Understanding the lignin effect and related structural parameters relevant to the recalcitrance of the plant cell wall and the individual and cooperative effects on enzymatic saccharification are vital for improving current processing and conversion methods for cellulosic biofuels. Data were collected from several pretreatment technologies (Hot-water, organo-solv, lime, lime-oxidant, dilute acid, and dilute acid-oxidant pretreatments) on cellulose ultrastructure, partial delignification followed by dilute acid pretreatment, dilute acid pretreatment of enzymatic isolated lignin, and melt rheology test of organo-solv lignin. Results showed minimal cellulose ultrastructural changes occurred due to lime and lime-oxidant pretreatments, which however especially at short residence time displayed relatively high enzymatic glucose yield. Dilute acid and dilute acid-oxidant pretreatments resulted in the largest increase in cellulose crystallinity, para-crystalline, and cellulose-Iβ allomorph content as well as the largest increase in cellulose microfibril or crystallite size. Organo-solv pretreatment generated the highest glucose yield, which was accompanied by the most significant increase in cellulose microfibril or crystallite size and decrease in relatively lignin contents. Lignin acted as a barrier which restricted cellulose crystallinity increase and cellulose crystallite growth during dilute acid pretreatment, and that partial delignification instead of complete lignin removal during DAP would benefit the increase of sugar yield. Furthermore, a deeper understanding of the structural change of lignin in the absence of cellulose-hemicellulose matrix during dilute acid pretreatment confirmed that delignification had the most beneficial effect in poplar, but for switchgrass was the xylan removal. In addition, investigation on the structural change of organo-solv lignin during melt rheology test indicated that high purity lignin isolated from plant biomass with the lowest S/G (syringyl/guaiacyl) ratios will exhibit superior processing performance characteristics to produce high-quality carbon fibers. These findings can aid both in the development of improved enzymes that contain activities to decompose recalcitrant structures and in the design of various processing conditions that efficiently convert specific biomass feedstock into sugars. They can also help in the design of new chemical modifications on lignin and innovative biosynthesis strategies for producing linear-fiber-forming lignin with high-performance.

<p>One of the great challenges facing society is to convert theglobal energy system to a sustainable process. Currently, 80%of the world´s energy is supplied through the combustionof fossil fuels. Not only are the fossil resources limited, theutilisation also increases the level of greenhouse gases in theatmosphere. The convertion to a sustainable energy system isproblematic since the technology needed to exploit mostnon-fossil energy sources is not yet fully developed, e.g.solar energy. Biofuel is an available renewable energy sourcewhich is already widely used in many countries. If an effectiveswitch-over from fossil fuels to biofuels is to be realised,biofuels must be viewed as a limited resource. Consequently, itis important that the handling, upgrading and utilisationprocesses involving biofuels are efficient so that itspotential can be fully exploited.</p><p>This thesis considers efficient biofuel utilisation andupgrading within the sawmill industry. The goal has been toanalyse not only the technical opportunities for energy savingsin the sawmill industry, but also to analyse the costeffectiveness and environmental impact of studied measures. Theheat demand of the sawmill industry is almost completelycovered by its own by-products; primarily bark, sawdust andwood chips. The increased demand and improved economic value ofwoody biofuels on the market is thus an incentive for thesawmill industry to place more focus on energy issues. Thesawmill industry also has a more or less constant heat loadover the year, which is a beneficial factor for integrationwith district heating networks, biofuel upgrading plants andcombined heat and power plants.</p><p>The conclusion of the study is that a variety of energyproducts such as heat, unrefined biofuel, pellets andelectricity can be efficiently produced in the sawmill industryand sold for profit to external customers. The payback periodsfor the proposed investments are moderate and both theemissions of volatile organic compounds and global CO2 aredecreased. Should the proposed measures be fully implemented atSwedish sawmills, about 2.8 TWh of biofuel could be savedannually, 0.5 TWh of waste heat could be sold as districtheating and 0.8 TWh of green electricity could be produced.Language: English</p><p><b>Keywords:</b>Sawmill industry, energy efficiency, heatrecovery, integration, biofuel, upgrading, district heating,fuel pellets, CHP, VOC, CO2</p>

