Layer-by-Layer assembled cobalt ferrite nanoparticles for chemical sensing

Ultra-thin (thicknesses of 50-90 nm) nanocomposite films of cobalt ferrite nanoparticles (np-CoFe2O4, 18 nm in diameter) and polyelectrolytes (doped polyaniline-PANI, poly-3,4-ethylenedioxy thiophene: polystyrene sulfonic acid-PEDOT:PSS, and sulfonated lignin-SL) are assembled layer-by-layer onto interdigitated microelectrodes aiming at to create novel nanostructured sensoactive materials for liquid media chemical sensors. The nanocomposites display a distinctive globular morphology with nanoparticles densely-packed while surrounded by polyelectrolytes. Due to the presence of np-CoFe2O4 the nanocomposites display low electrical conductivity according to impedance data. On the other hand, this apparent shortcoming turns such nanocomposites much more sensitive to the presence of ions in solution than films made exclusively of conducting polyelectrolytes. For example, the electrical resistance of np-CoFe2O4/PEDOT:PSS and PANI/SL/np-CoFe2O4/SL architectures has a 10-fold decrease when they are immersed in 20 mmol. L-1 NaCl solution. Impedance spectra fitted with the response of an equivalent circuit model suggest that the interface created between nanoparticles and polyelectrolytes plays a major role on the nanocomposites electrical/dielectrical behavior. Since charge transport is sensitive to nanoparticle-polyelectrolyte interfaces as well as to the physicochemical conditions of the environment, the np-CoFe2O4-based nanocomposites can be used as sensing elements in chemical sensors operated under ac regime and room temperature.

Dielectric properties of cobalt ferrite nanoparticles in ultrathin nanocomposite films

Multilayered nanocomposite films (thickness 50-90 nm) of cobalt ferrite nanoparticles (np-CoFe2O4, 18 nm) were deposited on top of interdigitated microelectrodes by the layer-by-layer technique in order to study their dielectric properties. For that purpose, two different types of nanocomposite films were prepared by assembling np-CoFe2O4 either with poly(3,4-ethylenedioxy thiophene):poly(styrene sulfonic acid) or with polyaniline and sulfonated lignin. Despite the different film architectures, the morphology of both was dominated by densely-packed layers of nanoparticles surrounded by polyelectrolytes. The dominant effect of np-CoFe2O4 was also observed after impedance spectroscopy measurements, which revealed that dielectric behavior of the nanocomposites was largely influenced by the charge transport across nanoparticle-polyelectrolyte interfaces. For example, nanocomposites containing np-CoFe2O4 exhibited a single low-frequency relaxation process, with time constants exceeding 15 ms. At 1 kHz, the dielectric constant and the dissipation factor (tan ᵟ) of these nanocomposites were 15 and 0.15, respectively. These values are substantially inferior to those reported for pressed pellets made exclusively of similar nanoparticles. Impedance data were further fitted with equivalent circuit models from which individual contributions of particle's bulk and interfaces to the charge transport within the nanocomposites could be evaluated. The present study evidences that such nanocomposites display a dielectric behavior dissimilar from that exhibited by their individual counterparts much likely due to enlarged nanoparticle- polyelectrolyte interfaces.

Risposte di resistenza a batteri fitopatogeni di importanti specie coltivate indotte da molecole segnale di diversa natura

A modern management of crop protection should be based on integrated control programmes, including the use of environmentally safe products. Antagonistic/beneficial bacteria and resistance inducers may have a great potential in the prophylaxis of diseases caused by common and quarantine pathogens. This work was carried out to confirm the ability of the known strain IPV-BO G19 (Pseudomonas fluorescens) against fire blight (Erwinia amylovora), as well as to evaluate their efficacy against southern bacterial wilt of tomato (Ralstonia solanacearum) and grapevine crown gall (Agrobacterium vitis). A virulent strain of R. solanacearum race 3 was inhibited by the antagonist on plate. When the pathogen was inoculated 48 h after their application to the root apparatus of tomato plants grown in a climatic chamber, bacterial wilt progression rate was clearly reduced. Moreover the defence response evoked by IPV-BO G19 was studied in tomato plants by monitoring the transcription of genes codifying for three PRs as PR-1a, PR-4, PR-5 and for an intracellular chitinase using multiplex RT-PCR and Real Time RT-PCR. In two field trials during 2005 and 2006, the strain IPV-BO G19 was compared with biofungicides and some abiotic elicitors to protect actively growing shoots of pear scions against fire blight. In both trials, IPV-BO G19 plus Na-alginate gave a high level of protection, three weeks after wound inoculation with E. amylovora. In pear leaf tissues treated with the antagonistic strain IPV-BO G19, catalase, superoxyde dismutase and peroxidise activity was evaluated as markers of induced resistance. The IPV-BO G19 strain was compared with other bioagents and resistance inducers to prevent grapevine crown gall under glasshouse and vineyard conditions.

