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073 Image analysis for quantification of fungal biomass and differentiation
Abstracts in this section are from papers presented at:
IFAC SYMPOSIUM ON MODELING AND CONTROL OF BIOTECHNICAL PROCESSES 1992 Keystone, Colorado, USA, 29 March - 2 April 1992 Full papers appear in the Proceedings volume, to which the page numbers relate, published by IFAC and available from Pergamon Press. (ISBN: 0 08 041710 8)
070 Monitoring and Control of Mammalian Cell D.R. Omstead, K.K. Frame, E.I. Tsao, G.P. Price, J.C. Glass, M.A. Bohn, K.P. McNamee, J.A. Reese, M.J. Munster, D. Jain, pp 21-28
067 Monitoring and Control of Microbial Fermentations P.M. Salmon, B.C. Buckland, pp 1-6 Robust process monitoring techniques are necessary to ensure consistent performance of industrial fermentations. The initiation of each process step (such as inoculation, induction, or feeding) should not follow a predetermined schedule, but should be based on measured parameters that reflect the state of the fermentation. The analysis of samples is especially important because most in situ probes provide information only about physical conditions within a fermentor, rather than about the condition of the microorganisms. Analysis of vent-gas samples using mass spectrometry has become indispensable, and the use of robotics systems for on-line sampling and analysis of broth samples is currently under development.
Mammalian cell cultures for commercial products, predominantly vaccines, have traditionally been produced in small laboratory vessels without significant process monitoring and control. The focus has recently evolved into areas such. as robotic control of roller-bottlebased processes. For many newer productions currently under clinical evaluation, the use of stirred fermentation vessels is more common. The n'end is toward advanced off-line monitoring, allowing better manual control of cell culture processes. The use of on-line biomass monitoring is also coming into use. This paper summarizes the status and trends of industrial on-line and off-line analysis for mammalian cell cultures and describes the expected future opportunities.
071 NAD(P)H and F420 Fluorescence Monitoring in Anaerobic Digestion J.M. Owens, P. Pullammanappallil, D.P. Chynoweth, S.A. Svoronos, pp 29-33
068 The Use of Infrared Spectroscopic Techniques in Monitoring and Controlling Bioreactors Kun Yu, J.A. Philips, pp 7-13
Anaerobic digesters are frequently upset by changes in the feed strength, changes in environmental conditions and toxic feed components. These can cause reactor failures and lead to lengthy and cosily start-ups. Realtime on-line measurements are needed to assess the state of the digester and guide effective process con~'ol strategies. This paper presents initial investigations of the response of fluorescence probes for the coenzymes NAD(P)H and F420 to common causes of digester upsets, namely an increase in loading, a decrease in loading, and toxin (phenol) addition.
The relative advantages and disadvantages of various spectroscopic techniques for identifying and measuring concentrations of dilute, multicomponent mixtures are reviewed. A more specific background on infrared spectroscopy is provided. The characteristics of nearand mid-infrared spectra of several microbial, mammalian and plant cell cultures are documented. Some of the factors critical to quantifying chemical species in fermentation samples, from the perspective of on-line implementation for control, are considered.
072 On-Line Monitoring of Volatile Compounds in Alcoholic Fermentations
069 On-Line Monitoring of the Physiological State of the Cell Population in a Bioreactor T. Yoshida, K.B. Konstantinov, W. Ruenglertpanyakul, R.M. Matanguihan, pp 15-20
M.N. Pons, T. Cacjot, A. Groboillot,
C. Mathis, M. Miiller, S. Schutze, pp 35-40 The use of a gas membrane sensor connected to a gas chromatograph or mass spectrometer is described to monitor the concentration of volatile compounds produced during alcoholic fermentations in which the type and concentration of natural substrates, the inoculation rates and the temperature were varied.
The concept of the physiological state control of bioreactor systems using a knowledge-based approach is discussed from the viewpoint of plant structure. The structure of the bioreactor system is elucidated, and the various state variables, especially physiological state variables, are classified. As an example of work on sensor development, an application of a di-electric measurement method is investigated. This will possibly provide a powerful tool for detecting the physiological state of cells such as a viability index. A multi-mode cascade of neural networks with a shape-analysis module has been designed for the estimation of unmeasurable variables and identification of the physiological state of the cell population.
