Investigation of Laundering and Dispersion Approaches for Silica and Calcium Phosphosilicate Nanoparticles Synthesized in Reverse Micelles

Open Access
Tabakovic, Amra
Graduate Program:
Materials Science and Engineering
Doctor of Philosophy
Document Type:
Date of Defense:
June 10, 2015
Committee Members:
  • James Hansell Adair, Dissertation Advisor
  • Gary Lynn Messing, Committee Member
  • Gail Lynn Matters, Committee Member
  • Christine Dolan Keating, Committee Member
  • Nanoparticles
  • nanoparticle laundering
  • nanotechnology
  • biomedicine
  • drug delivery
Nanotechnology, the science and engineering of materials at the nanoscale, is a booming research area with numerous applications in electronic, cosmetic, automotive and sporting goods industries, as well as in biomedicine. Composite nanoparticles (NPs) are of special interest since the use of two or more materials in NP design imparts multifunctionality on the final NP constructs. This is especially relevant for applications in areas of human healthcare, where the use of dye or drug doped composite NPs is expected to improve the diagnosis and treatment of cancer and other serious illnesses. Since the physicochemical properties of NP suspensions dictate the success of these systems in biomedical applications, especially drug delivery of chemotherapeutics, synthetic routes which offer precise control of NP properties, especially particle diameter and colloidal stability, are utilized to form a variety of composite NPs. Formation of NPs in reverse, or water-in-oil, micelles is one such synthetic approach. However, while the use of reverse micelles to form composite NPs offers precise control over NP size and shape, the post-synthesis laundering and dispersion of synthesized NP suspensions can still be a challenge. Reverse micelle synthetic approaches require the use of surfactants and low dielectric constant solvents, like hexane and cyclohexane, as the oil phase, which can compromise the biocompatibility and colloidal stability of the final composite NP suspensions. Therefore, appropriate dispersants and solvents must be used during laundering and dispersion to remove surfactant and ensure stability of synthesized NPs. In the work presented in this dissertation, two laundering and dispersion approaches, including packed column high performance liquid chromatography (HPLC) and centrifugation (sedimentation and redispersion), are investigated for silver core silica (Ag-SiO2) and calcium phosphosilicate (Caw(HxPO4)y(Si(OH)zOa)b.cH2O, CPS) composite NP suspensions synthesized in a cyclohexane/ polyoxyethylene (5) nonylphenylether (Igepal® CO-520) /water reverse micelle system. In Chapter 3, an HPLC laundering and dispersion approach for Ag-SiO2 composite NPs is studied. The deposition and detachment behavior of 5 w/w 3- aminopropyltriethoxy silane (APTES) dispersed Ag-SiO2 suspensions on 1 w/o, 5 w/o and 15 w/o APTES treated SiO2 column stationary phase microspheres is investigated with respect to final % mass yields for collected NP suspensions. Field emission scanning electron microscopy (FESEM) studies confirm multilayer deposition of Ag-SiO2 NPs during loading and retention on the column during laundering stages of the process independent of w/o APTES treatment on stationary phase microspheres, with individual deposited Ag-SiO2 NPs observed to be in the 20-30 nm diameter range. Patchy deposition patterns and irreversible agglomeration are also confirmed via FESEM, as fractions of loaded Ag-SiO2 NPs remain attached on the 1 w/o, 5 w/o and 15 w/o APTES stationary phases after completion of the collecting stage of the process. Column experiments monitored by UV –visible spectroscopy, along with % mass yield calculations, suggest that the use of 5 w/o and 15 w/o APTES treated stationary phases increases overall Ag-SiO2 NP % mass yield in the final collected suspensions when compared to 1 w/o APTES treated materials when using 70:30 ethanol:water (by volume) at operational pH 6 as mobile phase solvent mixture during the collecting stage. This is in agreement with the prediction that an increasing amine surface coverage on SiO2 stationary phase microspheres will result in increased electrostatic repulsive interactions during the collecting stage of the process. A modified DLVO theory is used to calculate total interaction energies (kT) and attachment efficiencies (in %) for each of the three stages of the packed column HPLC process, but is shown not to be in agreement with experimental observations due to low attachment efficiencies (%)calculated for the laundering stage of the process. In Chapter 4, a packed column HPLC process, which is used to launder and disperse CPSNPs synthesized in cyclohexane/Igepal® CO-520/water reverse