Synthesis and characterization of bioresorbable calcium phosphosilicate nanocomposite particles for fluorescence imaging and biomedical applications .

摘要

Organically doped calcium phosphosilicate nanoparticles (CPSNPs) were developed and characterized, driven by the need for non-toxic vectors for drug delivery and fluorescence biological imaging applications. In particular, advancement in drug delivery for the chemotherapeutic treatment of cancers is required to increase drug efficacy and improve patient quality of life. Additionally, brighter and more photostable fluorophores are needed to meet demands for improved sensitivity and experimental diversity, which may lead to improvements in early detection of solid tumors and advancement in understanding of biological processes. A literature survey on the state of the field for nanoparticle based biological fluorescence imaging and drug delivery is presented in Chapter 1. Chapter 2 focuses on the characterization techniques used in this work.;The development and optical characterization of 20-40 nm diameter, citrate functionalized Cy3 amidite doped calcium phosphosilicate nanoparticles (Cy3 CPSNPs) for in vitro fluorescence imaging is outlined in Chapters 3 and 4, respectively. In particular, sodium citrate was used to functionalize the surface and provide electrosteric dispersion of these particles. CPSNPs stabilized with sodium citrate routinely exhibited highly negative zeta potentials greater than -25 mV in magnitude. Furthermore, the fluorescence quantum yield of the encapsulated fluorophore was improved by more than 4.5-fold when compared to the unencapsulated dye. The bioimaging and drug delivery capability of CPSNPs was explored. Cy3 CPSNPs dissolved quickly in the acidic environment experienced during endocytosis, releasing the encapsulated fluorophore. This is consistent with solution phase experiments that show the particles are dissolved at pH 5. CPSNPs loaded with fluorescein and a hydrophobic growth inhibitor, ceramide C6, proved the ability to simultaneously image and delivery of the hydrophobic drug to cells in vitro.;Chapter 5 examined the colloidal stability of citrate and polyethylene glycol (PEG) functionalized CPSNPs in 70 volume % ethanol/30% water, both experimentally using TEM and theoretically using DLVO and polymeric steric dispersion theories. There are three basic mechanisms for colloidal stability for macroscopic suspensions (i.e., for particulate diameters down to ∼100nm), metastable electrostatic in which some finite degree of agglomeration continuously takes place because a finite energy barrier against agglomeration; and electrosteric and steric mechanisms in which infinitely high potential energy barriers toward agglomeration are present leading to thermodynamically stable suspensions. One of the fundamental issues addressed in this chapter was whether the mechanisms of electrosteric or steric dispersion, based on relatively large adsorbed polyelectrolytes for macroscopic size particulates, scales with particles in the range of ∼40 nm diameter such that a small, charged organic molecule such as citrate provides the thermodynamic colloidal stability of electrosteric mechanisms as suggested by preliminary theoretical calculations. The particle diameter-number distributions for as-prepared and after drying (at 25°C) and redispersion were used as metrics for thermodynamic colloidal stability. How efficiently particles redispersed after drying and reintroduction into the 70:30 ethanol:water solvent was used as the primary metric for whether the metastable electrostatic mechanism or thermodynamically stable electrosteric or steric approaches were responsible for the robust dispersion experimentally observed in the colloids. These experiments found that, even with the thin electrosteric layer provided by the adsorbed citrate, particles were electrosterically dispersed, and were unagglomerated when dried under argon and redispersed.;Preliminary work outlining the synthesis and characterization of silver core, calcium phosphosilicate shell nanoparticles for surface plasmon coupled emission and metal enhanced fluorescence applications is discussed in Chapter 6. Thin (2-5 nm) calcium phosphosilicate shells were formed around agglomerated silver cores in the presence of 8-Methoxypyrene-1,3,6-trisulfonic acid trisodium salt (MPTS). Calcium phosphosilicate shells were consistently formed after 72 hours in the presence of 5x10-5 M CaCl2, 3x10 -5 M Na2HPO4, 3x10-6 M Na 2SiO3, and silver core nanoparticles prepared by citrate reduction in aqueous solution. However, the particles synthesized were agglomerated, resulting in a loss of the plasmon resonance peak, and the shells prepared were not thick enough to provide sufficient separation of the fluorophore from the surface to prevent quenching and allow plasmon resonance enhanced fluorescence. (Abstract shortened by UMI.)