On-line High Performance Liquid Chromatography Measurements of Extracellular Metabolites in an Aerobic Batch Yeast (Saccharomyces cerevisiae) Culture
2011
Tohmola, Niina, University of Helsinki, Helsinki, Finland | Ahtinen, Jouni, Medicel Oy, Keilaranta, Espoo, Finland | Pitkanen, Juha-Pekka, Medicel Oy, Keilaranta, Espoo, Finland | Parviainen, Ville, University of Helsinki, Helsinki, Finland | Joenvaara, Sakari, Medicel Oy, Keilaranta, Espoo, Finland | Hautamaki, Mika, Medicel Oy, Keilaranta, Espoo, Finland | Lindroos, Peter, Medicel Oy, Keilaranta, Espoo, Finland | Makinen, Jarno, Medicel Oy, Keilaranta, Espoo, Finland | Renkonen, Risto, University of Helsinki, Helsinki, Finland
We constructed a bioprocess environment enabling automatic sampling from a bioreactor combined with a compact on-line high performance liquid chromatography (HPLC) unit. This setup allowed us to measure extracellular glucose, ethanol, glycerol, and acetate concentrations automatically at 5 min intervals during the cultivation. This environment also provides mechanical measurement of the optical density (OD) of cells and enables us to collect and store (-35℃) samples for further off-line analyses. Among the available devices, the performance of the sampling-analysis unit is by far the best with regard to speed and number of analytes. Both the sampling and analysis phases are easily controlled by software; thus, providing a unique environment to perform various bioprocess activity tasks, whether they would be cell line screening or optimisation of conditions for growth and productivity. Complex research set-ups can be created and continuous automated measurements empower long-term cultivations with a time series. We provide evidence for the applicability of this environment by performing three comparable batch cultivations with Saccharomyces cerevisiae yeast and show that both the on-line sampling and analysis modes produce reliable data for further use in the monitoring and controlling of bioprocesses. On-line data provided new insight into the dynamics of the diauxic shift during aerobic glucose batch cultivation. When cell growth and carbon dioxide production ceased for the first time during the diauxic shift, acetate accumulation and consumption of the remaining glucose below 0.15 g/L continued to occur for 1 h. At the same time, glycerol and ethanol began to be consumed. Samples were also collected during cultivation for later analysis of intracellular metabolites and to collect more valuable information about metabolism.
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