The Role Of Dissolved Oxygen Probes In Bioreactors

Dissolved oxygen (DO) probes are key components in bioreactors, ensuring optimal conditions for cell growth and biochemical reactions. These specialized sensors measure and monitor the oxygen levels within the bioreactor, providing the user with real-time data to maintain ideal oxygen levels. Proper oxygen levels are vital for maximizing cell health, yield, and productivity in bioprocesses such as cell culture and fermentation. Effective oxygen control not only improves bioreactor performance but also reduces risks such as oxygen depletion or oversaturation. Therefore, dissolved oxygen probes are indispensable tools for high-quality and efficient bioproduction in bioreactors. 

Dissolved oxygen (DO) is an important parameter that determines the success of bioprocesses within bioreactors. Whether that be cultivating mammalian cells, producing biochemical substances, or fermenting microorganisms to brew beer, controlling and maintaining DO levels is fundamental. However, these microbial strains and cell lines have specific oxygen level limits, demanding precise monitoring and control systems. 

What Is A Bioreactor?

A bioreactor is a system designed to create an optimal and controlled environment for biological and chemical reactions, facilitating the growth of microorganisms, cells, tissues, and algae. Widely used across industries like biotechnology, pharmaceuticals, food production, and wastewater treatment, bioreactors are valued for their efficiency, cost-effectiveness, and precise control over growth conditions. 

Why Is Oxygen Important In Bioreactors?

Different fermentation processes and cultures require specific oxygen conditions. 

Aerobic Cultures

These cultures are essential for mammalian cell cultures and many microbial cultures including E. coli. Aerobic cultures generally need a DO concentration above 30%. This high DO level supports robust cell growth and metabolism, leading to a productive production process. 

Anaerobic Fermentation

As the name suggests, this process occurs when there is no oxygen. Anaerobic microbes that thrive in oxygen-free environments that are also important for producing chemicals include lactic acid, ethanol, and acetic acid. Accurately measuring DO levels helps prevent any unintended oxygen interference, which could disrupt chemical production.

Microbial Fermentation 

This process involves lower DO concentrations and is commonly used in bacterial fermentation such as producing probiotics and cultivating lactic acid bacteria. The low DO concentration is key for microbial activities. 

Dissolved Oxygen Control In Bioreactors

To achieve optimal culture growth and high productivity, maintaining accurate DO levels is vital. The growing cells inside a bioreactor continuously consume oxygen, which demands a robust dissolved oxygen control system. A typical system involves many components that work together, including: 

• Dissolved oxygen sensors: At the heart of the bioreactor is one of the most important pieces of equipment. Dissolved oxygen probes measure the DO level/concentration in real time. When connected to the control software, DO probes act as the feedback mechanism for monitoring and maintaining stable oxygen levels.

• Impellers: These agitation devices are important in mixing the culture and gases evenly to ensure a uniform distribution of oxygen through the bioreactor chamber.
• Gas spargers: These introduce gases like air or oxygen into the bioreactor to adjust the DO level. In anaerobic environments/processes, gases such as nitrogen are used to displace the oxygen in the bioreactor.
• Control software: This is crucial for receiving data from the dissolved oxygen sensors and adjusting the gas composition, agitation, and gas flow to keep DO levels within the required range. 

Dissolved Oxygen Control 

Efficient DO control requires deep understanding and the optimization of key parameters. 

DO Cascades

A DO cascade is a control strategy that defines the sequence of actions the bioreactor takes to maintain stable oxygen levels. In a typical cascade system, various actuators like impellers and gas spargers are adjusted in a pre-defined order based on the DO readings. 

For example, an increase in agitation might be the first step to raise DO levels, followed by increased airflow, and lastly, the introduction of pure oxygen if needed.

Gas Sparger Types

Spargers play a critical role in controlling DO levels by introducing gases into the bioreactor. The type of sparger can significantly influence gas transfer efficiency:

• Porous Spargers: Create fine bubbles, which provide a larger surface area for gas transfer and are effective for oxygenation in aerobic cultures.

• Open-Pipe Spargers: Produce larger bubbles, which rise more quickly and are ideal for stripping gases like carbon dioxide in some fermentation processes.

Selecting the right sparger type depends on the specific needs of the bioprocess, such as whether fine control over DO levels or rapid degassing is required.

Gassing Techniques

The composition and flow of gases introduced into the bioreactor significantly affect the DO environment:

• A balanced mixture of oxygen and nitrogen is commonly used for standard aerobic cultures.
• Pure Oxygen Injection: Employed when high oxygen levels are needed rapidly. It’s typically used in high-density cultures where oxygen demand is high.
• Nitrogen Sparging: Used to reduce oxygen levels in anaerobic conditions or for stripping dissolved gases.

Effective DO control relies on a nuanced understanding of how different gases interact with the bioreactor’s microbial environment.

