<p>  </p> <p>As monoclonal antibodies (mAbs) become a more important part of the pharmaceutical industry, the need for better quicker analysis of then will also increase. To do this, better stationary phases specifically designed for mAbs need to be developed to analyze the quality of mAbs by their critical control attributes. The three main critical control attributes are size, charge, and glycosylation. This work focuses mainly on the development of stationary phases to analyze critical control attributes in size and charge through using a non-porous silica base and surface confined atom transfer radical polymerization to grow improved stationary phases that minimize undesired interactions and maximize desired interactions.</p>

>Magister Scientiae - MSc / Studies involving the use of natural clays such as bentonite, montmorillonite and natural zeolite clinoptilolite in water treatment have been reported. Researchers suggested cost effective processes, such as ion-exchange and adsorption for the removal of heavy metals from waste waters by using naturally occurring and synthetic materials. The current study investigated application of natural adsorbents in brine treatment. Brines are hypersaline waters generated in power stations and mining industries rich in Mg2+, K+, Ca2+,Na+, so,': cr and traces of heavy metals, thus there is a need for these brines to be treated to recover potable water and remove problematic elements. Natural adsorbents have been successfully used in waste water treatment because of their high surface area and high adsorptive properties when they are conditioned with acid or base. The natural adsorbents used in this study were obtained from Ecca Holdings company (Cape bentonite mine) Western Cape in South Africa, comprising bentonite clay and natural zeolite (clinoptilolite) and another clinoptilolite sample was obtained from Turkey. These adsorbents were investigated in their natural and pretreated form for removal of toxic elements in brine water. The pretreatment was aimed at removing Na+, K+, Ca2+, Mg2+ from the clinoptilolite as well as the bentonite and replacing these cations with the H+ cation to activate the materials. The cation exchange capacity (CEC) of natural zeolite from South Africa was found to be 2.14 meq/ g, Turkish Clinoptilolite was 2.98 meq/ g while South African bentonite was 1.73 meq/g. at 25°C using ammonium acetate (pH 8.2) method. Characterization of these natural adsorbents was done prior to pretreatment and after the treatment. ICP-AES analysis was used for determination of toxic elements in brines before and after sorption. The morphology of clays was characterized by X-ray diffraction (XRD), Brunauer Emmett Teller (N2-BET) and Scanning electron microscopy (SEM) for confirmatory purposes and X-ray Fluorescent spectroscopy (XRF) was used for the composition analysis of the natural adsorbent. The results from batch experiments prior to pretreatment of the natural adsorbents showed that these natural adsorbents contained Mg2+, K+, Ca2+, Na+ in their structures as charge balancing cations, thus needed pretreatment to remove the cations. The natural adsorbents were pre-treated with 0.02M HCI. After the pretreatment of natural adsorbents it was possible to enhance the percentage removal of the major cations from brine, and the Na+ and Mg2+ removal achieved (86 % and 85% respectively) from brine was more than C02+ (70% ) the SC was the adsorbent one that gave highest removal of cations in the brines. Trace elements removal was high with Cu2+and Zn2+ being the highest of toxic elements in brine. The optimum contact for the toxic element removal was found to be 30 min for the Turkish clinoptilolite and 1 hr for the South African clinoptilolite and South African bentonite clay. Leaching of Ae+ and Si4+ during adsorption was also investigated and it was found that less than 1 ppm of A13+ and Si4+ were leached into the solution during adsorption experiments indicating that these materials were stable. The investigation of pH showed that natural adsorbents did not perform well at low pH of 4 and 6. The adsorbents were able to work efficiently at the natural pH of 8.52 of the brine solution. These results show that natural adsorbents hold great potential to remove cationic major components and selected heavy metal species from industrial brine wastewater. Heterogeneity of natural adsorbents samples, even when they have the same origin, could be a problem when wastewater treatment systems utilizing natural clinoptilolite and bentonite are planned to be developed. Therefore, it is very important to characterize the reserves fully in order to make them attractive in developing treatment technologies.

In this study we have investigated the frequency dependence of bilayer lipid membranes for a series of glycerylmonoolein/ n-decane bilayers in various aqueous ionic solutions containing the ionophore valinomycin. Reliable values of membrane capacitance and conductance were obtained over the frequency range 0.2 - 200 KHz using an automatic balancing bridge under the control of a microprocessor unit. The admittance data was then normalized and curve-fitted to produce relaxation times and amplitudes from which the kinetic rate parameters, as deduced from a single slab dielectric membrane model, were calculated. These ac experimental rate constants were then compared with those obtained from charge-pulse relaxation methods.

