A simple method to monitor and control methanol feeding of Pichia pastoris fermentations using mid-IR spectroscopy |
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Authors: | Schenk Jonas Marison Ian W von Stockar Urs |
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Institution: | Laboratory of Chemical and Biochemical Engineering (LGCB), Station 6-CH J2 503, Ecole Polytechnique Fédérale de Lausanne (EPFL), CH-1015 Lausanne, Switzerland. |
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Abstract: | 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. |
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