The application of gas liquid chromatography to the simultaneous analysis of sugars and sugar phosphates in biological samples /|nby David J. LeBlanc. -- 260 St. Catharines [Ont. : s. n.],

A study was undertaken' to determine the applicability of gas liquid chromatography to the simultaneous analysis of sugars and sugar phosphates from biological samples. A new method of silylation involving dimethylsulfoxide, hexamethyldisilazane, trimethylchlorosilane and cyclohexane (1:0.2:0.1:1) which rapidly silylated sugars and sugar phosphates was developed. Subsequent chromatography on a 5% SE-52 column gave good resolution of the sugar and sugar phosphate samples. Sugar phosphates decomposed during chromatography and were lost at the 7 x 10-3 ~mole level. Acidic ethanol extraction of yeast samples revealed background contamination from the yeast sample, the culture medium and the silylation reagents which would further limit the level of detection obtainable with the glc for sugars in biological samples to the 3 x 10-4 ~mole level.

Icewine fermentation by Saccharomyces cervisiae : fundamental stress responses and comparative fermentation dynamics

Please consult the paper edition of this thesis to read. It is available on the 5th Floor of the Library at Call Number: Z 9999.5 B63 P54 2007

The osmoadaptive response of the wine yeast Saccharomyces cerevisiae K1-V1116 during icewine fermentation

The adapted metabolic response of commercial wine yeast under prolonged exposure to concentrated solutes present in Icewine juice is not fully understood. Presently, there is no information regarding the transcriptomic changes in gene expression associated with the adaptive stress response ofwine yeast during Icewine fermentation compared to table wine fermentation. To understand how and why wine yeast respond differently at the genomic level and ultimately at the metabolic level during Icewine fermentation, the focus ofthis project was to identify and compare these differences in the wine yeast Saccharomyces cerevisiae KI-Vll16 using cDNA microarray technology during the first five days of fermentation. Significant differences in yeast gene expression patterns between fermentation conditions were correlated to differences in nutrient utilization and metabolite production. Sugar consumption, nitrogen usage and metabolite levels were measured using enzyme assays and HPLC. Also, a small subset of differentially expressed genes was verified using Northern analysis. The high osmotic stress experienced by wine yeast throughout Icewine fermentation elicited changes in cell growth and metabolism correlating to several fermentation difficulties, including reduced biomass accumulation and fermentation rate. Genes associated with carbohydrate and nitrogen transport and metabolism were expressed at lower levels in Icewine juice fermenting cells compared to dilute juice fermenting cells. Osmotic stress, not nutrient availability during Icewine fermentation appears to impede sugar and nitrogen utilization. Previous studies have established that glycerol and acetic acid production are increased in yeast during Icewine fermentation. A gene encoding for a glycerollW symporter (STL1) was found to be highly expressed up to 25-fold in the i Icewine juice condition using microarray and Northern analysis. Active glycerol transport by yeast under hyperosmotic conditions to increase cytosolic glycerol concentration may contribute to reduced cell growth observed in the Icewine juice condition. Additionally, genes encoding for two acetyl CoA synthetase isoforms (ACSl and ACS2) were found to be highly expressed, 19- and II-fold respectively, in dilute juice fermenting cells relative to the Icewine juice condition. Therefore, decreased conversion of acetate to acetyl-CoA may contribute to increased acetic acid production during Icewine fermentation. These results further help to explain the response of wine yeast as they adapt to Icewine juice fermentation. ii

Synthesis of isotope-labelled methoxypyrazine compounds as internal standards and quantitative determination of aroma methoxypyrazines in water and wines by solid-phase extraction with isotope dilution-GC-MS /

An efficient way of synthesizing the deuterium labelled analogues of three methoxypyrazine compounds: 2-d3-methoxy-3-isopropylpyrazine, 2-d3-methoxy-3- isobutylpyrazine, and 2-d3-methoxy-3-secbutylpyrazine, has been developed. To confirm that the deuterium labels had been incorporated into the expected positions in the molecules synthesized, the relevant characterization by NMR, HRMS and GC/MS analysis was conducted. Another part of this work involved quantitative determination of methoxypyrazines in water and wines. Solid-phase extraction (SPE) proved to be a suitable means for the sample separation and concentration prior to GC/MS analysis.Such factors as the presence of ethanol, salt, and acid have been investigated which can influence the recovery by SPE for the pyrazines from the water matrix. Significantly, in this work comparatively simple fractional distillation was attempted to replace the conventional steam distillation for pre-concentrating a sample with a relatively large volume prior to SPE. Finally, a real wine sample spiked with the relevant isotope-labelled methoxypyrazines was quantitatively analyzed, revealing that the wine with 10 beetles per litre contained 138 ppt of 2-methoxy-3-isopropylpyrazine. Interestingly, we have also found that 2-methoxy-3-secbutylpyrazine exhibits an extremely low detection limit in GC/MS analysis compared with the detection limit of the other two methoxypyrazines: 2- methoxy-3-isopropylpyrazine and 2-methoxy-3-isobutylpyrazine.