<p>This thesis investigates, describes and understands the extensive process of technology commercialization. What stages there are, important aspects and implications. It is structured as a case analysis of project CHRISGAS development. CHRISGAS is a Swedish project, based in Värnamo, developing the technique of direct gasification of biomass to fuels.</p><p>The work has its origin in the debate of the imminent climate changes, where society needs to reduce its reliance on fossil fuels. The automotive sector (particularly transport) is significantly reliant. However, current attempts to transition to biofuels have not completely succeeded. New, efficient technologies must be commercialized, and the technology of wood gasification is said to be particularly promising for launching the next generation of biofuels.</p>

Maximizing enzymatic hydrolysis performance can be achieved through the combination of experimental work and modeling. The present work utilizes an enzymatic hydrolysis model based on reaction kinetics, Langmuir adsorption isotherms, and product inhibition of enzymes (β-glucosidase, cellobiohydrolase, and endoglucanase). The model was developed from a 10% w/w corn stover system. Glucose yield sensitivity to changes in parameter values was assessed and linked to biomass and enzyme characteristics. A commercial enzyme cocktail (CEC) was subsequently characterized by FPLC and gel electrophoresis to identify key enzymes/activities, and the CEC was used in the enzymatic hydrolysis of 20% w/w steam-exploded hardwood. The model was applied to experimental data from the enzymatic hydrolysis of the steam-exploded hardwood, which provided characteristic reaction rate and inhibition parameters consistent with cellulose and xylan hydrolysis. These model-based analyses enhanced understanding of hydrolysis at commercially relevant solids loadings, while identifying pathways to improve enzyme cocktails and enhance biomass conversion.

This thesis covers work carried out on algae bioreactors as a tertiary treatment process for wastewater treatment. The process was primarily assessed by the removal of Phosphorus and Nitrogen as an alternative to chemical and bacterial removal. Algal bioreactors would have the added advantage of carbon sequestration and a by-product in the energy rich algal biomass that should be exploited in the existing AD capacity. Laboratory scale bioreactors were run (4.5-30L) using the secondary treated final effluent from the local Loughborough sewage works. In a preliminary series of experiments several different bioreactor designs were tested. These included both batch feed and continuous flow feed configurations. The bioreactors were all agitated to keep the algal cells in suspension. The results demonstrated that the most effective and easy to operate was the batch feed process with the algal biomass by-product harvested by simple gravitational settling. Experiments also compared an artificial light source with natural light in outdoor experiments. Outdoor summer light produced greater growth rates but growth could not be sustained in natural UK winter light. Light intensity is proportional to productivity and algae require a minimum of around 97W/m2 to grow, an overcast winter day (the worst case scenario) was typically around 78W/m2, however this was only available for a few hours per day during Nov-Jan. The process would be better suited to areas of the world that receive year round sunlight. It was shown that phosphorus could be totally removed from wastewater by the algae in less than 24 hours depending on other operating variables. With optimisation and addition of more carbon, a HRT of 10-12 hours was predicted to achieve the EU WFD / UWWTD standard. It was further predicted that the process could be economically and sustainably more attractive than the alternatives for small to medium sized works. Biomass 3 concentrations of between 1-2g/L were found to best achieve these removals and produce the fastest average growth rates of between 125-150mg/L/d. The uptake rates of phosphorus and nitrogen were shown to be dependent on the type of algae present in the bioreactor. Nitrogen removal was shown to be less effective when using filamentous bluegreen algae whilst phosphorus removal was almost completely stopped compared to unicellular green algae that achieved a nitrogen uptake of 5.3mg/L/d and phosphorus uptake of 8mg/L/d. Soluble concentrations of Fe, Ni and Zn were also reduced by 60% in the standard 10 hours HRT. The predominant algae were shown to depend largely on these concentrations of phosphorus and nitrogen, and the strain most suited to that specific nutrient or temperature environment dominated. Nutrient uptake rates were linked to algal growth rates which correlated with the availability of Carbon as CO2. CO2 was shown to be the limiting factor for growth; becoming exhausted within 10 hours and causing the pH to rise to above 10.5. The literature showed this was a common result and the use of CO2 sparging would more than double performance making this process a good candidate for waste CO2 sequestration. Heat generated from combustion or generators with exhaust CO2 would also be ideal to maintain a year round constant temperature of between 20-25°C within the bioreactors. A number of possible uses for the algal biomass generated were examined but currently the most feasible option is wet anaerobic co-digestion. Further economic analysis was recommended on the balance between land area and complementary biomass generation for AD. It was also suggested given the interest as algae as a future fuel source, the process could also be adapted for large scale treatment and algal biomass production in areas of the world where land was available.

