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Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

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65 records · Page 4
All sourcesNASA NTRSDOE OSTI
DOE OSTI2022

Understanding the structure and composition of recalcitrant oligosaccharides in hydrolysate using high-throughput biotin-based glycome profiling and mass spectrometry

Novel Immunological and Mass Spectrometry Methods for Comprehensive Analysis of Recalcitrant Oligosaccharides in AFEX Pretreated Corn Stover. Lignocellulosic biomass is a sustainable alternative to fossil fuel and is extensively used for developing bio-based technologies to produce products such as food, feed, fuel, and chemicals. The key to these technologies is to develop cost competitive processes to convert complex carbohydrates present in plant cell wall to simple sugars such as glucose, xylose, and arabinose. Since lignocellulosic biomass is highly recalcitrant, it must undergo a combination of thermochemical treatment such as Ammonia Fiber Expansion (AFEX), dilute acid (DA), Ionic Liquid (IL) and biological treatment such as enzyme hydrolysis and microbial fermentation to produce desired products. However, when using commercial fungal enzymes during hydrolysis, only 75–85% of the soluble sugars generated are monomeric sugars, while the remaining 15–25% are soluble recalcitrant oligosaccharides that cannot be easily utilized by microorganisms. Previously, we successfully separated and purified the soluble recalcitrant oligosaccharides using a combination of charcoal and celite-based separation followed by size exclusion chromatography and studies their inhibitory properties on enzymes. We discovered that the oligosaccharides with higher degree of polymerization (DP) containing methylated uronic acid substitutions were more recalcitrant towards commercial enzyme mixtures than lower DP and neutral oligosaccharides. Here, we report the use of several complementary techniques that include glycome profiling using plant biomass glycan specific monoclonal antibodies (mAbs) to characterize sugar linkages in plant cell walls and enzymatic hydrolysate, matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) using structurally-informative diagnostic peaks offered by negative ion post-secondary decay spectra, gas chromatography followed by mass spectrometry (GC–MS) to characterize oligosaccharide sugar linkages with and without derivatization. Since oligosaccharides (DP 4–20) are small, it is challenging to mobilize these molecules for mAbs binding and characterization. To overcome this problem, we have applied a new biotin-coupling based oligosaccharide immobilization method that successfully tagged most of the low DP soluble oligosaccharides on to a micro-plate surface followed by specific linkage analysis using mAbs in a high-throughput system. This new approach will help develop more advanced versions of future high throughput glycome profiling methods that can be used to separate and characterize oligosaccharides present in biomarkers for diagnostic applications.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Enhanced 2'-Fucosyllactose production by engineered Saccharomyces cerevisiae using xylose as a co-substrate

2'-Fucosyllactose (2'-FL), a human milk oligosaccharide with confirmed benefits for infant health, is a promising infant formula ingredient. Although Escherichia coli, Saccharomyces cerevisiae, Corynebacterium glutamicum, and Bacillus subtilis have been engineered to produce 2'-FL, their titers and productivities need be improved for economic production. Glucose along with lactose have been used as substrates for producing 2'-FL, but accu- mulation of by-products due to overflow metabolism of glucose hampered efficient production of 2' -FL regardless of a host strain. To circumvent this problem, we used xylose, which is the second most abundant sugar in plant cell wall hydrolysates and is metabolized through oxidative metabolism, for the production of 2'-FL by engi- neered yeast. Specifically, we modified an engineered S. cerevisiae strain capable of assimilating xylose to pro- duce 2' -FL from a mixture of xylose and lactose. First, a lactose transporter (Lac12) from Kluyveromyces lactis was introduced. Second, a heterologous 2'-FL biosynthetic pathway consisting of enzymes Gmd, WcaG, and WbgL from Escherichia coli was introduced. Third, we adjusted expression levels of the heterologous genes to maximize 2'-FL production. The resulting engineered yeast produced 25.5 g/L of 2'-FL with a volumetric productivity of 0.35 g/L∙h in a fed-batch fermentation with lactose and xylose feeding to mitigate the glucose repression. Interestingly, the major location of produced 2'-FL by the engineered yeast can be changed using different culture media. While 72% of the produced 2'-FL was secreted when a complex medium was used, 82% of the produced 2'-FL remained inside the cells when a minimal medium was used. In conclusion, as yeast extract is already used as food and animal feed ingredients, 2'-FL enriched yeast extract can be produced cost-effectively using the 2'-FL-accumu- lating yeast cells.

