Considerations for osmolality measurement under elevated pCO(2): Comparison of vapor pressure and freezing point osmometry

Osmolality increases with pCO(2) in bioreactors with pH control, and it has been shown that osmolality compensation by decreasing the basal NaCl conce ntration partially :mitigates the adverse effects of elevated pCO(2) on ani mal cell growth, protein production, and glycosylation. Thus, measurement o f osmolality is important for a complete characterization of the culture en vironment under elevated pCO(2). However, osmolality measurement may be com promised by CO2 evolution. Freezing point depression and vapor pressure dep ression osmometry were directly compared for the measurement-bf osmolality in samples at elevated pCO(2) (up to 250 mmHg) and at a variety of pH value s (6.7-7.5). More:extensive degassing may be expected with the vapor pressu re osmometer due to the smaller sample volume and-larger surface area emplo yed. However, both types of osmometer yielded similar results for all pCO(2 ), and pH values studied. Moreover, the measured values agreed with osmolal ity values calculated using a semiempirical model. Further analysis showed that, while sample degassing may result in a large decrease in pCO(2), ther e is little associated decrease in osmolality. The great majority of total CO2 in solution is present as bicarbonate (HCO3-). Although a small amount of HCO2 is converted to CO2 to compensate for CO2 evolution, further deplet ion of HCO3- is inhibited by the associated increase in-medium pH and by th e need for HCO3- to maintain charge neutrality in solution. This explanatio n is consistent with the observed similarity in osmolality values for the t wo types of osmometer. It was also observed that osmolality did not change in samples that were frozen at -20 degrees C for up to 1 year. (C) 2000 Joh n Wiley & Sons, Inc.