The continued increase in the demand for fossil fuels combined with their ever dwindling supply has prompted the search for a suitable alternative fuel. The research contained within this dissertation seeks to increase the lipid content of cellular feedstocks, improve extraction efficiencies of lipids, and understand the pathways involved in the production of fatty alcohols and triacylglycerides from microbial feedstocks. As part of this research the diatom, Cheatoceros gracilis, was grown at small and large scale to determine optimal growing conditions. No apparent nutrient stress trigger was required to initiate the accumulation of the biodiesel precursor triacylglyceride, unlike other documented algal strains. A follow-up to this project demonstrated that the microalga C. gracilis may utilize light intensity as a trigger for lipid production. A major difficulty in the production of biofuels from microorganisms is the expensive process of dewatering, drying, and extracting the lipid compounds from the cells. As part of this research, a process has been developed that allows for lipid extraction to occur in the presence of water at a point as low as 2 percent solids or 98 percent water. This process utilizes a single organic solvent that mixes well with microbial lipids, but poorly with water allowing for efficient extraction of lipids and fast solvent to water separation. This process greatly decreases the cost of the microbial biofuels production associated with the removal of water from cell slurries. Triacylglycerides and fatty alcohols are oleochemicals that are commonly used in industrial, pharmaceutical, and consumable processes. A predicted fatty acyl CoA reductase enzyme was cloned into an E. coli vector, expressed, characterized and shown to be active as a dual reductive enzyme reducing a fatty acyl CoA to its respective fatty alcohol, constituting the first enzyme of this type discovered in a bacterium. The process of triacylglyceride production in microbes is fairly well understood; however, the process that regulates this production has not yet been fully explored. As part of this research, the model yeast organism, Yarrowea lipolytica, is utilized to identify essential genes for citrate transport that if removed could result in increasing triacylglyceride production in vivo.

Liquid Biofuels mainly Bioethanol and biodiesel are the main replacement for fossil fuels in the current world. But there are questions and concerns about the present biofuels production, mainly when it comes to matter of sustainability. In this thesis paper,Strategic Life Cycle Management data along with Life Cycle Analysis data has been used to analyze the sustainable biofuels condition in Sweden and the Netherlands. Data also has been collected through the interview from different stakeholders in Sweden and the Netherlands.

Life cycle assessment is an important tool to evaluate the impact of 2nd generation lignocellulosic ethanol, and its potential greenhouse gas (GHG) emissions benefits relative to gasoline. The choice of feedstock, process technology, and co-products may affect GHG emissions and energy metrics. Co-products may improve both the financial and environmental performance of the biorefinery. 26 well-to-wheel models of future lignocellulose-to-ethanol pathways were constructed, considering corn stover, switchgrass, and poplar feedstocks, three pre-treatment technologies, four co-product options, and the use of ethanol in a light-duty vehicle. Model results showed that all pathways with lignin pellet co-production had significantly lower net GHG emissions relative to gasoline and corresponding pathways producing only electricity. Pathways co-producing xylitol had at least 66% greater GHG emission reductions relative to pathways co-producing only lignin pellets. All feedstock/pretreatment/co-product combinations led to GHG reductions of at least 60%, meeting the threshold stipulated under the Energy Independence and Security Act.

Life cycle assessment is an important tool to evaluate the impact of 2nd generation lignocellulosic ethanol, and its potential greenhouse gas (GHG) emissions benefits relative to gasoline. The choice of feedstock, process technology, and co-products may affect GHG emissions and energy metrics. Co-products may improve both the financial and environmental performance of the biorefinery. 26 well-to-wheel models of future lignocellulose-to-ethanol pathways were constructed, considering corn stover, switchgrass, and poplar feedstocks, three pre-treatment technologies, four co-product options, and the use of ethanol in a light-duty vehicle. Model results showed that all pathways with lignin pellet co-production had significantly lower net GHG emissions relative to gasoline and corresponding pathways producing only electricity. Pathways co-producing xylitol had at least 66% greater GHG emission reductions relative to pathways co-producing only lignin pellets. All feedstock/pretreatment/co-product combinations led to GHG reductions of at least 60%, meeting the threshold stipulated under the Energy Independence and Security Act.