micelles and dispersed with 50 w/w citrate, is investigated. Similar to laundering approach reported in Chapter 3 for the Ag-SiO2 system, it consists of a loading, laundering and collecting stage, where solvent mixtures of varying dielectric constants are used as mobile phases to control colloidal interactions and support either deposition or detachment of CPSNPs from the packed column SiO2 stationary phase microspheres. FESEM studies on stationary phase samples recovered after loading and laundering stages confirm multilayer deposition of CPSNPs, indicating favorable conditions for attachment. SEM/EDS analysis of fractions collected during each stage of the process confirms removal of surfactant and spectator ions during the loading and laundering stages of the process, while presence of Ca, P and Si is confirmed for the NP suspensions collected during the collecting stage of the process. The dielectric constant of the mobile phase solvent mixture is found to influence the deposition and detachment of CPSNPs, as confirmed via modified DLVO theory calculations for the loading, laundering and collecting stages of packed column HPLC process. As expected, the DLVO calculations show that CPSNP removal is dependent on particle diameter, with smaller CPSNPs (< 20 nm) most likely lost during the loading and laundering stages of the process. Based on modified DLVO theory, high (~100%) attachment efficiencies are predicted for the loading and laundering stages, indicating that CPSNPs are deposited and retained on the column media. For the collecting stage of the process, detachment is predicted due to low attachment efficiencies calculated based on DLVO theory interaction energy curves. In Chapter 5, the development of two liquid chromatography tandem mass spectrometry (LC-MS/MS) techniques for the measurement of 5-fluorouracil (5-FU) dopant in CPSNP suspensions is presented and discussed. The first LC-MS/MS technique is limited due to a lack of validation for accuracy and precision, while the second, improved technique is successfully employed to determine the dependence of 5-FU dopant concentrations in CPSNP suspensions based on key synthetic parameters, including: micelle exchange time, addition of 5-FU drug to microemulsion A versus microemulsion B during synthesis, and scaling up of synthetic volume from 1x to 3x. Based on 5-FU concentrations (nM) measured by the improved LC-MS/MS method, a micelle exchange time of 2 minutes with the addition of the drug to microemulsion A is determined to be sufficient for adequate drug encapsulation for the typical 1x volume CPSNP synthetic procedure. This synthetic approach is then used in all studies reported in the Chapter 6. In Chapter 6, the effects of two laundering and dispersion approaches on the physicochemical properties, including particle diameter and morphology, drug encapsulation efficiencies and residual surfactant concentrations of final 5-FU doped CPSNP suspensions are studied. A centrifugation (sedimentation and redispersion) laundering and dispersion approach is developed and compared to a packed column HPLC laundering and dispersion approach where ~200 µm soda lime silicate glass spheres are utilized as the column packing material. Particle morphology and state of dispersion are assessed via transmission electron microscopy (TEM) and compared for 5-FU doped and ghost CPSNP suspensions laundered via either centrifugation or packed column HPLC. Suspensions are confirmed to have a spherical morphology and appear well dispersed independent of laundering method. Analyses of lognormal diameter distributions by number indicate that the doping of CPSNPs with 5-FU during CPSNP particle formation results in an increase in overall particle diameter independent of laundering method for the 50 w/w citrate CPSNP suspensions. Post-laundering secondary functionalization with polyethyelene glycol (PEG) and gastrin 10 (g10) is found to improve the colloidal stability of the final 5-FU doped CPSNP suspensions. Encapsulation efficiencies (%) determined from 5-FU dopant concentrations (nM) measured using LC-MS/MS and are low (< 1%), indicating that the synthetic approach reported here needs to modified to improve capture of 5-FU molecules in the CPSNP matrix. Based on optical density (OD) measurements via UV-visible spectroscopy, the residual surfactant concentrations for both 5-FU-CPSNP-citrate and ghost-CPSNP-citrate suspensions laundered by centrifugation are approximately an order of magnitude lower when compared to packed column HPLC laundered CPSNP suspensions (10-5 M for centrifugation versus 10-4 M for packed column HPLC), suggesting that multiple laundering cycles are necessary to improve surfactant removal using the packed HPLC approach presented here.