Rising demands for both food and energy have lifted up the idea of producing renewable biofuels from bioenergy grasses that can companion with associative N2ixing bacteria. Associative N2ixing bacteria can partially fulfill the N requirements of bioenergy grasses enabling successful application for marginal lands. The overall objective of this study was to measure the potential N2ixation of bacteria associated with three bioenergy grasses, giant miscanthus (Miscanthus x giganteus), switchgrass (Panicum virgatum; ‘Alamo’) and energycane (hybrid of Saccharum spontaneum and S. officinarum; HO 02-147) as well as identifying N2ixing bacteria associated with these energy grasses and applying these isolates to grasses to increase biomass yield. It was hypothesized that three different energy grasses, giant miscanthus, switchgrass, and energycane have different capabilities to support the growth of associative N2ixing bacteria, and thus exhibit different rate and quantities of N2ixation. The dynamics of delta15Nair in the root-zones of three perennial bioenergy grasses, biomass yield and nitrogen derived from (%Ndfa) were obtained. Greater potential for N2ixation was identified in energycane associated plant system compared to switchgrass and giant miscanthus. Sorghum bicolor (M81-E) was used to calculate the %Ndfa. Moreover, a 15N2 enriched greenhouse study was carried out to estimate and compare the contribution of ANF to support field data. Greenhouse data further confirmed that energycane associated bacteria has greater potential N2ixation compared to giant miscanthus and switchgrass. Composition and diversity of N2ixing bacteria associated as endophytes and in the rhizosphere community of these three energy grasses were identified in marginal lands. Diverse N2ixing bacteria were observed to be associated with different grasses and Azospirillum sp. was identified in energycane. The effect of N2ixing bacteria on biomass was studied by inoculating three energy grasses with the bacteria isolated from energycane. Increased root lengths of giant miscanthus were observed upon inoculation. However, no other changes in biomass yield or shoot lengths were observed in three bioenergy grasses. Further research is needed to quantify endophytic and rhizosphere N2ixing bacteria and to determine their role in association in an effort to increase the biomass yield of other grasses and reduce N fertilizer inputs.

Biorefineries for production of fuels, chemicals, or materials, can bean important contribution to reach a fossil-free economy. Large-scaleforest-based biorefineries are not yet cost competitive with their fossil counterparts and it is important to identify biorefinery supply chain configurations with good economic, CO2, and biomass performance if biorefineries are to be a viable alternative to the fossil refineries. Several factors influence the performance of biorefinery supply chains,e.g. type of conversion process, geographical localisation, and produc-tion capacity. These aspects needs to be analysed in conjunction to identify biorefineries with good supply chain performance. There ares everal approaches to improve the performance of biorefineries, wheree.g. integration with other industries can improve the economic perfor-mance by utilisation of excess heat and by-products. From a Swedish perspective the traditional forest industry is of interest as potential host industries, due to factors such as by-product availability, opportunity for heat integration, proximity to other biomass resources, and their experience in operating large-scale biomass supply chains. The objectives of this work were to investigate how different supply chain configurations influence the economic, biomass, and CO2 perfor-mance of thermochemical biorefineries integrated with forest industries,as well as methods for evaluating those supply chains. This work shows that there is an economic benefit for integration with the traditional forest industry for thermochemical biorefineries.This is especially true when the biorefinery concept can replace cur-rent old industrial equipment on site which can significantly improvethe economic performance of the biorefinery, highlighting the role the Swedish forest industry could play to reach a cost efficient large-scale implementation of lignocellulosic biorefineries. The cost for biomass is a large contributor to the total cost of biore-fineries and for traditional techno-economic evaluations, the biomass prices are considered as static variables. A large-scale biorefinery will likely have an impact on the biomass market, which could lead to both changes in the biomass price, as well as changed biomass demand for other industries. A framework where this is accounted for was intro-duced, combining a techno-economic perspective for evaluating the sup-ply chain performance, with a market model which identifies changes in biomass price and allocation due to the increased biomass competition. The biorefinery performance can be determined from several per-spectives and system boundaries, both from a plant-level and a national perspective. To facilitate a large-scale introduction of biorefineries and  maximise the benefit from their implementations, there is a need to identify biorefinery concepts with high performance considering severa system boundaries, which has been explored in this work.