The Domon Family of Plant Plasma Membrane B-Type Cytochromes: Heterologous Expression, Biochemical Characterization and Physiological Roles in Vivo

The DOMON domain is a domain widespread in nature, predicted to fold in a β-sandwich structure. In plants, AIR12 is constituted by a single DOMON domain located in the apoplastic space and is GPI-modified for anchoring to the plasma membrane. Arabidopsis thaliana AIR12 has been heterologously expressed as a recombinant protein (recAtAIR12) in Pichia pastoris. Spectrophotometrical analysis of the purified protein showed that recAtAir12 is a cytochrome b. RecAtAIR12 is highly glycosylated, it is reduced by ascorbate, superoxide and naftoquinones, oxidised by monodehydroascorbate and oxygen and insensitive to hydrogen peroxide. The addition of recAtAIR12 to permeabilized plasma membranes containing NADH, FeEDTA and menadione, caused a statistically significant increase in hydroxyl radicals as detected by electron paramagnetic resonance. In these conditions, recAtAIR12 has thus a pro-oxidant role. Interestingly, AIR12 is related to the cytochrome domain of cellobiose dehydrogenase which is involved in lignin degradation, possibly via reactive oxygen species (ROS) production. In Arabidopsis the Air12 promoter is specifically activated at sites where cell separations occur and ROS, including •OH, are involved in cell wall modifications. air12 knock-out plants infected with Botrytis cinerea are more resistant than wild-type and air12 complemented plants. Also during B. cinerea infection, cell wall modifications and ROS are involved. Our results thus suggest that AIR12 could be involved in cell wall modifying reactions by interacting with ROS and ascorbate. CyDOMs are plasma membrane redox proteins of plants that are predicted to contain an apoplastic DOMON fused with a transmembrane cytochrome b561 domain. CyDOMs have never been purified nor characterised. The trans-membrane portion of a soybean CyDOM was expressed in E. coli but purification could not be achieved. The DOMON domain was expressed in P. pastoris and shown to be itself a cytochrome b that could be reduced by ascorbate.

Pretrattamenti termo-meccanici ed enzimatici su scarti dell'industria enologica per la produzione di Biogas

A fundamental assumption for by-product from winery industy waste-management is their economic and commercial increase in value. High energetic value recovery from winery industry is an attractive economic solution to stimulate new sustainable process. Approach of this work is based about physic and biological treatment with grape stalks and grape marc to increase polysaccharides components of cell wall and energetic availability of this by-products. Grape stalks for example have a high percentage of lignin and cellulose and can’t be used, whitout pretreatment, for an anaerobic digestion process. Our findings show enzymatic and thermo-mechanical pre-treatments in combined application for optimise hydrolytic mechanism on winemaking wastes which represents 0,9 milion ton/year in Italy and on straw, cereal by-products with high lignin content. A screening of specifically industrial enzymatic complex for the hydrolysis lignocellulosic biomass were tested using the principal polysaccharides component of the vegetal cells. Combined thermo-mechanical and enzymatic pretreatment improve substrates conversion in batch test fermentation experiment. The conservation of the grape stalks, at temperature above 0°C, allow the growth of spontaneus fermentation that reduce their polysaccharides content so had investigated anarobic condition of conservation. The other objective of this study was to investigate the capability of a proprietary strain of L.buchneri LN 40177 to enhance the accessibility of fermentable forage constituents during the anaerobic conservation process by releasing the enzyme ferulate esterase. The time sequence study by batch tests showed that the L. buchneri LN-40177 inoculated grape stalk substrate was more readily available in the fermenter. In batch tests with grape stalk, after mechanical treatment, the L. buchneri LN41077 treated substrate yielded on average 70% more biogas per kg/DM. Thermo-mechanical, enzymatic and biological treatment with L. buchneri LN-40177 can increase the biogas production from low fermented biomasses and the consequent their useful in anaerobic biodigesters for agro-bioenergy production.