2′ -fucosyllactose↗
DOE OSTI2020

Fast growing high-yield wheat and canola for efficient nutrient recycling systems

Wheat and canola are two major rotation crops in the U.S. each offering advantageous traits for the agricultural value chain. Canola, a dicot rotation crop, generally offers high seed oil content. Wheat, a monocot rotation crop, offers more cellulose and other nutritious carbohydrates. As rotation crops, traits such as rapid growth rates, increased harvest yields, and high quality (digestibility, deconstructability) are desirable for food, biorefinery, and also nutrient recycling for crop rotation. Improvement of such traits has been the goal of numerous biotechnology efforts. However, alternative biotechnology techniques are typically constitutive and when applied to commercial crops have resulted in net negative consequences. For example, efforts to improve quality by lowering lignin biosynthesis have resulted in plants with poor structural integrity and diminished mass at maturity. With supports by SBIR programs from DOE and USDA, JBEI/LBNL and AFINGEN previously demonstrated engineered healthy switchgrass and alfalfa which grew with 20-30% more fermentable sugar release, 25% less lignin, faster growth, and more biomass yields than control plants. In this CRADA project supported by NASA STTR program, AFINGEN again collaborated with JBEI/LBNL and transferred the simple tissue-targeting technology from the demonstrated switchgrass and alfalfa to the two major rotation crops wheat and canola for efficient nutrient recycling systems.

09 BIOMASS FUELS↗
DOE OSTI2024

Effect of Temperature and pH on Microbial Communities Fermenting a Dairy Coproduct Mixture

Organic-rich industrial residues can serve as renewable feedstocks for the generation of useful products by microbial fermentation. We investigated fermenting communities enriched in a mixture of ultra-filtered milk permeate (UFMP) and acid whey from cottage cheese (CAW), two dairy coproducts rich in lactose. To evaluate how operational pH and temperature affect microbial communities and fermentation products, we operated 12 bioreactors for 140 days, each fed a 1:1 mixture of UFMP and CAW at either 35 °C or 50 °C and at either a pH of 4.8 or 5.5. The bioreactors operated at a pH of 4.8 resulted in the incomplete conversion of lactose, while those operated at a pH of 5.5 consistently fermented lactose, primarily into lactic, acetic, and hexanoic acids. The metagenomic analyses revealed that microbial communities obtained at a pH of 5.5 were dominated by lactic acid-producing organisms. Additionally, an inverse relationship was found between the abundance of chain elongating organisms and lactic acid accumulation, with 50 °C reducing the abundance of these organisms and enhancing lactic acid yields. We conclude that the pH and temperature are important determinants of the fermentation of dairy coproducts with a pH of 5.5 and 50 °C yielding the most promising results for lactic acid production. Additional research is required to better understand the factors affecting functional consistency of the process.

Biotechnology & Applied Microbiology↗
DOE OSTI2026

Data for Rewiring Yeast Metabolism for Producing 2,3-Butanediol and Two Downstream Applications: Techno-Economic Analysis and Life Cycle Assessment of Methyl Ethyl Ketone (MEK) and Agricultural Biostimulant Production

