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A series of medium-chain-length poly(3-hydroxyalkanoates) (MCL-PHAs) with enriched 3-hydroxynonanoate (HN) content (up to 95.8 mol% compared to 68.6 mol% without acrylic acid) or 3-hydroxyoctanoate (HO) content (up to 97.5 mol% compared to 88.0 mol% without acrylic acid) was produced in continuous culture by co-feeding fatty acid and glucose plus inhibiting fatty acid β-oxidation using acrylic acid. Using a similar feeding strategy in fed-batch fermentation, similar monomeric compositions but a higher biomass concentration and PHA content could be obtained. However, at a lower growth rate (0.15 h-1 vs. 0.25 h-1), the biomass concentration and PHA content could be greatly enhanced from 17.1 to 71.4 g L-1 and from 64.4 to 75.5%, respectively, while the HN content decreased slightly from 92.2 to 88.9 mol%. PHAs produced under acrylic acid inhibition possessed improved physical properties including a higher melting point, faster crystallization rate, and greater tensile strength at break and Young’s modulus. Two recovery methods were developed for the recovery of MCL-PHA from Pseudomonas putida KT2440. One applied acetone extraction which was capable of recovering all the PHA from the cells with a purity of 91.6% and no detectable polymer molecular weight loss using Soxhlet extraction. Further purification was achieved by redissolving in acetone and reprecipitating in cold methanol. The other method used sodium hydroxide to solubilize the non-PHA cellular material. PHA purity of about 85% was obtained from a biomass containing 65.6% PHA after treatment with 0.2 N NaOH at 22 ± 1oC for 2 h or with 0.1 N at 80 ± 1oC for 15 min. However, a treatment at 22 ± 1oC followed by a second NaOH treatment at 80 ± 1oC resulted in higher PHA purity (94.7%) with a recovery efficiency of 88%. Under these conditions, NaOH digestion had a negligible effect on PHA molecular weight. / Thesis (Ph.D, Chemical Engineering) -- Queen's University, 2010-08-30 22:44:44.501

Streptococcus sanguinis is a gram-positive facultative anaerobe that is indigenous to the oral cavity and a primary colonizer of the oral cavity. It serves as a tether for the attachment of several oral bacteria that colonize the tooth surface, form dental plaque, and cause periodontal disease. Previous experiments with streptococcal strains have suggested that cellular chain morphology of streptococci may influence the competitiveness, susceptibility to phagocytosis, acidurance, and aggregation of the bacterium. The purpose of this study was to systematically determine gene functions that contribute to cellular chain length morphology in the SK36 strain of S. sanguinis. Gene functions for 2048 mutants were elucidated along with Clusters of Orthologous Groups (COG) functions that may be related to or regulate chain formation and morphology. The COG functions with high ratios of genes involved with chain length morphology per number of total non-essential mutant COG functions were in the following order: Cell division and Chromosome Separation, Defense Mechanisms, and Signal Transduction Mechanisms, and Cell Motility and Secretion. Examination of gene annotations of the 326 mutants involved with chain morphology suggests that cellular chain length is dependent on cell wall division and septation, peptidoglycan synthesis, and cell wall mobility. Some of the genes that contribute to chain length properties may be co-regulated which may suggest that chain length phenotypes are a transcriptionally regulated property that further studies may confirm.

