04 Fakultät Energie-, Verfahrens- und Biotechnik
Permanent URI for this collectionhttps://elib.uni-stuttgart.de/handle/11682/5
Browse
606 results
Search Results
Item Open Access Construction of robust Escherichia coli strains for large-scale production(2022) Ziegler, Martin; Takors, Ralf (Prof. Dr.-Ing.)The biotechnical production of many fine chemicals, proteins or pharmaceuticals depends on large-scale microbial cultivations. Due to limited mixing, heterogeneities in process relevant parameters such as nutrient concentrations arise in such fermentations. Escherichia coli (E. coli) is a model organism frequently used in the biotechnological industry. If E. coli is cultivated under heterogeneous conditions, biological reactions of the microorganism result in reduced process performance. Since large-scale fermentations are not economically feasible in academic settings, scale-down reactors that mimic aforementioned heterogeneities are used to investigate heterogenous fermentations. Previous studies in scale-down reactors unraveled that, depending on the process strategy, the unstable supply of a limiting primary carbon or nitrogen source such as glucose or ammonium is one of the underlying causes of process performance loss. Low concentrations of glucose or ammonium elicit the stringent response as a biological starvation reaction which comprises extensive transcriptional reactions. In the first project that contributes to this thesis, the regulatory and transcriptional reactions of the strains E. coli MG1655 and E. coli SR to repeated exposure to ammonium starvation zones were examined in a scale-down reactor. The scale-down reactor followed a two-compartment approach and consisted of a stirred tank reactor and a plug-flow reactor simulating passage through a starvation zone. E. coli SR is a strain with modulated stringent response. It was observed that short-term starvation stimuli do not trigger this regulatory program in E. coli SR and the transcriptional reaction was noticeably reduced. Long-term adaptation of the strain to repeated cycles of limitation and starvation also clearly differed from E. coli MG1655. Despite lack of the stringent response, E. coli SR showed no deficits in the assimilation of the limiting ammonium or in biomass yield on ammonium. In the second project of this thesis, a series of deletion strains with robust phenotype against glucose starvation zones were constructed. Candidate genes were identified and successively removed from the genome of E. coli MG1655 by Recombineering. The fundamental growth parameters of the strains were determined in shaking flask fermentations and no noticeable differences compared to E. coli MG1655 were found. Chemostat cultivations in a scale-down reactor with glucose as the limiting nutrient source revealed that the final strain of the deletion series, E. coli RM214, had a significantly lower maintenance coefficient under heterogeneous conditions than E. coli MG1655. Moreover, in an exemplary heterologous protein productionscenario E. coli RM214 rhaB- pJOE4056.2_tetA proved to be more robust to heterogeneities and showed a significantly higher product yield than E. coli MG1655 rhaB- pJOE4056.2_tetA. In the third project of this thesis, the production of pyruvate in E. coli MG1655 by inhibition of pyruvate dehydrogenase through CRISPR interference was investigated. A central goal was to achieve the stable production in nitrogen-limited conditions. For this, different target sequences in the operon pdhR-aceEF-lpd were tested and the strains cultivated in shaking flask fermentations. All tested target sequences were generally suitable to trigger the accumulation of pyruvate. Combined CRISPR interference against two target sequences did not lead to an increased pyruvate yield in most cases. In addition, the strains E. coli MG1655 pdCas9 psgRNA_aceE_234 and E. coli MG1655 pdCas9 psgRNA_aceE_234_pdhR_329 were characterized in two phase fermentations in lab-scale reactors. The initial phase was an unlimited exponential growth phase and was followed by an ammonium-limited production phase. E. coli MG1655 pdCas9 psgRNA_aceE_234 only produced pyruvate during the exponential phase, and reuptake of pyruvate occurred in the second phase. In contrast, E. coli MG1655 pdCas9 psgRNA_aceE_234_pdhR_329 stably produced pyruvate during the exponential and the ammonium-limited phase and is a potential chassis strain for the growth-decoupled production of pyruvate derived bioproducts. The overarching research issues of the projects were the characterization of strains in heterogeneous conditions and the development of new strategies to improve their performance. The collected data leads me to conclude that the construction of robust microbial strains for large-scale applications is both expedient and feasible. Tailored genetic modifications are the method of choice to achieve this goal. Furthermore, suitable genetic constructs offer promising possibilities for the stable growth-decoupled production of chemicals in nitrogen-limited conditions.Item Open Access Messung und Modellierung der effektiven Wärmeleitfähigkeit von Dämmstoffschüttungen für vakuumwärmegedämmte Warmwasserspeicher(2022) Lang, Stephan; Spindler, Klaus (apl. Prof. Dr.