Homogenisation and oxygen transfer rates in large agitated and sparged animal cell bioreactors: Some implications for growth and production

Abstract

Because of concern for cell damage, very low agitation energy inputs have been used in industrial animal cell bioreactors, typical values being two orders of magnitude less than those found in bacterial fermentations. Aeration rates are also very small. As a result, such bioreactors might be both poorly mixed and also unable to provide the higher oxygen up-take rates demanded by more intensive operation. This paper reports experimental studies both of K( L ) a and of mixing (via pH measurements) in bioreactors up to 8 m(3) at Wellcome and of scaled down models of such reactors at Birmingham. Alongside these physical measurements, sensitivity of certain cell lines to continuously controlled dO(2) has been studied and the oxygen up-take rates measured in representative growth conditions. An analysis of characteristic times and mixing theory, together with other recent work showing that more vigorous agitation and aeration can be used especially in the presence of Pluronic F-68, indicates ways of improving their performance. pH gradients offer a special challenge.

Reactor Engineering in Large Scale Animal Cell Culture

“…However, as long ago as 1968, Kilburn and Webb (1968) showed that Pluronic F 68 (a block co-polymer of propylene oxide and ethylene oxide) was effective in decreasing cell damage. Such effectiveness was subsequently shown in many of the above studies, involving both stirred (Oh et al 1989(Oh et al , 1992Kioukia et al 1996;Nienow et al 1996) and bubble column bioreactors (Handa-Corrigan et al 1989;Kioukia et al 1992;Meier et al 1999).…”

Control of pH in large-scale, free suspension animal cell bioreactors: Alkali addition and pH excursions

“…Thus, this paper is relevant to both pH reduction due to CO 2 and to pH homogeneity and covers the experimental simulation of pH excursions as a result of alkali addition. It complements recent flow visualization of such phenomena (Nienow et al, 1996), the geometry and operating conditions here being very similar and based on the industrial-scale 8-m 3 bioreactors used by The Wellcome Foundation (now Glaxo Wellcome) (Langheinrich et al, 1998).…”

“…Aeration rates, because of the low oxygen demand of the current cell lines, are also very small (Nienow et al, 1996). Thus, the typical large animal cell bioreactor (Nienow et al, 1996) of 2-m diameter, T 200 , and of 8-m 3 in volume, V, when operated up to a liquid height, H (m), of H ‫ס‬ 1.3T, was agitated by a Rushton turbine of diameter, D (m), of 0.225T which is very small compared with the usual configuration of D ‫ס‬ 0.33T. The impeller was also positioned rather low in the vessel with a clearance, C (m), from the base C ‫ס‬ 0.225T rather than C ‫ס‬ 0.33T which is usually used and the agitator speed was a maximum of 1 s −1 if used continuously.…”

“…To study the development of pH excursions during the sparging of the gas mixture, in addition to the concept of constant vvm for assessing the desired flow rate, it is interesting to consider simulating the metabolism associated with CO 2 evaluation. The latter was calculated from the specific oxygen utilisation rate (SOUR) of NS0 cells which was 8.4 × 10 −17 mol cell −1 s −1 for a concentration, C C of 2 × 10 12 cells m −3 (Nienow et al, 1996). Since the ratio between the CO 2 accumulation rate (CAR mol cell −1 s −1 ) and OUR is around unity (Bonarius et al, 1995) the corresponding superficial gas velocity can be calculated from…”

“…Not only will these systems require a greater supply of nutrients (for example, glucose and glutamine as well as oxygen), they will also put greater demands on maintaining the homogeneity of the cells environment in the bioreactor. In addition to the well-known process limitation caused by an insufficient oxygen supply (Lin et al, 1993), media homogenization has also recently been shown to be of great importance (Kiss et al, 1994;Nienow et al, 1996;Langheinrich et al, 1998). A further important problem associated with the application of higher cell density systems and homogeneity is the reduction in pH, caused mainly by the increased carbon dioxide accumulation.…”