Browsing by Subject "Bioremediation"
Now showing 1 - 8 of 8
Results Per Page
Sort Options
Item 3D Printed Biocatalytic Silica Hydrogel Flow-Through Reactor For Atrazine Degradation(2017-06) Han, RyanOne of the most heavily used herbicides in the world, atrazine, provides a serious environmental challenge that we face presently. Atrazine has been consistently applied to farms due to its proven ability to remove broadleaf weeds, allowing for increased yields of corn crops, which is necessary to feed an ever-growing world population. However advantageous the use of atrazine is, toxic effects have been identified when humans ingest atrazine. Also, the high mobility of atrazine during run-off events after application to fields allows atrazine to be easily incorporated into water systems around agricultural land, creating a large-scale health and environmental problem as the increased atrazine concentrations negatively impact human health when ingested as well as ecological disturbances when affecting local algal communities. The presented work investigates the application of 3D printing as an approach to solving this significant problem. We hypothesize that with direct-write 3D printing of biologically active, printed materials to perform the bioremediation of atrazine, may enhance bioremediation capacity compared to conventional methods by utilizing the near limitless rapid design flexibility intrinsic to 3D printing to allow fabrication of structures with high surface area to volume ratio (SA:V), yielding lower diffusion length scales that allow improved encapsulated biocatalyst usage. We introduce a novel 3D printing method to produce application specific complex 3D geometries from a sol-gel based silica material with encapsulated biocatalysts. To produce a bioactive material with the incorporation of biocatalysts, silica hydrogel formed through a sol-gel process was used as the ink base. E. coli genetically engineered to overexpress the AtzA enzyme, which degrades the toxic herbicide atrazine to the non-toxic compound hydroxyatrazine, were encapsulated within the silica-based ink. This process leverages the strong mechanical properties, high chemical transport properties, and biocompatibility of the silica base material along with the full material customization, precision in spatial deposition, and design flexibility intrinsic to the 3D printing process to overcome obstacles that hinder the use of bioactive materials within conventional 3D printers (material constraints and biologically deadly processing). The developed 3D printer ink was characterized in terms of gelation kinetics, mechanical properties, cell distribution, and degradation capability. Results confirmed that the 3D printed AtzA biocatalysts sustained biodegradation ability through the removal of atrazine and production of hydroxyatrazine through batch reactor experiments. High SA:V geometries produced through 3D printing also showed improved degradation efficiency by encapsulated biocatalysts. This allowed for an advantage over previously presented work because by providing high SA:V structures, the atrazine did not have to diffuse over long length scales until it was biotransformed within a bacterial cell. Structures with low SA:V were shown to decrease in degradation efficiency because as the atrazine concentration gradient decreased, only the cells closer to the surface would perform the biotransformation of atrazine, the cells located more centrally would not contribute to the degradation. Therefore, with a decrease in diffusion length to all encapsulated biocatalysts, the overall function of the encapsulated population as the concentration of atrazine dropped would be improved over past methods. Additionally, a flow-through bioreactor was designed, simulated, and experimentally tested. ANSYS reaction-flow simulations were completed to determine experimental flow rates necessary to positively identify atrazine degradation in the flow-through bioreactor. Finally, atrazine degradation was proven in flow-through experiments at an inlet flowrate of 1 ml/min. Observed atrazine degradation equated to 15 ± 5% of overall inlet concentration atrazine. Through this work, we have shown as a proof of concept that 3D printed silica-encapsulated biocatalysts sustain the function to degrade an environmental pollutant. This work may be expanded further via the incorporation of multiple types of biocatalysts encapsulated in an organized pattern (multiple different 3D printer inks printed in a designed pattern) that enhances biotransformation and transport of products between the multiple biocatalysts. In addition, this work may be applied to advance fields where complex geometries of encapsulated biocatalysts are necessitated, which may include the fields of pharmaceutical, medical, environmental, and materials science.Item Adaptive Expansion of Biodegradation by Pseudomonas Putida F1(2017-06) Chan, BaoPseudomonas putida F1 (PpF1) catabolizes aromatic compounds via benzoate dioxygenase, phenylacetyl-CoA epoxidase, p-cymene-monooxygenase, and toluene dioxygenase-mediated pathways, and the latter is shown here to be highly flexible, supporting growth on previously untested aromatic