-Ing.)Das Ziel dieser Arbeit ist es, hinsichtlich Dämmwirkung und Kosten geeignete schüttfähige Wärmedämmstoffe für doppelwandige Warmwasserspeicher mit Vakuumwärmedämmung zu identifizieren und eine Vorhersage über die effektive Wärmeleitfähigkeit in Abhängigkeit von Luftdruck und Temperatur treffen zu können. Zu diesem Zweck werden expandierte Perlite unterschiedlicher mittlerer Korngrößen und Mischungen aus expandierten Perliten und pyrogener Kieselsäure untersucht. Schwerpunkt der Untersuchungen ist die Ermittlung der effektiven Wärmeleitfähigkeit bei unterschiedlichen Luftdrücken und Temperaturen. Die effektive Wärmeleitfähigkeit wird in einer eigens entwickelten Versuchsanlage, nach einem stationären Messprinzip, bei Luftdrücken zwischen 0,001 mbar und Atmosphärendruck von (960 ± 20) mbar sowie bei Probenmitteltemperaturen zwischen -5 °C und 90 °C bestimmt. Die maximale relative Messunsicherheit der Versuchsanlage beträgt < 8 % bei den geringsten und < 3 % bei den höchsten gemessenen effektiven Wärmeleitfähigkeiten. Reine feinkörnige expandierte Perlite mit Dichten der Schüttungen ≥ 182 kg/m³ erreichen bei Luftdrücken ≤ 0,1 mbar und allen gemessenen Probenmitteltemperaturen die geringsten effektiven Wärmeleitfähigkeiten. Bei einer Probenmitteltemperatur von 48 °C liegen diese bei ≤ 4,84 mW/(m·K). Mischungen aus einem vergleichsweise grobkörnigen expandierten Perlit mit einer sehr feinkörnigen und feinporigen pyrogenen Kieselsäure können hingegen, bei technisch einfacher zu handhabenden Luftdrücken von > 1 mbar, z. T. geringere effektive Wärmeleitfähigkeiten erreichen als reine expandierte Perlite. Mischungen dieser Komponenten werden in dieser Arbeit erstmals in Form einer losen Schüttung bzgl. ihrer effektiven Wärmeleitfähigkeit untersucht. Bei einer Probenmitteltemperatur von 48 °C werden in diesem Luftdruckbereich effektive Wärmeleitfähigkeiten dieser Mischungen von > 12,17 mW/(m·K) gemessen. Anhand der gemessenen effektiven Wärmeleitfähigkeiten sowie weiterer Stoffeigenschaften, werden vollständig prädiktive analytische Modelle der effektiven Wärmeleitfähigkeit entwickelt. Das Modell für Mischungen gilt für ein ausgewähltes Stoffpaar, während das Modell für expandierte Perlite für beliebige ungemahlene expandierte Perlite verwendbar ist. Es handelt sich nach Kenntnis des Autors um das erste vollständig prädiktive analytische Modell der effektiven Wärmeleitfähigkeit für expandierte Perlite, welches auch die Kopplung von Gas- und Festkörperwärmeleitung berücksichtigt und für welches lediglich drei einfach und kostengünstig zu messende Größen bestimmt werden müssen. Diese Größen sind der volumengewichtet gemittelte Korndurchmesser, die mittlere Korndichte sowie die Dichte der Schüttung des expandierten Perlits. Aus den Messwerten der effektiven Wärmeleitfähigkeit wird ein Zusammenhang von volumengewichtet gemitteltem Korndurchmesser zur Kopplung von Gas- und Festkörperwärmeleitung deutlich, der für das Modell für expandierte Perlite verwendet wird. Mit den Modellen ist es nun möglich, ohne entsprechende Messungen, die effektive Wärmeleitfähigkeit von Mischungen aus einem expandierten Perlit und einer pyrogenen Kieselsäure sowie für beliebige ungemahlene expandierte Perlite, mit zufriedenstellender bis hoher Genauigkeit vorherzusagen.Item Open Access Development of novel bispecific antibodies for cancer therapy targeting the receptor tyrosine kinases HER4 and EGFR(2024) Kühl, Lennart; Kontermann, Roland E. (Prof. Dr.)In this study, novel mono- and bispecific antibodies targeting the ErbB receptor family members EGFR and HER4 were investigated. Dual targeting of EGFR and HER4 by a bispecific, tetravalent antibody comprising a novel, antagonistic HER4-targeting antibody showed inhibition of proliferation and migration for a HB-EGF-stimulated ovarian cancer cell line. No inhibitory effects in a breast cancer cell line expressing EGFR and HER4 indicated