Die Entwicklung der Mesofauna eines Weinberges im nordöstlichen Rheinhessen im Vergleich zum Lennebergwald und weiteren Agrarflächen

ZusammenfassungrnrnrnDer Köderstreifentest, die Auswertung der Minicontainer und die Erfassung der Bodenlebewesen mit Hilfe der Bodenstechkerne ergeben zusammen eine gut standardisierte Methode zur Darstellung und Beurteilung der Mesofauna. Unter der Vorraussetzung gleicher abiotischer Faktoren ist es problemlos möglich, selbst unterschiedliche Standorte wie Agrarflächen, Weinberge und Waldböden vergleichend zu untersuchen.rnrnAuf den verschiedenen Versuchsflächen des Laubenheimer Weinberges gelingt es deutlich zu zeigen, wie wichtig eine naturnahe Begrünung für den Boden ist. Dies betrifft nicht nur die Entwicklung der Humusschicht und damit die Bodenlebewesen, sondern auch die Schaffung von Kapillaren und Poren, die durch schwere landwirtschaftliche Geräte im Rahmen der Bo-denverdichtung reduziert werden. Erosionserscheinungen kommen vollständig zum Stillstand. Das Ökosystem Boden sollte auch so gut wie keine Belastung durch Herbizide, Insektizide und Pestizide erfahren. Ähnliches gilt auch für agrarisch genutzte Flächen. rnrnDer Lennebergwald als Naherholungsregion von Mainz ist besonders schützenswert, da dieser durch intensiven Immissionseintrag aufgrund der Nähe zu den Autobahnen und durch die Eutrophierung über die Haustiere stark belastet wird. Die immer größere Ausdehnung des Siedlungsgebietes und die damit verbundene steigende Anzahl an Waldbesuchern, die durch Verlassen der vorgegebenen Wege den Boden zerstören, gefährden zusätzlich das Ökosystem.rnrnÜber Sinn und Zweck einer Flurbereinigung zu diskutieren ist hier nicht angebracht. Aus bo-denkundlicher Sicht ist sie nicht zu befürworten, da hiermit alle bodenbewahrenden Maßnah-men ignoriert werden. Wichtig ist es, bei den Landwirten Aufklärungsarbeit zu leisten, was bodenschonende und bodenweiterentwickelnde Bearbeitungsmethoden bedeuten. Mit Hilfe sachgemäßer Aufklärung und richtiger Umsetzung kann durch Begrünungsmaßnahmen der zum Teil sehr stark strapazierte Boden erhalten, gefördert und auf lange Sicht stabilisiert wer-den.rnrnAufgrund der festgestellten Tatsachen wurde ab 2008 auf eine flächige Dauerbegrünung um-gestellt, so dass es auch in den unbegrünten Rebzeilen zu einer Bodenverbesserung kommen kann. Mit großer Wahrscheinlichkeit dürfte diese schneller voranschreiten, da die Mesofauna von den benachbarten begrünten Rebzeilen einwandern kann. rnDie Mesofauna landwirtschaftlich genutzter Flächen und Waldgebiete kann, obwohl extrem unterschiedlich, miteinander verglichen werden.rnrnBrachflächen und Waldgebiete lassen sich aufgrund der unberührten Bodenstrukturen sogar gut miteinander vergleichen. Temperatur- und Niederschlagsverhältnisse müssen dabei über-einstimmen. Die Azidität der jeweiligen Böden gilt es zu berücksichtigen, da verschiedene Tiergruppen damit unterschiedlich umgehen. Collembolen bevorzugen neutrale Böden, wäh-rend Acari als Räuber mit den Lebewesen in sauren Böden besser zurechtkommen. Die Streu-auflage ist dabei von großer Bedeutung.rnrnIm Rahmen von Bearbeitungsmaßnahmen kommt es durch jeglichen Maschineneinsatz zu ei-ner mehr oder weniger starken Veränderung der Bodenstruktur und somit auch der darin le-benden Mesofauna. Bis sich diese erholt hat, steht meist schon die nächste Bodenbewirtschaf-tung an. Die Bodenverdichtung spielt auch eine Rolle. Bei herkömmlichem Ackerbau ist eine Fruchtfolge mit eingeschalteter Brache oder Gründüngung mit Klee oder Luzerne angebracht, um die Mesofauna nicht zu stark zu strapazieren. Organische Düngegaben leicht abbaubarer Streu sind deutlich zu bevorzugen gegenüber sehr zellulose- und ligninhaltigen Pflanzenresten. Die Einbringung von Stoppeln nach Aberntung von Getreidefeldern ist sinnvoll, solange dabei nicht zu tief in die Bodenstruktur eingegriffen wird (ZIMMER 1997).rnrnIm Rahmen der Sonderkultur Wein, bei der eine Bodenbearbeitung aus den aufgezeigten Gründen eigentlich nicht notwendig wäre, sind Dauerbegrünungsmaßnahmen generell von Nutzen: der Erosion wird vorgebeugt, die Bodenfeuchte konstant gehalten, der anfallende Mulch als Gründüngung genutzt. Dies sind alles entscheidende Faktoren, die die Meso- und Makrofauna fördern. Nur die Bodenverdichtung durch schweres Gerät, wie Schlepper und Vollernter, sind für den Boden nicht förderlich (HEISLER 1993, EHRENSBERGER 1993). Nie-derdruckreifen und Verringerung der Befahrung sind geeignete Gegenmaßnahmen. rnrnEntgegen landläufiger Winzermeinung, stellen die Pflanzen einer Begrünung eigentlich keine Konkurrenz für die Weinstöcke dar. Die Vorteile einer Begrünung sind nicht nur die Förde-rung der einheimischen Flora in ihrem standortgerechten Artenreichtum, sondern auch Ver-vielfältigung von Meso- und Makrofauna aufgrund der dadurch mehr anfallenden und ein-zuarbeitenden leicht abbaubaren Streu (GRIEBEL 1995).rn