Isolation and partial characterization of host cell wall surface glycoproteins: their involvement in agglutination, attachment and appressorium formation by piptocephalis virginiana

Cell surface proteins obtained by alkaline extraction from isolated cell walls of Mortierella pusilla and M. candelabrum, host and nonhost, respectively, to the mycoparasite, Piptocephalis virginiana, were tested for their ability to agglutinate mycoparasite spores. The host cell wall protein extract had a high agglutinating activity (788 a.u. mg- t ) as compared with the nonhost extract (21 a.li. mg- t ). SDS-polyacrylamide gel electrophoresis of the cell wall proteins revealed four protein bands, a, b, c, and d (Mr 117, 100, 85 and 64 kd, respectively) at the host surface, but not at the nonhost surface, except for the faint band c. Deletion of proteins b or c from the host cell wall protein extract significantly reduced its agglutinating activity. Proteins band c, obtained as purified preparations by a series of procedures, were shown to be two glycoproteins. Carbohydrate analysis by gas chromatography demonstrated that glucose and Nacetylglucosamine were the major carbohydrate components of the glycoproteins. It was further shown that the agglutinating activity of the pure preparation containing both band c was 500-850 times that of the single glycoproteins, suggesting the involvement of both glycoproteins in agglutination. The results suggest that the glycoproteins band c are the two subunits of agglutinin present at the host cell surface. The two glycoproteins band c purified from the host cell wall protein extract were further examined after various treatments for their possible role in agglutination, attachment and appressorium formation by the mycoparasite. Results obtained by agglutination and attachment tests showed: (1) the two glycoprotein-s are not only an agglutinin responsible for the mycoparasite spore agglutination, but may also serve as a receptor for the specific recognition, attachment and appressorium formation by the mycoparasite; (2) treatment of the rnycoparasite spores with various sugars revealed that arabinose, glucose and N-acetylglucosamine inhibited the agglutination and attachment activity of the glycoproteins, however, the relative percentage of appressorium formation was not affected by the above sugars; (3) the two glycoproteins are relatively stable with respect to their agglutinin and receptor functions. The present results suggest that the agglutination and attachment may be mediated directly by certain sugars present at the host and mycoparasite cell surfaces while the appressorlum formation may be the response of complementary combinations of both sugar and protein, the two parts of the glycoproteins at the interacting surfaces of two fungi.

Geochemistry and petrogenesis of the McElroy and Larder Lake assemblages, Abitibi Greenstone Belt, Northeastern Ontario

The McElroy and Larder Lake assemblages, located in the southern Abitibi Greenstone Belt are two late Archean metavolcanic sequences having markedly contrasting physical characteristics arid are separated from one another by a regional fault. An assemblage is an informal term which describes stratified volcanic and/or sedimentary rock units built during a specific time period in a similar depositional or volcanic setting and are commonly bounded by faults, unconformities or intrusions. The petrology and petrogenesis of these assemblages have been investigated to determine if a genetic link exists between the two adjacent assemblages. The McElroy assemblage is homoclinal sequence of evolved massive and pillowed fl.ows, which except for the basal unit represents a progressively fractionated volcanic pile. From the base to the top of the assemblage the lithologies include Fe-tholeiitic, dendritic flows; komatiite basaltic, ultramafic flows; Mg-tholeiitic, leucogabbro; Mg-tholeiitic, massive flows and Fe-tholeiitic, pillowed flows. Massive flows range from coarse grained to aphanitic and are commonly plagioclase glomerophyric. The Larder Lake assemblage consists of komatiitic, Mg-rich and Fe-rich tholeiitic basalts, structurally disrupted by folds and faults. Tholeiitic rocks in the Larder Lake assemblage range from aphanitic to coarse grained massive and pillowed flows. Komatiitic flows contain both spinifex and massive textures. Geochemical variability within both assemblages is attributed to different petrogenetic histories. The lithologies of the McElroy assemblage were derived by partial melting of a primitive mantle source followed by various degrees of crystal fractionation. Partial melting of a primitive mantle source generated the ultramafic flows and possibly other flows in the assemblage. Fractionation of ultramafic flows may have also produced the more evolved McElroy lithologies. The highly evolved, basal, dendritic flow may represent the upper unit 3 of a missing volcanic pile in which continued magmatism generated the remaining McElroy lithologies. Alternatively, the dendritic flows may represent a primary lava derived from a low degree (10-15%) partial melt of a primitive mantle source which was followed by continued partial melting to generate the ultramafic flows. The Larder Lake lithologies were derived by partial melting of a komatiitic source followed by gabbroic fractionation. The tectonic environment for both assemblages is interpreted to be an oceanic arc setting. The McElroy assemblage lavas were generated in a mature back arc setting whereas the Larder Lake lithologies were produced during the early stages of komatiitc crust subduction. This setting is consistent with previous models involving plate tectonic processes for the generation of other metavolcanic assemblages in the Abitibi Greenstone Belt.