that successful dual targeting does not solely rely on target expression. The complexity of HER4 with its isoforms and their different signaling properties makes HER4 a challenging cancer target that needs further in-depth research. To overcome resistances based on escape mutations located in the epitopes of clinically approved antibodies, novel antagonistic EGFR-targeting antibodies binding to a different epitope were developed. This epitope was mapped to domain III of EGFR and binding to clinically relevant EGFR ectodomain mutations resulted in inhibition of EGFR signaling in stable cell lines used as test systems. Favorable activities in comparison to clinically approved antibodies regarding inhibition of EGFR signaling and proliferation were observed for cancer cell lines expressing the EGFR wildtype. Bispecific T-cell engagers can lead to a T-cell mediated target cell killing independent of intracellular downstream signaling in the cancer cell. One challenge for the applicability of T-cell engagers in solid tumors is to keep the balance between T-cell mediated tumor cell killing and severe side-effects caused by a systemic activation of the immune system. Studies on eleven different eIg-based formats for EGFR-binding T-cell engagers showed that valency, geometry, and size influenced their activity profile. Furthermore, one bivalent and one trivalent, bispecific format were investigated for two novel EGFR-targeting moieties. As these molecules bind to clinically relevant escape mutations located in the ectodomain of EGFR, they are expected to show activity in patients with an acquired resistance to approved EGFR-targeting antibodies. These molecules led to a robust T-cell mediated cytotoxicity of cancer cells expressing EGFR. Additionally, benefits regarding an EGFR-level dependent cytotoxicity were observed for reduced binding to EGFR. An initial in vivo study using surrogate molecules in a syngeneic mouse model showed reduction of tumor growth and prolonged survival for treatment with a trivalent, bispecific T-cell engager comprising a novel EGFR-binding moiety. Taken together, beneficial effects of the novel molecules may contribute to improved therapies for patients with both pre-existing and acquired resistances to EGFR-targeting antibodies.Item Open Access Enhanced processing of regrind as recycling material in single-screw extruders(2021) Thieleke, Philipp; Bonten, ChristianRegrind processing poses challenges for single-screw extruders due to the irregularly shaped particles. For grooved feed zones, the output is lessened by the reduction of bulk density in comparison to virgin material. Simultaneously, the melt temperature increases, reducing the extruder’s process window. Through experimental investigations on a test stand, a novel feed zone geometry (nominal diameter 35 mm) is developed. It aligns the regrind’s specific throughput with that of virgin material. The regrind processing window is essentially increased. As the solids conveying in the novel feed zone cannot be simulated with existing methods, numerical simulations using the discrete element method are performed. Since plastic deformation occurs in the novel feed zone geometry, a new hysteresis contact model is developed. In addition to spheres, the regrind and virgin particles are modeled as superquadrics to better approximate the irregular shape. The new contact model’s simulation results show excellent agreement with experimental compression tests. The throughput of the extruder simulations is considerably underestimated when using spheres to represent the real particles than when using irregularly shaped superquadrics. Corresponding advantages can be seen especially for virgin material.Item Open Access Global potentials and costs of synfuels via Fischer-Tropsch process(2023) Buchenberg, Patrick; Addanki, Thushara; Franzmann, David; Winkler, Christoph; Lippkau, Felix; Hamacher, Thomas; Kuhn, Philipp; Heinrichs, Heidi; Blesl, MarkusThis paper presents the potentials and costs of synthetic fuels (synfuels) produced by renewable energy via PEM water electrolysis and the subsequent Fischer-Tropsch process for the years 2020, 2030, 2040, and 2050 in selected countries across the globe. The renewable energy potential was determined by the open-source tool pyGRETA and includes photovoltaic, onshore wind, and biomass. Carbon dioxide is obtained from biomass and the atmosphere by direct air capture. The potentials and costs were determined by aggregating minimal cost energy systems for each location on a state level. Each linear energy