Applications of self assembling nanoemulsion (SANE) techniques in bioprocessing and quantum dots (QDs)

This dissertation investigates applications of self assembling nanoemulsion techniques to increase adherent cell density and enhance medium requirement in bioreactors and methods to reduce quantum dot toxicity. It includes three studies, which are described in chapter 3 through chapter 5. Study 1 (Chapter 3) explores the impact of a dextran containing nanoemulsion-based nanocarrier on anchorage-dependent cells densities in a bioreactor. Cellulose-derived microcarriers were used as controls as they have been successfully applied to enhance the growth of anchorage-dependent cells maintained especially in bioreactors. Our studies utilized self-assembling nanoemulsions (SANE) formed by a modified phase inversion temperature (PIT) process to produce dextran oil and surfactant-containing nanocarriers having mean particle sizes of 26--30 nm compared to microcarriers which were greater than 6000 nm. Our results demonstrated that dextran-containing nanocarriers supported up to 10 fold greater cell density, and increased lactate concentration in the medium by 12%, protein lystate concentration from cells by 83% and greater glucose concentration by 59%, compared with cells cultured in the presence of an equal concentration of microcarriers. Therefore, nanocarriers with increased numbers of dextran molecules, developed in these studies may be useful to further increase the production of anchorage-dependent animal cell-derived products or production of mass cell growth for other applications.;Study 2 (Chapter 4) investigated the possibility of reducing media volume to maintain cell viability, by utilizing the same dextran-based nanocarrier prepared from a self-assembling nanoemulsion (SANE) method. Our results showed that at the same 60 mL volume of media, cell viability after day 3 was 6 fold greater in CHO cells exposed to dextran-containing nanocarriers compared to cellulose-based microcarriers. When CHO cells were exposed to 60 mL of media containing dextran-based nanocarriers compared to 100 mL of media for cellulose-microcarriers, at day 6, cell density was 7 fold greater similarly, the concentration of proteins obtained from cells at day 6 was nearly 3 fold greater for CHO cells exposed to dextran-containing nanocarriers compared to the cellulose-based microcarriers. Nanocarriers had 59% greater glucose concentration, used as a measure of the polymer dextran and cellulose content levels in the nanocarriers and microcarriers, respectively. Therefore, nanocarriers with increased numbers of dextran molecules, developed in these studies may be useful to further optimize media volume requirements for maximum cell growth.;Study 3 (Chapter 5) involves the characterization of encapsulating Cadmium Selenide quantum dots using Self Assembling Nanoemulsions as a method to reduce QD toxicity (SANE). Although, nanometer-scale semiconductors QDs have attracted widespread interest in medical diagnosis and treatment, many can have intrinsic toxicities, especially those composed of CdSe, associated with their elemental composition. Using our self-assembling nanoemulsion (SANE) formulations which we have previously reported to be composed of non-toxic components, i.e., such as vegetable oil, surfactant and water, we hypothesized that their appropriate utilization would reduce the toxicity of QDs by encapsulating the CdSe QDs in our (SANE) system using a modified phase inversion temperature (PIT) method. SANE encapsulation of the QDs did not alter their emission wavelength of 600 nm, which remained unchanged during the encapsulation process. In contrast, the zeta potential of encapsulated QDs was reduced from -30 mV to -6.59mV, which was previously reported to be associated with beneficial properties (increased bioavailability and efficacy) for SANE-encapsulated bioactives such as pharmaceuticals. Relative to the number of cells in untreated controls (3.30+/-0.01 X 104 cells/mL), the viability of HeLa cells exposed for 48 hours to un-encapsulated CdSe QDs at a concentration of 115 microg/ml was reduced by 77% (7.5+/-0.2 X 103 cells/mL) (p<0.05). In contrast, the number of viable HeLa cells following exposure to SANE-encapsulated CdSe QDs vs. un-encapsulated CdSe QDs at the same concentration was 3.02+/-0.11 X 104 cells/mL or a 307% increase in number of viable cells (p<0.05). When the dose of CdSe QDs was increased to 230 microg/mL, the number of viable HeLa cells after exposure to the unencapsulated CdSe QDs was 5.3+/-0.1 X 103 cells/mL compared to untreated controls (p<0.05). In contrast, at the same increased concentration (230 microg/mL) of unencapsulated CdSe QDs, the number of viable HeLa cells following exposure to SANE-encapsulated CdSe QDs was 2.90+/-0.11 X 104 cells/mL or a 448% increase in the number of viable cells (p<0.05). Exposure of HeLa cells to a nanoblank, (nanoemulsion without QDs), showed no significant effect on cell viability (3.21+/-0.08 X 104 cells/mL) compared to untreated cells (3.30+/-.0.01 X 104 cells/mL. In conclusion, application of our SANE technology for encapsulating QDs increases the viability of cells exposed to CdSe QDs while maintaining the original emission wavelength, and therefore may be applied to reduce QD toxicity.