Development of a small-scale bioreactor: application to in vivo NMR measurement.
A perfused bioreactor allowing in vivo NMR measurement was developed and validated for Eschscholtzia californica cells. The bioreactor was made of a 10-mm NMR tube. NMR measurement of the signal-to-noise ratio was optimized using a sedimented compact bed of cells that were retained in the bioreactor by a supporting filter. Liquid medium flow through the cell bed was characterized from a mass balance on oxygen and a dispersive hydrodynamic model. Cell bed oxygen demand for 4 h perfusion required a minimal medium flow rate of 0.8 mL/min. Residence time distribution assays at 0.8-2.6 mL/min suggest that the cells are subjected to a uniform nutrient environment along the cell bed. Cell integrity was maintained for all culture conditions since the release of intracellular esterases was not significant even after 4 h of perfusion. In vivo NMR was performed for (31)P NMR and the spectrum can be recorded after only 10 min of spectral accumulation (500 scans) with peaks identified as G-6P, F-6P, cytoplasmic Pi, vacuolar Pi, ATP(gamma) and ADP(beta), ATP(alpha) and ADP(alpha), NADP and NDPG, NDPG and ATP(beta). Cell viability was shown to be maintained as (31)P chemical shifts were constant with time for all the identified nuclei, thus suggesting constant intracellular pH.
Gmati
Dorra
D
Bio-P2 Research Unit, Department of Chemical Engineering, Ecole Polytechnique de Montréal, P.O. Box 6079, Centreville Station, Montréal, Québec, Canada.
Chen
Jingkui
J
Jolicoeur
Mario
M
D001426:Bacterial Proteins / Q000032:analysis* / Q000378:metabolism*; D019149:Bioreactors; D018929:Cell Culture Techniques / Q000295:instrumentation* / Q000379:methods; D049109:Cell Proliferation; D004867:Equipment Design; D019544:Equipment Failure Analysis; D031675:Eschscholzia / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy / Q000295:instrumentation* / Q000379:methods; D010100:Oxygen / Q000378:metabolism; D010865:Pilot Projects
D001426:Bacterial Proteins; D010100:Oxygen