alkenes, esters, amides, alcohols, amines, and multi-ring compounds. P. putida F1 is a highly studied model aromatic hydrocarbon oxidizer that grows on relatively few mono-substituted benzenes despite its genome encoding a toluene dioxygenase enzyme that oxidizes more than 150 compounds. While toluene dioxygenase and toluene dihydrodiol dehydrogenase oxidize many mono-substituted benzene ring compounds, further enzymatic processing may be inhibited, leading to accumulation of the respective catechol or 2-hydroxy-6-oxo-2,4-dienoate intermediate. The present study demonstrated that P. putida F1 can undergo adaption leading to a more expanded growth range of mono-substituted benzene ring compounds than had been previously demonstrated. Studies with well-characterized mutant derivatives of P. putida F1 and growth on expected metabolites of the toluene dioxygenase pathway indicated that the newly described metabolism was dependent on the Tod pathway enzymes. This study also revealed the interplay between the Tod pathway enzymes and catabolism of the aromatic acids liberated by TodF. TodABCDEF processing of allylbenzene and 1-phenylethanol liberates 3-butenoic acid and lactic acid, respectively, both of which support growth of P. putida F1.Item The effect of carbon inputs on microbial community structure and function: the role of fermentation processes in groundwater.(2009-12) Nelson, Denice KarenCarbon inputs to groundwater aquifers include intentional applications, as in bioremediation practices, and unintentional spills. The addition of carbon to an aquifer environment promotes the growth of a diverse and complex microbial community capable of generating several fermentation products, including some regulated compounds and methane, an explosive gas. This dissertation focuses on the fermentative community that develops in response to carbon application in an aquifer environment. Research was conducted to specifically examine 1) how fermentation processes affect partitioning of trichloroethene (TCE), a common groundwater contaminant, 2) the extent that continuous or pulsed carbon inputs affect microbial community structure and function, and 3) how an ethanol-based fuel (E85) stimulates fermentation processes, including methane generation, and the effect of ethanol toxicity on plume longevity. Remediation of groundwater plume source areas is challenging because lingering contaminants are often present as non-aqueous phase liquid (NAPL) and sorbed mass, and therefore difficult to remove via biodegradation or other commonly used remedial methods. Experimental results indicated that enhanced dissolution of TCE NAPL was possible through the addition and/or subsequent fermentation of a dilute molasses solution. Two mechanisms were responsible for the enhanced dissolution of NAPL; the addition of fresh molasses increased TCE solubility (>200%), thereby increasing the concentration gradient and subsequent mass transfer of NAPL to the dissolved phase, and mixing NAPL with fermented molasses solution significantly increased the surface area of the NAPL through formation of an emulsion, thereby increasing the mass flux of NAPL to the dissolved phase. In addition, the fermented liquid may have also decreased the soil partitioning coefficient (Kd) of TCE, indicating that enhanced transfer of sorbed mass to the aqueous phase could also occur in the presence of fermented molasses. These results can be used to optimize remedial systems to increase NAPL and sorbed-mass dissolution and are therefore important, particularly when bioremediation is used to polish residual source zones. The addition of organic carbon to a groundwater aquifer for biostimulation purposes promotes the growth of a diverse fermentative community as well as organisms targeted for contaminant degradation. Engineered carbon application systems commonly include either a continuous low dose of carbon, or periodic high doses of carbon. Experimental results indicated that a monthly pulse of a high dose (10% by volume) of molasses generated several fermentation products at high levels following each application, while a continuous feed of low molasses solution (0.4%) reached steady-state in 130 days, after which no further detection of fermentation products occurred. Methane generation in both systems was similar, indicating that methane production was not affected by the carbon addition strategy. Significant shifts in both Eubacteria and Archaea community structures were observed after carbon introduction, with the greatest changes correlating to the higher concentrations of carbon provided by the pulsed system. The total quantity of bacteria and methanogens was higher along the pulsed-fed column compared to the continuously-fed system. The continuously-fed column exhibited greater biofouling behavior. Taken together, biofouling did not appear to be a result of biomass quantity, rather a function of community structure. In summary, the method of carbon introduction (pulsed high-dose versus continuous low-dose) can result in significantly different community structures, functions, and densities of indigenous organisms. These data suggest that systems can be engineered to control fermentation product generation and biofouling