system was modelled and optimised by the optimisation framework urbs. The analysis focused on decentralised and off-grid synthetic fuels’ production. The transportation costs were roughly estimated based on the distance to the nearest maritime port for export. The distribution infrastructure was not considered since the already-existing infrastructure for fossil fuels can be easily adopted. The results showed that large amounts of synthetic fuels are available for EUR 110/MWh (USD 203/bbl) mainly in Africa, Central and South America, as well as Australia for 2050. This corresponds to a cost reduction of more than half compared to EUR 250/MWh (USD 461/bbl) in 2020. The synfuels’ potentials follow the photovoltaic potentials because of the corresponding low levelised cost of electricity. Batteries are in particular used for photovoltaic-dominant locations, and transportation costs are low compared to production costs.Item Open Access Findings from measurements of the electric power demand of air compressors(2021) Hummel, Ulf; Radgen, Peter; Ülker, Sercan; Schelle, RalphThe compressed air electric ratio (CAER) describes the ratio of the real electric power demand to the nominal mechanical power of an air compressor. The CAER is an important indicator as the electric power demand of air compressors varies throughout its operation dependent on compressor technology, pressure ratio, and free air delivery. The nameplate power of the compressor drive motor is not sufficient for evaluating the electric power demand; therefore, the CAER plays an important role in assessing the electric operating power demand. In this paper, results from measurements of fixed speed and variable speed (VFD) compressors are presented with the analysis of key influencing factors of the CAER. The data show that the pressure ratio of operating pressure to the maximum design outlet pressure has the largest impact on the CAER. For VFD compressors, the CAER is represented as a linear function dependent on the respective load. Fixed and variable speed compressors’ CAERs are always dependent on the load condition. In idle condition, the CAER was measured to be 0.2. In full load condition with a pressure ratio of 0.6, the CAER averages at a value of 0.87, meaning a 90 kW compressor at 0.6 pressure ratio draws 78.3 kW electric power.Item Open Access Membrane electrode assembly for water electrolysis(2023) Nguyen, Thi Hai Van; Friedrich, K. Andreas (Prof. Dr. rer. nat.)Maintaining a sufficient energy supply while minimizing the impact on the environment and climate is one of the greatest social and scientific challenges of our times. There are various fields of research and technological developments in the context of global warming and limitless growing energy demand, and the focus of this PhD programme is on artificial photosynthesis, more specifically on the assembly of Membrane electrode assembly for water electrolyzer part. Mimicking photosynthesis in a scheme to trap solar energy in chemical bonds (fuels) is a scientific and technological challenge. Having a cost-effective and reliable process stays one of the main limitations in order to achieving the long-term goal of this approach. In this work, within the European eSCALED project, the elaboration of Membrane Electrode Assembly (MEA) for water electrolysis by introducing new materials and low-cost fabrication methods was investigated.Item Open Access Multi-criteria comparison of energy and environmental assessment approaches for the example of cooling towers(2022) Wenzel, Paula M.; Radgen, PeterCooling towers remove economically or technically unusable heat using considerable amounts of electricity and, in many cases, water. Several approaches, which vary in methodology, scope, and level of detail, are used for environmental evaluations of these cooling systems. Although the chosen approach has a significant impact on decisions made at the plant level, no methodology has yet been standardized for selecting the approach that best serves the objectives of the evaluation. Thus, this paper provides comparison criteria for the systematic selection of suitable evaluation methods for cooling towers and classifies how the methods score in this respect. These criteria, such as ‘life cycle thinking’, ‘inventoried physical quantities’, ‘temporal resolution’, ‘formalization’, and ‘data availability’, are grouped by overall evaluation objectives such as ‘thoroughness’, ‘scientific soundness’, and ‘usability’. Subsequently, these criteria were used to compare material flow analysis, energy analysis, environmental