Interest in using woody biomass from forestlands for energy production has reemerged in recent years. In Kentucky, bioenergy has great potential to help reduce reliance on fossil fuels. However, questions still remain about economic and other social effects associated with forest-based bioenergy production. This study investigates some of the economic implications of harvesting woody biomass for bioenergy production alongside traditional forest products in Kentucky. Results show that forest-based bioenergy can increase financial return to nonindustrial private forest owners. This study also investigates social impacts and drivers of forest-based bioenergy in Kentucky. Results indicate that a variety of issues will have to be dealt with in order for bioenergy production from forestlands to be viable in Kentucky.

Growing concerns about climate change and the inevitable depletion of fossil fuel resources have led to an increased focus on renewable energy technologies and reducing GHG emissions. Limiting the atmospheric level of CO2 is essential to prevent the most damaging effects of climate change. Among renewable energy resources, biomass combustion has the largest potential to contribute to global energy demands, however it is considered to be a carbon neutral solution and so only limits CO2 concentrations rather than reducing them. Through pyrolysis rather than combustion, biomass can lead to carbon negative liquid, gaseous and solid fuels while also offering a route for long term carbon storage in the form of biochar. Biochar is a carbonaceous material which has shown potential for improving soil fertility, reducing GHG emissions and most importantly long term C storage in the environment. However many questions still remain unanswered with regard to biochar, especially the influence that process conditions can have on its performance in soil as well as any potential trade-offs between soil amendment, C sequestration and heat/power generation. This thesis is therefore focused on assessing the influence that process conditions and feedstock selection have on biochar properties related to carbon stabilisation, improving soil fertility (functional properties) as well as the distribution of energy amongst the pyrolysis co-products. To achieve this, a systematic set of biochar samples was produced, using a wide range of pyrolysis parameters (highest treatment temperature (HTT), heating rate, residence time, carrier gas flow rate and feedstock type), and analysed for physicochemical and functional properties. Pyrolysis HTT consistently showed a dominant influence on determining the final yields and properties of biochar, while the effect of other production parameters was varied. In this thesis the candidate first studied the effect that process conditions had on the long term stability of biochar, as an important indicator of its ability to sequester carbon. While increasing the HTT resulted in a decrease in biochar yield, overall the yield of stable-C increased with temperature. This meant that by applying a higher HTT during pyrolysis a higher C sequestration potential for biochar was achieved. Next to be examined was the influence that process conditions had on other functional properties (labile-C yield, biochar pH, extractable nutrients and cation exchange capacity (CEC)) was then examined. The labile-C yield of biochar decreased with increasing HTT due to the release of volatile matter, while the CEC and concentration of extractable nutrients tended to be higher in biochar produced at 450oC rather than greater HTTs. Biochar pH was also highly alkaline at elevated HTT. This indicated that while high HTT favoured C sequestration and biochar pH, lower HTT may be more favourable for other functional properties. Furthermore by assessing the mass and energy distribution amongst the solid, liquid and gaseous fractions, it was possible to determine the energy balance of the process and through this evaluate the trade-off between the C sequestration potential of biochar and the energy output of the liquid and gas fractions. As the severity of pyrolysis was raised, the total energy stored within the liquid and gaseous co-products increased at the expense of the energy content of biochar, therefore increasing the available energy output of the system and reducing the energy lost when using biochar for carbon storage rather than for bioenergy. This also demonstrated that the pyrolysis process could be fine-tuned to increase the amount of stored C while also improving the heat/power generation of the system. The higher energy content of the gas stream at elevated HTT was also seen to contain sufficient energy to sustain the pyrolysis process, which would free up the solid and liquid fractions for higher value applications while reducing the necessity for external fuel sources. Finally, the data set was used to produce statistical models enabling the prediction of biochar stable-C yield as well as the heating value of biochar. The results of this thesis therefore demonstrate that through applying high HTT the potential energy output of the pyrolysis system can be increased while producing a biochar product with high C sequestration potential and positive functional properties for soil amendment. Due to potential trade-offs, the final choice of process conditions and feedstock would then be made based on the specific requirements of a selected site for biochar application. Understanding the influence that production conditions have on the functional properties of biochar as well as the energy balance of the system is critical to developing specifically engineered bespoke biochar, be it for agricultural use, carbon storage, energy generation or combinations of the three.