Rising concerns for sustainability and global climate change have driven the development of sustainable production pathways for biofuels and chemicals from lignocellulosic biomass via integrated biological and chemical processes. We constructed an engineered Saccharomyces cerevisiae capable of producing 2,3-butanediol (2,3-BDO) from glucose without accumulating ethanol and glycerol, which hinder downstream processing of 2,3-BDO, through extensive metabolic reprogramming. Specifically, we introduced heterologous 2,3-BDO biosynthetic enzymes and deleted the major isozymes of ethanol and glycerol biosynthetic enzymes. In addition, we introduced an NAD+ regenerating Pyruvate-Malate (PM) cycle and enhanced the NAD+ regenerating capability of the PM cycle to resolve the redox imbalance from the deletion of ethanol and glycerol production pathways. The resulting engineered yeast produced 109.9 g/L of 2,3-BDO with a productivity of 1.0 g/L/h and a yield of 0.36 g/g glucose in a fed-batch fermentation. We also conducted techno-economic analysis (TEA) and life cycle assessment (LCA) of the production of methyl ethyl ketone (MEK) through catalytic dehydration of 2,3-BDO. A TEA based on the experimental results indicated that the minimum product selling price (MPSP) was estimated to be $1.90/kg. Regarding cradle-to-grave LCA, 100-year global warming potential (GWP100) and fossil energy consumption (FEC) were found to be 0.37 kg CO2 eq/kg and 3.1 MJ/kg, respectively. These results demonstrated the feasibility of cost-competitive and sustainable bio-based MEK production via yeast fermentation. In addition, we explored the possibility of using the fermentation broth containing 2,3-BDO as a biostimulant inducing drought tolerance in plants. As a result, the yeast 2,3-BDO fermentation broth can induce drought tolerance in Arabidopsis thaliana without a complicated purification process.

Economics↗
DOE OSTI2024

Soil application of high-lignin fermentation byproduct to increase the sustainability of liquid biofuel production from crop residues

When digestates from anaerobic digestion of crop residues are added to soil, a considerable body of information indicates that soil organic carbon (SOC) levels are comparable to those when crop residues are left in the field. This occurs although the amount of digestate added to soil is diminished by digestion and implies that digestion increases the proportion of carbon inputs stabilized as SOC. Here we examine the likelihood and implications of these features being manifested for soil application of high lignin-fermentation byproduct (HLFB) from liquid biofuel production. We show that steady-state SOC levels are much less sensitive to crop residue removal with HLFB return than without it, and provide an example supporting the feasibility of foregoing process energy and coproduct revenue when HLFB is returned to the soil. Informed by this review and analysis, we expect with moderate confidence that long-term SOC levels for soils amended with HLFB from some liquid cellulosic biofuel processes will not be substantially lower than those occurring when crop residues are left in the field. We have high confidence that the economically optimum rate of fertilizer nitrogen (N) application and N 2 O emissions will be lower at most sites for HLFB return to the soil than if crop residues were left in the field. We estimate that the per hectare N demand for processing crop residues to liquid biofuels is about a third of the per hectare demand for crop production, giving rise to an opportunity to use N twice and thereby realize cost savings and environmental benefits. These observations support but do not prove the hypothesis that a ‘win-win’ is possible wherein large amounts of liquid biofuel feedstock can be obtained from crop residues while improving the economics and sustainability of food and feed production. A research agenda aimed at exploring and testing this hypothesis is offered.

09 BIOMASS FUELS↗
DOE OSTI2020

Microalgae Biomass Production for Utilization of CO2 and Mitigation of Greenhouse Gas Emissions