The development of enzymatic biosensors and enzymatic biofuel cells (EBFCs) has been a significant area of research for decades. Enzymatic catalysis can provide for specific, reliable sensing of target analytes as well as the continuous generation of power from physiologically present fuels. However, the broad implementation of enzyme-based devices is still limited by low operational/storage stabilities and insufficient power densities. Approaches to improving upon these limitations have focused on the optimization of enzyme activity and electron transfer kinetics at enzyme-functionalized electrodes. Currently, such optimization can be performed through enzyme structural engineering, improvement of enzyme immobilization methodologies, and fabrication of advantageous electrode materials to enhance retained enzyme activity density at the electrode surface and electron transfer rates between enzymes and an electrode. In this work, varying electrode materials were studied to produce an increased understanding on the impacts of material properties on resulting biochemical, and electrochemical performances upon enzyme immobilization and an additional method of electroactive enzyme-based optimization was developed through the use of polymer-based protein engineering (PBPE). First, graphene/single-wall carbon nanotube cogels were studied as supports for membrane- and mediator-free EBFCs. The high available specific surface area and porosity of these materials allowed the rechargeable generation of a power density within one order of magnitude of the highest performing glucose-based EBFCs to date. Second, two additional carbon nanomaterial-based electrode materials were fabricated and examined as EBFC electrodes. Graphene-coated single-wall carbon nanotube gels and gold nanoparticle/multi-wall carbon nanotube-coated polyacrylonitrile fiber paddles were utilized as electroactive enzyme supports. The performance comparison of these three materials provided an increased understanding of the impact of material properties such as pore size, specific surface area and material surface curvature on enzyme biochemical and electrochemical characteristics upon immobilization. Third, PBPE techniques were applied to develop enzyme-redox polymer conjugates as a new platform for enzymatic biosensor and EBFC optimization. Poly(N-(3-dimethyl(ferrocenyl) methylammonium bromide)propyl acrylamide) (pFcAc) was grown directly from the surface of glucose oxidase (GOX) through atom-transfer radical polymerization. Utilization of the synthesized GOX-pFcAc conjugates led to a 24-fold increase in current generation efficiency and a 4-fold increase in EBFC power density compared to native GOX. GOX-pFcAc conjugates were further examined as working catalysts in carbon paper-based enzymatic biosensors, which provided reliable and selective glucose sensitivities and allowed a systematic analysis of sources of instability in enzyme-polymer conjugate-based biosensors and EBFCs. The knowledge gained through these studies and the in-depth characterization of an additional layer of optimization capacity using PBPE could potentially enhance the progress of enzymatic biosensor and EBFC development.

