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. 2023 Nov 6:11:1264787.
doi: 10.3389/fbioe.2023.1264787. eCollection 2023.

Evaluation of an external foam column for in situ product removal in aerated surfactin production processes

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Evaluation of an external foam column for in situ product removal in aerated surfactin production processes

Chantal Treinen et al. Front Bioeng Biotechnol. .

Abstract

In Bacillus fermentation processes, severe foam formation may occur in aerated bioreactor systems caused by surface-active lipopeptides. Although they represent interesting compounds for industrial biotechnology, their property of foaming excessively during aeration may pose challenges for bioproduction. One option to turn this obstacle into an advantage is to apply foam fractionation and thus realize in situ product removal as an initial downstream step. Here we present and evaluate a method for integrated foam fractionation. A special feature of this setup is the external foam column that operates separately in terms of, e.g., aeration rates from the bioreactor system and allows recycling of cells and media. This provides additional control points in contrast to an internal foam column or a foam trap. To demonstrate the applicability of this method, the foam column was exemplarily operated during an aerated batch process using the surfactin-producing Bacillus subtilis strain JABs24. It was also investigated how the presence of lipopeptides and bacterial cells affected functionality. As expected, the major foam formation resulted in fermentation difficulties during aerated processes, partially resulting in reactor overflow. However, an overall robust performance of the foam fractionation could be demonstrated. A maximum surfactin concentration of 7.7 g/L in the foamate and enrichments of up to 4 were achieved. It was further observed that high lipopeptide enrichments were associated with low sampling flow rates of the foamate. This relation could be influenced by changing the operating parameters of the foam column. With the methodology presented here, an enrichment of biosurfactants with simultaneous retention of the production cells was possible. Since both process aeration and foam fractionation can be individually controlled and designed, this method offers the prospect of being transferred beyond aerated batch processes.

Keywords: Bacillus; aerated fermentation processes; downstream processing; foam fractionation; in-situ product removal; surfactin.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision.

Figures

FIGURE 1
FIGURE 1
Schematic representation of overfoaming in aerated surfactin production processes using a foam trap. (A) Foam is building up in the headspace of the bioreactor, causing overflow; (B) Culture broth is mainly located in the foam trap, leaving the bioreactor almost empty as a result of severe foaming during the process [bioreactor figure adapted from Hoffmann et al. (2020)].
FIGURE 2
FIGURE 2
Integrated foam fractionation in aerated fermentation processes and experimental overview. A schematic representation of a bioreactor system with an external foam column is seen in the centre (bioreactor figure adapted from Hoffmann et al. (2020)).
FIGURE 3
FIGURE 3
Image of the external foam column to illustrate the design and principle of operation. The foam column is connected to the bioreactor in this depiction. Due to the size, the lower part, which contains the foam generator column, and the upper drainage column were photographed separately. However, the two parts are connected at the point where the images were merged. The white arrows indicate the flow direction of the foam and the recirculated liquid.
FIGURE 4
FIGURE 4
Exemplary images of foam behavior in the lower foam generator part of the foam column. Gas and liquid flow were co-current. (A) Small and stable foam bubbles which allowed for sampling. Photograph was taken during cultivation with surfactin producer Bacillus subtilis JABs24; (B) Foam bubbles were too large and burst quickly. Photograph was taken during cultivation with non-surfactin producer Bacillus subtilis 168; (C) Foaming was not possible, foam generator was filled with cultivation broth. Photograph was taken during cultivation with non-surfactin producer Bacillus subtilis 168.
FIGURE 5
FIGURE 5
Time course of bioreactor batch cultivations using Bacillus spp. Exemplary processes are shown, each representing one biological replicate with (A) strain Bacillus subtilis 168 until t = 36 h; (B) Bacillus subtilis JABs24, Replicate 1 until t = 60 h; (C) Bacillus subtilis JABs24 with integrated foam fractionation (ISPR), Replicate 1 until t = 60 h. Given are the cell growth as OD600 (black cross), the consumption of the carbon source glucose (gray square) and the consumption of the nitrogen source ammonium (black triangle) over the cultivation time. Solid lines indicate a dynamic curve fit that is either sigmoidal or logistic with 4 parameters. Curve fit of ammonia for Bacillus subtilis 168 does not include time-point t = 9 h. The dashed lines, however, do not represent a fit and are only integrated for simplified visualization.
FIGURE 6
FIGURE 6
Time-course of cell dry weight (CDW) and surfactin production during exemplary batch cultivations. (A) Reference process with Bacillus subtilis JABs24 without foam fractionation; (B,C) Foam column process as proof of principle with Bacillus subtilis JABs24 and integrated foam fractionation (ISPR). For the latter, (B) concentrations in the culture broth; (C) concentrations in the foamate.
FIGURE 7
FIGURE 7
Time-course of biomass and surfactin enrichment for exemplary batch cultivations. (A) Biomass enrichment; (B) Surfactin enrichment. Represented are the calculated values for the proof of principle process, employing Bacillus subtilis JABs24 with integrated foam fractionation (ISPR).

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Grants and funding

The author(s) declare financial support was received for the research, authorship, and/or publication of this article. This study was financially supported by the German Research Foundation (DFG), Project Number 365166982. CT further is a member of the “BBW ForWerts” graduate program funded by the Ministry of Science, Research and Arts (MWK) of Baden-Württemberg, Germany.