Sampling-free investigation of microbial carbon source preferences on renewable feedstocks via online monitoring of oxygen transfer rate.

The transition towards sustainable bioprocesses requires renewable feedstocks to reduce dependency on finite resources. While plant-based feedstocks offer significant potential, their complex composition poses new challenges. The microorganisms often exhibit polyauxic growth when presented with multiple carbon sources simultaneously, consuming them in a distinct order according to their carbon source preferences.

High-sensitivity real-time monitoring of pH and respiration activity unveils metabolic dynamics in shake flask cultures.

Erlenmeyer shake flasks are widely used during the first steps of bioprocess development. Despite their broad application in academia and industry, shake flasks usually lack standardized and user-friendly online monitoring techniques. In this work, the pH and Respiratory Activity MOnitoring System (pH-RAMOS) for the non-invasive online measurement of the oxygen transfer rate (OTR), carbon dioxide transfer rate (CTR), and pH in up to eight parallel shake flasks under sterile conditions is presented.

Impact of hydromechanical stress on CHO cells' metabolism and productivity: Insights from shake flask cultivations with online monitoring of the respiration activity.

The hydromechanical stress is a relevant parameter for mammalian cell cultivations, especially regarding scale-up processes. It describes the mechanical forces exerted on cells in a bioreactor. The maximum local energy dissipation rate is a suitable parameter to characterize hydromechanical stress.

Avoiding overflow metabolite formation in Komagataella phaffii fermentations to enhance recombinant protein production.

Background: Komagataella phaffii (K. phaffii), formerly known as Pichia pastoris, is a widely utilized yeast for recombinant protein production. However, due to the formation of overflow metabolites, carbon yields may be reduced and product recovery becomes challenging.

Unraveling the impact of pH, sodium concentration, and medium osmolality on Vibrio natriegens in batch processes.

Background: Vibrio natriegens, a halophilic marine γ-proteobacterium, holds immense biotechnological potential due to its remarkably short generation time of under ten minutes. However, the highest growth rates have been primarily observed on complex media, which often suffer from batch-to-batch variability affecting process stability and performance. Consistent bioprocesses necessitate the use of chemically defined media, which are usually optimized for fermenters with pH and dissolved oxygen tension (DOT) regulation, both of which are not applied during early-stage cultivations in shake flasks or microtiter plates.

Monitoring corn stover processing by the fungus Ustilago maydis.

A key aspect of sustainable bioeconomy is the recirculation of renewable, agricultural waste streams as substrates for microbial production of high-value compounds. One approach is the bioconversion of corn stover, an abundant maize crop byproduct, using the fungal maize pathogen Ustilago maydis. U.

Phosphate limitation enhances malic acid production on nitrogen-rich molasses with Ustilago trichophora.

Background: An important step in replacing petrochemical products with sustainable, cost-effective alternatives is the use of feedstocks other than, e.g., pure glucose in the fermentative production of platform chemicals.

Diffusion-driven fed-batch fermentation in perforated ring flasks.

Purpose: Simultaneous membrane-based feeding and monitoring of the oxygen transfer rate shall be introduced to the newly established perforated ring flask, which consists of a cylindrical glass flask with an additional perforated inner glass ring, for rapid bioprocess development.

Methods: A 3D-printed adapter was constructed to enable monitoring of the oxygen transfer rate in the perforated ring flasks. Escherichia coli experiments in batch were performed to validate the adapter.

Understanding exopolysaccharide byproduct formation in Komagataella phaffii fermentation processes for recombinant protein production.

Background: Komagataella phaffii (Pichia pastoris) has emerged as a common and robust biotechnological platform organism, to produce recombinant proteins and other bioproducts of commercial interest. Key advantage of K. phaffii is the secretion of recombinant proteins, coupled with a low host protein secretion.

Risk-Based Selection of Environmental Classifications for Biopharmaceutical Operations.

This article details a risk-based methodology designed to assign environmental classifications to the different operations in biopharmaceutical facilities manufacturing non-sterile (low bioburden) drug substance. Generally, environmental conditions for active pharmaceutical ingredient manufacture are established based on previous experiences or expectations or on extrapolated interpretations of current good manufacturing practices guidelines. Improvements in equipment design and operation, especially the use of closed systems, allow certain process steps to take place in controlled environment areas rather than in classified clean rooms.

Adaptive laboratory evolution of and to enhance anthranilate tolerance.

Microbial bioproduction of the aromatic acid anthranilate (-aminobenzoate) has the potential to replace its current, environmentally demanding production process. The host organism employed for such a process needs to fulfil certain demands to achieve industrially relevant product levels. As anthranilate is toxic for microorganisms, the use of particularly robust production hosts can overcome issues from product inhibition.

Developing the biofacility of the future based on continuous processing and single-use technology.

To maintain or strengthen their market position, biopharmaceutical producers have to adapt their production facilities to a drastically changed market environment. Contrary to currently used large scale batch-wise operated production facilities, where stainless steel equipment is widely applied, small scale and flexible production processes are desired. Consequently, the concept of the "biofacility of the future" has been developed, which combines the attributes fast, flexible, small, inexpensive and sustainable.

The identification of enzyme targets for the optimization of a valine producing Corynebacterium glutamicum strain using a kinetic model.

The enzyme targets for the rational optimization of a Corynebacterium glutamicum strain constructed for valine production are identified by analyzing the control of flux in the valine/leucine pathway. The control analysis is based on measurements of the intracellular metabolite concentrations and on a kinetic model of the reactions in the investigated pathway. Data-driven and model-based methods are used and evaluated against each other.

Modeling metabolic networks in C. glutamicum: a comparison of rate laws in combination with various parameter optimization strategies.

Background: To understand the dynamic behavior of cellular systems, mathematical modeling is often necessary and comprises three steps: (1) experimental measurement of participating molecules, (2) assignment of rate laws to each reaction, and (3) parameter calibration with respect to the measurements. In each of these steps the modeler is confronted with a plethora of alternative approaches, e. g.

Monitoring and modeling of the reaction dynamics in the valine/leucine synthesis pathway in Corynebacterium glutamicum.

The intracellular concentrations of the valine and leucine pathway intermediates in a Corynebacterium glutamicum strain were measured during a transient state. The data were obtained by performing a glucose stimulus-response experiment with the use of a rapid sampling device and advanced mass spectrometry. The glucose stimulus resulted in a 3-fold increase in the intracellular pyruvate concentration within less than a second, demonstrating the very fast interactions in metabolic networks.