Abstract
Many efforts have been invested in the development of culture strategies for animal cell culture process. Several strategies have been studied: batch, fed-batch and perfusion cultures [1]. For implementation of those strategies, a monitoring system for automated, controlled and optimised processes based on simple measurement could be of great interest. Oxygen is a key substrate in animal cell metabolism and its consumption is thus a parameter of great interest for bioprocess monitoring and control. The application of the OUR (Oxygen uptake rate) is investigate here. The main advantages of OUR is that correlates well with the physiological state of cells and also for the prediction of viable cell concentration [2-4]. Different methods for the oxygen uptake rate (OUR) determination in animal cell cultivation have been developed: Dynamic estimation in the liquid phase, the global mass balance in the gas phase, and the Stationary liquid mass balance. Dynamic estimation has a considerable disadvantage because of disturbances suffered by the growing cells because of the necessary variations of the DO concentration. Gas phase balancing has several advantages; knowledge of the kL·a value is not necessary, and yields a higher density of accurate data. However, it has not historically been widely used due to the need for complex and expensive instrumentation like mass spectrometers and extremely accurate DO control systems. The Stationary liquid mass balance method offers minimum cell stress and greatest imation accuracy, but still needs for a significant investment in mass flow controllers as well as some additional instrumentation to determine the oxygen’s molar fraction in the gas phase. Background This works offers a simplified embodiment of the Stationary liquid mass balance method for the continuous estimate of the OUR estimation by means of the use of inexpensive proportional valves and the monitoring of their control signals is introduced and compared with the Global mass balance and Dynamic methods. As far said electrovalves can be considered to be linear, it can be demonstrated through the Mean value theorem for integrals that the mass balance equation for the liquid phase can be expressed as a first order differential equation:
Highlights
The simplified implementation of the liquid stationary mass balance method consisted on bypassing the Biostat B-plus system native DO control loop based on ON-OFF valves, in order to use two PWM (Pulse Width Modulation) controlled proportional valves to regulate the inlet flow of the supply gasses (Air/O2, N2), Figure 1-D
The results obtained for every experiment/estimation method displayed analogous oxygen uptake rate (OUR) graphs
When the remaining glucose was not enough to feed the current cell concentration a secondary metabolic path was activated and the lactate previously produced started being consumed by the cells
Summary
As far said electrovalves can be considered to be linear, it can be demonstrated through the Mean value theorem for integrals that the mass balance equation for the liquid phase can be expressed as a first order differential equation: dCL dt = kL · a · Where: CL [mol/l]: DO concentration in the liquid phase C*L [mol/l]: DO concentration in equilibrium with the gas phase kL·a [1/h]: Mass transfer coefficient a [ ]: PWM control signal or duty cycle considering the DO concentration control loop error signal for a given constant set point. The previous equation (i) can be rewritten as a function of the control loop parameters and a new expression of the OUR proportional to the duty cycle and the mass transfer coefficient can be proposed: de = dCsLp - dCL.
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