Substrate gradients in bioreactors: origin and consequences

Gradients of glucose in time and space are shown in a 30 m³ cultivation of Saccharomyces cerevisiae grown in minimal medium to a cell density of 20 gl^–1. The fed-batch concept was used with glucose as the limiting component which was fed continuously to the process. As the mean glucose concentration declined throughout the process, the level of glucose was at all times different in three sampling ports (bottom/middle/top) of the reactor. These gradients were furthermore shown to depend on the feed position. This means that if the feed was supplied in the relatively stagnant mixing zone above the top impeller, the gradients were more pronounced than by feed in the well mixed bottom impeller zone. A rapid sampling system was constructed, and continuous glucose samples of every 0.15 s were analysed from a point of the reactor. Fifty samples were collected with this system, but the amount and frequency is possible to change. The results of these series show a variance of the glucose concentration where at one stage, a peak appeared of a relative difference in concentration of 40 mgl^–1. The pattern of these rapid glucose fluctuations was shown to depend on the turbulence level at the location of the feed. It was shown, that the fluctuations were more pronounced when the feed was localised in a relatively stagnant area than in the well-mixed impeller area, where the deviation from the mean was negligible. The fluid flow, in the impeller (gassed and ungassed) and bulk area (ungassed) of the reactor, was characterised by turbulence measurements using thermal anemometry. These types of areas resembles well the different areas of sampling as mentioned above. The turbulent frequencies in these areas were in the range of 10^–1 to 10⁴ Hz with the highest amplitudes at low frequencies. The spectra depicts a uniform time scale for all zones, especially at the low frequencies. The dominance of low frequency, high amplitude flow variations and the observed short-time oscillations in substrate concentration support the hypothesis of substrate transport over fairly long distances without substantial mixing both in the impeller, but especially, in the bulk zone of the reactor.Simulations with an integrated CFD and biokinetic model were performed. The predictions of the glucose gradients of this model were compared to measurements.This project was supported by a grant from the Nordic Programme on Bioprocess Engineering under the auspices of NI, the Nordic Fund for Technology and Industrial Development and NUTEK, The Swedish Board for Technical and Industrial Development. This investigation was possible to perform due to the support from Statoil Biosentrum in Stavanger, Norway, that provided their pilot plant. Especially the cooperation with Lars Aasberg and Andreas Rag is greatly acknowledged.