title: Decreased pCO(2) accumulation by eliminating bicarbonate addition to high cell-density cultures.
abstract: High-density perfusion cultivation of mammalian cells can result in elevated bioreactor CO(2) partial pressure (pCO(2)), a condition that can negatively influence growth, metabolism, productivity, and protein glycosylation. For BHK cells in a perfusion culture at 20 x 10(6) cells/mL, the bioreactor pCO(2) exceeded 225 mm Hg with approximate contributions of 25% from cellular respiration, 35% from medium NaHCO(3), and 40% from NaHCO(3) added for pH control. Recognizing the limitations to the practicality of gas sparging for CO(2) removal in perfusion systems, a strategy based on CO(2) reduction at the source was investigated. The NaHCO(3) in the medium was replaced with a MOPS-Histidine buffer, while Na(2)CO(3) replaced NaHCO(3) for pH control. These changes resulted in 63-70% pCO(2) reductions in multiple 15 L perfusion bioreactors, and were reproducible at the manufacturing-scale. Bioreactor pCO(2) values after these modifications were in the 68-85 mm Hg range, pCO(2) reductions consistent with those theoretically expected. Low bioreactor pCO(2) was accompanied by both 68-123% increased growth rates and 58-92% increased specific productivity. Bioreactor pCO(2) reduction and the resulting positive implications for cell growth and productivity were brought about by process changes that were readily implemented and robust. This philosophy of pCO(2) reduction at the source through medium and base modification should be readily applicable to large-scale fed-batch cultivation of mammalian cells.
authors.affiliation: Research and Development, Process Sciences, Bayer HealthCare, Pharmaceutical Division, 800 Dwight Way, Berkeley, California 94710, USA. chetan.goudar.b@bayer.com
authors.forename: Chetan T
authors.initials: CT
authors.lastname: Goudar
authors.forename: Ricaredo
authors.initials: R
authors.lastname: Matanguihan
authors.forename: Edward
authors.initials: E
authors.lastname: Long
authors.forename: Christopher
authors.initials: C
authors.lastname: Cruz
authors.forename: Chun
authors.initials: C
authors.lastname: Zhang
authors.forename: James M
authors.initials: JM
authors.lastname: Piret
authors.forename: Konstantin B
authors.initials: KB
authors.lastname: Konstantinov
mesh_terms: D000818:Animals; D019149:Bioreactors; D002245:Carbon Dioxide / Q000378:metabolism*; D018929:Cell Culture Techniques / Q000379:methods; D002460:Cell Line / Q000378:metabolism; D049109:Cell Proliferation; D008322:Mammals; D010313:Partial Pressure; D010477:Perfusion
chemical_list: D002245:Carbon Dioxide