The use of windows of operation as a bioprocess design tool

Bioprocess design problems are frequently multivariate and complex. However, they may be visualised by a graphical representation of the design constraints and correlations governing both the process and system under consideration, namely windows of operation. Windows of operation exist at all stages of process design and find use both in the identification of key constraints from limited information, and also, with more detailed knowledge, the sensitivity of a process to design or operating changes. In this way windows of operation may be used to help understand and optimise a bioprocess design. In this paper the formulation, development and application of windows of operation is discussed for a range of biological processes including fermentation, protein recovery and biotransformation.UCL is the Biotechnology and Biological Sciences Research Council's Interdisciplinary Research Centre for Biochemical Engineering and the Council's support is gratefully acknowledged.     

Selection of bioprocess simulation software for industrial applications

Two commercially available, process-simulation software packages (Aspen Batch Plus v1.2, Aspen Technology, Inc., Cambridge, Massachusetts, and Intelligen SuperPro v3.0, INTELLIGEN, INC., Scotch Plains, Ner Jersey) are evaluated for use in modeling industrial, biotechnology processes. Software is quantitatively evaluated by Kepner-Tregoe Decision Analysis (Kepner and Tregoe, 1981). This evaluation shows that Aspen Batch Plus v1.2 (ABP) and Intelligen SuperPro v3.0 (ISP) can successfully perform specific simulation tasks but do not provide a complete model of all phenomena occurring within a biotechnology process. Software is best suited to provide a format for process management, using material and energy balances to answer scheduling questions, explore equipment change-outs, and calculate cost data. The ability of simulation software to accurately predict unit operation scale-up and optimize bioprocesses is limited. To realistically evaluate the software, a vaccine manufacturing process under development at Merck & Company is simulated. Case studies from the vaccine process are presented as examples of how ABP and ISP can be used to shed light on real-world processing issues.     

Haemolysis,a tool to evaluate mechanical stress to cells in bioprocess equipment

Summary Suspensions of fragile sheep erythrocytes were used to compare the levels of mechanical stress in two different crossflow filter units of small pilot scale. The recycling pumps were identified as the critical components mainly responsible for haemolysis. The level of cell damage was largely determined by the pump type and the backpressure applied to increase the transmembrane pressure in the filter.     

Development of a versatile computer integrated control system for bioprocess controls

A general approach is described for the implementation of a networked multi-unit computer integrated control system. The use of data acquisition hardware and graphical programming tools alleviates tedious programming and maintains potency and flexibility. One application of the control system, the control of a mammalian cell perfusion culture based on a key nutrient glucose concentration, was demonstrated. The control system offers customized user interface for all process control parameters and allows the flexibility for continued improvement and implementation of new tailored functions. The temperature, pH, dissolved oxygen and glucose level were accurately controlled. This revised version was published online in June 2006 with corrections to the Cover Date.     

Development of a software tool for in silico simulation of Escherichia coli using a visual programming environment

This study describes the development of a software tool, EcoSim, to assist users in implementing quantitative in silico simulation easily. It consists of four parts: extracellular environment and constraints setting mode, table for optimal metabolic flux distribution and chart for changes of substrate concentration, dynamic flux distribution viewer and dynamic hierarchical regulatory network viewer. Representation of a hierarchical regulatory network was constructed with defined modeling symbols and weight in the central Escherichia coli metabolism. All programming procedures for EcoSim were accomplished in a visual programming environment (LabVIEW). To illustrate quantitative in silico simulation with EcoSim, this program was performed on E. coli using glucose and acetate as carbon sources. The simulation results were in agreement with the experimental data obtained from the literature. EcoSim can be used to assist biologists and engineers in predicting and interpreting dynamic behaviors of E. coli under a variety of environmental conditions.     

