(603c) Online Monitoring of Biomass in Pichia Pastoris Fermentation Using Capacitance-Based Sensors

In fermentation processes, biomass is a critical parameter and a key variable for optimization. Current biomass assays are performed off-line making them time consuming and limited in sample resolution. Thus, there is a clear advantage to transitioning to continuous on-line monitoring. Capacitance-based sensors were evaluated for the continuous monitoring of Pichia pastoris fermentations. Five different strains were successfully measured over the course of 14 fermentations using a dual frequency measurement technique. Permittivity was measured at a high frequency of 10 MHz and a characteristic frequency of 2.0 MHz. Regression analysis was performed on the data from multiple Pichia strains and resulted in a linear correlation between the on-line permittivity measurements and the off-line measurements of biomass as defined by dry cell weight. The regression coefficients were different when the carbon source was changed from glycerol to methanol. However, the linear relationship between dry cell weight and permittivity was preserved for each carbon source. Using these regression parameters and online permittivity measurements, dry cell weight was predicted within ± 5%. The accuracy of the prediction suggests that the on-line assay expands the resolution of off-line dry cell weight measurements with less effort, while providing greater insight into the process.

Going beyond the ability for on-line monitoring of biomass, capacitance sensors enable process control strategies based on biomass measurements. Currently, most Pichia pastoris methanol feeding strategies are feed-forward in that the profile is pre-determined (exponential, linear). However, biomass increase is not always as expected and feed-back correction mechanisms are required. The use of the capacitance signal to control methanol feeding rates to achieve specific biomass growth profiles has also been explored and results from these studies will be presented. Overall, the capacitance-based biomass sensors offer on-line fermentation monitoring and control capabilities aligned with Process Analytical Technology (PAT) initiatives from FDA and other regulatory agencies.