Microalgal cultivation processes for production of foods, feeds, fuels, fertilizers and other bioproducts and wastewater treatment are being considered for reducing CO2 and other greenhouse gas emissions. Major differences between microalgae and higher plant biomass cultivation are include their potential for much higher productivities, their higher content of major (N-P-K) and minor nutrients, and the challenge of harvesting such microscopic plants. Most critical, microalgal mass cultures, unlike higher plants, currently require fertilization with concentrated sources of CO2. As practically all of the C fixed into algal biomass will be re-emitted into the atmosphere in short order, algae processes do not sequester carbon. Their potential for CO2 emissions reductions must thus be based on comparisons with current technologies for the production of competing products, such as biofuels or animal feeds, or in wastewater treatment. Further, due to economic limitations to flue gas transport (3 to 12 % CO2), as well as highly variable diurnal and seasonal CO2 utilization and limited land and water availability near most CO2 sources, only a small fraction of waste CO2 emissions will be directly utilizeable for microalgae biomass production. However, for long-distance transport, flue gas CO2 capture by chemical processes would greatly increase the costs of flue gas utilization. More concentrated, sources of CO2 from refineries, fertilizer, chemical plants, and fermentations are available, but are also very limited relative to the quantities required for commodity production. Microalgae can also be cultivated on organic wastes, which provide both nutrients (N, P, K, etc.) and organic and inorganic carbon for algal growth, reducing greenhouse gas emissions compared to conventional treatment processes. To maximize the potential for algae biomass production and CO2 utilization, direct capture of CO2 from air will be required. This could be accomplished with the algal cultivation process itself in large open-raceway ponds, through chemical and biological enhancements of CO2 transfer into algal cutures. The relative productivities, economics, greenhouse gas balances, and resource potentials of these alternative CO2 sources for large-scale production of microalgae will be reviewed.

09 BIOMASS FUELS↗
DOE OSTI2023

Respiration is essential for aerobic growth of Zymomonas mobilis ZM4

ABSTRACT Zymomonas mobilis is an alpha-proteobacterium that is a promising platform for industrial scale production of biofuels due to its efficient ethanol fermentation and low biomass generation. Z. mobilis is aerotolerant and encodes a complete respiratory electron transport chain, but the benefit of respiration for growth in oxic conditions has never been confirmed, despite decades of research. Growth and ethanol production of wild-type Z. mobilis is poor in oxic conditions indicating that it does not benefit from oxidative phosphorylation. Additionally, in previous studies, aerobic growth improved significantly when respiratory genes were disrupted ( ndh ) or acquired point mutations ( cydA and cydB ), even if respiration was significantly reduced by these changes. Here, we obtained clean deletions of respiratory genes ndh and cydAB , individually and in combination, and showed, for the first time, that deletion of cydAB completely inhibited O 2 respiration and dramatically reduced growth in oxic conditions. Both respiration and aerobic growth were restored by expressing a heterologous, water-forming NADH oxidase, noxE . Oxygen can have many negative effects, including formation of reactive oxygen species (ROS) or directly inactivating oxygen sensitive enzymes. Our results suggest that the effect of molecular oxygen on enzymes had a greater negative impact on Z. mobilis than formation of ROS. This result shows that the main role of the electron transport chain in Z. mobilis is reducing the intracellular concentration of molecular oxygen, helping to explain why it is beneficial for Z. mobilis to use electron transport chain complexes that have little capacity to contribute to oxidative phosphorylation. IMPORTANCE A key to producing next-generation biofuels is to engineer microbes that efficiently convert non-food materials into drop-in fuels, and to engineer microbes effectively, we must understand their metabolism thoroughly. Zymomonas mobilis is a bacterium that is a promising candidate biofuel producer, but its metabolism remains poorly understood, especially its metabolism when exposed to oxygen. Although Z. mobilis respires with oxygen, its aerobic growth is poor, and disruption of genes related to respiration counterintuitively improves aerobic growth. This unusual result has sparked decades of research and debate regarding the function of respiration in Z. mobilis . Here, we used a new set of mutants to determine that respiration is essential for aerobic growth and likely protects the cells from damage caused by oxygen. We conclude that the respiratory pathway of Z. mobilis should not be deleted from chassis strains for industrial production because this would yield a strain that is intolerant of oxygen, which is more difficult to manage in industrial settings.