Growing concerns regarding the impact of fossil fuel use upon the environment and the cost of production have led to a growth in the interest of obtaining energy from biomass. 1st and 2nd generation biomass types, however, are often criticised for their high energy requirements and environmental impacts. Algal biomass is considered a 3rd generation biomass which does not require arable land for cultivation, typically has a high productivity and can be converted to a wide variety of energy carriers. Despite research on the concept of producing energy from algal biomass dating back to the 1960s there has been limited commercial development and the environmental advantages are still in doubt. This thesis investigated the potential of algal biomass as a source of bioenergy feedstock by considering the cultivation and processing of localised species of algae and applying life cycle assessment (LCA) methodology to algal biofuel production systems. Experiments were conducted to examine the productivity of a wild algal species in wastewater and the potential recoverable bioenergy yields. The LCA studies drew together data from external studies, commercial databases, industrial reports and experimental work to assess the environmental impacts and the energy balance for each system considered. The thesis investigated the generation of biofuel from both freshwater algal biomass and marine algal biomass. For both cases, the current state of research was examined and the gaps determined. Existing studies suggest the high intensity of microalgal biomass production (fertiliser requirements, high energy harvesting) greatly reduces the overall sustainability. Part of this thesis therefore investigated the possibility of a low input system of microalgal cultivation. A recommended approach was suggested using local species cultivated in wastewater as the nutrient source and a conversion strategy based on the characteristics of the dominant species. The practicality and effectiveness of cultivating and processing locally grown algal biomass under low input conditions was determined by experiments that were conducted in the laboratory. Algal biomass was collected locally and cultivated in the laboratory using agricultural effluent as the nutrient source. The productivity of the algae was monitored alongside the uptake of nutrients. The effluent provided a good media for the cultivation of the wild algae and the nitrogen and phosphorous loading of the effluent was reduced by as much as 98% for NH4+ and 90% for PO4³-. The algal biomass was also tested for its potential as a feedstock for bioethanol production as well as biochar alongside pyrolysis oils and gases. Compared to alternative biomass types tested, the algal biomass appeared to be a good candidate for bioethanol production providing a 38% recovery of bioethanol. The biomass appeared a less favourable substrate for energy recovery from pyrolysis but this process could be considered for carbon biofixation. The sustainability of incorporating microalgal cultivation in wastewater treatment was tested by conducting a life cycle assessment of a large scale system. The life cycle assessment used Haifa wastewater treatment plant in Israel as a case study. The study compared algal cultivation with energy recovery to conventional nutrient removal (A2O process) for enhanced nutrient removal within the wastewater treatment plant. It was found that the use of algal ponds for nutrient removal compared favourably to conventional treatment under specific conditions. These conditions were: the algal biomass is converted to both biodiesel and biogas and the algal biomass is converted to biodiesel, bioethanol and biogas. In these cases the energy balance was greater and the global warming potential and eutrophication potential were less. The conventional nutrient removal was, however, found to be the better method in terms of the acidification potential. Despite being the favourable method of nutrient removal the cultivation and processing of algae relies upon several key assumptions: high year round growth of algae, no contamination and access to a high land area for the cultivation ponds. The sustainability of recovering bioenergy from the cultivation of macroalgae was also tested. A life cycle assessment was conducted investigating the energy return on investment and six environmental impacts for three cultivation methods and three process streams to convert the biomass to bioenergy. Cultivation and processing in Chile was used as a case study due to the depth of knowledge and availability of data. The cultivation scenarios were: bottom cultivation of Gracilaria chilensis, the long line cultivation of Gracilaria chilensis and the long line cultivation of Macrocystis pyrifera. The processing streams were: bioethanol, biogas and both bioethanol and biogas. Most of the data used in the life cycle assessment was obtained from studies conducted in Chile and from communication with local fisherman. It was found that the bottom cultivation of Gracilaria chilensis and conversion to bioethanol and biogas produced the best energy return on investment (2.95) and was most beneficial in terms of the environmental impacts considered. Alternative circumstances were also considered which included new research (untested on a large scale) related to the value used for productivity and conversion of the biomass. This analysis indicated that an EROI of 10.3 could be achieved for the long-line cultivation of Macrocystis pyrifera and conversion to bioethanol and biogas alongside very limited environmental impacts. This result relies, however, upon favourable assumptions that have not yet been proven on a large scale. The work conducted in this thesis highlights the potential of recovering energy from algal biomass. The experimental work and life cycle analysis of freshwater algal cultivation demonstrates the importance of using wastewater treatment as added value to the system. Maximising energy recovery by using a combination of conversion techniques was also shown to be key in providing the most sustainable solution. The sustainability of energy produced from macroalgae was established as being preferable to several conventional energy sources. Innovative methods to improve the system were also shown to greatly enhance the concept.