073 Image Analysis for Quantification of Fungal Biomass and Differentiation C.R. Thomas, G.C. Paul, C.A. Kent, H.L. Packer, E. Keshavarz-Moore, M.D. Lilly, pp 41-44 Fully-automatic image analysis methods are described for the measurement of vacuolation in filamentous fungi
733
734
Abstracts
growing in dispersed form in submerged cultures. For chrysogenum biomass, volumes of cytoplasmic and vacuolised or degenerated regions can be converted successfully into dry weight estimations. The vacuolation can be characterised in detail for physiological studies. Although off-line, the method is rapid enough to be considered for use as a process control tool. Links between cytoplasmic content and antibiotic production are being sought. The method might be used to provide data for structured models of fungal fermentations, and for control of product formation in those fermentations.
PeniciUium
074 Modelling of Filamentous Microorganisms J. Nielsen, pp 45-52 Based on reported mechanisms for filamentous growth, a simple morphologically structured growth model is set up. The model is based on a structuring of the biomass into three types of cells: apical cells, subapical cells, and hyphal cells. The model can describe the morphology of filamentous microorganisms, e.g. the hyphal growth unit, and it is compared with experimental data for the morphology of Geotrichum candidum grown in a submerged culture.
075 Structured Segregated Model for Optimal Profit Control of Baker's Yeast Production J.Q. Yuan, K.H. Bellgardt, pp 53-58 In fed-batch production of baker's yeast the sugar flow profile must regard the Crabtree-Effect and the oxygen transfer capacity of the reactor to avoid loss of yield. This paper describes optimal fed-batch control strategies which also consider the final product quality. Optimization is based on a three-part mathematical process model: the structured growth model, the agedistribution model and the cost model. New parts of the model are presented and simulation results are compared with experimental data. The optimum operation is then investigated. Unlike usual control trajectories, consideration of the quality criterion leads to a pulsed control of substrate flow. The model was verified in laboratory-scale experiments.
076 Modelling the Cyathus Striatus Fermentation Process: Comparison of Three Methods N.A. Jalel, V. Hags, A.R. Mirzal, R. Boeker, A. Munack, J.R. Leigh, pp 59-64 In controlling a fermentation process, the most difficult aspect is that of finding a suitable model. This paper describes three different approaches to modelling the Cyathus Striatus fermentation process. First, conventional biotechnological wisdom has been used to develop a set of equations whose parameters have been determined numerically. Second, a linear multivariable model has been fitted to experimentally obtained data. The equations have been integrated to generate a set of time-varying equations. Thirdly, neural nets are trained on experimental data. The approaches are compared in regard to their utility in state estimation and in trajectoryfollowing control.
077 An Analysis of the Impact of Controlled Variables Selection on the Operation of Anaerobic Digestion Processes M. Perrier, D. Dochain pp 65-70 Different evaluated process. Chemical
adaptive nonlinear controller designs are for the operation of the anaerobic digestion The selected controlled variables are the Oxygen Demand, propionate concentration,
and dissolved hydrogen concentration. The manipulated variable is the dilution rate. Although the responses are model-specific, the methodology can be applied to an actual lab scale or industrial scale bioreactor. The study reveals the intrinsic behaviour of the balances between the assumed bacterial species. The methodology demonstrates the usefulness of advanced control •techniques to get a better understanding of a process, even if the controller designs are based on incomplete knowledge.
078 Issues in the Optimization, Estimation and Control of Fed-Batch Bioreactors Using Tendency Models P. Tsobanakls, S.H. Lee, J.A. Phillips, C. Georgakls, pp 71-76 The tendency modeling approach for optimization, estimation, and control is applied to bioreaetors. This approach uses a "tendency" or "grey" model based on the available fundamental knowledge of the process and emphasizes the incorporation of new process data such as the power, the oxygen transfer capacity, and the rheological characteristics of the broth. The techniques is demonstrated on simulation studies and preliminary experimental results of penicillin fermentation. The model can be updated periodically, utilizing data collected both on-line and off-line. The updated model can be used off-line to develop optimal feeding policies. On-line, a Kalman filter can utihze the tendency model to estimate poorly, infrequently, or unmeasured states.
