Soft sensor based on 2D‐fluorescence and process data enabling real‐time estimation of biomass in Escherichia coli cultivations

In bioprocesses, specific process responses such as the biomass cannot typically be measured directly on-line, since analytical sampling is associated with unavoidable time delays. Accessing those responses in real-time is essential for Quality by Design and process analytical technology concepts. Soft sensors overcome these limitations by indirectly measuring the variables of interest using a previously derived model and actual process data in real time. In this study, a biomass soft sensor based on 2D-fluorescence data and process data, was developed for a comprehensive study with a 20-L experimental design, for Escherichia coli fed-batch cultivations. A multivariate adaptive regression splines algorithm was applied to 2D-fluorescence spectra and process data, to estimate the biomass concentration at any time during the process. Prediction errors of 4.9% (0.99 g/L) for validation and 3.8% (0.69 g/L) for new data (external validation), were obtained. Using principal component and parallel factor analyses on the 2D-fluorescence data, two potential chemical compounds were identified and directly linked to cell metabolism. The same wavelength pairs were also important predictors for the regression-model performance. Overall, the proposed soft sensor is a valuable tool for monitoring the process performance on-line, enabling Quality by Design.