Browsing by Subject "Microalgae"
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Item Artificial Leaf for Biofuel Production and Harvesting: Transport Phenomena and Energy Conversion(2013-08) Murphy, Thomas Eugene; Berberoglu, HalilMicroalgae cultivation has received much research attention in recent decades due to its high photosynthetic productivity and ability to produce biofuel feedstocks as well as high value compounds for the health food, cosmetics, and agriculture markets. Microalgae are conventionally grown in open pond raceways or closed photobioreactors. Due to the high water contents of these cultivation systems, they require large energy inputs for pumping and mixing the dilute culture, as well as concentrating and dewatering the resultant biomass. The energy required to operate these systems is generally greater than the energy contained in the resultant biomass, which precludes their use in sustainable biofuel production. To address this challenge, we designed a novel photobioreactor inspired by higher plants. In this synthetic leaf system, a modified transpiration mechanism is used which delivers water and nutrients to photosynthetic cells that grow as a biofilm on a porous, wicking substrate. Nutrient medium flow through the reactor is driven by evaporation, thereby eliminating the need for a pump. This dissertation outlines the design, construction, operation, and modeling of such a synthetic leaf system for energy positive biofuel production. First, a scaled down synthetic leaf reactor was operated alongside a conventional stirred tank photobioreactor. It was demonstrated that the synthetic leaf system required only 4% the working water volume as the conventional reactor, and showed growth rates as high as four times that of the conventional reactor. However, inefficiencies in the synthetic leaf system were identified and attributed to light and nutrient limitation of growth in the biofilm. To address these issues, a modeling study was performed with the aim of balancing the fluxes of photons and nutrients in the synthetic leaf environment. The vascular nutrient medium transport system was also modeled, enabling calculation of nutrient delivery rates as a function of environmental parameters and material properties of the porous membrane. These models were validated using an experimental setup in which the nutrient delivery rate, growth rate, and photosynthetic yield were measured for single synthetic leaves. The synthetic leaf system was shown to be competitive with existing technologies in terms of biomass productivity, while requiring zero energy for nutrient and gas delivery to the microorganisms. Future studies should focus on utilizing the synthetic leaf system for passive harvesting of secreted products in addition to passive nutrient delivery.Item Bioprocessing of Microalgae for Bioenergy and Recombinant Protein Production(2013-07-31) Garzon Sanabria, Andrea JThis dissertation investigates harvesting of marine microalgae for bioenergy and production of two recombinant proteins for therapeutic applications in Chlamydomonas reinhardtii. The first study describes harvesting of marine microalgae by flocculation using aluminum chloride (AlCl_3), natural polymer chitosan, and synthetic cationic polymers. Harvesting and concentration process of low concentration microalgae cultures ranging from 1 to 2 g dry weight per liter was affected by algogenic organic matter (AOM), ionic strength, cell concentration, polymer charge density, and media pH. Marine microalgae flocculation was greatly affected by the presence of AOM independently of the flocculant chemistry. Presence of AOM demanded extra flocculant dosage i.e., 3-fold of AlCl3, 7-fold of highly charged synthetic cationic polymer, and 10-fold of chitosan. Flocculant dosage required for > 90 % flocculation efficiency in the presence of AOM was 160 mg/L, 50 mg/L, and 20 mg/L when using AlCl_3, chitosan, and best (more efficient) synthetic polymer respectively. The high-ionic strength of saline water did not have a significant effect on flocculation efficiency when using AlCl_3. However, to achieve efficient algal biomass removal, application of highly-charged synthetic polymers was required to overcome the presence of electrolytes. The best synthetic cationic polymer tested herein, which achieved greater than 90 % flocculation efficiency at 20 mg/L dosage, was a polymer with 99 % cationic charge density. Cell concentration also affected flocculant dosage requirement; low density cultures (10^6 cells/mL) required 6-fold greater dosages than cultures grown until early stationary phase (10^7 cells/mL). The second study addresses cultivation, extraction and purification challenges of two complex recombinant proteins, an immunotoxin molecule (MT51) and malaria vaccine antigen (Pfs25) produced in the chloroplast of C. reinhardtii. Main challenges identified were i) low