A simplified implementation of the stationary liquid mass balance method for on‐line OUR monitoring in animal cell cultures

Abstract

BACKGROUND Compared with other methods, the stationary liquid mass balance method for oxygen uptake rate (OUR) determination offers advantages in terms of estimation accuracy and reduction of stress. However, the need for sophisticated instrumentation, like mass flow controllers and gas analysers, has historically limited wider implementation of such a method. In this paper, a new simplified method based on inexpensive valves for the continuous estimation of OUR in animal cell cultures is evaluated. The determination of OUR values is based on accurate operation of the dissolved oxygen (DO) control loop and monitoring of its internal variables. RESULTS The method developed was tested empirically in 2 L bioreactor HEK293 batch cultures. OUR profiles obtained by a dynamic method, global mass balance method and the developed simplified method were monitored and compared. The results show how OUR profile obtained with the proposed method better follows the off‐line cell density determination. The OUR estimation frequency was also increased, improving the method capabilities and applications. The theoretical rationale of the method was extended to the sensitivity analysis which was analytically and numerically approached. CONCLUSIONS The results showed the proposed method to be not only cheap, but also a reliable alternative to monitor the metabolic activity in bioreactors in many biotechnological processes, being a useful tool for high cell density culture strategies implementation based on OUR monitoring. © 2017 Society of Chemical Industry

The Effect of the Expression of the Antiapoptotic BHRF1 Gene on the Metabolic Behavior of a Hybridoma Cell Line

“…This method is based on interrupting the air inflow to the bioreactor for a few minutes; then, the dissolved oxygen concentration decreases due to the respiratory activity of the cells and to the oxygen desorption from the liquid phase to the bioreactor headspace. The specific methodology used in this work is detailed in the literature [31,32].…”

Non-disruptive method to estimate the volumetric oxygen mass transfer coefficient (kLa) throughout a fermentation

“…OTR is most often the rate-limiting step due to the low solubility of oxygen in aqueous solutions, such as fermentation media [5,6]. To determine the OTR, it is necessary to know the volumetric mass transfer coefficient, k L a, which subsequently controls OUR [7]. Yet, current methods to obtain k L a have significant limitations, 1) not amenable to real-time measurement; 2) contributes to DO stress; 3) low time resolution; or 4) requires inlet and off-gas mass spectrometry equipment [7][8][9][10].…”

“…To determine the OTR, it is necessary to know the volumetric mass transfer coefficient, k L a, which subsequently controls OUR [7]. Yet, current methods to obtain k L a have significant limitations, 1) not amenable to real-time measurement; 2) contributes to DO stress; 3) low time resolution; or 4) requires inlet and off-gas mass spectrometry equipment [7][8][9][10]. Further, k L ais sensitive to many culture parameters, such as antifoam additions, bioreactor shape, cell density, media characteristics, and stir speed.…”

“…The k L acoefficient is determined once the gas flow is restored and the DO increases with time. As the dynamic method is disruptive, k L a is infrequently determined in production vessels due to the risk of culture loss [7]. The global mass balance method subtracts the inlet and outlet oxygen concentrations assessed by off gas analyzers [7].…”

“…As the dynamic method is disruptive, k L a is infrequently determined in production vessels due to the risk of culture loss [7]. The global mass balance method subtracts the inlet and outlet oxygen concentrations assessed by off gas analyzers [7]. The stationary liquid mass balance method uses the difference between the liquid oxygen saturation and measured DO.…”