Wheat grain quality and consumption is influenced by its constituents structure and concentrations. In the first part of the dissertation, six Canadian bread wheat cultivars; four (CDC Teal, AC Superb, AC Barrie, AC Splendor) belonging to the Canada Western Red Spring (CWRS), and two (AC Foremost, and AC Crystal) to the Canada Prairie Spring Red (CPSR) market classes were characterized for the relationship between their starch constituents and starch in vitro enzymatic hydrolysis. CPSR cultivars with relatively longer amylopectin chains of DP 37-45, reduced chain lengths of DP 15-18, and a low volume percent of small C-type starch granules, had reduced starch in vitro enzymatic hydrolysis rates. In the second part of the dissertation, near-isogenic wheat lines differing at the Waxy locus were analyzed for the influence of genome-specific granule-bound starch synthase I (GBSSI/Waxy; Wx-A, Wx-B, Wx-D) on starch composition, structure and starch in vitro enzymatic hydrolysis. Amylose concentration was more severely affected in genotypes with GBSSI missing from two genomes (double nulls) than from one genome (single nulls) of wheat, indicating dosage dependent amylose synthesis. Subtle differences in amylopectin chain length distribution were observed among non-waxy, partial and completely waxy starches, suggesting a non-limiting role of genome-specific GBSSI for amylopectin synthesis. A suppressive role of Wx-D on the short chain phenotype of wheat amylopectin was observed. In addition, Wx-D increased the volume percentage of large A-type starch granules and reduced starch hydrolysis index. Thus, among the waxy isoproteins, Wx-D might be the major contributor for reducing the rate of in vitro starch enzymatic hydrolysis in wheat. In the third part of the dissertation, endosperm starch’s physicochemical properties and structure during grain development in wheat waxy-null genotypes were analyzed. The study was conducted with pure starch isolated from wheat grains at 3-30 days post anthesis (DPA), at three day intervals. Changes in amylopectin structure were observed until 12 DPA, suggesting the formation of a basic amylopectin skeleton by this stage. A differential influence of waxy isoproteins on amylopectin structure formation has been suggested, with Wx-B and Wx-D affecting short glucan chains of DP 6-8 at 3 and 6 DPA, Wx-A being effective at 9 and 12 DPA, and Wx-D affecting DP 18-25 chains from 18-30 DPA.

In vivo production of medium-chain-length poly(3-hydroxyalkanoates) (MCL-PHA) containing a side chain carboxyl group from azelaic acid (AzA), a nine-carbon α,ω-dicarboxylic acid, was investigated using Pseudomonas citronellolis DSM 50332 in a phosphate (P)-limited chemostat. Co-feeding with nonanoic acid (NA) and inhibition of β-oxidation with acrylic acid (AA) were strategies that were used to stimulate the incorporation of carboxylated monomers, but both were unsuccessful. P. citronellolis DSM50332 was capable of growing on AzA as a sole source of carbon and energy, indicating that enzymes in β-oxidation utilized AzA and its derivatives. However, the MCL-PHA produced from AzA comprised 3-hydroxyoctanoate (C8) and 3-hydroxydecanoate (C10) monomers, which was consistent with precursor supplied via the de novo fatty acid biosynthesis pathway. This evidence suggests that one or more of 3-ketoacyl-CoA reductase (FabG), enoyl-CoA hydratase (PhaJ) and PHA synthase (PhaC) of this organism do not have the low specificity required to utilize a carboxylated substrate. Future work involving mutations may broaden the substrate specificity of these key enzymes to overcome this obstacle. Two-stage high-cell density carbon (C)-limited chemostat cultivation of P. putida KT2440 was examined for MCL-PHA production from nonanoic acid (NA) at high intracellular polymer content and volumetric productivity. Growth conditions stimulating good PHA production were first established in single-stage chemostat, which yielded 63.1 wt% PHA containing 90 mol% C9 units and a productivity of 1.52 g L-1h-1 at a dilution rate of 0.30 h-1. This productivity was higher than any value reported in literature for continuous MCL-PHA production systems and comparable to the upper range of fed-batch results. Two-stage production yielded promising results, notably the increase in polymer content from the first to second stage. However, complications involving foaming and an unexplained decline in PHA content adversely affected system performance. The best PHA content and overall productivity were 58.5 wt% and 0.76 g L-1h-1, respectively. Nonetheless, the results demonstrate the potential to achieve high PHA content without the need for pure oxygen at high dilution rates, warranting further investigation focusing on the optimization of growth conditions. / Thesis (Master, Chemical Engineering) -- Queen's University, 2011-12-19 15:48:21.808