transgene expression level, ii) proteolytic instability of MT51 immunotoxin, and iii) aggregation of Pfs25 antigen. Optimal expression and accumulation of Pfs25 antigen required growing C. reinhardtii cultures to late exponential phase (10^6 cells/mL) and inducing transgene expression for 24 h at a photon irradiance of 120 ?mol/m^2s.Item Developing Optimal Growth Parameters for the Green Microalgae Nannochloris oculata and the Diatom Nitzschia sp. for Large scale Raceway Production(2011-10-21) Luedecke, Phillip RyanMicroalgae produce large quantities of lipids that can be used for biofuel feedstock. The goal of this project was to determine the effect of several engineering and management parameters on the productivity of microalgae cultivated in large, outdoor facilities. The specific objectives were focused on the effects of inoculation ratios; the effects of light, temperature, and culture depth on growth; and the minimum circulation velocity necessary to maintain growth and minimize settling in open ponds. Microalgae must first be cultured in smaller quantities before the raceway is inoculated for optimized growth. Concentration ratios are defined as the ratio of the volume of microalgae inoculum to the volume of new growth media. The microalgae species used was Nannochloris oculata (UTEX #LB 1998). Inoculation ratios studied varied from 1:1 to 1:32 and were grown in 500 mL Erlenmeyer flasks. The study found that 1:16 and 1:32 were too dilute, while the 1:8 concentration had the largest growth rate. Determination of the effects of temperature, light intensity, and cultivation depth is critical to maintaining healthy cultures. Excess light intensity can result in photoinhibition and temperatures above the maximum growing tolerance can have detrimental effects. These factors can affect growth and evidence suggests an interaction that exacerbates these effects. In an outdoor culture there are few practical control variables other than pond depth. As cultivation depth increases, the algae undergo "selfshading" and the increased cultivation volume hinders temperature changes. Scaled raceway ponds were maintained at 10.16 cm (4 inch) and 13.97 cm (5.5 inch) depths. The species used was Nannochloris oculata and it was found to grow best at 785 micromol m?? s??m^-2 s^-1, 20?C, and 10.16 cm. Diatoms are attractive because of high growth rates, faster lipid production, and greater cell density. The latter promotes rapid settling once mixing has stopped. Because of the silica cell wall composition, diatoms are believed to be more susceptible to shear forces which can result in lysis. Determining the natural settling rate to the minimum channel velocity relationship in cultivation ponds was the objective. No flocculants/coagulants were added which created a case of "natural" settling. Four pennate Nitzschia sp. and one centric diatom were tested in a jar tester. There was no significant difference in settling times between the species. The mean settling time was 4.55 cm min?? and the minimum channel velocity was determined to be 10.12 cm min??.Item The effect of 2E,4E-decadienal on lipid-related gene expression in Phaeodactylum tricornutum(2013-08) Beck, Emily Christine; Mehdy, Mona Cynthia, 1955-Microalgae have been proposed as a potential feedstock for biofuel production, and as a result, interest in the biology of these organisms has intensified. These organisms also synthesize lipids that are vital to human health and nutrition. Stress has been shown to have an effect on lipid composition and gene expression in microalgae, but many studies have focused on the effects of abiotic stressors. The purpose of this study was to investigate the effect of biotic stress on lipid-related gene expression in Phaeodactylum tricornutum, a model species of microalgae. The source of biotic stress used in this study was 2E,4E/Z-decadienal, a diatom-derived oxylipin that has been shown to function as a stress signal among diatoms. Real-time RT-qPCR analysis revealed that expression of a patatin-like phospholipase was significantly decreased in decadienal-treated cultures as compared to a solvent control. The expression of a delta-9 desaturase gene believed to be responsible for production of 16:1 fatty acids was increased by a factor of 12. FabI, a gene involved in fatty acid biosynthesis, and PtD5a, which codes for an ER-localized desaturase, were both down-regulated in cells exposed to decadienal. However, changes in expression were only shown to be significant for the patatin-like phospholipase gene. Increased expression of the delta-9 desaturase gene may be a protective mechanism against infection from pathogens, since 16:1 fatty acids have been shown to have antibacterial properties. Regulation of membrane desaturation may also serve to stabilize photosynthetic membranes during times of stress. The down-regulation of the phospholipase gene was