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There is increasing concern regarding alternative, sustainable energy sources, such as biofuels, to replace declining oil reserves. The abundance of lignocellulosic biomass makes it the only imaginable resource that can potentially substitute a substantial portion of the fossil fuels we use today, but current methods for producing biofuels from non-food crops are cost intensive and not economically viable. Synthetic biology provides several potential approaches for developing biologically mediated processes for the conversion of lignocellulosic biomass into biofuels. Such systems are based on engineered microbes that produce enzymes for catalysing the conversion of cellulose into fermentable sugars and subsequently into high value products. Effective degradation of cellulose requires multiple classes of enzyme working together. In naturally occurring cellulose degrading microbes, bioconversion is catalysed by a battery of enzymes with different catalytic properties. However, naturally occurring cellulases with multiple catalytic domains seem to be rather rare in known cellulose-degrading organisms. Using synthetic biology approaches, seven cellulases with multiple catalytic domains were engineered and tested to determine the usefulness of such chimeric enzymes to replace cloning of multiple enzymes for biomass conversion. Catalytic domains were taken from Cellulomonas fimi endoglucanases CenA, CenB and CenD, exoglucanase Cex, and β-glucosidase, Cfbglu as well as Cytophaga hutchinsonii cellodextrinase CHU2268. All fusions retained both catalytic activities of the parental enzymes. To investigate the benefits of fusion, Citrobacter freundii NCIMB11490 was transformed with either fused or non-fused enzymes and cultured with cellulose blotting papers as main carbon source. Cells expressing fusions of Cex with CenA or CenD reproducibly showed higher growth than cells expressing non-fused versions, as well as more rapid physical destruction of paper. The opposite was observed for the other combinations. Comparing two different Cex and CenA fusions, CxnA2, which contains two carbohydrate binding modules (CBMs), degraded filter paper faster and led to better growth than CxnA1, which contains only one CBM. It was observed that CxnA1 was exported to the supernatant of E. coli and C. freundii cultures, as also seen for Cex and CenA, although there is no clear biological mechanism for this. Monitoring of growth using colony counts is laborious, but the use of optical density is not possible for cellulose-based cultures as it is affected by the insoluble cellulose particles. The SYBR Green I/propidium iodide live/dead staining protocol was therefore evaluated for growth measurements and was found to allow rapid measurements of large numbers of samples. In conclusion, these studies have demonstrated a simple and useful method for making chimeric proteins from libraries of multiple parts. The results demonstrate that use of fusion proteins can improve biomass conversion in vivo, and could potentially reduce the necessity for cloning of multiple enzymes and improve product yields. A simple and effective method for monitoring growth of bacteria in turbid cultures using a fluorimeter has also been developed.

Over-exploitation of fossil fuels coupled with increasing pressure to reduce carbon emissions are prompting a transition from conventional petrochemical feedstocks to sustainable and renewable sourced carbon. The use of lignocellulosic biomass as a feedstock for integrated biorefining is of current high interest, as separation into its component parts affords process streams of cellulose, hemicellulose and lignin, each of which can serve as a starting point for the production of biobased chemicals and fuels. Given the large number of potential sources of lignocellulosic feedstocks, the biorefinery will need to adapt to the supplies available over a normal growing season. Of particular importance is the lignin fraction, as its conversion to chemicals and materials to allow economic viability of the operation. Previous work has demonstrated that organosolv fractionation effectively separates lignocellulosic biomass into its component parts. In this project, we investigated the use of organosolv technology for separating mixtures of lignocellulosic feedstocks to isolate pure lignin. Mixtures of switchgrass (Panicum virgatum), southern yellow pine (Pinus taeda L.), and hybrid poplar (Populus spp.) were separated using organosolv fractionation. Experiments were performed by heating the feedstock mixtures at 150oC in a 3.5 L flow-through reactor with a ternary, one-phase solvent mixture of methyl isobutylketone (MIBK), ethanol (EtOH) and water (H2O) in a wt% ratio of 16/34/50, and containing sulfuric acid as a catalyst. The impact of different process variables was examined by experimental design (‘Design of Experiments’) to minimize the number of experimental runs using a balanced approach in the response surface to maximize inference. The process variables included two different runtimes (60, 120 min), two different sulfuric acid levels (0.05, 0.15 M), and four different wt% feedstock ratios for switchgrass/pine/poplar ([10/10/80], [10/80/10], [80/10/10], [33/33/33]). After completion of the initial experimental matrix, four additional center-points were carried out using a 90 min runtime, and 0.1 M acid level to validate the results for each of the four feedstock ratios. The dependent factors were lignin yield, lignin purity, and cellulose purity. Response surface methodology (RSM) was used to evaluate the impact of the process variables and to determine optimization settings for the process.

