Three studies were conducted to determine the effects of roasting on ruminal degradability of sunflower seeds and the effects of feeding roasted sunflower seeds on ruminal fermentation, nutrient digestibility and milk yield and composition of dairy cows. Experimental treatments were a control diet with no added sunflower seed, a raw sunflower seed diet and a roasted sunflower seed diet. Sunflower seed diets contained 6% fat whereas the control diet contained 3% fat. In study one, two ruminally fistulated cows were used in a randomized complete block design to determine the effects of roasting on ruminal degradation of sunflower seeds. In the second study, three ruminally cannulated lactating Holstein cows were used in a 3 x 3 Latin square experiment to determine the effects of dietary treatments on ruminal fermentation and total tract nutrient utilization. In the last study, three primiparous and six multiparous Holstein cows were used in three 3 x 3 Latin squares to determine the effects of dietary treatments on milk yield and composition.

The removal of multiple substrates in a defined mixed culture process was investigated in the treatment of brewery wastewater. The study was conducted using both batch and a semi-continuous reactor system called self-cycling fermentation. Batch experiments were conducted using a synthetic brewery wastewater containing glucose, ethanol and maltose. Activated sludge from a municipal wastewater treatment plant was acclimatized in the synthetic brewery wastewater. The microbes capable of degrading this wastewater were analyzed by a combination of microscopy, spread plating, and Polymerase Chain Reaction-Denaturing Gradient Gel Electrophoresis (PCR-DGGE) and identified as Acinetobacter sp., Enterobacter sp. and Candida sp. From the pure culture batch experiments, it was determined that Enterobacter could degrade glucose and maltose but no ethanol, while Acinetobacter and Candida could degrade all three carbon sources. In mixed culture batch experiments, Enterobacter was dominant in degrading the sugar concentrations to levels permissible for Acinetobacter to degrade ethanol. PCR-DGGE was found to be effective in identifying the dominant species but selective carbon source plating was required to determine viability and track the population dynamics. Kinetic experiments were carried out in a semi-continuous, self-cycling fermentation process using the defined mixed culture in media containing glucose and various initial concentrations of ethanol and maltose. The overall rate of substrate removal was attributable to both the suspended culture and the biofilm formed during the process. A rate expression was developed for this system for the range of substrate concentrations tested. The data indicated that substrate removal by the suspended culture was a function of only the biomass concentration. However, substrate removal by the biofilm was found to be limited to the surface cells and determined to be a function of substrate concentration only.

Production of poly(3-hydroxybutyrate) (PHB) was done in a cyclone bioreactor using various culture methods; including batch (lab and pilot scale) fed-batch and self-cycling fermentation with and without starvation periods. Alcaligenes eutrophus DSM 545 was used to accumulate about 87% (wt/wt) PHB to a total of 6.2 g L$ sp{-1}$ PHB at the end of a 48 hour batch. Similar pilot scale experiments contained a maximum of 96% (wt/wt) PHB with 4.9 g L$ sp{-1}$ accumulated. Fed-batch culture of A. eutrophus produced 96% (wt/wt) PHB with a final PHB concentration of 22.2 g L$ sp{-1}$ after 54 h. Self-cycling fermentation (SCF) production of PHB resulted in an average of 35% (wt/wt) PHB without starvation periods with production rates reaching 0.24 g L$ rm sp{-1} hr sp{-1}.$ With starvation periods of 4, 6 and 8 h extended on the cycle times, production of PHB decreased except in the 8 hour starvation period which was 59% (wt/wt). However, the rates of production all decreased to below 0.13 g $ rm L sp{-1} h sp{-1}$ as the lengths of the starvation periods were increased.

Self-cycling fermentation (SCF) control was applied to a semi-continuous, aerobic, fixed-film reactor for the treatment of simulated and real brewery wastewater. The simulated wastewater was treated at 25°C. After approximately 3 hrs, 86% to 96% of the soluble BOD (SBOD) was removed, producing an effluent with a SBOD between 169 mg/L and 412 mg/L. The treatment of real brewery wastewater was undertaken at 25°C and 35°C, and was operated under ammonia-nitrogen deficient condition. 83% to 92% of the total BOD (TBOD) was removed after 3 hrs at 25°C, and after 1.5 to 2 hrs at 35°C. The treated effluent produced had a TBOD between 120 and 438 mg/L. The suspended solids in the treated effluent contributed between 63% and 71% of the TBOD. Hence, better treatment efficiency would have been possible if an efficient clarifier was installed, or the suspended solids were removed from the brewery wastewater prior to treatment.

