In many practical applications involving surfactants, achieving defoaming without affecting interfacial activity is a challenge. In this study, the antifoaming performance of REP-type block polymer nonionic surfactant C12EOmPOn was determined, and molecular dynamics simulation method was employed to investigate the molecular behaviors of surfactants at a gas/water interface, the detailed arrangement information of the different structural segments of the surfactant molecules and the inter-/intra-interactions between all the structural motifs in the interfacial layer were analyzed systematically, by which the antifoaming mechanisms of the surfactants were revealed. The results show that the EO and PO groups of REP-type polyether molecules are located in the aqueous phase near the interface, and the hydrophobic tails distribute separately, lying almost flat on the gas/water interface. The interaction between the same groups of EOs and POs is significantly stronger than with water. REP block polyethers with high polymerization degrees of EO and PO are more inclined to overlap into dense layers, resulting in the formation of aggregates resembling \"oil lenses\" spreading on the gas/water interface, which exerts a stronger antifoaming effect. This study provides a smart approach to obtaining efficient antifoaming performance at room temperature without adding other antifoam ingredients.

Foam is generated in mammalian cell cultures by excessive agitation or gas sparging. This occurs particularly in cultures that generate recombinant proteins at high cell concentrations. Three antifoam agents were tested for their compatibility with antibody-producing Chinese hamster ovary (CHO) cells. One agent (antifoam 204) was completely inhibitory to growth at a concentration of 10 ppm, one agent (antifoam C) showed partial inhibition and a third (antifoam SE-15) showed no inhibition at this concentration. A novel foam image analyzer (LabCam) was used to evaluate two antifoams (C and SE-15) for their ability to dissipate foam generated in cell culture media by enhanced agitation. The presence of antifoam in the media reduced significantly the foam layer that was generated and this was shown to be rapidly dissipated in the presence of 10 ppm SE-15. The antifoams were also tested for foam dissipation in cultures of CHO cells at >106 cells/mL. Supplementation of the cultures with SE-15 resulted in dissipation of foam generated by excessive gas sparging within 2 min. Under equivalent conditions 75% of foam dissipated in the presence of antifoam C, within 2 min but there was a residual foam layer up to 25 min. This study showed the value of an optical monitoring system (LabCam) for measuring foam generation and dissipation in a bioreactor to assess the efficiency of antifoam agents to reduce foam in a bioreactor. This has the potential for use as a control system that could be designed for continuous monitoring and foam control in a mammalian cell bioprocess.

Bioreactors are the operative backbone, for example, for the production of biopharmaceuticals, biomaterials in tissue engineering, and sustainable substitutes for chemicals. Still, the Achilles\' heel of bioreactors nowadays is the aeration which is based on intense stirring and gas sparging, yielding inherent drawbacks such as shear stress, foaming, and sterility concerns. We present the synergistic combination of simulations and experiments toward a membrane stirrer for the efficient bubble-free aeration of bioreactors. A digital twin of the bioreactor with an integrated membrane-module stirrer (MemStir) was developed with computational fluid dynamics (CFD) studies addressing the determination of fluid mixing, shear rates, and local oxygen concentration. Usability of the MemStir is shown in a foam-free recombinant production process of biosurfactants (rhamnolipids) from glucose with different strains of Pseudomonas putida KT2440 in a 3-L vessel and benchmarked against a regular aerated process. The MemStir delivered a maximal oxygen transfer rate (OTRmax ) of 175 mmol L-1 h-1 in completely foam-free cultivations. With a high space-time yield (STY) of 118 mgRL L-1 h-1 during a fed-batch fermentation, the effectiveness of the novel MemStir is demonstrated. Simulations show the generic value of the MemStir beyond biosurfactant production, for example, for animal cell cultivation.

Foot-and-mouth disease virus (FMDV) is endemic in many parts of the world. Vaccination is an important control measure, limits viral spread, and can help to eradicate the disease. However, vaccination programs are cost-intensive because of the short shelf life of vaccines and the need for frequent re-vaccination. Animal-component-free (ACF) or chemically defined media (CDM) at high cell densities are a promising approach for the production of inexpensive high-quality vaccines, but the occurrence of cell density effects has been reported for various virus-cell systems in vaccine production. For FMDV, the use of CDM or ACF media for vaccine production has not been studied and no information about cell density effects is available. This work describes the propagation of FMDV in ACF or in CDM. Cells were grown at increasing cell densities and either 100% media exchange or addition of 30% fresh media was performed before infection with FMDV. Increasing cell densities reduced the viral titer and increased yield variability in all media except BHK300G. This effect can be mitigated by performing a 100% media exchange before infection or when using the controlled environment of a bioreactor. The media composition and also a fragile relationship between virus and cell metabolism seem to be causal for that phenomenon.