I started my microbiological research when I embarked on a PhD at Essex University England in 1978-1981 mainly on microbial interactions in saltmarsh sediments and environment. I continued for a couple of years work as a postdoctoral research officer for two years 1981-1982 on the same topic. I joined the Kuwait Institute for Scientific Research in 1983 and immediately diverted to interest in microbial biotechnology, single cell proteins, wastewater, treatment, microbial bioactives and biosurfactant production and applications. I continued these lines of research and initiated these areas when I joined Ulster University for a short period 1990-1991 and then as an honorary researcher 1992-1996 and finally taking a fulltime position in 1997 which is continued to date. I was involved in the initiation of several areas of research at UU among which the main and most prolific was on biosurfactant and related research and applications. I have been working with biosurfactants for 35 years as I realized the significance of this research field and its future potentials which compelled me to persevere in this area while establishing an international network of collaborators and scientists interested in working on biosurfactants. The main question I had was figuring the reason why some microorganisms produce biosurfactants? The answer has not been simple or straight forward, as these compounds appear to have several functions linked to the existence of these microorganisms. One of the apparent functions in microorganism that use hydrocarbon substrates for growth or to exist in oily substrates is to make these substrates available for them to metabolise. Other roles include motility in viscous environments or for the purpose of controlling the quorum sensing mechanisms of cells, which adjust gene expressions depending on cell density or on their surrounding environment. Evidence of biosurfactants playing an important role in biofilm formation and maintenance and disruption of mature biofilms integrity, to sustain channels for nutrient and gaseous exchanges also exist. All the above-hypothesised functions of biosurfactants have been put forward for some bacteria. The reasons why several yeasts, however, produce large quantities of biosurfactants remain not fully explained. One explanation for yeasts is the type of environment they grow in which may have waxes or high molecular weight hydrophobic molecules. Interest in biosurfactants mainly emerged during the eighties and nineties and reached a critical stage when commercial exploitation started by international companies which aimed to explore using them as ingredients in some of their products rather than using chemical surfactants to meet their sustainability initiative and green programs. Interest started mounting rapidly as and their beneficial outcomes reached a stage to be dependent on successfully tailoring these compounds for specific applications and exploring production at a price that makes them attractive alternatives to chemical surfactants. Many issues remain to be investigated before largescale exploitation for many biosurfactants becomes possible. These include cost of production, achievable yields, and safety issues for some of the producing strains. Many biosurfactants have already been used as ingredients in several commercial products. I have always believed that there are no major impediments to the use of biosurfactants in a wide range of products and applications which includes benefits to the environmental, uses in food, biofuel, biomedical, pharmaceutical, and sustainability issues. Some are already slowly happening and expected to increase significantly as the drive towards enhancing the green credentials and sustainability of many product become essential to expand and target specific sectors. This commercial interest in biosurfactants has been reflected by the number and diversity of Industrial Companies and European Union-funded research projects that I have become involved with and that have major roles in biosurfactants research. These include, projects funded by Unilever Company (5 partners) to use them in their cleaning and sanitary products, the EU FP7 project KILL.SPILL (32 partners), which examined the remediation of marine oil spills, the EU FP7 BIOSURFING (10 partners) project, which focuses on the production of 'new to nature' sophorolipids with applications in the food and pharmaceutical sectors; and the Horizon 2020 MARISURF project (12 partners), which pioneered the search for new biosurfactants from marine bacteria for a range of potential applications. All these projects have many industrial partners supporting and exploring possibilities of using these compounds in their products or services. The contribution of my research and published work on biosurfactants, bioactives and microbial biotechnology in general had advanced the knowledge and interest and met many research and industrial needs. Perhaps the most significant of my achievement was in collaborating with Unilever which commenced in 2006 supported by funding from the then Department of Trade and Industry (DTI) for a multi partner project of total value £1.9 million that aimed to design and manufacture sustainable products. These collaborations extended the scope of biosurfactant application to included 2 CAST studentships, Post-doctoral visiting researchers and a second project funding through the Technology Strategy Board (TSB) in 2012 for a second major multiple partner project of around £1.0 million. These collaborations resulted in Unilever filing 9 patent applications and us publishing 10 collaborative scientific papers in international journal in collaboration with Oxford University. The results of this collaborations and drive into this area has culminated in Unilever partnering with Evoniks (A German speciality chemicals company with a biosurfactant production facility) and launching their green cleaning ingredients. This innovative ingredient was recently introduced to the market in the summer of 2019 as Unilever's 'Quix' dishwashing brand in Chile which was the first time a rhamnolipid biosurfactant was used in a household cleaning product anywhere in the world. The whole journey of this research work has produced several doctoral and postdoctoral researchers all working locally and within the international arena. I am gratified that I have become one of the prominent scientists in the field and was invited in Sept. 2019 to give the opening keynote plenary lecture entitled "Microbial Biosurfactant; State of the Art" in the first ever conference on Biosurfactants organized by the University of Hohenheim in Stuttgart, Germany. I also am scheduled for the Keynote address for the 2nd Biosurfactant conference in Stuttgart-Hohenheim in September 2022 with the title "A Journey into Biosurfactants Research and Development for Over Three Decades".