A platform for dynamic simulation and control of movement based on OpenSim and MATLAB

Numerical simulations play an important role in solving complex engineering problems and have the potential to revolutionize medical decision making and treatment strategies. In this paper, we combine the rapid model-based design, control systems and powerful numerical method strengths of MATLAB/Simulink with the simulation and human movement dynamics strengths of OpenSim by developing a new interface between the two software tools. OpenSim is integrated with Simulink using the MATLAB S-function mechanism, and the interface is demonstrated using both open-loop and closed-loop control systems. While the open-loop system uses MATLAB/Simulink to separately reproduce the OpenSim Forward Dynamics Tool, the closed-loop system adds the unique feature of feedback control to OpenSim, which is necessary for most human movement simulations. An arm model example was successfully used in both open-loop and closed-loop cases. For the open-loop case, the simulation reproduced results from the OpenSim Forward Dynamics Tool with root mean square (RMS) differences of 0.03° for the shoulder elevation angle and 0.06° for the elbow flexion angle. MATLAB's variable step-size integrator reduced the time required to generate the forward dynamic simulation from 7.1s (OpenSim) to 2.9s (MATLAB). For the closed-loop case, a proportional-integral-derivative controller was used to successfully balance a pole on model's hand despite random force disturbances on the pole. The new interface presented here not only integrates the OpenSim and MATLAB/Simulink software tools, but also will allow neuroscientists, physiologists, biomechanists, and physical therapists to adapt and generate new solutions as treatments for musculoskeletal conditions.     

Automated image processing as an analytical tool in cell cryopreservation for bioprocess development

The continuous availability of cells with defined cell characteristics represents a crucial issue in the biopharmaceutical and cell therapy industry. Here, development of cell banks with a long-term stability is essential and ensured by a cryopreservation strategy. The strategy needs to be optimized for each cell application individually and usually comprises controlled freezing, storage at ultra-low temperature, and fast thawing of cells. This approach is implemented by the development of master and working cell banks. Currently, empirical cryopreservation strategy development is standard, but a knowledge-based approach would be highly advantageous. In this article, we report the development of a video-based tool for the characterisation of freezing and thawing behaviour in cryopreservation process to enable a more knowledge-based cryopreservation process development. A successful tool validation was performed with a model cryopreservation process for the β-cell line INS-1E. Performance was evaluated for two working volumes (1.0 mL and 2.0 mL), based on freezing-thawing rates (20 °C to − 80 °C) and cell recovery and increase of biomass, to determine tool flexibility and practicality. Evaluation confirmed flexibility by correctly identifying a delay in freezing and thawing for the larger working volume. Further more, a decrease in cell recovery from 0.94 (± 0.14) % using 1.0 mL working volume to 0.61 (± 0.05) % using a 2.0 mL working volume displays tool practicality. The video-based tool proposed in this study presents a powerful tool for cell-specific optimisation of cryopreservation protocols. This can facilitate faster and more knowledge-based cryopreservation process development

Graphical abstract

In this study, a video-based analytical tool was developed for the characterisation of freezing and thawing behaviour in cryopreservation process development. Evaluation of the practicality and flexibility of the developed tool was done based on a scale-up case study with the cell line INS-1E. Here, the influence of sample working volume on process performance was investigated. Increasing the volume from 1to 2 mL led to a delay in freezing and thawing behaviour which caused cell recovery loss. We believe that the developed tool will facilitate more directed and systematic cryopreservation process development.

     

Dynamic metabolic modeling for a MAB bioprocess

Production of monoclonal antibodies (MAb) for diagnostic or therapeutic applications has become an important task in the pharmaceutical industry. The efficiency of high-density reactor systems can be potentially increased by model-based design and control strategies. Therefore, a reliable kinetic model for cell metabolism is required. A systematic procedure based on metabolic modeling is used to model nutrient uptake and key product formation in a MAb bioprocess during both the growth and post-growth phases. The approach combines the key advantages of stoichiometric and kinetic models into a complete metabolic network while integrating the regulation and control of cellular activity. This modeling procedure can be easily applied to any cell line during both the cell growth and post-growth phases. Quadratic programming (QP) has been identified as a suitable method to solve the underdetermined constrained problem related to model parameter identification. The approach is illustrated for the case of murine hybridoma cells cultivated in stirred spinners.     

