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Uncoupling Foam Fractionation and Foam Adsorption for Enhanced Biosurfactant Synthesis and Recovery.

ABSTRACT: The production of biosurfactants is often hampered by excessive foaming in the bioreactor, impacting system scale-up and downstream processing. Foam fractionation was proposed to tackle this challenge by combining in situ product removal with a pre-purification step. In previous studies, foam fractionation was coupled to bioreactor operation, hence it was operated at suboptimal parameters. Here, we use an external fractionation column to decouple biosurfactant production from foam fractionation, enabling continuous surfactant separation, which is especially suited for system scale-up. As a subsequent product recovery step, continuous foam adsorption was integrated into the process. The configuration is evaluated for rhamnolipid (RL) or 3-(3-hydroxyalkanoyloxy)alkanoic acid (HAA, i.e., RL precursor) production by recombinant non-pathogenic Pseudomonas putida KT2440. Surfactant concentrations of 7.5 g_RL/L and 2.0 g_HAA/L were obtained in the fractionated foam. 4.7 g RLs and 2.8 g HAAs could be separated in the 2-stage recovery process within 36 h from a 2 L culture volume. With a culture volume scale-up to 9 L, 16 g RLs were adsorbed, and the space-time yield (STY) increased by 31% to 0.21 gRL/L·h. We demonstrate a well-performing process design for biosurfactant production and recovery as a contribution to a vital bioeconomy.

SUBMITTER: Blesken CC

PROVIDER: S-EPMC7766737 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature

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Uncoupling Foam Fractionation and Foam Adsorption for Enhanced Biosurfactant Synthesis and Recovery.

Blesken Christian C CC Strümpfler Tessa T Tiso Till T Blank Lars M LM

Microorganisms 20201218 12

The production of biosurfactants is often hampered by excessive foaming in the bioreactor, impacting system scale-up and downstream processing. Foam fractionation was proposed to tackle this challenge by combining in situ product removal with a pre-purification step. In previous studies, foam fractionation was coupled to bioreactor operation, hence it was operated at suboptimal parameters. Here, we use an external fractionation column to decouple biosurfactant production from foam fractionation, ...[more]

PMID: 33353027

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Project description:Rhamnolipids are among the glycolipids that have been investigated intensively in the last decades, mostly produced by the facultative pathogen Pseudomonas aeruginosa using plant oils as carbon source and antifoam agent. Simplification of downstream processing is envisaged using hydrophilic carbon sources, such as glucose, employing recombinant non-pathogenic Pseudomonas putida KT2440 for rhamnolipid or 3-(3-hydroxyalkanoyloxy)alkanoic acid (HAA, i.e., rhamnolipid precursors) production. However, during scale-up of the cultivation from shake flask to bioreactor, excessive foam formation hinders the use of standard fermentation protocols. In this study, the foam was guided from the reactor to a foam fractionation column to separate biosurfactants from medium and bacterial cells. Applying this integrated unit operation, the space-time yield (STY) for rhamnolipid synthesis could be increased by a factor of 2.8 (STY = 0.17 gRL/L·h) compared to the production in shake flasks. The accumulation of bacteria at the gas-liquid interface of the foam resulted in removal of whole-cell biocatalyst from the reactor with the strong consequence of reduced rhamnolipid production. To diminish the accumulation of bacteria at the gas-liquid interface, we deleted genes encoding cell-surface structures, focusing on hydrophobic proteins present on P. putida KT2440. Strains lacking, e.g., the flagellum, fimbriae, exopolysaccharides, and specific surface proteins, were tested for cell surface hydrophobicity and foam adsorption. Without flagellum or the large adhesion protein F (LapF), foam enrichment of these modified P. putida KT2440 was reduced by 23 and 51%, respectively. In a bioreactor cultivation of the non-motile strain with integrated rhamnolipid production genes, biomass enrichment in the foam was reduced by 46% compared to the reference strain. The intensification of rhamnolipid production from hydrophilic carbon sources presented here is an example for integrated strain and process engineering. This approach will become routine in the development of whole-cell catalysts for the envisaged bioeconomy. The results are discussed in the context of the importance of interacting strain and process engineering early in the development of bioprocesses.

Project description:Biosurfactants are eco-friendly surface-active molecules recommended for enhanced oil recovery techniques. In the present study, a potential lipopeptide (biosurfactant) encoding the iturin A gene was synthesized from Bacillus aryabhattai. To improvise the yield of the lipopeptide for specific applications, current research tends toward engineering and expressing recombinant peptides. An iturin A gene sequence was codon-optimized, amplified with gene-specific primers, and ligated into the pET-32A expression vector to achieve high-level protein expression. The plasmid construct was transformed into an E. coli BL21 DE3 host to evaluate the expression. The highly expressed recombinant iturin A lipopeptide was purified on a nickel nitrilotriacetic acid (Ni-NTA) agarose column. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) revealed that the purity and molecular mass of iturin A was 41 kDa. The yield of recombinant iturin A was found to be 60 g/L with a 6.7-fold increase in comparison with our previously published study on the wild strain. The approach of cloning a functional fragment of partial iturin A resulted in the increased production of the lipopeptide. When motor oil was used, recombinant protein iturin A revealed a biosurfactant property with a 74 ± 1.9% emulsification index (E24). Purified recombinant protein iturin A was characterized by mass spectrometry. MALDI-TOF spectra of trypsin digestion (protein/trypsin of 50:1 and 25:1) showed desired digested mass peaks for the protein, further confirming the identity of iturin A. The iturin A structure was elucidated based on distinctive spectral bands in Raman spectra, which revealed the presence of a peptide backbone and lipid. Recombinant iturin A was employed for enhanced oil recovery through a sand-packed column that yielded 61.18 ± 0.85% additional oil. Hence, the novel approach of the high-level expression of iturin A (lipopeptide) as a promising biosurfactant employed for oil recovery from Bacillus aryabhattai is not much reported. Thus, recombinant iturin A demonstrated its promising ability for efficient oil recovery, finding specific applications in petroleum industries.

Dataset Information

Uncoupling Foam Fractionation and Foam Adsorption for Enhanced Biosurfactant Synthesis and Recovery.

Publications

Uncoupling Foam Fractionation and Foam Adsorption for Enhanced Biosurfactant Synthesis and Recovery.

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