09 BIOMASS FUELS↗
DOE OSTI2023

Butyric Acid Production from Delignified Corn Stover Using Thermophilic Bacterial Co-Cultures

Butyric acid (BA) is a valuable platform chemical in the food and pharmaceutical industries, and it is also a potential precursor for the production of biobutanol and sustainable aviation fuels (SAFs). BA is mainly synthesized from petroleum; thus, cost-effective, and sustainable alternatives for its production are attracting the interest of several sectors. The present work proposes a solids-to-acids bioprocess to produce BA from corn stover utilizing a co-culture that consists of two thermophilic bacteria, Clostridium thermocellum, a well-known efficient degrader of insoluble and oligomeric cellulosic substrates, and Clostridium thermobutyricum, a highly efficient BA producer from monomeric sugars. After initial proof of concept experiments, a series of optimization studies were carried out to evaluate the process limits of this co-culture. First, the co-cultivation of both microorganisms at different inoculum sizes in deacetylated and mechanically refined corn stover (DMR) was evaluated. No significant differences were found on the solids deconstruction and carbohydrates utilization among all the treatments. In addition, BA production was similar under all conditions, ranging between 2.1 and 2.4 g/L. Next, the deconstruction, and BA production capabilities of the co-culture at increased DMR solids contents (3, 4.5, and 6% (w/v)) were tested. Although no difference was observed in the solids deconstruction and carbohydrates utilization among treatments, BA production increased concomitantly to solids loading; the maximum values observed were 2.8, 3.6, and 6.2 g/L in the 3, 4.5, and 6% treatments, respectively. In fact, BA production did not reach its absolute maximum in the 6% treatment after 160 h of fermentation. Lastly, a fed-batch experiment was performed to investigate the co-culture capabilities and possible system constraints to achieve higher BA titers. Data on substrate modifications and product formation as well as on the growth of each microorganism will be shown. Results from this work demonstrate a promising bioprocess approach for the production of BA from lignocellulosic biomass using a thermophilic bacterial co-culture.

bacteria↗
DOE OSTI2026

Data for Engineering and Evolution of Yarrowia lipolytica for Producing Lipids from Lignocellulosic Hydrolysates

Yarrowia lipolytica , an oleaginous yeast, shows promise for industrial fermentation due to its robust acetyl-CoA flux and well-developed genetic engineering tools. However, its lack of an active xylose metabolism restricts the conversion of cellulosic sugars to valuable products. To address this, metabolic engineering, and adaptive laboratory evolution (ALE) were applied to the Y. lipolytica PO1f strain, resulting in an efficient xylose-assimilating strain (XEV). Whole-genome sequencing (WGS) of the XEV followed by reverse engineering revealed that the amplification of the heterologous oxidoreductase pathway and a mutation in the GTPase-activating protein gene (YALI0B12100g) might be the primary reasons for improved xylose assimilation in the XEV strain. When a sorghum hydrolysate was used, the XEV strain showed superior xylose consumption and lipid production compared to its parental strain (X123). This study advances our understanding of xylose metabolism in Y. lipolytica and proposes effective metabolic engineering strategies for optimizing lignocellulosic hydrolysates.

Hydrolysate↗
DOE OSTI2025

A taxogenomic view of the genus Torulaspora : an expansion from ten to twenty-two species

The yeast genus Torulaspora (subphylum Saccharomycotina, family Saccharomycetaceae) is mostly known from its type species, T. delbrueckii, a frequent colonizer of wine and sourdough bread fermentations. The genus currently contains 10 species that are typically found in various natural terrestrial environments in temperate and tropical climates. Here we employ taxogenomic analyses to investigate a large collection of Torulaspora strains obtained in multiple surveys we carried out in Asia, Australasia, North America, South America, and Europe, and to which we added several strains maintained in culture collections. Our analyses detected twelve novel species that are formally described here, thereby more than doubling the species diversity of Torulaspora. We also sketch a genotype-phenotype map for the genus and show how the complex relationship between key genes and the physiological traits they control both between and within species. This remarkable increase in the number of species in the genus Torulaspora highlights how limited the current inventory of fungal taxa is. It also shows how integrated taxogenomic approaches can foster the assessment of species circumscriptions in fungi.

59 BASIC BIOLOGICAL SCIENCES↗