Entwicklung eines Thermodruckhydrolyse-Verfahrens zur Vorbehandlung von Weizenstroh ohne vorherige mechanische Zerkleinerung für die Produktion von Polyhydroxybuttersäure

In der vorliegenden Arbeit wurde der nachwachsende Rohstoff Weizenstroh für die Produktion des Biopolymers Polyhydroxybuttersäure genutzt. Als Lignocellulose enthält Weizenstroh einen hohen Anteil an Glucose und Xylose in Form von Cellulose und Hemicellulose. Eine Gewinnung ist aufgrund der komplexen Struktur mit Lignin als dritte Hauptkomponente nur durch eine Vorbehandlung möglich. Hierzu wurde ein thermochemisches Vorbehandlungsverfahren im halbtechnischen (125 l Reaktor) und technisch (425 l Reaktor) Maßstab mit verdünnter Salpetersäure (bis 1 % v/v) etabliert und hinsichtlich verschiedener Versuchsparameter (Behandlungstemperatur, Säure-Konzentration, etc.) optimiert. Auf eine mechanische Vorbehandlung wurde verzichtet. Danach erfolgte eine enzymatische Hydrolyse der vorbehandelten Biomasse. Der PHB-Produzent Cupriavidus necator DSM 545 wurde eingesetzt, um aus den freigesetzten Zuckern PHB zu synthetisieren. rnDurch die Optimierung der Vorbehandlung konnten bis zu 90 % der Glucose und 82 % der Xylose nach der enzymatischen Hydrolyse aus dem Stroh als Monomere und Oligomere freigesetzt werden. Außerdem wurde eine erfolgreiche Überführung des Vorbehandlungsprozesses in den 425 l Reaktor demonstriert. In den gewonnenen Zucker-Hydrolysaten konnten hohe Zelldichten und PHB-Gehalte mit bis zu 38 % erreicht werden. Eine vorherige kostenintensive Reinigung der Hydrolysate war nicht nötig. Zusätzlich konnte gezeigt werden, dass die Reststoffe nach der enzymatischen Hydrolyse, Zellkultur und PHB-Extraktion ausreichendes Potential für eine Biogas-Produktion besitzen. rn

No adverse effect of genetically modified antifungal wheat on decomposition dynamics and the soil fauna community - a field study