<p> When a polymer system is strained below its glass transition temperature, T9 , deformed regions called crazes may be formed that have a characteristic extension ratio, λ. Examining the regions of deformed material, which are almost visible to the naked eye, and measuring λ with the use of atomic force microscopy gives information about the space between entanglements along a polymer chain, which is a truly molecular quantity. In this work we present the results of experiments in which entanglements in high molecular weight polystyrene (PS) samples have effectively been swelled by diluting the network with low molecular weight PS. We find that these experiments not only tell us how the molecular weight of a polymer can affect its contribution to the polymer network, but also give an entirely new method of determining the entanglement molecular weight, Me, of a polymer system. </p> / Thesis / Master of Science (MSc)

β-glucan is a major component of the fungal cell wall consisting of (1→3)-β linked glucose polymers with (1→6)-β linked side chains. The published classical isolation procedure of β-glucan from Saccharomyces cerevisiae is expensive and time-consuming. Thus, the aim of this research was to develop an effective procedure for the extraction of glucans. We have developed a new method for glucan extraction that will be cost effective and will maintain the native structure of the glucan. The method that we developed is 80% faster and utilizes 1/3 of the reagents compared to the published classical method. Further, the method developed increases the yield from 2.9 % to 10.3 %. Our new process has a branching frequency of 18.4 down from 197 and a side chain of 5.1 up from 2.5. The data indicate a more preserved native structure of isolated glucans.

Freshwater ecosystems are some of the most threatened on the planet. Efforts to conserve, restore, or otherwise manage large rivers and the services they provide are hindered by limited understanding of the functional dynamics of these systems. This shortcoming is especially evident with regard to trophic structure and energy flow. In this study I use natural abundances of carbon and nitrogen isotopes to examine patterns of energy flow and food-chain length of large-river food webs characterized by different landscape-scale hydrologic features. Ten locations along an approximately 500 km stretch of the Upper Paraná River Basin, Brazil, provided the setting for this work. Carbon derived from C3 plants and phytoplankton were the dominant energy sources across all webs, but relative contributions differed among landscape types (low-gradient river, high-gradient river, river stretches downstream of reservoirs, and reservoirs). Increases in food chain length corresponded with higher relative importance of phytoplankton derived carbon, likely due to size-structured effects of the phytoplankton-zooplankton-secondary consumer trophic link. River impoundment corresponded with decreased ecological and economic efficiency of fisheries production, an important ecosystem service provided by many tropical rivers.

