Fast “Feast/Famine” cycles for studying microbial physiology under dynamic conditions: a case study with Saccharomyces cerevisiae

Microorganisms are constantly exposed to rapidly changing conditions, under natural as well as industrial production scale environments, especially due to large-scale substrate mixing limitations. In this work, we present an experimental approach based on a dynamic feast/famine regime (400 s) that leads to repetitive cycles with moderate changes in substrate availability in an aerobic glucose cultivation of Saccharomyces cerevisiae. After a few cycles, the feast/famine produced a stable and repetitive pattern with a reproducible metabolic response in time, thus providing a robust platform for studying the microorganism’s physiology under dynamic conditions. We found that the biomass yield was slightly reduced (−5%) under the feast/famine regime, while the averaged substrate and oxygen consumption as well as the carbon dioxide production rates were comparable. The dynamic response of the intracellular metabolites showed specific differences in comparison to other dynamic experiments (especially stimulus-response experiments, SRE). Remarkably, the frequently reported ATP paradox observed in single pulse experiments was not present during the repetitive perturbations applied here. We found that intracellular dynamic accumulations led to an uncoupling of the substrate uptake rate (up to 9-fold change at 20 s.) Moreover, the dynamic profiles of the intracellular metabolites obtained with the feast/famine suggest the presence of regulatory mechanisms that resulted in a delayed response. With the feast famine setup many cellular states can be measured at high frequency given the feature of reproducible cycles. The feast/famine regime is thus a versatile platform for systems biology approaches, which can help us to identify and investigate metabolite regulations under realistic conditions (e.g., large-scale bioreactors or natural environments).info:eu-repo/semantics/publishedVersio

CFD-simulations of mixing and bioconversions in gassed stirred tank reactors

Rührkesselreaktoren kommen in zahlreichen chemischen und biotechnischen Produktionsprozessen zur Anwendung, für biotechnische Stoffumwandlungen sind sie sogar der wichtigste Standardapparat. Die Methoden der numerischen Strömungssimulation (Computational Fluid Dynamics, CFD) liefern heute die Grundlage für detaillierte Untersuchungen der lokalen und zeitabhängigen Vorgänge bei Stoffumsetzungen in der turbulenten Zweiphasenströmung im Rührkesselreaktor. Im Rahmen der vorliegenden Arbeit wurden Methoden zum Einsatz von CFD-Simulationen bei der Analyse und Vorausberechnung biotechnischer Stoffumsetzungen in Rührkesselreaktoren entwickelt und auf exemplarische biotechnische Stoffumsetzungen angewendet. Bei der Modellierung wurde ein Mittelweg zwischen Komplexität und praktischer Anwendbarkeit gewählt, der auf der Entkopplung der Simulation von Fluiddynamik und Stoffumsetzungen beruht. Dadurch wurde die dynamische Simulation der Stoffbilanzen auf stationären Strömungsfeldern ermöglicht. Zunächst wurde das Durchmischungsverhalten in verschiedenen Reaktorkonfigurationen mit Einfach- und Mehrfachrührern anhand der Simulation von Pulsexperimenten untersucht. Simulationen der während der aeroben Fermentation von Saccharomyces cerevisiae auftretenden Substratverteilungen zeigen exemplarisch den erheblichen Einfluss der Konzentrationsgradienten auf das Ergebnis von im Zulaufverfahren durchgeführten Bioprozessen. Als Einflussfaktoren wurden die Rührerkombination im Mehrfachrührersystem sowie Rührerdrehzahl und Belüftungsrate betrachtet, wobei insbesondere die Wahl einer geeigneten Rührerkombination entscheidend ist. Es wird gezeigt, wie die unerwünschte Nebenproduktbildung durch die Berechnung eines optimalen Zulaufortes des Substrats minimiert werden kann. Zusätzlich wurden auf der Basis zweiphasiger, mit einem algebraic-slip-Ansatz berechneter Strömungsfelder die zugehörigen Verteilungen der Gelöstsauerstoffkonzentration simuliert.Stirred tank reactors (STR) are widely used in chemical and biotechnical industries to carry out a variety of process operations. For biotechnical conversion processes, the STR is the most important type of reactor. Today, the methods of Computational Fluid Dynamics (CFD) provide the tools required for detailed simulations of conversion processes in stirred tank reactors. The primary purpose of this work was to develop methods for applying the CFD simulations to analysis, design, and scale-up of biotechnical conversion processes in aerated stirred tank reactors, taking into account some reasonable simplifying model assumptions. Thus, a trade-off between maximum complexity and practical applicability was chosen. Momentum balance equations and material balance equations were simulated separately under the assumption of neglegible effects of mass transfer and conversion on the flow field. This allowed the use of stationary flow fields for dynamic simulations of conversion processes. Pulse experiments for the determination of mixing properties of different single-impeller and multi-impeller stirred tanks were simulated. Simulations of distributions of the carbon and energy source during fermentations of the yeast Saccharomyces cerevisiae elucidated the influence of concentration gradients on yield and undesired byproduct formation of a typical fed-batch process. The effect of different impeller combinations, impeller speeds and gassing rates was studied. The choice of an appropriate impeller combination is crucial for axial mixing and process performance. Undesired byproduct formation could be minimized by calculating an optimum substrate feeding position. Corresponding distributions of dissolved oxygen concentration were simulated based on two phase flow fields and local rates of mass transfer gas/liquid. Two phase flow was modeled in an Eulerian way with an algebraic-slip approach

Die Liquiditaetsdisposition der Kreditinstitute: unter besonderer Beruecksichtigung ihrer Abhaengigkeit von der Geldpolitik der Zentralbank und ihrer Bedeutung fuer diese

Available from Bibliothek des Instituts fuer Weltwirtschaft, ZBW, Duesternbrook Weg 120, D-24105 Kiel A 192630 / FIZ - Fachinformationszzentrum Karlsruhe / TIB - Technische InformationsbibliothekSIGLEDEGerman

Impact of mixing imperfections on yeast bioreactor performances: Scale-down reactor concept and related experimental tools

peer reviewedA method combining environmental data extracted from the dissolved oxygen profile of a fed-batch bioreactor and a dynamic discrete Markov chain model has been presented in order to give more insight about the glucose and dissolved oxygen fluctuations experienced by the microorganisms during cultivation in heterogeneous bioreactor. The fed-batch cultivation of Saccharomyces cerevisiae has been performed in a well-mixed and a partitioned scale-down reactor (SDR). The analysis of the environmental sequences has shown extended time lengths for the glucose availability and depletion sequences in the case of the SDR under a DO-controlled fed-batch culture. The Markov chain model developed in this work is able to capture the stochastic environmental events, i.e. in our case the environmental states experienced by the microorganisms crossing the tubular part of the SDR. The simulation results show clearly an extension of the starvation periods in the case of the culture performed in the SDR. The simulations have been performed at the single cells level allowing future improvements of our model and notably in the context of the population segregation phenomena occurring in fed-batch cultures. As a perspective, flow cytometry has been presented as a high-throughput analytical tool for the investigation of yeast physiology at the single cell level and in process-related conditions

Food sharing dataset

Excel table with raw data of common marmosets behavior during food sharing experiment