The cultivation of genetically modified (GM) plants has raised several environmental concerns. One of these concerns regards non-target soil fauna organisms, which play an important role in the decomposition of organic matter and hence are largely exposed to GM plant residues. Soil fauna may be directly affected by transgene products or indirectly by pleiotropic effects such as a modified plant metabolism. Thus, ecosystem services and functioning might be affected negatively. In a litterbag experiment in the field we analysed the decomposition process and the soil fauna community involved. Therefore, we used four experimental GM wheat varieties, two with a race-specific antifungal resistance against powdery mildew (Pm3b) and two with an unspecific antifungal resistance based on the expression of chitinase and glucanase. We compared them with two non-GM isolines and six conventional cereal varieties. To elucidate the mechanisms that cause differences in plant decomposition, structural plant components (i.e. C:N ratio, lignin, cellulose, hemicellulose) were examined and soil properties, temperature and precipitation were monitored. The most frequent taxa extracted from decaying plant material were mites (Cryptostigmata, Gamasina and Uropodina), springtails (Isotomidae), annelids (Enchytraeidae) and Diptera (Cecidomyiidae larvae). Despite a single significant transgenic/month interaction for Cecidomyiidae larvae, which is probably random, we detected no impact of the GM wheat on the soil fauna community. However, soil fauna differences among conventional cereal varieties were more pronounced than between GM and non-GM wheat. While leaf residue decomposition in GM and non-GM wheat was similar, differences among conventional cereals were evident. Furthermore, sampling date and location were found to greatly influence soil fauna community and decomposition processes. The results give no indication of ecologically relevant adverse effects of antifungal GM wheat on the composition and the activity of the soil fauna community.

Bioremediation with White-Rot Fungi at Fisherville Mill: Analyses of Gene Expression and Number 6 Fuel Oil Degradation

Extracellular enzymes that white-rot fungi secrete during lignin decay have been proposed as promising agents for oxidizing pollutants. We investigated the abilities of the white-rot fungi Punctularia strigosozonata, Irpex lacteus, Trichaptum biforme, Phlebia radiata, Trametes versicolor, and Pleurotus ostreatus to degrade Number 6 fuel oil in wood sawdust cultures. Our goals are to advise bioremediation efforts at a brownfield redevelopment site on the Blackstone River in Grafton, Massachusetts and to contribute to the understanding of decay mechanisms in white-rot fungi. All species tested degraded a C10 alkane. When cultivated for 6 months, Irpex lacteus, T. biforme, P. radiata, T. versicolor and P. ostreatus also degraded a C14 alkane and the polycyclic aromatic hydrocarbon phenanthrene. Gene expression analyses of P. strigosozonata indicate differential gene expression in the presence of Number 6 oil and on pine and aspen sawdust.

Factors Influencing Spatial Variability in Nitrogen Processing in Nitrogen-Saturated Soils

Nitrogen (N) saturation is an environmental concern for forests in the eastern U.S. Although several watersheds of the Fernow Experimental Forest (FEF), West Virginia exhibit symptoms of Nsaturation, many watersheds display a high degree of spatial variability in soil N processing. This study examined the effects of temperature on net N mineralization and nitrification in N-saturatedsoils from FEF, and how these effects varied between high N-processing vs. low N-processingsoils collected from two watersheds, WS3 (fertilized with [NH4]2SO4) and WS4 (untreated control). Samples of forest floor material (O2 horizon) and mineral soil (to a 5-cm depth) were taken from three subplots within each of four plots that represented the extremes of highest and lowest ratesof net N mineralization and nitrification (hereafter, high N and low N, respectively) of untreated WS4 and N-treated WS3: control/low N, control/high N, N-treated/low N, N-treated/high N. Forest floor material was analyzed for carbon (C), lignin,and N. Subsamples of mineral soil were extractedimmediately with 1 N KCl and analyzed for NH4+and NO3– to determine preincubation levels. Extracts were also analyzed for Mg, Ca, Al, and pH. To test the hypothesis that the lack of net nitrification observed in field incubations on the untreated/low N plot was the result of absence ofnitrifier populations, we characterized the bacterial community involved in N cycling by amplification of amoA genes. Remaining soil was incubated for 28 d at three temperatures (10, 20, and30°C), followed by 1 N KCl extraction and analysis for NH4+ and NO3–. Net nitrification was essentially 100% of net N mineralization for all samples combined. Nitrification rates from lab incubation sat all temperatures supported earlier observations based on field incubations. At 30°C, rates from N- t reated/high N were three times those of N-treated/low N. Highest rates were found for untreated/high N (two times greater than those of N-treated/high N), whereas untreated/low N exhibited no net nitrification. However, soils exhibitingno net nitrification tested positive for presence of nitrifying bacteria, causing us to reject our initial hypothesis. We hypothesize that nitrifier populations in such soil are being inhibited by a combination of low Ca:Al ratios in mineral soil and allelopathic interactions with mycorrhizae of ericaceous species in the herbaceous layer.

