Datensatz

Zusammenfassung

A quantitative understanding of intracellular processes, especially fluxes and metabolic regulation can help to optimize biotechnological production processes. Metabolic Flux Analysis is applied to quantify intracellular fluxes from extracellular concentration measurements and biomass growth. For correct calculation of biomass fluxes, the biomass composition has to be characterized. In a population of adherent cells, the cell size and the biomass composition are changing during time course of cultivation. In this work the biomass composition, focusing on protein, lipid, DNA and RNA content, of the adherent growing MDCK cell line was characterized during growth in different culture systems. Cultivations in six-well plates showed strong variations in biomass composition. In the attachment- and lag-phase the content of the most intracellular biomass components increased rapidly together with the cell size more than two-fold. Afterwards, an exponential decrease could be observed for the cell size and the contents of the most intracellular components. The ratios of protein, DNA and RNA to cell dry weight decreased while the lipid content was increasing. The growth behavior of the cells in a bioreactor cultivation using MicroHexTM microcarriers as attachment surface showed big differences compared to cultures in six-well plates. While ratios of DNA and RNA to cell dry weight were comparable, the lipid content was nearly doubled and the protein content was decreased. Presumably, the differences can be related to (1) the segmented growth area that leads to an uneven distribution of cells on microcarriers and (2) hydrodynamic forces that lead to an altered cell physiology (the maximum cell size was smaller in bioreactor cultures). The functional relation between biomass components and the cell volume distribution was analyzed using multiple linear regression. For both cultivation systems high functional relations could be observed (R2>0.9), which allows rapid estimation of the biomass components. However, the reconstruction of the cell volume from the size distribution showed that the arrangement of the distribution could be further optimized.