The act of decision making involves a choice amongst tradeoffs. In agriculture this is no different. This dissertation is composed of four papers that examine the tradeoffs being made across different agricultural decision making processes. The first two papers examine the tradeoffs made at the individual producer level while the last two papers examine tradeoffs made at the national policy level. The first paper investigates farmer attitudes towards how a hypothetical set of production practices referred to as "]conservation agriculture" will affect yield, labor use, erosion, and cost in two communities of Bolivar province, Ecuador. By evaluating the tradeoffs producers are willing to make when choosing to adopt such practices, changes in producer welfare associated with adoption may be identified. These measures can assist in identifying constraints to adoption and aid in extension and policy outreach development. The second paper aims to gain a better understanding of the dynamic relationship between farmers and food buyers. This issue is examined from the perspective of small-scale specialty crop producers who are currently or are considering marketing their products into wholesale food markets. With a focus on farms in Virginia and North Carolina, this study seeks to identify key contract characteristics and buyer attributes which are valued by small-scale specialty crop producers; quantify tradeoffs small-scale specialty crop producers are willing to make between buyer attributes and contract characteristics when establishing a new contractual relationship; and determine the factors influencing these tradeoffs. The third and fourth papers examine the demands that U.S. biofuel production has placed on domestic nutrient fertilizer production. A key argument in favor of domestic biofuel production is that it is a renewable path towards energy independence. However the inputs used in the production of biofuel feedstock, primarily fertilizer nutrients, are anything but renewable. These two papers add to the discussion surrounding biofuel policies by asking an important question that has not received the attention it deserves: "What about the non-renewable inputs (e.g., nutrient fertilizers) that go into producing the inputs (e.g., corn) used for biofuel production?"

The low efficiency of collection, storage and transportation in the switchgrass supply chain has hindered the commercialization of a switchgrass-based biofuel industry, even given its ecological and environmental advantages in carbon sequestrate, soil quality, water use, and pollution pressure. Thus, designing a switchgrass-based supply chain balancing both environmental and economic performance is important to expedite the development of the cellulosic biofuel industry to meet the national energy plan. The objectives of this study are to 1) determine economic cost and multiple environmental outcomes in feedstock supply chains and 2) identify the relation between the economic and environmental performances. The first paper considers three objectives: minimization of economic cost, greenhouse gas (GHG) emissions, and soil erosions. The second paper focuses on the relation between economic cost and abated greywater footprint for industrialized supply of cellulosic biofuel in west Tennessee. The improved augmented epsilon method and compromise solution method were applied to high-resolution spatial data to determine the optimal placement of the feedstock supply chains. Results in the first paper indicated that land change into switchgrass production is crucial to both plant-gate cost and environmental impact of feedstock supply. Converting croplands to switchgrass incurred higher opportunity cost from land use change but stored more soil carbon and generated less soil erosion. Tradeoffs in higher feedstock costs with lower GHG emissions and lower soil erosion on the frontier were captured. Soil erosion was found more cost effective criterion than GHG emission in general. The compromise solution location for the conversion facility generated at 63% increase in feedstock cost but improved the environmental impact in lowering 27 % GHG emission and decreasing soil erosion by 70 times lower in the feedstock supply chain compared with cost minimization location. Results in the second paper showed that tradeoff between feedstock costs and greywater footprint was mainly associated with the changes of land use, while ambient water quality condition was also influential to the selection of feedstock production area. The average imputed cost of lowering grey water footprint in the most preferred feedstock supply chain in west Tennessee was $0.94 m-3 [per cubic meter].