Biological materials : Part A. tuning LCST of raft copolymers and gold/copolymer hybrid nanoparticles and Part B. biobased nanomaterials

The research described in this dissertation is comprised of two major parts. The first part studied the effects of asymmetric amphiphilic end groups on the thermo-response of diblock copolymers of (oligo/di(ethylene glycol) methyl ether (meth)acrylates, OEGA/DEGMA) and the hybrid nanoparticles of these copolymers with a gold nanoparticle core. Placing the more hydrophilic end group on the more hydrophilic block significantly increased the cloud point compared to a similar copolymer composition with the end group placement reversed. For a given composition, the cloud point was shifted by as much as 28 °C depending on the placement of end groups. This is a much stronger effect than either changing the hydrophilic/hydrophobic block ratio or replacing the hydrophilic acrylate monomer with the equivalent methacrylate monomer. The temperature range of the coil-globule transition was also altered. Binding these diblock copolymers to a gold core decreased the cloud point by 5-15 °C and narrowed the temperature range of the coil-globule transition. The effects were more pronounced when the gold core was bound to the less hydrophilic block. Given the limited numbers of monomers that are approved safe for in vivo use, employing amphiphilic end group placement is a useful tool to tune a thermo-response without otherwise changing the copolymer composition. The second part of the dissertation investigated the production of value-added nanomaterials from two biorefinery “wastes”: lignin and peptidoglycan. Different solvents and spinning methods (melt-, wet-, and electro-spinning) were tested to make lignin/cellulose blended and carbonized fibers. Only electro-spinning yielded fibers having a small enough diameter for efficient carbonization ( Peptidoglycan (a bacterial cell wall material) was copolymerized with poly-(3-hydroxybutyrate), a common polyhydroxyalkanoate produced by bacteria with the objective of determining if a useful material could be obtained with a less rigorous work-up on harvesting polyhydroxyalkanoates. The copolyesteramide product having 25 wt.% peptidoglycan from a highly purified peptidoglycan increased thermal stability by 100-200 °C compared to the poly-(3-hydroxybutyrate) control, while a less pure peptidoglycan, harvested from B. megaterium (ATCC 11561), gave a 25-50 °C increase in thermal stability. Both copolymers absorbed more moisture than pure poly-(3-hydroxybutyrate). The results suggest that a less rigorously harvested and purified polyhydroxyalkanoate might be useful for some applications.

THE POTENTIAL OF INDUSTRIAL WASTE AND AGRICULTURAL FEEDSTOCK TOWARDS SUSTAINABLE BIOFUELS PRODUCTION: TECHNO-ECONOMIC AND ENVIRONMENTAL IMPACT PERSPECTIVES