surprising, since the release of fatty acids from membrane lipids for oxylipin production is a common response to stress. It is recommended that this experiment be improved upon and expanded in order to determine whether the results obtained are reproducible and how these changes in gene expression correlate with physiological effects.Item Effect of temperature, dissolved inorganic carbon and light intensity on the growth rates of two microalgae species in monocultures and co-cultures(2014-05) Almada-Calvo, Fernando; Kinney, Kerry A.; Katz, Lynn E.The enormous biodiversity of microalgae as well as their high photosynthetic rates can be exploited for a wide variety of applications including the production of high value chemicals, nutraceuticals, aquaculture feed, and most recently, biofuels. Regardless of the application, the productivity of the microalgae culture must be optimized in order to make the systems economically feasible. One environmental factor that greatly affects the productivity of mass cultivation systems is temperature since it can be prohibitively expensive to control in outdoor systems. Temperature affects microalgae growth rates both directly by its effect on metabolic rates, and indirectly, by changing the bioavailability of the inorganic carbon present in solution. In the first part of this research, the effects of dissolved inorganic carbon (DIC) concentration (varied by sparging CO₂-enriched air) and temperature on the growth of a model microalga species (Nannochloris sp., UTEX LB1999) were investigated in a turbidostat bioreactor. The results indicate that increasing DIC concentration yields higher microalgae growth rates up to an optimum value (around 3 mM for Nannochloris sp.) but higher concentrations actually inhibited growth. Since increasing the temperature decreases the DIC concentration for a given gas pCO₂, it is necessary to adjust the pCO₂ to maintain the target DIC concentration in the optimal range for growth. In the next phase of the research, the effect of average light intensity (Gav) and temperature on the growth rate of two microalgae species (Nannochloris sp., UTEX1999. and Phaeodactylum tricornutum, UTEX646) was investigated. Growth rates were measured over a range of average light intensities and temperatures using a turbidostat bioreactor. A multiplicative model was developed to describe growth as a function of both average light intensity and temperature. In the third phase of this research, both microalgae species were grown together to explore the effects of temperature fluctuations on the population dynamics of the co-culture. It was observed that Nannochloris was inhibited by the presence of P. tricornutum in the medium, probably due to the excretion of secondary metabolites into the medium that affected Nannochloris growth (allelopathic effects). The temperature and average light intensity model developed under monoculture conditions was modified to incorporate the allelopathic effects observed. The resulting model provided a reasonable fit to the dynamic behavior of a Nannochloris/P. tricornutum co-culture subjected to temperature variations in chemostat experiments.Item High-throughput Microfluidic Screening Platforms for Microalgae Study(2014-12-15) Kim, Hyun SooMicroalgae have been envisioned as a future source of renewable energy. Both fossil fuel depletion and environmental concern have drawn more interest in microalgal biofuels, but the production cost of these biofuels are not yet economically competitive. Significant improvements such as development of better performing microalgal strains, optimization of culture conditions, and better understanding of microalgal biology are required for commercial viability. To resolve these limitations, massively parallel studies are needed, however, current microalgae culture systems are lack of high-throughput screening capabilities, and thus not suitable for the parallel studies. Here, three different high-throughput microfluidic microalgae screening platforms have been developed, each of which addresses major bottlenecks towards economically feasible microalgal biofuel. The first platform, a high-throughput microfluidic photobioreactor array has been developed to investigate the effect of different culture conditions on microalgal growth and oil production. This platform can provide up to 64 different culture conditions on-chip, such as combinations of different light intensities, light cycles, and culture media/chemical compositions. Single cell/colony trapping sites in culture compartments allowed for long-term analysis of microalgal growth and oil production with single cell/colony resolution. The light conditions that induced 1.8-fold higher oil accumulation over the typically used culture conditions were successfully identified. The second platform, as a microalgae library screening tool, a high-throughput microfluidic single-cell screening