, understanding of the many factors that influence the interaction of biomolecules, specifically peptides, with metal surfaces remains limited. Understanding of the relationship between peptide sequence and resulting binding affinity and configurations would allow predictive design of peptides to achieve desired peptide/metal interface characteristics. Here, we measured the kinetics and thermodynamics of binding on a Au surface for a series of peptide sequences designed to probe specific sequence and context effects. For example, context effects were explored by making the same mutation at different positions in the peptide and by rearranging the peptide sequence without changing the amino acid content. The degree of peptide-surface contact, predicted from advanced molecular simulations of the surface-adsorbed structures, was consistent with the measured binding constants. In simulations, the ensemble of peptide backbone conformations showed little change with point mutations of the anchor residues that dominate interaction with the surface. Peptide-capped Au nanoparticles were produced using each sequence. Comparison of simulations with nanoparticle synthesis results revealed a correlation between the colloidal stability of the Au nanoparticles and the degree of structural disorder in the surface-adsorbed peptide structures for this family of sequences. These findings suggest new directions in the optimization and design of biomolecules for in situ peptide-based nanoparticle growth, binding, and dispersion in aqueous media.

Gold nanoparticles can serve as an ideal radiosensitizer for radiation therapy due to the high-atomic-number nature of gold and the increased tumor specificity in nanoparticle form. The degree of radiosensitization is highly dependent on both the local gold nanoparticle concentration in the tumor and the radiation source type. Previous Monte Carlo simulations have demonstrated that the gamma-ray energy spectrum of Ytterbium-169 is a strong candidate for a high dose rate brachytherapy implementation of gold nanoparticle-aided radiation therapy. Therefore, the current study focuses on the design of a high dose rate Ytterbium-169 source that would maximize dose enhancement during gold nanoparticle-aided radiation therapy; while meeting the practical constraints for the production of a clinically relevant brachytherapy source. Different encapsulation materials are studied in order to determine its effect on the dosimetric characteristics of the source. Specifically, the photon spectra, secondary electron spectra, and dose enhancement characteristics are calculated via Monte Carlo simulations to elucidate the effects on potential radiosensitization during gold nanoparticle-aided radiation therapy. Furthermore, this project involves a study into the modification of external x-ray beams from a Philips RT-250 orthovoltage x-ray machine in an attempt to match the dosimetric characteristics of the Ytterbium-169 brachytherapy source. This investigation will enable the production of an external beam that can serve as a good surrogate of an actual brachytherapy source and facilitate the pre-clinical investigation of gold nanoparticle-aided radiation therapy with Ytterbium-169.

Vaccination is one of the most effective public health interventions in the world, saving millions of lives and preventing the onset of debilitating diseases. With widespread emergence of multi-drug resistant pathogens, the importance of preventative medicine has become even more apparent. However, one of the limiting factors in developing novel vaccines that are both safe and highly immunogenic is the availability of adjuvant delivery systems licensed for human use. The purpose of this study was to investigate the role gold nanoparticles could play as an effective vaccine delivery system. A variety of coupling chemistries were explored for their ability to conjugate protein and polysaccharide antigens onto the surface of gold nanoparticles for the development of vaccines against a number of biologically important human pathogens including Y. pestis, B. mallei and S. pneumoniae. Retention of antigenicity and coupling efficiency of conjugated molecules was measured using characterisation techniques such as localised surface plasmon resonance and immunoblotting. Gold nanoparticle coupled antigens were then used to immunise mice and to measure the protective efficacy and the immunological response induced. The findings indicate antigen-specific immune responses are elevated when an antigen is coupled onto gold nanoparticles. Moreover, immunological data from nanoparticle coupled glycoconjugate vaccines against B. mallei and S. pneumoniae indicate the likely presence of a strong T cell immune response which is essential for providing immunological memory. Finally, an intracellular trafficking assay was carried out to identify some of the mechanisms that might be involved in uptake of gold nanoparticles into professional phagocytes. Confocal imaging of receptors associated with endosomal compartments revealed that gold nanoparticles may enter cells through multiple pathways. The findings reported in this study suggest that gold nanoparticles may be an excellent candidate for further investigation as a novel vaccine delivery system.