Monoclonal antibody production in commercial scale cell culture bioprocessing requires a thorough understanding of the engineering process and components used throughout manufacturing. It is important to identify high impact components early on during the lifecycle of a biotechnology-derived product. While cell culture media selection is of obvious importance to the health and productivity of mammalian bioreactor operations, other components such as antifoam selection can also play an important role in bioreactor cell culture. Silicone polymer-based antifoams were known to have negative impacts on cell health, production, and downstream filtration and purification operations. High throughput screening in micro-scale bioreactors provides an efficient strategy to identify initial operating parameters. Here, we utilized a micro-scale parallel bioreactor system to study an IgG1 producing CHO cell line, to screen Dynamis, ProCHO5, PowerCHO2, EX-Cell Advanced, and OptiCHO media, and 204, C, EX-Cell, SE-15, and Y-30 antifoams and their impacts on IgG1 production, cell growth, aggregation, and process control. This study found ProCHO5, EX-Cell Advanced, and PowerCHO2 media supported strong cellular growth profiles, with an IVCD of 25-35 × 106 cells-d/mL, while maintaining specific antibody production (Qp  > 2 pg/cell-d) for our model cell line and a monomer percentage above 94%. Antifoams C, EX-Cell, and SE-15 were capable of providing adequate control of foaming while antifoam 204 and Y-30 noticeably stunted cellular growth. This work highlights the utility of high throughput micro bioreactors and the importance of identifying both positive and negative impacts of media and antifoam selection on a model IgG1 producing CHO cell line. © 2017 The Authors Biotechnology Progress published by Wiley Periodicals, Inc. on behalf of American Institute of Chemical Engineers Biotechnol. Prog., 34:262-270, 2018.

The Brazilian sugarcane industry constitutes one of the biggest and most efficient ethanol production processes in the world. Brazilian ethanol production utilizes a unique process, which includes cell recycling, acid wash, and non-aseptic conditions. Process characteristics, such as extensive CO2 generation, poor quality of raw materials, and frequent contaminations, all lead to excessive foam formation during fermentations, which is treated with antifoam agents (AFA). In this study, we have investigated the impact of industrial AFA treatments on the physiology and transcriptome of the industrial ethanol strain Saccharomyces cerevisiae CAT-1. The investigated AFA included industrially used AFA acquired from Brazilian ethanol plants and commercially available AFA commonly used in the fermentation literature. In batch fermentations, it was shown that industrial AFA compromised growth rates and glucose uptake rates, while commercial AFA had no effect in concentrations relevant for defoaming purposes. Industrial AFA were further tested in laboratory scale simulations of the Brazilian ethanol production process and proved to decrease cell viability compared to the control, and the effects were intensified with increasing AFA concentrations and exposure time. Transcriptome analysis showed that AFA treatments induced additional stress responses in yeast cells compared to the control, shown by an up-regulation of stress-specific genes and a down-regulation of lipid biosynthesis, especially ergosterol. By documenting the detrimental effects associated with chemical AFA, we highlight the importance of developing innocuous systems for foam control in industrial fermentation processes.

Simethicone emulsion is used to regulate foaming in cell culture operations in biopharmaceutical processes. It is also a potential source of endotoxin contamination. The inactivation of endotoxins in dilute simethicone emulsions was assessed as a function of time at different steam temperatures using a Limulus amebocyte lysate kinetic chromogenic technique. Endotoxin inactivation from steam-heat treatment was fit to a four-parameter double exponential decay model, which indicated that endotoxin inactivation was biphasic, consisting of fast and slow regimes. In the fast regime, temperature-related effects were dominant. Transitioning into the slow regime, the observed temperature dependence diminished, and concentration-related effects became increasingly significant. The change in the Gibbs free energy moving through the transition state indicated that a large energy barrier must be overcome for endotoxin inactivation to occur. The corresponding Arrhenius pre-exponential factor was >>10(12) s(-1) suggesting that endotoxins in aqueous solution exist as aggregates. The disorder associated with the endotoxin inactivation reaction pathway was assessed via the change in entropy moving through the transition state. This quantity was positive indicating that endotoxin inactivation may result from hydrolysis of individual endotoxin molecules, which perturbs the conformation of endotoxin aggregates, thereby modulating the biological activity observed. Steam-heat treatment decreased endotoxin levels by 1-2 logarithm (log) reduction (LRV), which may be practically relevant depending on incoming raw material endotoxin levels. Antifoam efficiency and cell culture performance were negligibly impacted following steam-heat treatment. The results from this study show that steam-heat treatment is a viable endotoxin control strategy that can be implemented to support large-scale biopharmaceutical manufacturing.

In this paper we present briefly our current understanding of the modes of action of foam control agents (often termed \"defoamers\" or \"antifoams\"). After summarizing the background knowledge, reviewed in previous articles, the focus of the presentation is shifted to the antifoam studies from the last decade. The new experimental results, obtained by various research groups, are reviewed briefly to reveal the main mechanisms of antifoam action and the related key factors, governing the efficiency of the foam control agents. The role of the entry, spreading and bridging coefficients, of the entry barrier of the antifoam entities, and of the dynamics of surfactant adsorption is specifically discussed.