Electrochemical Impedance Spectroscopy (EIS) was used to develop a novel methodology for studying ionic interaction with lipids arranged in a lipid cubic phase (LCP). Studying different types of ions, both cations and anions, validated the method. A free-standing LCP membrane was formed between two cell compartments and impedance experiments were carried out in a 2-electrode setup to estimate dielectric properties of the membrane, exposed to the following electrolyte solutions at different concentrations: KCl, CsBr, CaCl2, MgCl2, CsCl, NaCl, NaOAc and NaTryptophan. Two different LCP were used in this setup, i.e: Monoloein/water and the ternary system of monoolein/dioleoylphosphatidylcholine/water (MO/DOPC/H2O). SAXRD measurements were performed to determine the space group of the cubic phase and confirm the stability of the LCP during measurements. Membrane resistances and capacitances were found from equivalent circuit fitting to the impedance data. The membrane resistance was shown to be related to ionic interaction with the lipid head group in the water channels of the LCP. Membrane capacitance were correlating to condensing and swelling effect of LCP due to the exposure of ions. The results correlated well with the SAXRD results and earlier published data. The results also indicate that these membranes become less permeable to ions as they increase in size as well as in charge or polarity.  Cyclic voltammetry was used to study the applications of a LCP for modification of the bioanode in a biofuel cell. The monoolein cubic phase was used to host Glucose oxidase (GOx) and a freely diffusing ferrocene carboxylate was used as mediator. The supported cubic phase had an intrinsic resistance in the same order of magnitude as the freestanding MO-LCP membrane as measured with EIS. / Elektrokemisk impedans spektroskopi har använts för att utveckla en ny metod för att studera joners växelverkan med lipider som bildat en kubisk fas. Olika typer av joner, både positiva och negativa, användes för att validera metoden. Ett fristående membran uppbyggt av en kubisk fas separerade två avdelningar i en elektrokemisk cell. Cellen fylldes med elektrolyt-lösningar och impedansmätningar kunde utföras mellan två platina elektroder placerade i vardera avdelning. Membranet exponerades för följande elektrolytlösningar av olika koncentration: KCl, CsBr, CaCl2, MgCl2, CsCl, NaCl, NaOAc and NaTryptofan. Två olika kubiska faser användes i denna uppställning, dvs: Monoloein/vatten och det ternära systemet monoolein/dioleoylfosfatidylkolin/vatten(MO/DOPC/H2O). Med hjälp av SAXRD kunde den kubiska fasens kristallstruktur bestämmas och dess stabilitet under mätningarna bekräftas. De dielektriska egenskaperna hos membranet bestämdes genom att anpassa impedansspektrat till en ekvivalent krets bestående av resistanser, kapacitanser och konstant-faselement. Membranresistansen visade sig vara relaterad till jonernas växelverkan med lipidhuvudgruppen i vattenkanalerna i kubiska fasen. Ju starkare växelverkan desto högre var resistansen. Membrankapacitansen kunde korreleras med kondenserande och uppsvällande effekter på kubiska fasen förorsakade av exponeringen till joner. Resultaten bekräftades av SAXRD mätningar och även tidigare publicerade data. Resultaten indikerar också tydligt att permeabiliteten hos membranet minskar med ökad jonstorlek, jonladdningoch polaritet hos jonen. Cyklisk voltammetri användes för att studera en tillämpning av kubiska fasen i en tänkt applikation som bioanod i en biobränslecell. Elektroden modifierades med en kubisk fas innehållande GOx och tillsammans med en fritt diffunderande ferrocen karboxylat som mediator, där oxidation av glukos studeras. Det visade sig att den kubiska fasen hade en resistans av samma storleksordning som det fristående membranet uppmätt med impedansspektroskopi.

Lignin, one of main components in the woody cell walls, is a complex heterogeneous biopolymer, which provides structural support and transportation of water in plants. It is highly recalcitrant to degradation (both chemically and environmentally) and protects cellulose from being degraded/hydrolyzed. Due to the structural complexity of native lignin, complete characterization and elucidation of lignin’s structure remains very challenging. The overarching goal of this work is to develop mass spectrometry based analytical methods to contribute to a better understanding of lignin structures. This dissertation will focus on the development and application of High-Resolution Accurate-Mass (HRAM) Mass Spectrometry (MS) as main analytical technique for studying lignin model compounds, including understanding the ionization behavior, studying corresponding fragmentation patterns and extracting structural information for structural elucidation eventually. Analytical methods were also developed to study the post-pretreatment products of the synthetic trimeric model compound using High-Performance Liquid Chromatography (HPLC) coupled with High-Resolution Accurate Mass (HRAM) Mass Spectrometry (MS). The first project of this dissertation focuses on mass spectral the characterization of lignin models from the in vitro oxidative coupling reactions. Three specific trimeric compounds were isolated and their ionization behaviors were investigated using HRAMMS via electrospray ionization (ESI). The reaction parameters of the in vitro oxidative coupling reaction were critical in alternating the linkage profiles of resulting dehydrogenation polymers (DHPs). Reaction parameters were tuned to obtain desired DHP linkages profile. Upon the isolation of three different trimeric compounds, a systematic comparison of ionization efficiency of three trimeric compounds was carried out using ESI-HRAM-MS under different ionization conditions. The second project was aimed to design a synthetic route for a lignin model compound that will be a good representation for native lignin during the pretreatment process. The model compound of interest has not been obtained previously through chemical synthesis. Due to the reactivity of cinnamyl alcohol, which contains the unsaturated side chain, this new synthesis strategy was developed based on the known aldol-type reaction route. A versatile synthesis procedure for preparation of β-O-4 oligomeric compounds was designed and implemented to include the most important functional groups (phenolic alcohol, aryl glycerol β-aryl ether bond and unsaturated side chain) in the resulting model compound. This new synthesis route also allowed incorporation of different monolignols. In the third project, Fenton chemistry was applied to a synthetic lignin model compound. Due to the non-specificity in the post pretreatment product profile, nontargeted analytical strategy was developed and applied to study the post-pretreatment products of the model compound using HPLC-HRMS. The results from this dissertation showed a significant difference in ionization behavior between three structurally different model compounds and indicated that primary structures of lignin compounds can largely affect corresponding electrospray ionization properties as well as fragmentation pattern. The work in this dissertation provides analytical techniques for non-targeted analysis of complex lignin samples and an insightful understanding of Fenton’s reaction pretreatment upon lignin model compound.

