A simple method to monitor and control methanol feeding of Pichia pastoris fermentations using mid-IR spectroscopy.
Mid-infrared FTIR spectroscopy is an efficient tool for the monitoring of bioprocesses, since it is fast and able to detect many compounds simultaneously. However, complex and time-consuming calibration procedures are still required, and have inhibited the spreading of these instruments. A simple and quick method to calibrate a FTIR instrument was developed for the control of fed-batch fermentations of the methylotrophic yeast Pichia pastoris. Based on the assumptions that (1) only substrate concentration may change significantly during a fed-batch process and (2) absorbance can be considered as proportional to concentration, a linear two-point calibration was implemented. Long-term instability of the instrument had to be addressed in order to get accurate results: two fixed points, on both sides of substrate absorbance peak, were used to perform on-line a linear correction of the signal drift. Fed-batch experiments at constant methanol (substrate) concentration ranging from 0.8 to 15gl(-1) were carried out. Off-line HPLC control analysis showed a good agreement with on-line FTIR data, with standard error of prediction values < 0.12gl(-1). Even though methanol acts both as carbon source and inducer of protein expression, no significant effect was observed on the level of protein expression in the recombinant strain used.
Schenk
Jonas
J
Laboratory of Chemical and Biochemical Engineering (LGCB), Station 6-CH J2 503, Ecole Polytechnique Fédérale de Lausanne (EPFL), CH-1015 Lausanne, Switzerland.
Marison
Ian W
IW
von Stockar
Urs
U
D019149:Bioreactors; D002138:Calibration; D005285:Fermentation; D000432:Methanol / Q000032:analysis* / Q000378:metabolism; D010843:Pichia / Q000378:metabolism*; D017550:Spectroscopy, Fourier Transform Infrared / Q000295:instrumentation / Q000379:methods
D000432:Methanol