Series of homo- and copolyimides containing controlled degrees of sulfonic acid ion conducting pendant groups have been synthesized from both phthalic (five-) and naphthalic (six-membered) dianhydrides and appropriate wholly aromatic diamines and heterocyclic analogues. The goal is to identify thermally and hydrolytically stable ion conducting polymers (ICP) suitable as proton exchange membranes, PEM, for fuel cells. The candidate ICP's have been synthesized and characterized for molecular weight, chemical composition, film forming properties, thermal transition behavior, boiling water stability, solvent solubility and water absorption and conductivity. Commercially available five-membered ring dianhydrides such as 6FDA, BPDA, and six-membered ring dianhydrides such as naphthalene tetracarboxylic dianhydride (NDA) have been used. High molecular weight five-membered ring polyimides were obtained from an equimolar ratio of diamines and dianhydride using a one-pot ester-acid procedure by initially converting the dianhydride to a diester-diacid derivative, followed by the reaction with sulfonated and unsulfonated aryl diamines. The sulfonated diamine monomer was allowed to oligomerize with the diester-diacid of the dianhydride for 2-3 hours, before unsulfonated diamine was charged into the reaction flask. The levels of sulfonation in the polymer backbones were controlled by varying the mole ratio of sulfonated diamine to unsulfonated diamine. For the six-membered ring polyimides, phenolic solvents, e.g. m-cresol, have been used. In general, 4,4′-diamino-biphenyl-2,2′-disulfonic acid (DPS) has been employed as the source of the sulfonated unit. The chemical compositions of both sulfonated and unsulfonated polyimides were obtained using ¹H-NMR and FT-IR. The sulfonic acid contents in both diamine monomers, as well as the sulfonated polyimides were also analyzed by acid-base potentiometric titration. In all cases, high inherent viscosity values and good film forming ability of the polymers were the key indications of high molecular weight. The viscosity values increased with an increase of sulfonation degree in the polymers. This increase of viscosity in these ionomers can be attributed to the increase of polymer chain aggregation with their increasing ionic character. Polymers were fabricated into membranes via solution casting or spin casting from DMAc or m-cresol in order to study film-forming properties. The solution cast dry films of the sulfonated polyimide membranes gave tough, ductile membranes and demonstrated moderate to high water absorption, which is necessary for PEM fuel cells. However, swollen films, in general, showed poor hydrolytic stability which resulted in brittle membranes. The solution-cast membranes were thermally analyzed to study the effect of the degree of sulfonation on the thermal properties of sulfonated polymers. All the thermograms of the sulfonated polyimide films exhibited a two-step degradation behavior. The first weight loss, observed between 300-400 °C, corresponds to desulfonation in the sulfonated block, and the second weight loss, observed for a temperature around 500 °C or above, corresponds to the polymer backbone degradation. The TGA thermograms indicated that the initial weight losses were steeper for polymers with higher sulfonation degrees. Furthermore, the weight loss temperature of sulfonated polyimides decreased and broadened with increasing sulfonation levels. However, the onset temperature of the first weight loss was independent of the degree of sulfonation. Weight loss data in TGA curves of the sulfonated polymers were used to calculate the degree of sulfonation. Experimental and theoretical values were in good agreement with each other. The sulfonated five-membered polyimide membranes were aged in an air-oven at increasing temperatures (30-220 °C) for 30 min and then titrated with TMAH using non-aqueous potentiometric titration. All the films that were aged up to 220 °C were still completely soluble in DMAc. Moreover, the sulfonic acid groups were unchanged. In addition, several new flexible sulfonated and unsulfonated diamines and bis(naphthalic anhydride) monomers containing phosphineoxide [-P(O)-] or sulfone [-S(O)₂ -] moieties in their structure have been synthesized and characterized with various analytical techniques. The structural design of naphthalic polyimides by incorporating bis(naphthalic anhydrides) was one approach to give a better solubility and processability of their related products. Development of an iterative approach for defining the optimum degree of sulfonation that will produce the highest ionic conductivity while still retaining other important properties such as flexibility, strength, hydrolytic stability has been a goal of this research and will be discussed in the thesis. / Ph. D.