Rising oil prices have been a concern for both developed and developing countries, especially in more recent years as it tends to have a crippling effect on production and transportation. Many countries have moved towards the development of fossil fuel alternatives as a means of achieving energy independence and achieving environmental targets (for example the Kyoto Protocol). Developments in both these types of energy markets (fossil fuel and renewable fuels) may impact Canadian Prairie agriculture. Most of Canadian prairie crops are exported. The Canadian prairies are land locked to some extent. The closest ocean access to the eastern portion of the prairies is the port of Churchill, but is closed during the winter season. Crops are therefore transported west through the Rocky Mountains or east through the Great Lakes to get to a port. This requires hundreds of kilometres of truck and rail transportation, which is fuel dependent. To a lesser extent, at the micro-level farmers depend on fossil fuels to operate machinery to facilitate efficient crop production. If oil prices continue on an upward trajectory, will farmers cropping behaviour change? Furthermore, the development of the bioethanol industry on the Canadian prairies has given wheat farmers another crop option. As oil prices increase, the price of ethanol increases as well. Also, demand is bolstered by renewable fuel standards and government tax exemptions or subsidies. This study seeks to put forward the notion that as oil prices increase, crop production and transportation costs also increase thereby reducing farmers’ gross margins. Also, ceteris paribus, as oil prices increase there will be an increased demand for, and an increase in the price of biofuels thereby increasing the price of biofuel feedstock. Higher feedstock prices are expected to increase the gross margins of farmers. Therefore higher oil prices drive increased crop competition between traditional cropping (cropping for food exports) and energy cropping. This thesis seeks to ascertain at what level of oil prices would farmers, in general, be willing to switch from producing wheat for traditional (hard/food wheat) purposes to bioenergy (soft/ biofuel wheat) cropping alternatives. Also under varying scenarios of oil price growth and government support to the biofuel industry, this thesis seeks to ascertain the impact of biofuel industry expansion on grain elevator pricing behaviour and the structure of the elevator industry, assuming elevators spatially compete with each other for farmers’ crops. An agent based model (ABM) is employed for this study. The model is selected over other types as the researcher wants to capture the increased complexity stemming from the competition between crops that belong to at least one distribution chain. Agent based networks allow for emergent behaviour that is obtained from the spatial competition of elevators. Finally, the agent based model allows for spatial heterogeneity in location of farmers in terms of soil quality and their proximity to an elevator, which affects crop productivity and transportation costs, respectively. The ABM (also called the FARMCHAIN model) is comprised of over 35000 farmer agents, 176 elevator agents, 6 canola crushing plant agents, 5 ethanol plant agents and 1 biodiesel plant agent located on the 20 census agricultural regions (CARs) of Saskatchewan. Farmers allocate land based on their expected gross margins. Farmers produce and truck crops to the designated distribution chain. Crops move through the chain and at every stage the associated costs are computed and apportioned to the farmer. At the end of the period, gross margins are computed and these gross margins are used in computing the expected gross margins for the subsequent period. It is found that real annual crude prices would have to be greater than $133 before farmers begin to switch to producing biofuel wheat (soft wheat) from food wheat (hard wheat). This would have to be approximately 30% higher than that of 2008 in which crude prices were at record levels. Also, if biofuel support is declining then it would take a considerably higher price to entice farmers, in aggregate, to switch.

Due to its availability, low-price, and higher degree of reduction than lignocellulosic sugars, glycerol has become an attractive carbon source for the production of fuels and reduced chemicals. However, this high degree of reduction of carbon atoms in glycerol also results in significant challenges in regard to its utilization under fermentative conditions. Therefore, in order to unlock the full potential of microorganisms for the fermentative conversion of glycerol into fuels and chemicals, a detailed understanding of the anaerobic fermentation of glycerol is required. The work presented here highlights a comprehensive experimental investigation into fermentative glycerol metabolism in Escherichia coli, which has elucidated several key pathways and mechanisms. The activity of both the fermentative and respiratory glycerol dissimilation pathways was found to be important for maximum glycerol utilization, a consequence of the metabolic cycle and downstream effects created by the essential involvement of PEP-dependent dihydroxyacetone kinase (DHAK) in the fermentative glycerol dissimilation pathway. The decoupling of this cycle is of central importance during fermentative glycerol metabolism, and while multiple decoupling mechanisms were identified, their relative inefficiencies dictated not only their level of involvement, but also implicated the activity of other pathways/enzymes, including fumarate reductase and pyruvate kinase. The central role of the PEP-dependent DHAK, an enzyme whose transcription was found to be regulated by the cyclic adenosine monophosphate (cAMP) receptor protein (CRP)-cAMP complex, was also tied to the importance of multiple fructose 1,6-bisphosphotases (FBPases) encoded by fbp, glpX, and yggF. The activity of these FBPases, and as a result the levels of fructose 1,6-bisphosphate, a key regulatory compound, appear to also play a role in the involvement of several other enzymes during fermentative glycerol metabolism including PEP carboxykinase. Using this improved understanding of fermentative glycerol metabolism as a platform, E. coli has been engineered to produce high yields and titers of ethanol (19.8 g/L, 0.46 g/g), co-produced along with hydrogen, and 1,2-propanediol (5.6 g/L, 0.21 g/g) from glycerol, demonstrating its potential as a carbon source for the production of fuels and reduced chemicals.