Two seemingly separate areas motivate this work. The first is the water scarcity, salinization, and drainage concerns that cause both environmental and private damages in arid regions throughout the world, including the San Joaquin Valley of California's Central Valley. The second is interest in producing bioenergy in an energetically, privately, and environmentally positive manner. These two branches intersect with the growth of highly energetic biofuel crops on marginal, or poor quality, land with saline drainage water as a form of Integrated Farm Drainage Management (IFDM). To analyze this intersection, this dissertation contains three chapters. The first paper consists of background information on the San Joaquin Valley, marginal land, biofuels, and drainage water, and an arithmetic estimation of potential Bermuda grass (Cynodon dactylon) bioenergy production as an IFDM crop. The second paper develops yield as a function of salinity, irrigation systems, irrigation timing, nitrogen, climate effects, and applied water. The functions, which are very flexible, are compared to other functions used in the literature and the results from a field experiment. These functions are used in the third paper, which develops a farm-level bio-economic optimization model of IFDM crops, including the biofuel crop, Brassica spp. These works show that growing biofuel crops, and other IFDM crops, on marginal lands in drainage-impacted regions can be privately beneficial and provide environmental benefits relative to traditional approaches.

High world food prices after 2006 concerned governments across the globe. Among various contributing factors, biofuels have received wide attention in recent years with more and more studies analyzing the implication of biofuel policies on global food supply and demand conditions. Relying on a model based upon U.S. biofuel policies, this paper studies the price links between crop and energy commodities induced by biofuel production, and simulates commodity price changes due to market and policy shifts. Results suggest price links exist between crop and energy commodities but more accurate measurement of commodity price changes induced by different market conditions will require the construction of a more detailed structural model.

The massive consumption of fossil fuels and associated environmental issues result in an increased interest in alternative resources such as biofuels. The renewable biofuels can be upgraded from bio-oils that are derived from biomass pyrolysis. Catalytic cracking and hydrodeoxygenation (HDO) are two most promising bio-oil upgrading techniques for biofuel production. Heterogeneous catalysts are essential for upgrading bio-oil into hydrocarbon biofuel. Although some progresses have been made, the cost and effectiveness of catalysts still remain challenges. The main objective of this study was to develop efficient heterogeneous catalysts for upgrading bio-oils into advanced hydrocarbon biofuel with low costs. In catalytic cracking, Ni/HZSM-5 and Co-Mo/HZSM-5 were used for upgrading bio-oils derived from prairie cordgrass (PCG) and alkali lignin pyrolysis in a two-stage reactor system. 12%Ni/HZSM-5 catalyst yielded the highest amount of gasoline fraction hydrocarbons (32.45%) for PCG bio-oil upgrading. 4%Mo- 2%Co/HZSM-5 catalyst showed a robust ability in the PCG catalytic cracking with highest yield of hydrocarbons at 41.08%. 1.27% NiO/HZSM-5 catalyst generated the highest biofuel yield at 27.5% and produced biofuel with the highest content of hydrocarbons at 69.4% in alkali lignin bio-oil upgrading. In hydrodeoxygenation (HDO), Ni based activated carbon catalysts including Ni/AC, Ni-Fe/AC, Ni-Mo/AC and Ni-Cu/AC, and Ni-Zn/Al2O3 catalysts with different Ni and/or Zn loading ratios were designed for HDO of PCG and pine sawdust (PSD) bio-oils. The Ni/AC catalysts produced the highest content of gasoline range hydrocarbons (C6-C12) at 32.63% in PCG biofuel product, while Ni-Mo/AC generated biofuel product with the highest content of gasoline blending alkyl-phenols at 38.41%. 15%Ni-5%Zn/Al2O3 catalyst generated the highest PSD biofuel yield at 44.64 wt.% and the highest hydrocarbon content at 50.12% in biofuel product. However, the upgrading cost of HDO is still high due to the high consumption of external hydrogen. In this paper, the hydrogen generated from cheap water using zinc hydrolysis for in-situ bio-oil HDO was firstly reported. The effect of different temperatures (200 °C, 250 °C and 300 °C) on PSD bio-oil HDO over Pd/C catalyst was investigated. The results show that 250 °C yielded biofuel with the highest heating value at 30.17 MJ/kg and the highest hydrocarbons content at 24.09%. In bio-oil HDO using zinc metal only, 400°C bio-oil upgrading process produced biofuel product with highest hydrocarbons content at 68.95%.