and selection platform has been developed to examine growth and oil production of various microalgal strains, followed by selective extraction of particular microalgae showing desired traits to off-chip reservoirs for further analysis. Single microalga was isolated and cultured, and its growth and oil accumulation were analyzed through 1024 single-cell trapping/culturing sites in the platform, where opening and closing of each trap can be individually controlled with integrated microfluidic control layers. By opening only a specific site out of the 1024 trapping sites, microalgae in particular trapping sites were selectively released and successfully collected off-chip. The third platform, a high-throughput droplet microfluidics-based microalgae screening platform has been developed to investigate the growth and the oil production of microalgal libraries with much higher throughput. Growth was characterized by encapsulating a single microalga into a droplet (functions as an independent bioreactor) and tracking its behavior over time. Oil production was also quantified through on-chip staining process, the key feature of the platform, where oil content in microalgae can be stained and measured through on-chip fluorescent tagging. Growth and oil accumulation under different culture conditions were successfully analyzed and compared, demonstrating the capability of the platform as a high-throughput screening tool. We have developed series of high-throughput microfluidic screening platforms for microalgae study, which provides the capabilities of analyzing microalgal growth and oil production under different culture conditions or among large numbers of microalgal library. The developed platforms will serve as powerful tools to accelerate research in addressing the limitations of microalgal biofuels as well as to significantly advance the current state of microalgal biofuel production.Item Nutritional Contribution of Phytoplankton to the Pacific White Shrimp Litopenaeus vannamei(2012-07-16) Sanchez Corrales, Dagoberto RaulThe goal of this study was to characterize the nutritional contribution of microalgae to white-legged shrimp and optimize fish meal (FM) and fish oil (FO) inclusion levels in their diets in the presence of microalgae. Phytoplankton composition was first determined in a typical Peruvian intensive commercial shrimp farm and in a semi-closed greenhouse-covered reservoir. A predominance of 76.3% cyanobacteria was observed for most of 9 months in all shrimp ponds. However, with the fertilization program in a reservoir tank, 60.7% diatoms and 22.8% cyanobacteria predominated. Thus, with the imposed fertilization regimen, the microalgae composition was manipulated to be different than that in commercial shrimp ponds. The microalgae composition was then evaluated along with different dietary levels of FM and squid meal (SM) in a feeding trial to evaluate the potential of phytoplankton to reduce FM and SM levels in shrimp feeds. Six diets were formulated to contain either 5, 10 or 20% SM combined with either 6.5 or 12% FM. Dietary effects on growth and survival were compared in a "clear-water system" (CWS) and a "green-water system" (GWS). Results suggest that 6.5% FM and 5% SM can be used as a cost-effective combination in feeds for shrimp. The effects of different dietary levels of FO and soybean lecithin (LT) on shrimp growth in CWS and GWS were evaluated in another feeding trial to determine if dietary phospholipids and phytoplankton increase the availability of essential fatty acids (EFAs) to shrimp. Six diets were formulated to contain 1, 2 or 3% FO combined with either 1 or 4% LT. Shrimp fed diets containing 1% LT and 1% FO in both systems had significantly lower weight gain and higher feed conversion ratio. Cephalothorax lipids and phospholipids were higher in shrimp fed diets containing 4% LT. Inclusion of 4% LT increased the availability of EFAs, and could contribute to reduce the FO in shrimp diets. The contribution of phytoplankton to shrimp weight gain, varied from 38.8 to 60.6%. This study demonstrated that cost-effective diets could be formulated with reduced inclusion levels of FM and FO considering the contribution of microalgae to the nutrition of shrimp.Item pH-induced flocculation/deflocculation process for harvesting microalgae from water(2014-08) Choi, Jin-Yong, Ph. D.; Kinney, Kerry A.; Katz, Lynn Ellen; Kinney, Kerry A.; Katz, Lynn E.Historically, the presence of microalgae (algae hereafter) in natural waters has been viewed as a nuisance due to its adverse impact on water quality. More recently, however, algae are being investigated as potential sources of biofuel and a range of natural products. These applications require the development of large-scale cultivation systems for mass production that include growth, harvesting, concentration, and product recovery components. While challenges still remain with