Challenges in meeting the mass transfer demands of high‐density perfusion cultures in single‐use bioreactors

Increasing protein therapeutics production by adopting intensified perfusion cultures places high demand on the mass transfer capabilities of single‐use bioreactors (SUB). Here, we compare these capabilities for four SUB designs against worst‐case O 2 transfer requirements for supporting Chinese hamster ovary (CHO) perfusion cultures. Impact of power dissipation, gas flow rate, O 2 enrichment, pCO 2 and sparge‐hole size on O 2 and CO 2 mass transfer (OTR and CTR) was characterized. Only one design delivered sufficient O 2 and removed sufficient CO 2 to support a worst‐case cell line at 100 × 10 6 viable cells/mL with a single‐sparger at both 50 and 1000 L scale. Larger‐scale versions of 3 designs were also tested. Performance at a 50‐L scale was not predictive of large‐scale performance (200 or 1000 L) for any design. Changes in the volume and composition of the gas phase during residence in the vessels were characterized. Substantial changes were found. Calculations of predicted culture pCO 2 were performed across the design space, assuming a respiratory quotient of 1. A response surface was constructed to predict culture pCO 2 and used to examine the consequences of various strategies for increasing OTR. Most strategies disproportionately increase OTR relative to CTR, leading to an increase in culture pCO 2 . The implications on SUB design and scale‐up for high‐intensity perfusion cultures are discussed.