Modelling of shear sensitive cells in stirred tank reactor using computational fluid dynamics

摘要

Animal cells are often cultured in stirred tank reactors. Having no cell wall, these animal cells are very sensitive to the fluid mechanical stresses that result from agitation by the impeller and from the rising and bursting of bubbles, which are generated within the culture medium in the stirred tank to supply oxygen by mass transfer to the cells. If excessive, these fluid mechanical stresses can result in damage/death of animal cells. Stress due to the rising and bursting of bubbles can be avoided by using a gas-permeable membrane, in the form of a long coiled tube (with air passing through it) within the stirred tank, instead of air-bubbles to oxygenate the culture medium. Fluid mechanical stress due to impeller agitation can be controlled using appropriate impeller rotational speeds. The aim of this study was to lay the foundations for future work in which a correlation would be developed between cell damage/death and the fluid mechanical stresses that result from impeller agitation and bubbling. Such a correlation could be used to design stirred-tank reactors at any scale and to determine appropriate operating conditions that minimise cell damage/death due to fluid mechanical stresses.Firstly, a validated CFD model of a baffled tank stirred with a Rushton turbine was developed to allow fluid mechanical stresses due to impeller agitation to be estimated. In these simulations, special attention was paid to the turbulence energy dissipation rate, which has been closely linked to cell damage/death in the literature. Different turbulence models, including the k-ε, SST, SSG-RSM and the SAS-SST models, were investigated.All the turbulence models tested predicted the mean axial and tangential velocities reasonably well, but under-predicted the decay of mean radial velocity away from the impeller. The k-ε model predicted poorly the generation and dissipation of turbulence in the vicinity of the impeller. This contrasts with the SST model, which properly predicted the appearance of maxima in the turbulence kinetic energy and turbulence energy dissipation rate just off the impeller blades. Curvature correction improved the SST model by allowing a more accurate prediction of the magnitude and location of these maxima. However, neither the k-ε nor the SST models were able to properly capture the chaotic and three-dimensional nature of the trailing vortices that form downstream of the blades of the impeller. In this sense, the SAS-SST model produced more physical predictions. However,this model has some drawbacks for modelling stirred tanks, such as the large number of modelled revolutions required to obtain good statistical averaging for calculating turbulence quantities. Taking into consideration both accuracy and solution time, the SSG-RSM model was the least satisfactory model tested for predicting turbulent flow in a baffled stirred tank with a Rushton turbine.In the second part of the work, experiments to determine suitable oxygen transfer rates for culturing cells were carried out in a stirred tank oxygenated using either a sparger to bubble air through the culture medium or a gas-permeable membrane. Results showed that the oxygen transfer rates for both methods of oxygenation were always above the minimum oxygen requirements for culturing animal cells commonly produced in industry, although the oxygen transfer rate for air-bubbling was at-least 10 times higher compared with using a gas-permeable membrane. These results pave the way for future experiments, in which animal cells would be cultured in the stirred tank using bubbling and (separately) a gas-permeable membrane for oxygenation so that the effect of rising and bursting bubbles on cell damage/death rates can be quantified. The effect of impeller agitation on cell damage/death would be quantified by using the gas permeable membrane for oxygenation (to remove the detrimental effects of bubbling), and changing the impeller speed to observe the effect of agitation intensity.In the third and final part of this work, the turbulent flow in the stirred tank used in the oxygenation experiments was simulated using CFD. The SST turbulence model with curvature correction was used in these simulations, since it was found to be the most accurate model for predicting turbulence energy dissipation rate in a stirred tank. The predicted local maximum turbulence energy dissipation rate of 8.9x10¹ m2/s3 at a rotational speed of 900 rpm was found to be substantially less than the value of 1.98x10⁵ m2/s3 quoted in the literature as a critical value above which cell damage/death becomes significant. However, the critical value for the turbulence energy dissipation rate quoted in the literature was determined in a single-pass flow device, whereas animal cells in a stirred tank experience frequent exposure to high turbulence energy dissipation rates (in the vicinity of the impeller) due to circulation within the stirred tank and long culture times. Future cell-culturing experiments carried out in the stirred tank of this work would aim to determine a more appropriate critical value for the turbulence energy dissipation rate in a stirred tank, above which cell damage/death becomes a problem.