An optimized and fast-growing microbial culture is a dynamic process, and ensuring that the equipment is suitable for meeting these challenges is necessary for project success. These cultures tend to have a high oxygen demand that the fermenter must be able to meet to sustain cell growth and desired productivity. The culture also generates significant amounts of heat that the fermenter must remove to maintain temperature control. To ensure success during actual process runs, it is important to understand oxygen supply and heat removal capabilities before operating the process in the fermenter. Water-based tests were developed to characterize heat removal and oxygen supply capacity to understand if any equipment modifications would be required to meet process requirements. This testing should be performed early in technology transfer facility fit activities to allow time for equipment modifications, if needed.Microbial media generally are batched into the fermenter and then steam sterilized-in-place (SIP). The SIP cycle is automated to ensure control within the required temperature range. Temperatures outside the acceptable range could result in insufficiently sterilized medium or overheated medium. Additionally, evaporation or condensation can occur during the SIP cycle. The change in fermenter weight should be characterized or eliminated during SIP operations to ensure the target medium concentration for optimal growth is achieved. After sterilization, the fermentation medium often is held at specified conditions until inoculation. Determining the evaporation rate during the medium hold is essential for achieving the correct medium concentration at the time of inoculation. To compensate for the evaporation rate, additional water can be added during media preparation, and adjustments to the air flow rate can be made to minimize evaporative losses. Water-based testing was developed to understand parameters around media SIP and hold conditions to ensure the target starting media concentration would be met.
Materials and Methods
Heat Removal Characterization
in which m is the mass of water in the fermenter (kg), C p is the heat capacity of water at 37 °C (4.181 kJ/kg/°C),1 T is the temperature of water in the fermenter (°C), t is time (h), and dT/dt is the rate of temperature change as the slope of water temperature versus time curve.
Strategies for maintaining temperature control involve equipment modifications such as: lowering chilled water or glycol temperature to increase the driving force for heat removal (dT/dt), ensuring jacket and coils have minimum resistance, and adding internal cooling coils to increase heat transfer surface area.
The fermenters were filled to different target weights and heated to approximately 75 °C. The temperature set point was changed to 10 °C, which resulted in a 100% cooling output on the temperature control loop. The rate of temperature change was estimated by the slope of the linear portion of the temperature-versus-time curve from 41 °C to 34 °C. This range encompasses the process temperature set point of 37 °C. The HTR was then calculated using Equation 1.