Biofuels are a promising solution to society's quest for sustainable energy. In the transportation sector, biodiesel is the leading alternative diesel fuel currently in use today. However, the current global and domestic production of biodiesel is far below the petro-diesel consumption and demand. To increase the availability of biodiesel in the market, new methods of biodiesel production must be developed to take advantage of the plentiful low quality waste derived feed stocks that currently present problems to biodiesel production using conventional methods. This research presents one new approach based upon using heterogeneous highly cross linked mixed bed solid phase catalysts to facilitate the production of biodiesel from feed stocks with high concentrations of free fatty acids (FFA). The performance of the heterogeneous mixed bed catalysts method developed in this research was evaluated and optimized for catalyst concentration and reaction duration while the mixing rate, reaction temperature, initial FFA composition of the feed stock and the alcohol-to-oil molar ratio were kept constant. The presented method reduces the FFA content of the starting feed stock while limiting the release of water into the reaction. Through experimentation, it was found that FFA removal with the mixed bed resin is due to ion exchange with the quaternary ammonium functional group and not catalysis to form esters. A model describing the heterogeneous processing method is presented. The outcome of this research is the development of a new processing method that can be used to create biodiesel from poor quality raw feed stock materials.

Decreasing fossil fuel reserves and environmental concerns necessitate a shift toward biofuels. However, the chemistry of many biomass to fuel conversion pathways remains to be thoroughly studied. The future of biorefineries thus depends on developing new pathways while optimizing existing ones. Here, potential chemicals are added to create a superstructure, then an algorithm is run to enumerate every feasible reaction stoichiometry through a mixed integer linear program (MILP). An optimal chemical reaction pathway, taking into account thermodynamic, safety, and economic constraints is then found through reaction network flux analysis (RNFA). The RNFA is first formulated as a linear programming problem (LP) and later recast as an MILP in order to solve multiple alternate optima through integer cuts. A graphical method is also developed in order to show a shortcut method based on thermodynamics as opposed to the reaction stoichiometry enumeration and RNFA methods. A hypothetical case study, based on the conversion of woody biomass to liquid fuels, is presented at the end of the work along with a more detailed look at the glucose and xylose to 2-mthyltetrahydrofuran (MTHF) biofuel production pathway.

As the global demand for energy rises, there are significant efforts to find alternative energy sources. In the United States (US), these efforts are primarily motivated by a desire to increase energy security and reduce the potential impacts on climate change caused by carbon dioxide emissions from the burning of fossil fuels. Biofuels are considered a potential partial solution, which are being encouraged through public policy. Cellulosic ethanol is a biofuel that is required in increasing amounts over time as part of the Renewable Fuel Standards. Thus, researchers are exploring the environmental impacts of using this biofuel on a large scale. This dissertation research performed an environmental evaluation using the Life Cycle Assessment technique on Bioenergy Sorghum, a crop which was specifically produced as an energy crop, used in a conversion process (MixAlco version 1) that can produce cellulosic ethanol. Results indicate that the conversion process is highly optimized with minimal environmental concerns. Analysis of the crop production, however, demonstrate that further investigation is warranted regarding the depletion of natural resources and emissions from the fertilizers and pesticides/herbicides, due to large scale production of energy crops. A new policy is proposed to support the sustainable, environmentally responsible development of cellulosic ethanol in the US.

This thesis describes a geographically- and temporally-resolved, integrated biological and engineering model that estimates algal biomass and lipid production under resource-limited conditions with hourly and county resolution. Four primary resources are considered in this model: sunlight, carbon dioxide, water, and land. The variation in quantity and distribution of these resources affects algae growth, and is integrated into the analysis using a Monod model of algae growth, solar insolation data, and published values for water, carbon dioxide, and land availability. Finally, lipid production is calculated by assuming oil content based on dry weight of the biomass. The model accommodates a range of growth and production scenarios, including water recycling, co-location with wastewater treatment plants and coal-fired generators, and photobioreactor type (open pond or tubular), among others. Results for every county in Texas indicate that between 86 million and 2.2 billion gallons of lipids per year can be produced statewide for the various growth scenarios. The analysis suggests that algal biomass and lipid production does indeed vary geographically and temporally across Texas. Overall, most counties are water-limited for algae production, not sunlight or carbon dioxide-limited. However, there are many nuances in biomass and lipid production by county. Counties in west Texas are typically not solar- or land-limited, but are constrained by either water or carbon dioxide resources. Consequently, counties in east Texas are limited by either water, or land (depending on the fraction of water recycling). Varying carbon dioxide concentration results in higher growth rates, but not always increased biomass and lipid production because of limitations of other resources in each county. / text