Le but de mon projet de doctorat est la recherche de chitosanases thermostables qui peuvent mener la réaction d'hydrolyse du chitosane à de hautes températures. La procédure mise au point pour isoler ces chitosanases était planifiée pour moduler l'effet antimicrobien du chitosane qui augmente avec son poids moléculaire. Les objectifs spécifiques de ce projet sont, mettre au point un nouveau dosage de Csn, purifier, caractériser et cloner le gène des chitosanases les plus thermostables sélectionnées et mettre au point un milieu de production de chitosanase. La première étape du projet est la recherche de nouvelles chitosanases thermostables, via un criblage ciblé de bactéries productrices de chitosanases. En effet, une nouvelle méthode d'enrichissement était utilisée par l'ajout de chitosane de différents poids moléculaires à notre source bactérienne, soit les composts. La deuxième étape, est la réalisation d'un dosage de l'activité chitosanase en utilisant le soluble-dyed Remazol Brillant Bleu-Chitosane (sRBB-C) qui a été mise au point pour détecter à grande échelle une activité chitosanase de manière facile et rapide. Enfin, la troisième étape est un test de thermostabilité en présence de substrat, appliqué à des chitosanases choisies, pour sélectionner les plus performantes à l'étape de la purification. Parmi le lot de chitosanases testées, la chitosanase notée Csn1794 s'est distinguée par sa thermostabilité à 70 degrés C, ainsi elle a été retenue pour des études plus approfondies. Les études biochimiques réalisées sur la Csn1794 après purification ont révélé qu'elle a un poids moléculaire de 40 kDa, un pH optimal de 4.8 et des K[indice inférieur m] et k[indices inférieurs cat] de 0.042 mg/ml et 7588 min[indices supérieurs -1] respectivement. Le temps de demi-vie de la Csn1794 en présence de chitosane est plus de 20 heures à 70 degrés C. L'activité de la Csn1794 varie légèrement avec le degré d'acétylation du chitosane, elle hydrolyse la carboxyméthyl-cellulose, mais pas la chitine. Le clonage du gène de la Csn1794 par génétique inverse a permis de déterminer sa séquence. Ce gène codé pour une protéine de 441 acides aminés. La Csn1794 appartient à la famille 8 des glycosides hydrolases (GH8). Le rang taxonomique de l'isolat produisant la Csn1794 a été déterminé par des méthodes classiques ainsi que par des tests de biologie moléculaire. Les résultats obtenus indiquent qu'il s'agit d'un isolat appartenant à une espèce non caractérisée appartenant au genre Paenibacillus qu'on a appelé Paenibacillus sp. 1794. Enfin, la méthode de plan d'expériences était utilisée pour mettre au point le milieu de production de la Csn1794. Les essais réalisés par les plans d'expériences Plackett-Burman ont permis non seulement de définir un milieu de base pour la production de la Csn1794, mais aussi les oligosaccharides et le sucrose se sont distingués comme facteurs à effet nettement positif sur la production de la Csn1794. Les essais par plans d'expérience Box-Hunter ont permis l'étude d'interactions entre les différents facteurs dont le niveau était déterminé par les plans Taguchi. Les résultats obtenus indiquent qu'en plus de milieu de base, l'ajout de 10g/l de glucosamine, 7g/l d'oligosaccharide et 4g/l de sucrose constitue la meilleure combinaison pour un milieu qui permet de produire une moyenne de 7U/ml de Csn1794 d'une manière constante. En conclusion, nous disposons d'une nouvelle chitosanase thermostable, facile à produire et à purifier, qui sera un outil adéquat pour l'application au niveau industriel. Ceci va non seulement permettre de mener le processus d'hydrolyse de chitosane à haute température, mais aussi d'utiliser de grandes concentrations de substrat sans que la viscosité ne devienne excessive. Au niveau de la recherche fondamentale, la Csn1794 peut nous apporter plus d'informations d'une part, sur la thermostabilité des enzymes et d'autre part, sur les enzymes de la famille GH8, notamment les chitosanases.

Lactobacillus panis PM1 is a novel microorganism isolated from thin stillage (TS), a major by-product resulting from bioethanol fermentation, and was selected as the focus of this thesis due to its ability to produce 1,3-propanediol (1,3-PDO) from glycerol. The purpose of this thesis was to understand the central and auxiliary metabolic pathways of L. panis PM1 and to metabolically-engineer strain PM1 based on the improved metabolic knowledge for industrial applications. The 16S rRNA sequence and carbohydrate fermentation pattern were used to classify L. panis PM1 as belonging to the group III lactobacilli; thus, strain PM1 exclusively fermented glucose to lactate, acetate, and/or ethanol, clearly suggesting that its primary metabolism occurred via the 6-phosphogluconate/phosphoketolase (6-PG/PK) pathway. In contrast to typical group III lactobacilli, for fructose fermentation, L. panis PM1 utilized both the 6-PG/PK and the Embden-Meyerhof pathways, showing distinct strain-specific characteristics (more lactate, less acetate, no mannitol, and sporadic growth). In the PM1 strain, auxiliary metabolic pathways governed end-product formation patterns along with central metabolism. Under aerobic conditions, a coupled NADH oxidase-NADH peroxidase system was a determinant for NAD+ regeneration and was regulated by oxygen availability; however, the accumulation of its major end-product, hydrogen peroxide, eventually resulted in oxidative stress. The citrate-to-succinate route was another important auxiliary pathway in L. panis PM1. This route was directly connected to central energy metabolism, producing extra ATP for survival during the stationary phase, and was regulated by the presence of citrate, acetate, and succinate and a transcriptional repressor (PocR). Lactobacilli panis PM1 produced 1,3-PDO via the glycerol reductive route; however, the absence of the glycerol oxidative route restricted the utilization of glycerol to solely that of electron acceptor. Lower ratio of glucose to glycerol, in combination with PocR, repressed the glycerol reductive route, resulting in less 1,3-PDO production. In an effort to metabolically engineer L. panis PM1, an artificial glycerol oxidative pathway was introduced, and the engineered PM1 strain successfully produced a significant amount of important platform chemicals, including 1,3-PDO, lactate, and ethanol, solely from TS. Overall, this thesis reveals the significant feasibility of utilizing L. panis PM1 for industrial fermentative applications.

Thesis (M.Sc.)(Hons)--University of Western Sydney, Hawkesbury,1998. / "Thesis submitted in partial fulfillment of the requirements for the Degree of Master of Science (Honours) in Food Science." Includes bibliographical references.

Zugl.: Aachen, Techn. Hochsch., Diss., 2007.

Thesis (M.Sc. (Pharmacy))--Mahidol University, 1984.

Thesis (M.Sc. (Microbiology)) -- Mahidol University, 1979. / Partial support by National Research Council.