Master of Agribusiness / Department of Agricultural Economics / Jason Bergtold / Markets for alternative fuels are emerging and are of great interest to both public and private companies, as well as government agencies looking to differentiate fuel sources to achieve improved and sustainable operational efficiencies. This creates a growing need for innovation and an increased supply of biofuel feedstocks for bioenergy options such as bio-jet fuel. This thesis aims to assess the logistical feasibility of producing oilseed bio feedstocks and the practicality of building new crush facilities specifically for bio-jet fuel production in Kansas. A logistical optimization model is built by applying data to estimate the potential Kansas supply of rapeseed as a possible feedstock option; transportation and facility costs associated with building; and proposed crushing facility sites, by considering the estimated demand for bio-jet fuel within Kansas. The developed optimization model determined that even average yields per acre and modest adoption rates by farmers willing to incorporate rapeseed into their crop rotations could provide enough feedstock to supply one or two crushing facilities, depending on a variety of additional factors, including bio-jet fuel demand in Kansas. Sensitivity analysis was performed on key model factors and determined that the most influential factor on both size and number of proposed crushing facilities was the market demand for bio-jet fuel. Ultimately, further research is required to better understand the actual market demand for bio-jet fuel within Kansas and how competition or supply supplementation of other bio feedstocks can affect the size or number of proposed crushing facilities. There are currently six oilseed crushing facilities operating in Kansas; although all are dedicated to soybean or sunflower seed. Further studies may find these sites as viable alternative options to building new crushing facilities for a separate type of feedstock.

The modernized world is over-consuming low-cost energy sources that strongly contributes to pollution and environmental stress. As a consequence, the interest for environmentally friendly alternatives has increased immensely. One such alternative is the use of solar energy and water as a raw material to produce biohydrogen through the process of photosynthetic water splitting. In this work, the relation between H2-production and photosynthesis in the green algae Chlamydomonas reinhardtii was studied with respect to three main aspects: the establishment of prolonged H2-production, the involvement of PSII in H2-production and the electron pathways associated with PSII during H2-production. For the first time, this work reveals that PSII plays a crucial role throughout the H2-producing phase in sulfur deprived C. reinhardtii. It further reveals that a wave-like fluorescence decay kinetic, before only seen in cyanobacteria, is observable during the H2-producing phase in sulfur deprived C. reinhardtii, reflecting the presence of cyclic electron flows also in green algae.

>Magister Scientiae - MSc / The yeast Saccharomyces cerevisiae is considered a good host used for heterologous protein production due to the organism’s microbial safety, rapid growth and eukaryotic post- translational processing. As a fermentative organism, S. cerevisiae is thus not only a useful platform for the production of biopharmaceuticals and industrial enzymes, but also a promising organism for second-generation biofuel production. Substantial effort has been focused on alleviating the many bottlenecks in recombinant gene expression, as well as in the secretory pathway to enhance heterologous protein titres. It was recently shown that highly active endocytosis could decrease the overall secreted protein titre in the supernatant. In this study, we aimed to block endocytotic and vacuolar complexes to ultimately disrupt, or impair, the endocytotic and vacuolar mechanisms of proteolysis and test the effect that this would have on secreted heterologous protein titres. This was accomplished by knocking out various genes involved in endocytosis and transforming the strains with genes encoding various hydrolases including β-glucosidase (Bgl), xylanase (Xyn2), endoglucanase (Eg2) and cellobiohydrylase (Cbh1). Our study demonstrated that genetic blocking of endocytotic mechanisms as well as vacuolar complexes could theoretically improve heterologous protein secretion in S. cerevisiae. Endoglucanase (Eg2) titres displayed improvement of 26% and 30% in strains which had the RVS161 and VRP1 genes deleted and xylanase titres displayed an improvement of 71% and 143% in strains with the END3 and SSA4 gene deletions. Several of the gene knockouts tested improved Xyn2 and Eg2 titres but the effect of the different gene targets varied widely. A double knock-out strain with deletions in CLC1 and RVS161 secreted 104% more Eg2 than its parental control strain on a per dry cell weight basis, a significant synergistic improvement. Other double knock-out strains displayed additive or similar activities when compared to their controls. Cbh1 secretion could not be improved through the gene deletions tested in our study and Bgl activity could not be measured in our transformants. These results demonstrate the different relationships of various heterologous proteins with various components of the secretion machinery and may also imply how endocytosis as well as vacuolar complexes affect the level of secreted protein.

