Growth and docosahexaenoic acid production performance of the heterotrophic marine microalgae Crypthecodinium cohnii in the wave‐mixed single‐use reactor CELL‐tainer

Abstract

The process development of docosahexaenoic acid (DHA) with the heterotrophic marine microalgae Crypthecodinium cohnii is challenging, since the oxygen demand of the cells is high and the cell itself sensitive to high shear forces. Furthermore, the marine medium, which comprises a high salt concentration, accelerates corrosion of steel typically applied in bioreactors. Single‐use bioreactors (SUBs) provide an alternative to common stainless steel stirred tank reactors (STR) for the development of bioprocesses with marine microorganisms, when the development stage does not allow for the application of special steel or the replacement of chloride ions in the medium. Among the readily‐available SUBs, the CELL‐tainer exhibits outstanding high oxygen transfer rates. A direct comparison of this SUB with a standard stirred tank bioreactor (STR) was performed for the cultivation of C. cohnii. The biomass and DHA concentrations were similar in both devices, while the biomass yield coefficient, related to the glucose consumption, was reduced by 35% and the DHA yield coefficient by 21% in the STR. The impact of shear stress on cell physiology was elucidated by flow cytometry, revealing a five times higher amount of cell debris in the STR and 14 times higher amount of cells with a low cell membrane potential as evaluated by bis‐(1,3‐dibutylbarbituric acid) trimethine oxonol staining. Foam and biofilm formation, which is a problem in the STR, did not occur in the SUB. The study underlines that the CELL‐tainer can replace the STR, ensuring sufficient oxygen transfer rates and decreasing cell damage and foam formation.

Separation of Heterotrophic Microalgae Crypthecodinium cohnii by Dielectrophoresis

“…The cell populations showed comparable results with previous studies in all cultivations ( Hillig et al, 2014 ; Marba-Ardebol et al, 2017 ; Niehaus, 2019 ). It is evident from Figure 2A that glucose was fully consumed on day 4 of the cultivation and that no major growth-supporting carbon source was available in the medium from that day on.…”

“…C. cohnii populations (strain PGM-1, ATCC 30772) were cultivated in sterile 100 ml Erlenmeyer flasks in M1 cultivation medium following a protocol of Hillig ( Hillig et al, 2014 ). 2.5 ml of preculture were diluted in 25 ml M1 medium for this purpose.…”

Characterization of the Metabolic Response of Streptomyces clavuligerus to Shear Stress in Stirred Tanks and Single-Use 2D Rocking Motion Bioreactors for Clavulanic Acid Production

“…In STRs, the increase of k L a implies vigorous stirring to favor mass transfer from gas to liquid phase by increasing the turbulence of the system, thus increasing shear stress on the medium and cells [27,28]. In contrast, wave and rocking-motion reactors are characterized by up to 100-fold lower shear forces compared to STRs [21,22,29] and especially 2D rocking-motion bioreactors provide higher k L a values compared to simple wave systems that are comparable to those obtained in STRs at high stirring speed [23][24][25].…”

“…In particular, 2D rocking-motion bioreactors, which create a wave or cause eddy formation at the vessel wall for the suction of air into the liquid phase, exhibited comparably higher gas mass transfer rates while mechanical shear forces are considerably lower than in stirred tank bioreactors. This leads to an altered micromorphology and subsequently different growth and product synthesis rates as observed in cultivations performed in the two-dimensional rocking-motion bioreactor concept CELL-tainer® [19,23,24]. In this regard, the filamentous fungus Aspergillus niger (A. niger) cultivated in this low shear-force environment, showed a similar macromorphology as in shake flask cultures, while large pellets were formed.…”

Editorial: Single‐use technology in biopharmaceutical manufacturing

“… prove that single‐use systems are advantageous for the rapid realization of vaccine productions up to biosafety level 3. Two research articles comparing product expressions in stirred and wave‐mixed single‐use bioreactors and three Research Articles describing the scaling‐up of stirred single‐use bioreactors follow. Minow et al.…”