Microbial biomass concentration is a key bioprocess parameter, estimated using various labor, operator and process cross-sensitive techniques, analyzed in a broad context and therefore the subject of correct interpretation. In this paper, the authors present the results of P. pastoris cell density estimation based on off-line (optical density, wet/dry cell weight concentration), in-situ (turbidity, permittivity), and soft-sensor (off-gas O₂/CO₂, alkali consumption) techniques. Cultivations were performed in a 5 L oxygen-enriched stirred tank bioreactor. The experimental plan determined varying aeration rates/levels, glycerol or methanol substrates, residual methanol levels, and temperature. In total, results from 13 up to 150 g (dry cell weight)/L cultivation runs were analyzed. Linear and exponential correlation models were identified for the turbidity sensor signal and dry cell weight concentration (DCW). Evaluated linear correlation between permittivity and DCW in the glycerol consumption phase (<60 g/L) and medium (for Mut⁺ strain) to significant (for Mut^S strain) linearity decline for methanol consumption phase. DCW and permittivity-based biomass estimates used for soft-sensor parameters identification. Dataset consisting from 4 Mut⁺ strain cultivation experiments used for estimation quality (expressed in NRMSE) comparison for turbidity-based (8%), permittivity-based (11%), O₂ uptake-based (10%), CO₂ production-based (13%), and alkali consumption-based (8%) biomass estimates. Additionally, the authors present a novel solution (algorithm) for uncommon in-situ turbidity and permittivity sensor signal shift (caused by the intensive stirrer rate change and antifoam agent addition) on-line identification and minimization. The sensor signal filtering method leads to about 5-fold and 2-fold minimized biomass estimate drifts for turbidity- and permittivity-based biomass estimates, respectively.

This contribution discusses the main challenges related to successful application of automatic control systems used to control specific growth rate in industrial biotechnological processes. It is emphasized that, after the implementation of basic automatic control systems, primary attention shall be paid to the specific growth rate control systems because this process variable critically affects the physiological state of microbial cultures and the formation of the desired product. Therefore, control of the specific growth rate enables improvement of the quality and reproducibility of the biotechnological processes. The main requirements have been formulated that shall be met to successfully implement the specific growth rate control systems in industrial bioreactors. The relatively easy-to-implement schemes of specific growth rate control systems have been reviewed and discussed. The recommendations for selection of particular control systems for specific biotechnological processes have been provided.

Docosahexaenoic acid (DHA) is one of the most important long-chain polyunsaturated fatty acids (LC-PUFAs), with numerous health benefits. Crypthecodinium cohnii, a marine heterotrophic dinoflagellate, is successfully used for the industrial production of DHA because it can accumulate DHA at high concentrations within the cells. Glycerol is an interesting renewable substrate for DHA production since it is a by-product of biodiesel production and other industries, and is globally generated in large quantities. The DHA production potential from glycerol, ethanol and glucose is compared by combining fermentation experiments with the pathway-scale kinetic modeling and constraint-based stoichiometric modeling of C. cohnii metabolism. Glycerol has the slowest biomass growth rate among the tested substrates. This is partially compensated by the highest PUFAs fraction, where DHA is dominant. Mathematical modeling reveals that glycerol has the best experimentally observed carbon transformation rate into biomass, reaching the closest values to the theoretical upper limit. In addition to our observations, the published experimental evidence indicates that crude glycerol is readily consumed by C. cohnii, making glycerol an attractive substrate for DHA production.

This article presents a comparative study on the development and application of two distinct adaptive control algorithms for biomass specific growth rate control in fed-batch biotechnological processes. A typical fed-batch process using Escherichia coli for recombinant protein production was selected for this research. Numerical simulation results show that both developed controllers, an adaptive PI controller based on the gain scheduling technique and a model-free adaptive controller based on the artificial neural network, delivered a comparable control performance and are suitable for application when using the substrate limitation approach and substrate feeding rate manipulation. The controller performance was tested within the realistic ranges of the feedback signal sampling intervals and measurement noise intensities. Considering the efforts for controller design and tuning, including development of the adaptation/learning algorithms, the model-free adaptive control algorithm proves to be more attractive for industrial applications, especially when only limited knowledge of the process and its mathematical model is available. The investigated model-free adaptive controller also tended to deliver better control quality under low specific growth rate conditions that prevail during the recombinant protein production phase. In the investigated simulation runs, the average tracking error did not exceed 0.01 (1/h). The temporary overshoots caused by the maximal disturbances stayed within the range of 0.025–0.11 (1/h). Application of the algorithm can be further extended to specific growth rate control in other bacterial and mammalian cell cultivations that run under substrate limitation conditions.

This article presents the development and application of a distinct adaptive control algorithm that is based on fuzzy logic and was used to control the specific growth rate (SGR) in a fed-batch biotechnological process. The developed control algorithm was compared with two adaptive control systems that were based on a model-free adaptive technique and gain scheduling technique. A typical mathematical model of recombinant Escherichia coli fed-batch cultivation process was selected to evaluate the performance of the fuzzy-based control algorithm. The investigated control techniques performed similarly when considering the whole process duration. The adaptive PI controller with fuzzy-based parameter adaptation demonstrated advantages over the previously mentioned algorithms—especially when compensating the deviations of the SGR. These deviations usually occur when the equipment malfunctions or process disturbances take place. The fuzzy-based control system was stable within the investigated ranges. It was determined that, regarding control quality, the investigated control algorithms are suited to control the SGR in a fed-batch biotechnological process. However, substrate feeding rate manipulation and limitation needs to be used. Taking into account the time needed to design and tune the controller, the developed controller is suitable for practical applications when expert knowledge is available. The proposed algorithm can be further adapted and developed to control the SGR in other cell cultivations while running the process under substrate limitation conditions.

An adaptive control system for the set-point control and disturbance rejection of biotechnological-process parameters is presented. The gain scheduling of PID (PI) controller parameters is based on only controller input/output signals and does not require additional measurement of process variables for controller-parameter adaptation. Realization of the proposed system does not depend on the instrumentation-level of the bioreactor and is, therefore, attractive for practical application. A simple gain-scheduling algorithm is developed, using tendency models of the controlled process. Dissolved oxygen concentration was controlled using the developed control system. The biotechnological process was simulated in fed-batch operating mode, under extreme operating conditions (the oxygen uptake-rate’s rapidly and widely varying, feeding and aeration rate disturbances). In the simulation experiments, the gain-scheduled controller demonstrated robust behavior and outperformed the compared conventional PI controller with fixed parameters.