Energy use and the environment are inextricably linked and form a key role in concerns over sustainability. All methods of energy production involve resource uncertainties and environmental impacts. A clear example of this is the use of fossil fuels which present three main problems, being: finite resources; significant contribution to environmental pollution; and reliance on imports. Hence there is a clear need to reduce the use of fossil fuels for energy. Bioenergy has the potential to both displace fossil fuels, and reduce the effect of climate change by sequestering carbon dioxide during the production of biomass. It is also possible that bioenergy can reduce the UK’s dependence on energy imports and boost the rural economy. This thesis provides an interdisciplinary assessment of bioenergy production in the UK. Due to the complexities of bioenergy systems several appraisal methods have been used. An initial study examined the barriers to and drivers for UK bioenergy development as a whole. It was found that for projects to be successful, bioenergy schemes need to be both economically attractive and environmentally sustainable. A biomass resource assessment was then completed using the South West of England as a case study. This demonstrates that bioenergy can make a useful contribution to the UK’s energy supply, due to the diverse range of biomass feedstocks currently available. However a range of barriers and constraints will need to be overcome if the UK is to reach its bioenergy potential. To assess the potential environmental impacts of bioenergy production different case studies were selected. Life cycle assessment is widely regarded as one of the best methodologies for the evaluation of burdens associated with bioenergy production. This was applied, alongside net energy analysis, to a small-scale biomass gasification plant which uses wood waste as a feedstock. As an alternative biomass source, the perennial energy crops Miscanthus and Willow were also assessed. Several different scenarios of biomass cultivation, transportation, and energy conversion were then compared, to assess the potential environmental impacts. Biomass gasification offers good potential for reducing fossil fuel use and climate change impacts. Nonetheless embodied energy in the construction phase can be high and other impacts such as particulate emissions, ecotoxicity and land use can be important. Therefore environmental benefits are maximised when both electricity and heat are utilised together, and when waste is used as feedstock. The ultimate applicability of biomass gasification is restricted by the quantity of feedstocks that can be made available for conversion. Perennial energy crops offer several advantages over annual crops including more positive energy balances and reduced agro-chemical inputs. However their cultivation needs to be carefully sited to avoid issues of land use change and the displacement of food crops. This study shows that each bioenergy production pathway needs to be assessed using a range of appraisal techniques, which include: biomass resource assessment, technical and economic feasibility, life cycle assessment and net energy analysis. It concludes that biomass gasification CHP offers an alternative to fossil fuel generation but more technical knowledge is required in the UK if it is to become widely used for biomass energy.

More deaths are caused every year by indoor air pollution than malaria, HIV/AIDS and tuberculosis combined. Cooking with traditional fuels such as charcoal and fuelwood with poor ventilation causes the single most important environmental health risk factor worldwide. It also contributes to environmental issues such as deforestation as traditional biomass fuels and cooking stoves are inefficient and requires large quantities of wood. This is especially critical in Africa where the largest regional population growth in the world is expected to occur. A solution to these issues was realized through fuel pellets and modern cooking stoves by Emerging Cooking Solutions, a company started by two Swedes and based in Zambia. The production of fuel pellets in Zambia is dependent on pine sawdust from small sawmills and is a declining source of raw material. However, other sources of biomass are available in Zambia such as pigeon pea stalk, an agricultural waste product, and sicklebush, an invasive tree species. If these species are viable for pelletization, the production of pellets can increase while reducing issues with sicklebush and promoting cultivation of pigeon pea. The aim of this work is to evaluate if pigeon pea stalk and sicklebush are viable to pelletize in Zambia and how the press is affected by the different raw materials. A pilot study is done at Karlstad University with a single unit press, hardness tester and soxhlet extractor to evaluate how the material constituents correlate to friction in the press channel and hardness of the pellets. The results of the pilot study provide support for full scale tests done in a pellet plant in Zambia. The normal production of pellets from pine sawdust is used as quality and production reference for the tests with pigeon pea stalk, sicklebush, and different mixes of the raw materials. The properties used to evaluate the quality of the pellets are hardness, durability, moisture content, bulk density, and fines. The press configuration is evaluated by logging the electricity consumption by the press motor, calculating the power and specific energy consumption from the logs, and observations during the tests. The results show that sicklebush, and mixes of sicklebush with pigeon pea stalk can produce pellets with better quality than the reference pine pellets. An interesting composition is a mix of 80% pigeon pea and 20% sicklebush that produces pellets with the best quality of all the tests. However, pellets produced from sicklebush and pigeon pea show a larger variation in hardness as compared to the reference pellets from pine sawdust. Mixing pigeon pea with pine reduces these variations but reduces the hardness of the pellets below the reference. The press struggles to process sicklebush and pigeon pea stalk with fluctuating power consumption that causes the motor to trip. The inhomogeneity of the materials in sicklebush and pigeon pea are identified to cause the issues in the press. Production improvements are discussed to facilitate the production of pigeon pea stalk and sicklebush pellets.