Oxygen transfer is a critical limitation in aerobic fermentations directly impacting cell growth, productivity and bioprocess efficiency. The addition of pressure to a bioreactor contributes to increasing the Oxygen Transfer Rate (OTR) and therefore gas solubility in the fermentation broth. More bioprocess scientists are turning to pressurized systems to increase the availability of dissolved gases and accelerate gas-dependent bioprocesses.

This application note demonstrates how operating at elevated pressure significantly enhances dissolved oxygen availability and OTR (k_La), using the Bioxplorer 400P high-pressure parallel bioreactor system. By increasing pressure, oxygen solubility in the fermentation broth improves, enabling faster and more efficient bioprocesses. The Bioxplorer 400P allows precise control of dissolved oxygen via pressure offering a powerful alternative to traditional agitation-based strategies.

k_La is the volumetric oxygen mass transfer coefficient, measuring the efficiency of oxygen transfer from bubbles to liquid in the bioreactor. In practical terms, k_La determines how quickly oxygen becomes available to cells, making it a critical parameter for aerobic fermentation performance, productivity, and scale-up.
Higher operating pressure increases oxygen solubility in the fermentation broth, which can enhance oxygen transfer
and improve overall process efficiency.
Previous studies using the H.E.L bench-top bioreactor systems (BioXplorer 100 & 400) have shown that a constant k_La can
be used as a basis for scale-up, as the values obtained can be accurately reproduced in conventional scale bioreactors
(Gill et al., 2008). In these studies, kLa is used as a measure of how well oxygen is transferred to the liquid under pressure.

The BioXplorer 400P is an automated 4-zone parallel pressure bioreactor system used for the discovery and optimization of novel bioprocesses.
Designed for applications ranging from syngas fermentation to cell-free enzymatically catalyzed processes, the BioXplorer 400P uses pressure and precise gas feed control to accelerate bioprocesses that depend on the availability of dissolved gases

Experiments were designed to evaluate the impact of pressure on oxygen transfer under controlled conditions.
• Reactor: 500 mL stainless steel reactor
• Working volume: 300 mL w/v dilute solution of sodium sulphite.
• Gas: 1 vvm (300 sccm) air gas blend containing H₂.
• Temperature: 37 °C.
• Stirring: 1500 rpm.
The BioXplorer 400P stainless steel pressure vessel was fitted with gas sparger, pH
probe, DO probe, overhead motor and magnetic drive with Ruston impeller (Figure 2).
The system was cycled between gassing with air and scrubbing with nitrogen with increasing pressure.

Figure 3 shows the dissolved oxygen (DO) profile of a reactor while being cycled between gassing with air and
with nitrogen. DO in the liquid increases as pressure increases within the reactor.

The gradient of the DO profile represents the rate of change of oxygen concentration, which is a measure of oxygen
flux (Figure 4). By using the fact that 100% saturation corresponds to an oxygen concentration of ~9 mg/L (at room
temperature), the corresponding oxygen flux profile (in grams per liter per minute) can be generated.

Pressure can be used to manipulate the oxygenation level. This would traditionally be achieved by manipulating stirrer speed. However, this is not an option for more fragile organisms, for example, algae, amoeba, etc., due to shear
forces that affect cell integrity. In the experiment, represented by the data below (Figure 5), a control loop has been
created that links a requested DO set point with the control output to a back-pressure regulator (BPR). The pressure
in the system will vary because of the automatic regulation of the BPR, which is not, however, explicitly manipulated.

Data was processed by H.E.L analysis tools. Here, each colored section represents a separate curve-fitting region. Markers represent measured data points. Each fitted curve yields a separate estimate of kLa. Each of the k_La values shows a high degree of reproducibility, with k_L a = 2.0 min-1 +/- 10% – i.e. independent of pressure.

In the first 250 minutes of the experimental run, the DO set point varied between 120 and 250%. These DO intervals
within the reactor were easily reached and maintained automatically with the help of the BPR, regulating pressures
between 0.4 and 2.5 bar. The DO set point was kept stable after 250 minutes, and the stability of DO was tested with
the addition of sodium sulphite, which removes oxygen from the liquid in an oxygen scavenging reaction. Once sodium sulphite was fed, the BPR effectively increased the pressure in the reactor to keep DO at the set point until the
end of the experiment.

According to Henry’s Law, the DO concentration (but not k_La) scales approximately linearly with oxygen partial pressure. In our high-pressure experiments, the DO value improved several-fold and showed a near-proportional relationship with applied pressure.
Operating a bioprocess at elevated pressure (e.g. up to 10 bar) using the BioXplorer 400P can therefore deliver substantial improvements in oxygen transfer. Achieving similar gains through k_La optimization alone would be significantly
more challenging, due to the influence of additional factors such as gas–liquid interfacial area, bubble size, and agitation. By using pressure instead of increased agitation, the risk of shear stress can be minimized, helping to protect sensitive cultures.
Our results demonstrate that applying pressure in the BioXplorer 400P bioreactors increases DO levels and enhances
gas transfer rates, providing a robust approach to improving yield, productivity, and biomass growth in bioprocesses.