Disenchantment with the Washington Consensus has led to an emphasis on growth diagnostics. In the case of Brazil, the literature suggests three main factors impeding growth: low domestic savings, a shortage of skilled workers, and a lack of investment in the country's transportation infrastructure. The unique contribution of this study is to show the inter-temporal implications of relaxing these constraints. We fit a multi-sector Ramsey model to Brazilian data, validate its fit to times data, and provide empirical insights into the economy's structural transformation to long-run equilibrium. Then, the sensitivity of these results to relaxing each of these three constraints is investigated in a manner that yields the same long-run level of wellbeing. Analytical concepts adapted from static trade theory are used to provide a detailed explanation of how the economy responds in transition growth to the relaxation of these impediments. Addressing these factors clearly benefits the economy, but they do not launch the economy to a substantially higher growth path. In order to enhance energy security and independence, Brazil has supported the production and use of ethanol. Brazil's leadership in this market reveals complex inter-linkages between ethanol, sugarcane, sugar and fossil fuels. These sectors have been growing an average of 14% per year, while the country's growth rates have been very modest. This paper presents a theoretical framework for understanding the interaction between Brazil's economic growth and the evolution of these sectors as the economy transitions toward long-run equilibrium. Then, the sensitivity of these results is analyzed under two simulations; first, a reduction of the cost of financial intermediation (which the literature identifies as one of the factors affecting Brazil's growth), and second, an increase in ethanol prices by 2.6%, based on the expectation that biofuels' world demand is increasing.

Lignocellulosic ethanol offers a promising alternative to conventional fossil fuels. One among the major limitations in the lignocellulosic biomass hydrolysis is unavailability of efficient and environmentally biomass degrading technologies. Plantbased production of these enzymes on large scale offers a cost effective solution. Cellulases, hemicellulases including mannanases and other accessory enzymes are required for conversion of lignocellulosic biomass into fermentable sugars. β- mannanase catalyzes endo-hydrolysis of the mannan backbone, a major constituent of woody biomass. In this study, man1 gene encoding β-mannanase was isolated from Trichoderma reesei and expressed via the chloroplast genome. PCR and Southern hybridization analysis confirmed the site-specific transgene integration into the tobacco chloroplast genomes and homoplasmy. Transplastomic plants were fertile and set viable seeds. Germination of seeds showed inheritance of transgenes into the progeny without Mendelian segregation. Expression of the endo-β-mannanase gene for the first time in plants facilitated its characterization for use in enhanced lignocellulosic biomass hydrolysis. Gel diffusion assay for endo-β-mannanase showed the zone of clearance confirming functionality of chloroplast-derived mannanase. Endo-β-mannanase expression levels reached up to 25 units per gram of leaf (fresh weight). Chloroplastderived mannanase had higher temperature stability (40 °C to 70 °C) and wider pH optima (pH 3.0 to 7.0) than E.coli enzyme extracts. Plant crude extracts showed 6-7 fold iv higher enzyme activity than E.coli extracts due to the formation of disulfide bonds in chloroplasts, thereby facilitating their direct utilization in enzyme cocktails without any purification. Chloroplast-derived mannanase when added to the enzyme cocktail containing a combination of different enzymes yielded 20% more glucose equivalents from pinewood than the cocktail without mannanase. Our results demonstrate that chloroplast-derived mannanase is an important component of enzymatic cocktail for woody biomass hydrolysis and should provide a cost-effective solution for its diverse applications in the paper, oil, pharmaceutical, coffee and detergent industries.

The possibility of growing sugar beet Beta vulgaris as an energy crop has received a great deal of attention in New Zealand. This paper concentrates on two aspects of a sugar beet industry. The net foreign earnings of growing 10% of New Zealand's petrol demand. Implications of land use change by sugar beet as it substitutes for traditional agricultural products. The implications of land use change is examined in two ways. Firstly possible physical impacts of growing sugar beet. Secondly possible change in commodity prices as sugar beets substitutes for arable production. This study concludes with an examination of present and future policy toward ethanol from biomass in New Zealand with reference to United States policy measures. A linear programming model was used to evaluate on farm land use changes on cropping and pastoral systems, when sugar beet was grown as a cash crop. The types of land use change were then extrapolated to farm the basis of a national analysis. The aim of this national analysis was to outline the land required and the production that would be lossed to provide data for the net foreign earnings analysis. Net foreign earnings from growing 10% of New Zealand's petrol demand is some $39 million. The implications of land use change included possible physical damage of land resources as well as increased demand for resources such as water and fertilizer. Possible price changes in agricultural commodities when there is a change in land use, remain uncertain. This is an area that requires study. The magnitude of land use change would indicate some increase in commodity prices as a result of growing sugar beet. The results of the model show a 1:1.6 substitution rate of sugar beet to cash cropping land. The utilization of beet residue as stock feed enabled stock numbers to be maintained while growing sugar beet as a cash crop. Optimism regarding a saving in foreign exchange by growing some of New Zealand's fuel requirements, deserves close appraisal. However, planned sources of transport fuels for New Zealand exclude ethanol from biomass as a supplier for some considerable time to come.