This Ph.D. research is comprised of three major components; (i) Characterization study to analyze the composition of defatted corn syrup (DCS) from a dry corn mill facility (ii) Hydrolysis experiments to optimize the production of fermentable sugars and amino acid platform using DCS and (iii) Sustainability analyses. Analyses of DCS included total solids, ash content, total protein, amino acids, inorganic elements, starch, total carbohydrates, lignin, organic acids, glycerol, and presence of functional groups. Total solids content was 37.4% (± 0.4%) by weight, and the mass balance closure was 101%. Total carbohydrates [27% (± 5%) wt.] comprised of starch (5.6%), soluble monomer carbohydrates (12%) and non-starch carbohydrates (10%). Hemicellulose components (structural and non-structural) were; xylan (6%), xylose (1%), mannan (1%), mannose (0.4%), arabinan (1%), arabinose (0.4%), galatactan (3%) and galactose (0.4%). Based on the measured physical and chemical components, bio-chemical conversion route and subsequent fermentation to value added products was identified as promising. DCS has potential to serve as an important fermentation feedstock for bio-based chemicals production. In the sugar hydrolysis experiments, reaction parameters such as acid concentration and retention time were analyzed to determine the optimal conditions to maximize monomer sugar yields while keeping the inhibitors at minimum. Total fermentable sugars produced can reach approximately 86% of theoretical yield when subjected to dilute acid pretreatment (DAP). DAP followed by subsequent enzymatic hydrolysis was most effective for 0 wt% acid hydrolysate samples and least efficient towards 1 and 2 wt% acid hydrolysate samples. The best hydrolysis scheme DCS from an industry's point of view is standalone 60 minutes dilute acid hydrolysis at 2 wt% acid concentration. The combined effect of hydrolysis reaction time, temperature and ratio of enzyme to substrate ratio to develop hydrolysis process that optimizes the production of amino acids in DCS were studied. Four key hydrolysis pathways were investigated for the production of amino acids using DCS. The first hydrolysis pathway is the amino acid analysis using DAP. The second pathway is DAP of DCS followed by protein hydrolysis using proteases [Trypsin, Pronase E (Streptomyces griseus) and Protex 6L]. The third hydrolysis pathway investigated a standalone experiment using proteases (Trypsin, Pronase E, Protex 6L, and Alcalase) on the DCS without any pretreatment. The final pathway investigated the use of Accellerase 1500® and Protex 6L to simultaneously produce fermentable sugars and amino acids over a 24 hour hydrolysis reaction time. The 3 key objectives of the techno-economic analysis component of this PhD research included; (i) Development of a process design for the production of both the sugar and amino acid platforms with DAP using DCS (ii) A preliminary cost analysis to estimate the initial capital cost and operating cost of this facility (iii) A greenhouse gas analysis to understand the environmental impact of this facility. Using Aspen Plus®, a conceptual process design has been constructed. Finally, both Aspen Plus Economic Analyzer® and Simapro® sofware were employed to conduct the cost analysis as well as the carbon footprint emissions of this process facility respectively. Another section of my PhD research work focused on the life cycle assessment (LCA) of commonly used dairy feeds in the U.S. Greenhouse gas (GHG) emissions analysis was conducted for cultivation, harvesting, and production of common dairy feeds used for the production of dairy milk in the U.S. The goal was to determine the carbon footprint [grams CO2 equivalents (gCO2e)/kg of dry feed] in the U.S. on a regional basis, identify key inputs, and make recommendations for emissions reduction. The final section of my Ph.D. research work was an LCA of a single dairy feed mill located in Michigan, USA. The primary goal was to conduct a preliminary assessment of dairy feed mill operations and ultimately determine the GHG emissions for 1 kilogram of milled dairy feed.

Biochemical Conversions of Lignocellulosic Biomass for Sustainable Fuel-Ethanol Production in the Upper Midwest

Biofuels are an increasingly important component of worldwide energy supply. This research aims to understand the pathways and impacts of biofuels production, and to improve these processes to make them more efficient. In Chapter 2, a life cycle assessment (LCA) is presented for cellulosic ethanol production from five potential feedstocks of regional importance to the upper Midwest - hybrid poplar, hybrid willow, switchgrass, diverse prairie grasses, and logging residues - according to the requirements of Renewable Fuel Standard (RFS). Direct land use change emissions are included for the conversion of abandoned agricultural land to feedstock production, and computer models of the conversion process are used in order to determine the effect of varying biomass composition on overall life cycle impacts. All scenarios analyzed here result in greater than 60% reduction in greenhouse gas emissions relative to petroleum gasoline. Land use change effects were found to contribute significantly to the overall emissions for the first 20 years after plantation establishment. Chapter 3 is an investigation of the effects of biomass mixtures on overall sugar recovery from the combined processes of dilute acid pretreatment and enzymatic hydrolysis. Biomass mixtures studied were aspen, a hardwood species well suited to biochemical processing; balsam, a high-lignin softwood species, and switchgrass, an herbaceous energy crop with high ash content. A matrix of three different dilute acid pretreatment severities and three different enzyme loading levels was used to characterize interactions between pretreatment and enzymatic hydrolysis. Maximum glucose yield for any species was 70% oftheoretical for switchgrass, and maximum xylose yield was 99.7% of theoretical for aspen. Supplemental β-glucosidase increased glucose yield from enzymatic hydrolysis by an average of 15%, and total sugar recoveries for mixtures could be predicted to within 4% by linear interpolation of the pure species results. Chapter 4 is an evaluation of the potential for producing Trichoderma reesei cellulose hydrolases in the Kluyveromyces lactis yeast expression system. The exoglucanases Cel6A and Cel7A, and the endoglucanase Cel7B were inserted separately into the K. lactis and the enzymes were analyzed for activity on various substrates. Recombinant Cel7B was found to be active on carboxymethyl cellulose and Avicel powdered cellulose substrates. Recombinant Cel6A was also found to be active on Avicel. Recombinant Cel7A was produced, but no enzymatic activity was detected on any substrate. Chapter 5 presents a new method for enzyme improvement studies using enzyme co-expression and yeast growth rate measurements as a potential high-throughput expression and screening system in K. lactis yeast. Two different K. lactis strains were evaluated for their usefulness in growth screening studies, one wild-type strain and one strain which has had the main galactose metabolic pathway disabled. Sequential transformation and co-expression of the exoglucanase Cel6A and endoglucanase Cel7B was performed, and improved hydrolysis rates on Avicel were detectable in the cell culture supernatant. Future work should focus on hydrolysis of natural substrates, developing the growth screening method, and utilizing the K. lactis expression system for directed evolution of enzymes.