network analysis, life cycle inventory, life cycle assessment, environmental footprint methods, emergy analysis, exergy analysis, and the physical optimum method. In conclusion, material flow analysis is best suited for the analysis of cooling towers when impact assessment is not required; otherwise, life cycle assessment meets most of the defined criteria. Moreover, only exergy-based methods allow for the inclusion of volatile ambient conditions.Item Unknown A timed off-switch for dynamic control of gene expression in Corynebacterium glutamicum(2021) Siebert, Daniel; Altenbuchner, Josef; Blombach, BastianDynamic control of gene expression mainly relies on inducible systems, which require supplementation of (costly) inducer molecules. In contrast, synthetic regulatory circuits, which allow the timed shutdown of gene expression, are rarely available and therefore represent highly attractive tools for metabolic engineering. To achieve this, we utilized the VanR/PvanABK* regulatory system of Corynebacterium glutamicum, which consists of the transcriptional repressor VanR and a modified promoter of the vanABK operon (PvanABK*). VanR activity is modulated by one of the phenolic compounds ferulic acid, vanillin or vanillic acid, which are co-metabolized with d-glucose. Thus, gene expression in the presence of d-glucose is turned off if one of the effector molecules is depleted from the medium. To dynamically control the expression of the aceE gene, encoding the E1 subunit of the pyruvate dehydrogenase complex that is essential for growth on d-glucose, we replaced the native promoter by vanR/PvanABK* yielding C. glutamicum ΔPaceE::vanR-PvanABK*. The biomass yield of this strain increased linearly with the supplemented amount of effector. After consumption of the phenolic compounds growth ceased, however, C. glutamicumΔPaceE::vanR-PvanABK* continued to utilize the residual d-glucose to produce significant amounts of pyruvate, l-alanine, and l-valine. Interestingly, equimolar concentrations of the three phenolic compounds resulted in different biomass yields; and with increasing effector concentration, the product spectrum shifted from pyruvate over l-alanine to l-valine. To further test the suitability of the VanR/PvanABK* system, we overexpressed the l-valine biosynthesis genes ilvBNCE in C. glutamicum ΔPaceE::vanR-PvanABK*, which resulted in efficient l-valine production with a yield of about 0.36 mol l-valine per mol d-glucose. These results demonstrate that the VanR/PvanABK* system is a valuable tool to control gene expression in C. glutamicum in a timed manner by the cheap and abundant phenolic compounds ferulic acid, vanillin, and vanillic acid.Item Open Access Constitutive correlations for mass transport in fibrous media based on asymptotic homogenization(2023) Maier, Lukas; Kufferath-Sieberin, Lars; Pauly, Leon; Hopp-Hirschler, Manuel; Gresser, Götz T.; Nieken, UlrichMass transport in textiles is crucial. Knowledge of effective mass transport properties of textiles can be used to improve processes and applications where textiles are used. Mass transfer in knitted and woven fabrics strongly depends on the yarn used. In particular, the permeability and effective diffusion coefficient of yarns are of interest. Correlations are often used to estimate the mass transfer properties of yarns. These correlations commonly assume an ordered distribution, but here we demonstrate that an ordered distribution leads to an overestimation of mass transfer properties. We therefore address the impact of random ordering on the effective diffusivity and permeability of yarns and show that it is important to account for the random arrangement of fibers in order to predict mass transfer. To do this, Representative Volume Elements are randomly generated to represent the structure of yarns made from continuous filaments of synthetic materials. Furthermore, parallel, randomly arranged fibers with a circular cross-section are assumed. By solving the so-called cell problems on the Representative Volume Elements, transport coefficients can be calculated for given porosities. These transport coefficients, which are based on a digital reconstruction of the yarn and asymptotic homogenization, are then used to derive an improved correlation for the effective diffusivity and permeability as a function of porosity and fiber diameter. At porosities below 0.7, the predicted transport is significantly lower under the assumption of random ordering. The approach is not limited to circular fibers and may be extended to arbitrary fiber geometries.