Effect of hyperosmotic pressure on growth of recombinant Chinese hamster ovary cells and Erythropoietin (EPO) production was investigated. Cells were cultivated in batch modes at various osmolalities. When the osmolality increased from 314 to 463mOsm/Kg, specific EPO productivity $(q_p)$ was increased up to 1.6-fold but cell growth was inhibited.
EPO has a complex oligosaccharide structure that plays an important role in biological activity in vivo. To investigate the influence of hypoerosmotic pressure on the glycosylation, structural analysis of oligosaccharide was carried out. Recombinant human EPO was produced by CHO cells grown under various osmotic pressure and purified from culture supernatants by heparin-sepharose affinity column and immunoaffinity column. N-linked oligosaccharides were released enzymatically and isolated by paper chromatography. The isolated oligosaccharides were labeled with a fluorescent dye, 2-aminobenzamide and analyzed with MonoQ anion exchange chromatography and GlycosepN amide chromatography for the assignment of GU (glucose unit) value. Glycan analysis by HPLC that the relative amount of neutral (asialo) oligosaccharide was increased from 20.16% to 29.76% when osmolality was elevated. In case of tetrasialylated glycan, total relative amount was decreased from 9.41% to 1.46%.