Effects of Ethylene on Secondary Xylem Formation in Arabidopsis thaliana

Ethylene has myriad roles as a plant hormone, ranging from senescence and defending against pathogen attacks to fruit ripening and interactions with other hormones. It has been shown to increase cambial activity in poplar, but the effect on wood formation in Arabidopsis hypocotyl has not previously been studied. The Auxin-Regulated Gene involved in Organ Size (ARGOS), which increases organ size by lengthening the time for cell division, was found to be upregulated by ethylene. We tested the effect of ethylene treatment at 10 and 100 µM ACC on three genotypes of Arabidopsis, Col0 (wild-type), an ARGOS deficient mutant (argos), and ein3-1, an ethylene insensitive mutant. ARGOS expression analysis with qPCR indicated that ACC does induce ARGOS and ARGOS-LIKE (ARL) in the hypocotyl. As seen in poplar, ethylene also decreases stem elongation.Histochemical staining, showed that ethylene changes the way secondary xylem lignifies, causing gaps in lignification around the outer edge of secondary xylem. Our results also implied that ethylene treatment changes the proportion of secondary to total xylem, resulting in less secondary, whereas in poplar, ethylene treatment caused an increase.

Nutritive Value of the Crop Residues from bt-Corn Hybrids and Their Effects on Performance of Grazing Beef Cows

One non bt-corn hybrid (Pioneer 3489) and three btcorn hyrids (Pioneer 34RO7, Novartis NX6236, and Novartis N64-Z4) were planted in replicated 7.1-acre fields. After grain harvest, fields were stocked with 3 mature cows in midgestation to be strip-grazed as four paddocks over 126 days. Six similar cows were allotted to replicated drylots. All cows were fed hay as necessary to maintain a condition score of 5 on a 9-point scale. Cows were condition-scored biweekly and weighed monthly. Forage yield and weathering losses were determined by sampling one 4-m2 location per grazed or ungrazed paddock in each field with a minimum total of 2 locations of grazed or ungrazed forage per field. To measure forage selection during grazing, samples of grazed forage were collected from the rumen of one fistulated steer that grazed for 2 hours after ruminal evacuation. Non-bt-corn hybrids had greater (P<.05) infestation of corn borers in the upper stalk, lower stalk and ear shank than bt-corn hybrids. However, there were no differences in grain yields or dropped grain between hybrids. Crop residue dry matter, organic matter and in vitro digestible dry matter yields at the initiation of grazing did not differ between corn hybrids. Dry matter, organic matter and in vitro digestible dry matter losses tended (P<.10) to be greater from the NX6236 and N64-Z4 hybrids than from the 3489 and 34RO7 hybrids and were greater (P<.05) from grazed than non-grazed areas of the fields. At the initiation of grazing, dry matter concentrations of the crop residues from the NX6236 and N64-Z4 hybrids tended to be lower than those from the 3489 and 34RO7 hybrids. Crop residues from the NX6236 and N64-74 hybrids had lower concentrations of acid detergent fiber (P<.05) and acid detergent lignin (P=.07) and higher concentrations of in vitro digestible organic matter than the 3489 and 34RO7 hybrids. Over the grazing season, corn hybrid did not affect mean rates of change in forage composition. The concentration of in vitro digestible organic matter in forage selected by steers after two weeks of grazing did not differ. However, steers grazing corn crop residues consumed forage with higher (P<.05) concentrations of neutral detergent fiber, acid detergent fiber, and acid detergent insoluble nitrogen than steers fed hay. The acid detergent fiber concentration of forage selected by steers grazing the 3489 and N64-Z4 hybrids was lower (P < .05) than concentrations from the 34RO7 and NX6236 hybrids. In order to maintain similar body condition score changes, cows grazing crop residues from the 3489, 34RO7, NX6236, and N64-Z4 hybrids required 650, 628, 625, and 541 kg hay DM/cow compared with a hay requirement of 1447 kg hay DM/cow for cows maintained in a drylot.