The Importance Of Impellers 

Impellers not only promote efficient mixing but also influence oxygen transfer rates:

• Rushton Turbine Impellers: Provide high shear and are effective in creating turbulence, and enhancing oxygen dissolution.
• Marine Propeller Impellers: Offer gentle mixing, suitable for fragile mammalian cell cultures that could be damaged by excessive shear forces.

The choice of impeller directly impacts the oxygen transfer rate (OTR) and overall mixing efficiency, making it a critical consideration for DO control.

What Is 100% DO?

In bioprocessing, “100% DO” refers to the saturation point of oxygen in the medium, where the liquid can hold no more dissolved oxygen under specific conditions. It’s important to distinguish this from the absolute oxygen concentration, which varies with temperature, pressure, and salinity. Proper calibration of DO probes ensures that readings reflect the true oxygen availability for the cultured cells.

Types Of DO Sensors: Polarographic vs. Optical

Choosing the right DO sensor can significantly influence measurement accuracy and reliability. Here are two types of dissolved oxygen probes, mainly used in bioreactors:

• Polarographic DO Sensors: Utilize a membrane-covered electrode. These sensors are reliable and cost-effective, but they require regular maintenance and calibration.
• Optical DO Sensors: Use a fluorescence-based technique to measure oxygen. They offer high stability, low maintenance, and faster response times compared to polarographic sensors. However, they are often more expensive.

Optical sensors are becoming increasingly popular due to their robustness and reduced need for frequent calibration.

The Importance Of kLa & OTR In DO Control

Two critical parameters, kLa (volumetric mass transfer coefficient) and OTR (oxygen transfer rate) are essential for understanding and optimizing oxygen dynamics in bioreactors:

• kLa:  Reflects the efficiency of oxygen transfer from the gas phase to the liquid phase. A higher kLa indicates better oxygen dissolution, which is crucial for high-density cultures.
• OTR: Measures the rate at which oxygen is transferred to the culture medium. It’s influenced by factors such as agitation speed, gas flow rate, and the presence of surfactants.

By monitoring kLa and OTR, bioprocess engineers can fine-tune bioreactor conditions to optimize oxygen availability for cell growth.

Enhancing DO Regulation In Bioreactor Control Systems

Advanced bioreactor control systems, like those from Eppendorf, provide comprehensive solutions for DO management:

• Compatibility: These systems can work with both polarographic and optical DO sensors, offering flexibility based on the specific application.
• Customization: Users can set up detailed DO cascades, customizing agitation, gas sparging, and gassing strategies for precise control.
• Automation: Features like Auto Calibrate allow for quick and consistent calibration, enhancing reproducibility across batches.

Practical Applications: DO Monitoring Beyond Bioreactors

The importance of dissolved oxygen monitoring extends beyond traditional bioreactors:

• Dissolved oxygen in aquaculture: Maintaining optimal DO levels prevents fish suffocation and supports healthy aquatic environments.
• Dissolved oxygen in wastewater treatment: DO levels impact microbial activity needed for waste decomposition, affecting treatment efficiency.
• Dissolved oxygen in fermentation industries: From brewing to pharmaceutical production, DO control is crucial for consistent product quality and yield.

How To Avoid Common Errors In DO Measurement

Accurate DO readings are vital for maintaining optimal conditions. Here are some common sources of error and how to avoid them:

• Air Bubbles: These can interfere with sensor readings, leading to inaccurate data. Anti-bubble sensors or careful placement of probes can mitigate this issue.

• Fouling: Sensors can become coated with biomass or media, affecting readings. Regular cleaning and maintenance help maintain accuracy. We also recommend adding an anti-fouling probe screen.
  
• Calibration Drift: Frequent calibration is necessary to account for sensor drift over time, especially with polarographic sensors.

Care & Maintenance Tips For DO Probes

Proper maintenance of dissolved oxygen probes extends their lifespan and ensures accurate measurements:

• Calibration: Perform regular calibration to adjust for environmental changes. Daily calibration is recommended for high-precision bioreactor applications.
• Membrane Replacement: For galvanic sensors, change the permeable membrane every 4-8 weeks to maintain optimal performance.
• Cleaning: Keep sensors clean, especially in media-rich environments, to prevent fouling and maintain consistent readings.

Maintaining DO For Optimal Bioprocess Outcomes

To achieve high productivity and consistent results in bioprocessing, accurate DO measurement and control are non-negotiable. 

Utilizing advanced DO probes like those from Atlas Scientific provides the reliability needed for sophisticated bioreactor systems. By combining cutting-edge sensors with optimized DO control strategies, bioprocess engineers can ensure that their cultures thrive under ideal conditions.

Summing Up

As bioprocessing technologies evolve, the role of dissolved oxygen probes will continue to expand. With the increasing complexity of cell cultures and the demand for higher yields, precise DO control is more critical than ever. 

If you would like to learn more about dissolved oxygen in bioreactors or what dissolved oxygen would best suit your system, do not hesitate to contact the world-class team at Atlas Scientific.