The mining industry is a significant contributor to the Canadian economy. However, the mining activities can be detrimental to the environment due to the release of pollutants. Ammonia is one of the noxious and toxic contaminants associated with mining, ammonia contamination is created by the oxidizing agent in explosives. The explosives impacted mining wastewater (EIMWW) usually contains ammonia and other metal ions. The ammonia in EIMWW could harm the aquatic environment by the depletion of oxygen and its lethal toxicity to aquatic organisms. Before release to environment, EIMWW needs to be treated with an easy-to-operate method for ammonia removal at the remote mining sites. Ion-exchange (IE) with zeolite is an effective method for ammonia removal that is easy-to-operate, is not significantly impacted by cold temperature or toxicity effects. However, the traditional IE regeneration approach of using high concentration NaCl solutions creates a secondary polluting stream. Chlorine regeneration of ammonia-loaded zeolite appears to be a promising option, an evaluation of this option is the main topic of this thesis. This thesis includes three initiatives. The first is a set of multi-cycle batch loading-regeneration tests to assess the viability of ammonia removal with a commercial zeolite (SIR-600) for the treatment of a synthetic EIMWW (containing total ammonia nitrogen (TAN), K, and Ca) and to examine the performance of different ion-exchange regeneration solutions. The long-term TAN uptake of SIR-600 regenerated using a NaOCl (100 mg free Cl2/L) solution was 0.24 meq/g, which was approximately 20% lower than that after a NaCl regeneration. However, chlorine regeneration is promising because the selectivity of SIR-600 for TAN over Ca and K increased after the chlorine regeneration. To simulate recycling of the NaOCl regenerants, K and Ca were added to the NaOCl solution, it did not substantially affect the subsequent SIR-600’s ion uptake. This initiative represents a significant contribution since the earlier studies into chlorine regeneration did not investigate the impact of competing ions. The second initiative addressed concerns regarding the long-term integrity of SIR-600 arising from its exposure to high chlorine concentrations during the regeneration. The five-week long chlorine batch exposure tests with solutions of up to 1000 mg free Cl2/L showed that chlorine exposure did not significantly affect the SIR-600’s characteristics in terms of particle size distribution, surface area, FTIR spectra and ion uptake. Thus, SIR-600 has the potential for long-term use in field applications. The final initiative evaluated the feasibility of chlorine regeneration for continuous flow IE column systems used for ammonia removal from a synthetic EIMWW. Continuous flow column systems are important because these are the standard IE units used in full-scale applications. Multi-cycle column loading-regeneration tests were performed to compare the zeolite performance using a NaOCl (1000 ppm as free Cl2) solution with that using a 5% NaCl regeneration. The influence of loading duration was also assessed. The use of 6-hr loading cycles were shown to be preferable to 23-hr loading cycles because it had lower effluent concentrations and they could achieve higher overall TAN mass removals per unit time. After three operational cycles, the SIR-600 had similar TAN uptake performances (0.21 meq/g Vs. 0.21 meq/g) after NaOCl regeneration and after salt (NaCl) regeneration. This is in contrast to the lower TAN uptakes for the NaOCl regeneration in the batch tests, this indicates that batch tests are not always representative of full-scale applications. Compared to NaCl regenerated SIR-600, SIR-600 after NaOCl regeneration had a higher preference for TAN over Ca and K, which makes this type of regeneration very promising. Its only apparent limitation is that the NaOCl regeneration required a longer duration. During the NaOCl regeneration, the main mechanism appears to be the oxidation of ammonia to nitrogen gas and hydrogen ions, however the Na in the NaOCl solution also seems to have a role in the regeneration.

Nitrogen removal from municipal wastewater with application of Anammox process offers cost reduction, especially if it is combined with maximal use of organic content of wastewater for biogas production. In this study a new technology is proposed, which is based on ammonium concentration from municipal wastewater by ion exchange followed by biological removal of ammonium from the concentrated stream by partial nitritation/Anammox process. In experiments on ammonium concentration four the most common ion exchange materials were tested in packed bed columns, namely strong and weak acid cation exchange resins, natural and synthetic zeolites. Experiments with synthetic wastewaters with different content and municipal wastewater showed that strong acid cation resin is the most suitable for ammonium concentration from municipal wastewater due to its high exchange capacity and fast regeneration. Since NaCl was used for regeneration of ion exchange materials, spent regenerant had elevated salinity. Experiments with activity determination showed that both nitritation and Anammox bacteria are inhibited by NaCl, where effect on Anammox bacteria is more severe. Adaptation of partial nitritation/Anammox biomass was studied using two strategies of salinity increase and it was possible to adapt the biomass to NaCl content of 10-15 g/L. The technology was tested in batch mode using strong acid cation resin for ammonium concentration from pretreated municipal wastewater, and partial nitritation/Anammox biomass for nitrogen removal from concentrated stream. It was shown that it is possible to remove 99.9% of ammonium from wastewater with ion exchange while increasing concentration of ammonium in spent regenerant by 18 times. Up to 95% of nitrogen from spent regenerant was removed by partial nitritation/Anammox biomass in batch tests. Moreover, possibilities of integrati on of the technology into municipal wastewater treatment technology, challenges and advantages were discussed. / <p>QC 20140219</p>