behavior by manipulating the style of carbon application. Methane, however, will need to be controlled in either system. A spill of ethanol-based fuel will not only contaminate an aquifer, but will also serve as a food source to stimulate fermentative organisms that can generate potentially regulated compounds and create an environment conducive for production of explosive methane gas. Experimental results indicated that a continuous supply of a dilute ethanol-based fuel (E85) resulted in a profound shift in the community structure of Eubacteria and Archaea accompanied by the production of volatile fatty acids and butanol, a compound with a groundwater regulatory standard in Minnesota. Data also indicated that dissolved methane was produced at concentrations that could accumulate to an explosive level (>2 mg/L) in headspace. Quantitative polymerase chain reaction (qPCR) data showed a statistically significant increase in methanogenic populations, when compared to a control column. These results strongly correlated to areas of the column containing acetate, a breakdown product of ethanol. Toxicity data indicated that microbial growth was completely inhibited at approximately 6% (vol/vol) ethanol. These results suggest that even though ethanol is readily degradable, the core of an E85 spill may serve as a long-term source of contamination, and subsequent methane production, as it cannot be degraded until significant dilution has occurred. The research presented in this dissertation shows that the addition and subsequent fermentation of molasses can enhance the mass transfer of TCE, and that the style of carbon application affects the microbial community structure, density of biomass, and subsequent production of fermentation processes. Similarly, an input of E85 will result in the generation of fermentation products, some of which are regulated, and produce methane at levels that can potentially accumulate to explosive levels. This research furthers our understanding of the importance of fermentation processes resulting from carbon inputs to a groundwater environment. These results can be used to optimize bioremediation systems that incorporate carbon addition in order to manage fermentation product formation and biofouling impacts, and to mitigate potential human health hazards stemming from ethanol-based fuel spills through more accurate fate and transport modeling efforts.Item Engineering Biocatalytic And Living Materials(2019-05) Benson, JoeyProviding clean and safe water to the world’s growing human population in one of the major challenges of the 21st century. Increased human activity contaminates waters with toxic chemicals, making it unsafe for consumption. Bioremediation utilizing encapsulated bacteria has emerged as an efficient method for the removal of toxic chemicals from the environment, and can permanently remove a wide range of pollutants by breaking them down into simple compounds. In this presentation, biocatalytic materials were developed for water bioremediation utilizing encapsulated bacteria. Silica gels with encapsulated bacterial cells and spores were designed and optimized for cytocompatibility, porosity, and catalytic activity. As a case study, the gels were used to remove a hydrophobic herbicide from drinking water. First, the surface properties or the silica gel were optimized for biodegradation of the hydrophobic herbicide, by incorporating hydrophobic side chains into the gel. The hydrophobic groups rapidly removed the herbicide by adsorption, and the adsorbed herbicide was rapidly degraded by encapsulated cells, resulting in high biodegradation rates. Next, materials were engineered for the encapsulation, germination and outgrowth of bacterial spores. Encapsulation of the robust bacterial spores enabled a much wider range of synesis conditions, resulting in materials with better mechanical properties. After encapsulation, spores could be germinated by adding nutrients, causing them to grow into metabolically active cells. In addition, spores could be encapsulated in desiccated materials, enabling long term storage with a low risk of fowling. After the gels were designed and characterized, an emulsion system was developed for production of silica gel microspheres with encapsulated bacteria for use in a packed bed bioreactor, and the system was scaled up to produce 52 gallons of microspheres for field test. Finally, robust bionanocomposites with controlled morphologies were produced via the self-assembly and biosilification of fusion proteins. The scaffold forming protein EutM was genetically fused to four different silica biomineralization peptides, enabling control over the scaffold morphology and extent of biosilicification. At high silica concentrations, the proteins catalyzed the formation of a macroscale silica gel, and the microstructure and mechanical properties of the gel could be tuned by adjusted the protein concentration. The silica precipitating protein scaffolds developed in this work represent an ideal biomaterial for the fabrication of living materials because of their small size, self-assembling properties and their robustness.Item Fate and degradation of petroleum hydrocarbons in stormwater bioretention cells.