This thesis addresses the mechanisms involved in cellulosedegradation in general and Totally Chlorine Free (TCF) bleachingof pulp in particular. The thesis shows that the cellulosedegradation during high consistency ozone bleaching is explainedby free radical chain reactions. By simulation, it has been shown that the number, weight andviscosity average of liner polymer chain length can be used tocalculate the number of random scissions in a linear polymer ofany molecular weight distribution, provided that there is acalibrated Mark-Houwink equation. A model describing partialdegradation of molecular weight distributions of linear polymersmeasured with viscometry was developed and verifiedexperimentally. The model predicts viscometric measurement ofchemical cellulose degradation by a rapidly reacting reagent tobe strongly dependent on cellulose accessibility. The role of free radical reactions in cellulose degradationwas studied by varying the amount of ferrous ions and ozone addedto the cotton linters. The result was compared to the resultsobtained from cellulose of lower crystallinity (cellulose beads)by measuring average chain length. When a ferryl ion reacted withcotton linters in the presence of ozone, the very formation ofone glycosidic radical was more significant to degradation thanthe final step of forming one oxidised glycoside. The inefficientdegradation observed of the oxidation step is explainable by theamount of accessible glycosides being too small to influenceviscometry. The efficient degradation observed in associationwith the glycosidic radical formation is explained by initiationof free radical chain reactions that are propagated as long asthere is ozone in the system. As none of these phenomena werefound in the less crystalline cellulose, cellulose structureappears to be important for how free radical mediated cellulosedegradation develops. The theory of free radical chain reactions coupled withdiffusion suggests a concentric expansion of the chain reactionsoutwards from the initial site of radical formation duringozonation of carbohydrates. This was confirmed by demonstratingfree radical chain reactions spreading from a spot of initiationoutwards during ozonation of a filter paper, using a pH-indicatorto monitor acid formation. Furthermore, the interior and exteriorof cellulose fibres doped with initiator were shown to bepermeated by small holes after ozonation. Ethylene glycol was shown to improve the selectivity duringozone bleaching of oxygen bleached kraft pulp at pH 3. Optimalconditions were obtained at pH 3 for 25 wt% ethylene glycol. Theinfluence of ethylene glycol on selectivity is explained by aproportion of the free radical chain reactions being carried bythe ethylene glycol instead of the cellulose during ozonebleaching. The observations were summarised in the form of amodel where the observed degradations for pulp, bleached pulp andcotton fibres during both ozone bleaching and ethylene glycolassisted ozone bleaching were shown to agree with each other. From g-irradiation of ozonised aqueous solutions of alcohol,the rate constant of superoxide formation from the peroxylradical of methanol was estimated to be 10 s-1. Rate constants of the reactions between ozone andalkylperoxyl radicals were determined to be around 104M-1s-1. The possibility of the reaction betweenalkylperoxyl radicals and ozone contributing significantly tofree radical chain reactions during ozonation of carbohydratesand alcohols could therefore be ruled out. Cellulose, degradation, free radical, ozone, selectivity,ethylene glycol, alcohol, bleaching, kraft pulp, cotton linters,delignification, fibre, fibril, crystallinity, ferryl ion, freeradical chain reactions, TCF, viscometry, molecular weightdistributions, random scissions.

The research presented in this dissertation involves the synthesis, characterization, and application of novel anionic and cationic chiral molecular micelles in capillary electrophoresis (CE) for the separation of diverse chiral compounds. Chapter 1 presents brief overview of the surfactants, micelle polymer, CE and micellar electrokinetic chromatography (MEKC). Chapter 2 describes the simultaneous enantioseparation of eight single chiral center â-blockers using two novel leucine and isoleucine based polymeric surfactants. The simultaneous enantioseparation of multichiral center bearing â-blockers, nadolol and labetalol is described in chapter 3. A synergistic approach, using a combination of polysodium N-undecenoxycarbonyl-L-isoleucinate (poly-L-SUCIL) and sulfated â-CD showed dramatic enantioseparation of four stereoisomers of nadolol. On the other hand for labetalol, enantiomeric separation remains unaffected using the dual chiral selector system. Chapter 4 deals with the enantiomeric separation of the binaphthyl derivatives that was found to be influenced by pH, type and concentration of the background electrolyte as well as concentration of the polymeric surfactant. In chapter 5, characterization of five alkenoxy leucine-based surfactants with variations in chain length (C8-C11), polymerization concentration and degree of polymerization showed significant effects on the chiral resolution and efficiency of hydrophobic â-blockers. The synthesis and characterization of two positively charged amino acid derived chiral ionic liquids (ILs) and their corresponding polymers is presented in chapter 6. Chiral separation of two acidic analyte (difficult to resolve with anionic micelles) can be achieved with both monomers and polymers of ILs. In chapter 7, the synthesis and detailed characterization of three pH independent amino acids derived (L-leucinol, L-isoleucinol and L-valinol) sulfated chiral polymeric surfactants is presented. These chiral sulfated surfactants are thoroughly characterized and the morphological behavior of polymeric sulfated surfactants is revealed using cryogenic high-resolution electron microscopy. The work clearly demonstrates for the first time the superiority of chiral separation in MEKC coupled to mass spectrometry at low pH. Finally, in chapter 8, six amino acid derived chiral surfactants with carboxylate and sulfate head groups were compared for enantioseparation of broad range of structurally diverse racemic compounds at neutral and basic pH conditions.