Browsing by Subject "Bioreactor"

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Bioprocess exploitation of microaerobic auto-induction using the example of rhamnolipid biosynthesis in Pseudomonas putida KT2440
(2025) Grether, Jakob; Dittmann, Holger; Willems, Leon; Schmiegelt, Tabea; Benatto Perino, Elvio Henrique; Hubel, Philipp; Lilge, Lars; Hausmann, Rudolf; Grether, Jakob; Department of Bioprocess Engineering, Institute of Food Science and Biotechnology, University of Hohenheim, Fruwirthstr. 12, 70599, Stuttgart, Germany; Dittmann, Holger; Department of Bioprocess Engineering, Institute of Food Science and Biotechnology, University of Hohenheim, Fruwirthstr. 12, 70599, Stuttgart, Germany; Willems, Leon; Department of Bioprocess Engineering, Institute of Food Science and Biotechnology, University of Hohenheim, Fruwirthstr. 12, 70599, Stuttgart, Germany; Schmiegelt, Tabea; Department of Bioprocess Engineering, Institute of Food Science and Biotechnology, University of Hohenheim, Fruwirthstr. 12, 70599, Stuttgart, Germany; Benatto Perino, Elvio Henrique; Department of Bioprocess Engineering, Institute of Food Science and Biotechnology, University of Hohenheim, Fruwirthstr. 12, 70599, Stuttgart, Germany; Hubel, Philipp; Core Facility Hohenheim, Mass Spectrometry Core Facility, University of Hohenheim, Ottilie-Zeller-Weg 2, 70599, Stuttgart, Germany; Lilge, Lars; Department of Bioprocess Engineering, Institute of Food Science and Biotechnology, University of Hohenheim, Fruwirthstr. 12, 70599, Stuttgart, Germany; Hausmann, Rudolf; Department of Bioprocess Engineering, Institute of Food Science and Biotechnology, University of Hohenheim, Fruwirthstr. 12, 70599, Stuttgart, Germany
Background: In biomanufacturing of surface-active agents, such as rhamnolipids, excessive foaming is a significant obstacle for the development of high-performing bioprocesses. The exploitation of the inherent tolerance of Pseudomonas putida KT2440, an obligate aerobic bacterium, to microaerobic conditions has received little attention so far. Here low-oxygen inducible promoters were characterized in biosensor strains and exploited for process control under reduction of foam formation by low aeration and stirring rates during biosynthesis of rhamnolipids. Results: In this study, homologous promoters of P. putida inducible under oxygen limitation were identified by non-targeted proteomic analyses and characterized by fluorometric methods. Proteomics indicated a remodeling of the respiratory chain and the regulation of stress-related proteins under oxygen limitation. Of the three promoters tested in fluorescent biosensor assays, the promoter of the oxygen-sensitive cbb3-type cytochrome c oxidase gene showed high oxygen-dependent controllability. It was used to control the gene expression of a heterologous di-rhamnolipid synthesis operon in an auto-inducing microaerobic two-phase bioprocess. By limiting the oxygen supply via low aeration and stirring rates, the bioprocess was clearly divided into a growth and a production phase, and sources of foam formation were reduced. Accordingly, rhamnolipid synthesis did not have to be controlled externally, as the oxygen-sensitive promoter was autonomously activated as soon as the oxygen level reached microaerobic conditions. A critical threshold of about 20% oxygen saturation was determined. Conclusions: Utilizing the inherent tolerance of P. putida to microaerobic conditions in combination with the application of homologous, low-oxygen inducible promoters is a novel and efficient strategy to control bioprocesses. Fermentation under microaerobic conditions enabled the induction of rhamnolipid production by low oxygen levels, while foam formation was limited by low aeration and stirring rates.
The influence of growth rate-controlling feeding strategy on the surfactin production in Bacillus subtilis bioreactor processes
(2024) Hiller, Eric; Off, Manuel; Hermann, Alexander; Vahidinasab, Maliheh; Benatto Perino, Elvio Henrique; Lilge, Lars; Hausmann, Rudolf
Background The production of surfactin, an extracellular accumulating lipopeptide produced by various Bacillus species, is a well-known representative of microbial biosurfactant. However, only limited information is available on the correlation between the growth rate of the production strain, such as B. subtilis BMV9, and surfactin production. To understand the correlation between biomass formation over time and surfactin production, the availability of glucose as carbon source was considered as main point. In fed-batch bioreactor processes, the B. subtilis BMV9 was used, a strain well-suited for high cell density fermentation. By adjusting the exponential feeding rates, the growth rate of the surfactin-producing strain, was controlled. Results Using different growth rates in the range of 0.075 and 0.4 h-1, highest surfactin titres of 36 g/L were reached at 0.25 h-1 with production yields YP/S of 0.21 g/g and YP/X of 0.7 g/g, while growth rates lower than 0.2 h-1 resulted in insufficient and slowed biomass formation as well as surfactin production (YP/S of 0.11 g/g and YP/X of 0.47 g/g for 0.075 h-1). In contrast, feeding rates higher than 0.25 h-1 led to a stimulation of overflow metabolism, resulting in increased acetate formation of up to 3 g/L and an accumulation of glucose due to insufficient conversion, leading to production yields YP/S of 0.15 g/g and YP/X of 0.46 g/g for 0.4 h-1. Conclusions Overall, the parameter of adjusting exponential feeding rates have an important impact on the B. subtilis productivity in terms of surfactin production in fed-batch bioreactor processes. A growth rate of 0.25 h-1 allowed the highest surfactin production yield, while the total conversion of substrate to biomass remained constant at the different growth rates.
A robust fermentation process for natural chocolate-like flavor production with Mycetinis scorodonius
(2022) Rigling, Marina; Heger, Fabienne; Graule, Maria; Liu, Zhibin; Zhang, Chen; Ni, Li; Zhang, Yanyan
Submerged fermentation of green tea with the basidiomycete Mycetinis scorodonius resulted in a pleasant chocolate-like and malty aroma, which could be a promising chocolate flavor alternative to current synthetic aroma mixtures in demand of consumer preferences towards healthy natural and ‘clean label’ ingredients. To understand the sensorial molecular base on the chocolate-like aroma formation, key aroma compounds of the fermented green tea were elucidated using a direct immersion stir bar sorptive extraction combined with gas chromatography–mass spectrometry–olfactometry (DI-SBSE-GC-MS-O) followed by semi-quantification with internal standard. Fifteen key aroma compounds were determined, the most important of which were dihydroactinidiolide (odor activity value OAV 345), isovaleraldehyde (OAV 79), and coumarin (OAV 24), which were also confirmed by a recombination study. Furthermore, effects of the fermentation parameters (medium volume, light protection, agitation rate, pH, temperature, and aeration) on the aroma profile were investigated in a lab-scale bioreactor at batch fermentation. Variation of the fermentation parameters resulted in similar sensory perception of the broth, where up-scaling in volume evoked longer growth cycles and aeration significantly boosted the concentrations yet added a green note to the overall flavor impression. All findings prove the robustness of the established fermentation process with M. scorodonius for natural chocolate-like flavor production.