Monoclonal antibodies are large, complex biomolecules that can be difficult to characterize. Characterization is important because of the various post translational modifications that can occur during manufacturing, processing, and storage. Some modifications can lead to efficacy and safety issues and therefore are heavily monitored. A leading way to monitor various modifications is by using liquid chromatography. The high sensitivity, reproducibility, and ability to quantitate analytes makes it very attractive for monoclonal antibody characterization. The large molecular size of monoclonal antibodies (150 kDa) makes them challenging to separate efficiently and with high enough resolution to be helpful. New column technologies that would help improve protein separation efficiencies and slectivities would greatly help in this challenging process. In this thesis, three novel bonded phases are developed for the separation of monoclonal antibodies including a weak anion and cation exchanger (WAX, CEX) for the separation of charged species as well as a novel hydrophilic interaction chromatography (HILIC) for the separation of glycoforms. Column develop is achieved by optimizing selectivity and improving efficiency of separations by altering particle surface chemistry.

No description available.

The objective of this research is to further define the relationship between the charge state of insulin, and the self assembly properties of insulin and PEGylated insulin in solution. Polyethylene glycol (PEG) chains were covalently attached to insulin in order to evaluate their impact on insulin¿s systemic duration of action after pulmonary dosing. This thesis will focus on the assembly properties of the PEG-insulin and insulin, and also demonstrate how the charge state, which was modified by the covalent attachment of PEG, relates to different modes of behavior by anion and cation exchange chromatography. In addition, explain how modifying the assembly state extends to improving formulation properties of spray-dried insulin powders. This thesis is an investigation into the relationship of insulin¿s charge state controlled by pH and how the charge state affects the self assembly of insulin, especially when the zinc ion is removed. Ionic interaction is one of the major forces affecting insulin assembly. The theory that a change in the charge state of insulin could modulate the ionic interaction and reduce hexamer formation at alkaline conditions was investigated. Experiments were designed to measure the level of hexamer with light scattering, and the amount of hexamer was then correlated with the pH and zinc content of the solutions. The importance of the charge state of the monomer and its behavior extends to chromatography and purification modes as well. Specifically, the purification of various species of PEGylated insulin presents a challenge. By varying mobile phase pH which induces the charge to insulin, an ion exchange method demonstrated very high resolution and controllable interaction between the ion exchange media and the insulin derivatives. A highly accurate method for determining molecular weight and thus the average associated state of insulin in solution has been developed using the MALS (Multi-Angle Light Scattering). Insulin concentration, pH, and metal ion concentrations, were in pharmaceutically relevant ranges. The MALS method was developed to evaluate how the parameters above affect the self-assembly properties of insulin, and use the assembly properties to improve formulations of insulin or PEGylated insulin. To use the light scattering technique the dn/dc (change in refractive index with change in concentration) is required. During the method development, the dn/dc of insulin was measured at 690 nm, and a value of 0.185 mL/g based on theory was confirmed. A novel approach for preparing insulin powders with improved chemical stability, based on maintaining the dissociation of hexamers in solution during the spray drying process was developed. The mode presented here is to remove the zinc ions from solution, increase the pH from 6.6 to 7.8, and maintain a low concentration of insulin approximately 2 to 15 mg/mL. Each of these factors alone decreases the hexamer population in solution, but by combining all three factors, hexamers are driven to very low levels of equilibrium. The increased stability of the powders is predominately related to the decrease in covalent insulin dimer (CID). The data presented correlates a reduced hexamer population in the solution with lower levels of CID¿s in the dry powder compared to controls. The CID formation rate was reduced by 40% compared to a control.

<p> An optimization study of an intermittent-flow multistage fluidized ion exchange column was performed using a stochastic approximation method. A new type of downcomer, a fluid diode, was designed and employed to alleviate liquid bypassing through the downcomer. The well known ion exchange system, H⁺/Na⁺ exchange on Dowex 50W resin, was used in this work. </p> <p> The volumetric efficiency of the system was optimized with regard to certain column and diode parameters. A maximum volumetric efficiency of 71.8 hr⁻¹ was obtained for the following conditions: </p> <p> average liquid flowrate = 3661 ml/min ; </p> <p> resin flowrate = 56.1 gm/min ; </p> <p> plate spacing 11.43 cm ; </p> <p> lateral diode displacement= 0.794 cm. </p> <p> Experiments have shown that a 78.2% increase in volumetric efficiency was achieved by use of the fluidic diode downcomers. </p> / Thesis / Master of Engineering (ME)