Biofuel cells (BFCs) that utilize enzymes as catalysts represent a new sustainable and renewable energy technology. Numerous efforts have been directed to improve the performance of the enzymatic BFCs (EBFCs) with respect to power output and operational stability for further applications in portable power sources, self-powered electrochemical sensing, implantable medical devices, etc. This concept article details the latest advances about the EBFCs based on porous nanoarchitectures over the past 5 years. Porous matrices from carbon, noble metal, and polymer promote the development of EBFCs through the electron transfer and mass transport benefits. We will also discuss some key issues on how these nanostructured porous media improve the performance of EBFCs in the end.

Lignocellulosic biomass is a potential feedstock for production of sugars, which can be fermented into ethanol. The work presented in this thesis proposes some solutions to overcome problems with suboptimal process performance due to elevated cultivation temperatures and inhibitors present during ethanol production from lignocellulosic materials. In particular, continuous processes operated at high dilution rates with high sugar utilisation are attractive for ethanol fermentation, as this can result in higher ethanol productivity. Both encapsulation and membrane bioreactors were studied and developed to achieve rapid fermentation at high yeast cell density. My studies showed that encapsulated yeast is more thermotolerant than suspended yeast. The encapsulated yeast could successfully ferment all glucose during five consecutive batches, 12 h each at 42 °C. In contrast, freely suspended yeast was inactivated already in the second or third batch. One problem with encapsulation is, however, the mechanical robustness of the capsule membrane. If the capsules are exposed to e.g. high shear forces, the capsule membrane may break. Therefore, a method was developed to produce more robust capsules by treating alginate-chitosan-alginate (ACA) capsules with 3-aminopropyltriethoxysilane (APTES) to get polysiloxane-ACA capsules. Of the ACA-capsules treated with 1.5% APTES, only 0–2% of the capsules broke, while 25% of the untreated capsules ruptured within 6 h in a shear test. In this thesis membrane bioreactors (MBR), using either a cross-flow or a submerged membrane, could successfully be applied to retain the yeast inside the reactor. The cross-flow membrane was operated at a dilution rate of 0.5 h-1 whereas the submerged membrane was tested at several dilution rates, from 0.2 up to 0.8 h-1. Cultivations at high cell densities demonstrated an efficient in situ detoxification of very high furfural levels of up to 17 g L-1 in the feed medium when using a MBR. The maximum yeast density achieved in the MBR was more than 200 g L-1. Additionally, ethanol fermentation of nondetoxified spruce hydrolysate was possible at a high feeding rate of 0.8 h-1 by applying a submerged membrane bioreactor, resulting in ethanol productivities of up to 8 g L-1 h-1. In conclusion, this study suggests methods for rapid continuous ethanol production even at stressful elevated cultivation temperatures or inhibitory conditions by using encapsulation or membrane bioreactors and high cell density cultivations. / <p>Akademisk avhandling som för avläggande av teknologie doktorsexamen vid Chalmers tekniska högskola försvaras vid offentlig disputation den 4 april 2014, klockan 9:30 i KE-salen, Kemigården 4, Göteborg.</p>