respect to many of the processes involved in mass production, one of the most technically and economically challenging steps is harvesting the algae from dilute growth cultures, especially in systems where chemical additives are of concern either within the algae concentrate or the effluent water. For this reason, a pH-induced flocculation/deflocculation method using the hydroxides of alkali or alkaline earth metals (e.g., lime, caustic soda) is of particular interest for algae harvesting as Na, Ca and Mg are typically present in natural waters. The goal of this research was to determine the underlying mechanisms responsible for algae coagulation by magnesium and calcium and to evaluate the potential of these mechanisms for harvesting algae for a range of synthetic and field source water chemistries. In the first two phases of this research, the mechanisms for coagulation with magnesium and calcium were studied independently. A series of bench-scale experiments were designed to isolate the potential mechanisms of algae destabilization associated with each of these cations as a function of water chemistry, and microscopic analyses were performed to characterize the flocculated algae/precipitate mixtures. In the third phase of this research, removal of algae in field source waters was evaluated with respect to the underlying science elucidated in the previous phases. The results indicate that the dominant algae destabilization mechanism associated with magnesium shifts from Mg adsorption/charge neutralization to Mg(OH)₂[subscript (S)] precipitation-enhanced coagulation with increasing pH. Moreover, dissolved Mg²⁺ adsorption to the algae surface led to effective algae coagulation, while minimizing the mass of precipitated Mg(OH)₂[subscript (S)] . For Ca, this research identified the importance of the nucleation process (heterogeneous vs. homogeneous nucleation) on algae removal efficiency. Heterogeneous nucleation is a key factor for optimizing algae removal; thus, the degree of oversaturation with respect to CaCO₃[subscript (S)] is a crucial operating parameter. This research demonstrated that the algae harvesting process using pH-induced flocculation/deflocculation method can be optimized for a wide range of source waters if the water chemistry (e.g. pH, ion concentration, alkalinity, ionic strength) is properly incorporated into the system design.Item Technical Feasibility Study on Biofuels Production from Pyrolysis of Nannochloropsis oculata and Algal Bio-oil Upgrading(2013-12-02) Maguyon, MonetIncreasing environmental concerns over greenhouse gas emissions, depleting petroleum reserves and rising oil prices has stimulated interest on biofuels production from biomass sources. This study explored on biofuels production from pyrolysis of Nannochloropsis oculata and subsequent bio-oil upgrading by fractional distillation and zeolite upgrading. The extent of producing biofuels from N. oculata was initially investigated at various pyrolysis temperatures (400, 500 and 600^(0)C) at 100 psig. The distribution of the products significantly varied with temperature. Maximum char and gas yields were achieved at 400^(0)C (52% wt) and 600^(0)C (15% wt), respectively. Liquid production (aqueous and bio-oil) peaked at 500^(0)C (35% wt). The effect of temperature was also tested against product compositions and properties. Mass and energy conversion efficiencies were also estimated to be about 76% and 68%, respectively. The operating condition for maximum bio-oil production from N. oculata pyrolysis was subsequently determined. Optimum yield was achieved at 5400C and 0 psig where liquid yield was about 43% wt (23% wt bio-oil, 20% wt aqueous) while char and gas yields were about 32 and 12% wt, respectively. The bio-oil obtained has high carbon (72% wt) and hydrogen (10% wt) contents and high energy content (36 MJ/kg). Char and gas also contain considerable energy contents of about 20 MJ/kg and 21MJ/m3, respectively. Separation of the bio-oil and aqueous liquid product (ALP) components by fractional distillation was then investigated. Heavy distillates has the highest yield (75% wt), followed by light distillates (19% wt). Significant reduction in moisture contents and increase in heating values were observed in the bio-oil distillates. The ALP distillate obtained at 150-1800C was found to contain considerable amounts of acids, esters, amides and lactams and has heating value of about 24 MJ/kg. Finally, HZSM-5 upgrading was done at various temperatures and reaction times. Reaction temperature greatly affected product yields and upgraded bio-oil composition. The best operating condition was found to be 285^(0)C for 3.5 h, which can produce treated bio-oil with higher hydrocarbons (86%), lower oxygenated (3%) and lower nitrogenous (11%) components. Higher heating value (40 MJ/kg), high carbon (80% wt), and low oxygen (3% wt) contents were also achieved.