Integrating vitrinite reflectance, rock-eval pyrolysis, flourescence microscopy and palynology of the Athabasca oil sands, Kearl Lake area, northeastern Alberta

Three cores from the Kearl Lake Oil Sands area within the Athabasca deposit of northeastern Alberta have been analyzed to understand the thermal history of the McMurray and Clearwater formations of the Lower Cretaceous Mannville Group. The approach involves the integration of vitrinite reflectance (VR), Rock-Eval pyrolysis, fluorescence microscopy, and palynology. Mean VR varies between 0.21 and 0.43% Ro and indicates thermally immature levels equivalent to the rank of lignite to sub-bituminous coal. Although differing lithologies have influenced VR to some extent (i.e., coals and bitumen-rich zones), groundwater influence and oxidation seem not to have measurably altered YR. Rock-Eval analysis points to Type III/IV kerogen, and samples rich in amorphous organic matter (ADM) show little to no fluorescence characteristics, implying a terrestrial source of origin. Palynology reveals the presence of some delicate macerals but lack of fluorescence and abundant ADM suggests some degradation and partial oxidation of the samples.

Astringency and other oral sensations : biological sources of individual variation and association with food and beverage behaviour

Orosensory perception strongly influences liking and consumption of foods and beverages. This thesis examines the influence of biological sources of individual variation on the perception of prototypical orosensory stimuli, food liking, self-reported alcohol liking and consumption, and indices of health. Two orosensory indices were examined: propylthiouracil (PROP) responsiveness, a genetically-mediated index of individual variation associated with enhanced responsiveness to orosensory stimuli often expressed as PROP taster status (PTS); and thermal taster status (TTS), a recently reported index of orosensory responsiveness. Taster status in PTS and/or TTS confers greater responsiveness to most orosensory stimuli. Gender, age, ethnicity, and fungiform papillae (FP) density were not associated with orosensory responsiveness to tastants, an astringent, and a flavour. Unlike PROP responsiveness, FP density was not associated with TTS. Both PROP responsiveness and TTS were associated with increased responsiveness to orosensory stimuli, including temperature and astringency. For PROP, this association did not hold when stimuli were presented at cold or warm temperatures, which are ecologically valid since most foods and beverages are not consumed at ambient temperature. Thermal tasters (TTs), who perceive 'phantom' taste sensations with lingual thermal stimulation, were more responsive to stimuli at both temperatures than thermal non-tasters (TnTs). While PTS, TIS, and gender affected self-reported liking and consumption of some alcoholic beverages, gender associated with the greatest number of beverage types and consumption parameters, with males generally liking and consuming alcoholic beverages more than females. Age and gender were the best predictors of alcoholic beverageAiking and consumption. As expected, .. liking of bitter and fatty foods and cream was inversely related to PROP responsiveness. TTS did not associate with body mass index or waist circumference, and contrary to previous studies, neither did PROP responsiveness. Taken together, TnTs' greater liking of cooked fruits and vegetables and high alcohol, and astringent alcoholic beverages than TTs suggests differences between TTS groups may be driven by perceived temperature and texture. Neither an interaction between PTS and TTS nor a TTS effect on PROP responsiveness was observed, suggesting these two indices of individual variation exert their influences on orosensory perception independently.