Makers of bioprocess engineerings

Makers of bioprocess engineerings     

Control of substrate concentration in a continuous bioprocess

This paper is the first one presenting an application of the control law based on the ratio of measured and optimal values for a real fermentation pilot plant operated in continuous mode. A controller of this type takes into account the process variables and may be designed by using classical linear methods. A simple algorithm is applied for a real CSTR to control the substrate concentration. The results obtained confirm that this approach offers the possibility to combine simplicity and effectiveness in bioprocess control.     

Plasmid stability in a recombinant mammalian cell bioprocess

Summary A simple experimental method is devised to determine the fraction of plasmid-harboring cells in a bioprocess employing recombinant mammalian cells. The fraction of plasmid-harboring cells decreased as serum content in the growth medium decreased. The relatively higher increase in the generation time of the plasmid-harboring cell was primarily responsible for this decrease. The mathematical expression obtained for this fraction in terms of the two parameters, i.e. the generation time ratio and the plasmid-loss probability, could represent the experimental data extremely well. The numerical values of these parameters could show the inherent insight of the system. It was found that the data plot against time can draw us to a misleading conclusion of the absence of the effect of serum concentration.     

Biohydrogen production and bioprocess enhancement: a review

This paper provides an overview of the recent advances and trends in research in the biological production of hydrogen (biohydrogen). Hydrogen from both fossil and renewable biomass resources is a sustainable source of energy that is not limited and of different applications. The most commonly used techniques of biohydrogen production, including direct biophotolysis, indirect biophotolysis, photo-fermentation and dark-fermentation, conventional or "modern" techniques are examined in this review. The main limitations inherent to biochemical reactions for hydrogen production and design are the constraints in reactor configuration which influence biohydrogen production, and these have been identified. Thereafter, physical pretreatments, modifications in the design of reactors, and biochemical and genetic manipulation techniques that are being developed to enhance the overall rates and yields of biohydrogen generation are revisited.     

Development of tool use in a macaque and a gorilla

The development of the capacity to use a stick as a tool was tested in a macaque (Macaca fuscata) and a gorilla (Gorilla gorilla gorilla) infants that had previously shown to be able to use strings and supports as dragging tools. Subjects were tested between 15 and 38 months of age. Different levels of competence between the subjects emerged over testing. The macaque developed a stereotyped strategy to cope with the problem, only getting random successes, whilst the gorilla developed a flexible strategy and revealed to be able to mentally represent the solution of the problem. In fact, when not successful using the stick, the gorilla thought out an alternative strategy, choosing and adapting a new object to use it as a tool.     

Development of a high cell-density fed-batch bioprocess for the heterologous production of 6-deoxyerythronolide B in Escherichia coli

A robust high cell-density fed-batch bioprocess was developed for the heterologous production of 6-deoxyerythronolide B (6-dEB), the macrocyclic core of the antibiotic erythromycin, with a recombinant Escherichia coli strain expressing the 6-deoxyerythronolide B synthase (DEBS) from Saccharopolyspora erythraea. Initial evaluation of the E. coli strain in a 5-l bioreactor with the addition of exogenous propionate for polyketide biosynthesis resulted in a maximum cell density of 30 g l(-1) (OD600 approximately 60) and the production of 700 mg l(-1) of 6-dEB. Retention of the two plasmids harboring the heterologous genes was maintained between 90 and 100% even in the absence of antibiotic selection. However, the accumulation of excess ammonia in the culture medium was found to significantly decrease the productivity of the cells. Through optimization of the medium composition and fermentation conditions, the maximum cell density was increased by two-fold, and a final titer of 1.1 g l(-1) of 6-dEB was achieved. This represents an 11-fold improvement compared to the highest reported titer of 100 mg l(-1) with E. coli as the production host.