(2012-08) LeFevre, Gregory HallettThis dissertation describes the investigation of the fate of hydrocarbons in stormwater bioretention areas and those mechanisms that affect hydrocarbon fate in such systems. Seventy-five samples from 58 bioretention areas were collected and analyzed to measure total petroleum hydrocarbon (TPH) residual and biodegradation functional genes. TPH residual in bioretention areas was greater than background sites but low overall (<3 µg/kg), and well below either the TPH concentration of concern or the expected concentration, assuming no losses. Bioretention areas with deep-root vegetation contained significantly greater quantites of bacterial 16S rRNA genes and two functional genes involved in hydrocarbon biodegradation. Field soils were capable of mineralizing naphthalene, a polycyclic aromatic hydrocarbon (PAH) when incubated in the laboratory. In an additional laboratory investigation, a column study was initiated to comprehensively determine naphthalene fate in a simulated bioretention cell using a 14C-labeled tracer. Sorption to soil was the greatest sink of naphthalene in the columns, although biodegradation and vegetative uptake were also important loss mechanisms. Little leaching occurred following the first flush, and volatilization was insignificant. Significant enrichment of naphthalene degrading bacteria occurred over the course of the experiment as a result of naphthalene exposure. This was evident from enhanced naphthalene biodegradation kinetics (measured via batch tests), significant increases in naphthalene dioxygenase gene quantities, and a significant correlation observed between naphthalene residual and biodegradation functional genes. Vegetated columns outperformed the unplanted control column in terms of total naphthalene removal and biodegradation kinetics. As a result of these experiments, a final study focused on why planted systems outperform unplanted systems was conducted. Plant root exudates were harvested from hydroponic setups for three types of plants. Additionally, a solution of artificial root exudates (AREs) as prepared. Exudates were digested using soil bacteria to create metabolized exudates. Raw and metabolized exudates were characterized for dissolved organic carbon, specific UV absorbance, spectral slope, florescence index, excitation-emission matrices, and surface tension. Significant differences on character were observed between the harvested exudates and the AREs, as well as between the raw and metabolized exudates. Naphthalene desorption from an aged soil was enhanced in the presence of raw exudates. The surface tension in samples containing raw harvested exudates was reduced compared to samples containing the metabolized exudates. Plant root exudates may therefore facilitate phytoremediation by enhancing contaminant desorption and improving bioavailability. Overall, this resarch concludes that heavily planted bioretention systems are a sustainable solution to mitigating stormwater hydrocarbon pollution as a result of likely enhanced contaminant desorption, and improved biodegradation and plant uptake in such systems.Item An Interdisciplinary Geochemical and Genomics Approach to Understanding Fungal Selenium Transformations for the Bioremediation of Contaminated Waters(2021-07) Sabuda, MarySelenium (Se) is both a micronutrient required for most life and an element of environmental concern due to its toxicity in high concentrations. Se can be released into the environment through both natural and anthropogenic (human) activity, where it can exist as volatile or organic Se(-II), nanoparticulate Se(0), or aqueous Se(+IV/VI). Coal mining, processing, and burning can release high levels of Se to the environment, as selenium can easily substitute for sulfur, a main component of coal. Se is also useful in the medical field, where it has anticancer properties and Se(0) is an effective coating on medical devices. While most knowledge of biotic Se transformations is related to either anaerobic or aerobic bacterial processes, some common soil Ascomycota fungi can reduce Se under oxic conditions. These microeukaryotes readily transform elevated concentrations of this essential toxin from a bioavailable aqueous phase (Se(IV/VI)) to solid or volatile phases (Se(0/-II)), which is ideal for engineering efficient, cost-effective treatment strategies for Se-contaminated environments. Elucidating the geochemical and genetic mechanisms behind filamentous fungal Se transformation strategies will progress biotechnological applications for biogenic Se nanoparticles, and aid in a more complete understanding of Se biogeochemical cycling.Item Nutrient Transformations by Microorganisms for Novel Animal Feed Ingredients(2018-02) Barnharst, TannerThe world population will reach 9 billion people by 2050 or sooner and as a result, we must produce 70% more food than we currently are. This food challenge lends itself to many innovative solutions. In this thesis, the use of microorganisms to produce higher value animal feeds is examined. The current challenge in intensive aquaculture is to control the level of nutrient pollutants in the wastewater and provide sustainable sources of proteins for feed. A synthetic lichen type biofilm was developed to have the fungus Mucor indicus and the microalga Chlorella vulgaris grow together on a polymer matrix referred to as a “mycoalgae” biofilm. When the biofilm grows, it takes up phosphorus and nitrogen compounds and converts them to proteins and other cellular products. It cleans the water from nutrient pollutants as the algae are attached to the biofilm leaving purified water at the end of the process. Under 25 mg L-1 total ammonia-N (TAN) conditions, the biofilm reduced TAN to undetectable limits within 48 h with over 69% of the TAN reduction taking place by 24 h. The biofilm reduced levels of phosphate-P from 15 mg to undetectable limits within 24 h. Under the same conditions, 860 mg of dry mycoalgae biomass was generated at the end of the process on 16 cm2 of mesh and 100 ml of culture media. This process allows for easy harvesting of the algae with no energy intensive process of separating the algae from the supernatant. The generated biofilm is composed of two organisms that have been shown to positively aid fish health when included as a feed supplement. Secondary Fermentation of Corn Ethanol Co-Products for improved Amino Acid Qualities In 2016, 5.28 billion bushels of corn were used to produce about 14.79 billion gallons of ethanol in the United States. As a result, about 36 million tons of Dried Distillers Grains with Solubles (DDGS) were manufactured and fed to livestock 1. DDGS are a common feed supplement in cattle rations as it is inexpensive and has positive feeding characteristics. One of the drawbacks of DDGS is that it is lacking in in key amino acids such as tryptophan, arginine, and lysine. Historically these amino acids have been supplemented by external addition of feed grade amino acids to rations. The research carried out attempts to fortify DDGS with higher amounts of key amino acids through secondary fermentation of Wet Distillers Grains by fungi. When cultured on WDG the fungi consumes carbohydrates, which are unavailable to livestock, and converts the carbohydrates to proteinaceous biomass, which serves to close the amino acid gap in corn ethanol co-products. The fungi used are Generally Regarded as Safe (GRAS) and have been used to produce feed ingredients historically. Because of the research, more sustainable forms of animal feed will be produced due to the improved feeding value of the co-productsItem Smart Bioremediation Technology to Achieve High Sulfate Reduction in Mining Waters of NE Minnesota - Phase 1 Report(University of Minnesota Duluth, 2015-06) Hendrickson, David W; Hanson, Jeffrey JThere exists a significant need in northeastern Minnesota to provide a viable solution to the current challenge of maintaining existing iron ore and developing non-ferrous mining industries while simultaneously protecting watersheds from elevated aqueous sulfate concentrations that could prove detrimental to biota, especially wild rice. ”Smart Bioremediation Technology to Achieve High Sulfate Reduction in Mining Waters of NE Minnesota – Phase I” focuses on proof of concept early-stage development of a realistic solution to the aqueous sulfate issue potentially threatening Minnesota’s existing $3 billion/year ferrous mining industry as well as Minnesota’s projected $4 billion/year non-ferrous mining industry (Skurla, 2012). Initial funding was provided for this Phase I work by the Natural Resources Research Institute (NRRI) and an Innovation Grant from the Laurentian Vision Partnership through the East Range Joint Powers Board. The design of the technology included smart sensors and controls which enabled remote operation and monitoring of the pilot scale system. Solar panels mounted on the systems floating bioreactor modules provided DC power to operate embedded pumps, sensors, controls, and data transmitters. The system was designed to enable stand alone, year round remote operation in environmental conditions encountered in either operating or legacy mining operations across the U.S. The modular nature of the technical design allows for practical scale up to accommodate flow requirement needs for the mining industry. The robust system design combined biological sulfate reduction with remediation hydrogeology approaches to remove sulfur from mining impacted waters (Reinsel, 2015). Sulfate reducing bacteria (SRB) from local stream sediments were utilized to provide the sulfate reduction. Preliminary analytical results indicate that the smart bioremediation technology is capable of producing aqueous sulfate reduction in the mining waters flowing through the bioreactor systems. The Phase I project has provided a proof of concept design for remediation of sulfate in mining impacted waters. Additional studies (Phase II study and MN Drive study) are currently under way and will be delivered during summer, 2016. The purpose of these future studies is to deliver a smart technology bioremediation water treatment system that is capable of being commercialized and that can effectively decrease aqueous sulfate levels in impacted waters in a cost-effective manner to concentrations that can be further decreased by other technologies so that stringent aqueous sulfate concentrations can be achieved.