079 Substrate Control In Fedbatch Cultivations using a Model-Based Modification of a PI-Regulator P. Hagander, O. Hoist, pp 77-82 A fedbatch process shows exponential growth under ideal conditions. Good substrate concentration control requires that the regulator can track an exponentially growing feed demand. This paper proposes replacing an I-term in standard PI-control with a model-basexl observer for an exponentially growing demand. The regulator can now track the demand without error. Pseudomonas cepacia was grown on salicylate as sole carbon and energy source. On-line measurement of salicylate concentration was carried out using a filtration system. Introducing more instability into the controller requires attention to the anti-windup features. No such problems were found during the cultivations or in simulations of the effect of conceivable disturbances.
080 Can we Identify Biotechnical Processes? G. Bastin, L. Chen, V. Chotteau, pp 83-88 The identification problem of biotechnological processes is threefold: (i) determination of the number of biological reactions, (ii) identification of the underlying reaction network, (iii) identification of the kinetics. In most practical cases, these three parts of the identification problem can be completely deeoupled from one another.
081 Some Improvements in the Identification of Bioprocesses A. Munack, pp 89-94 In identification of dynamic models for bioteelmical processes, one is sometimes confronted with the situation, that models with different structure may be equally well fitted to the measured data. Some possibilities for experiment design in order to detect the most suitable model structure are presented and applied to a growth model of Cyathus striatus.
IFAC SYMPOSIUM ON MODELING AND CONTROL OF BIOTECHNICAL PROCESSES 1992 Keystone, Colorado, USA, 29 March - 2 April 1992 Full papers appear in the Proceedings volume, to which the page numbers relate, published by IFAC and available from Pergamon Press. (ISBN: 0 08 041710 8)
070 Monitoring and Control of Mammalian Cell D.R. Omstead, K.K. Frame, E.I. Tsao, G.P. Price, J.C. Glass, M.A. Bohn, K.P. McNamee, J.A. Reese, M.J. Munster, D. Jain, pp 21-28
067 Monitoring and Control of Microbial Fermentations P.M. Salmon, B.C. Buckland, pp 1-6 Robust process monitoring techniques are necessary to ensure consistent performance of industrial fermentations. The initiation of each process step (such as inoculation, induction, or feeding) should not follow a predetermined schedule, but should be based on measured parameters that reflect the state of the fermentation. The analysis of samples is especially important because most in situ probes provide information only about physical conditions within a fermentor, rather than about the condition of the microorganisms. Analysis of vent-gas samples using mass spectrometry has become indispensable, and the use of robotics systems for on-line sampling and analysis of broth samples is currently under development.
Mammalian cell cultures for commercial products, predominantly vaccines, have traditionally been produced in small laboratory vessels without significant process monitoring and control. The focus has recently evolved into areas such. as robotic control of roller-bottlebased processes. For many newer productions currently under clinical evaluation, the use of stirred fermentation vessels is more common. The n'end is toward advanced off-line monitoring, allowing better manual control of cell culture processes. The use of on-line biomass monitoring is also coming into use. This paper summarizes the status and trends of industrial on-line and off-line analysis for mammalian cell cultures and describes the expected future opportunities.
071 NAD(P)H and F420 Fluorescence Monitoring in Anaerobic Digestion J.M. Owens, P. Pullammanappallil, D.P. Chynoweth, S.A. Svoronos, pp 29-33
068 The Use of Infrared Spectroscopic Techniques in Monitoring and Controlling Bioreactors Kun Yu, J.A. Philips, pp 7-13
Anaerobic digesters are frequently upset by changes in the feed strength, changes in environmental conditions and toxic feed components. These can cause reactor failures and lead to lengthy and cosily start-ups. Realtime on-line measurements are needed to assess the state of the digester and guide effective process con~'ol strategies. This paper presents initial investigations of the response of fluorescence probes for the coenzymes NAD(P)H and F420 to common causes of digester upsets, namely an increase in loading, a decrease in loading, and toxin (phenol) addition.
The relative advantages and disadvantages of various spectroscopic techniques for identifying and measuring concentrations of dilute, multicomponent mixtures are reviewed. A more specific background on infrared spectroscopy is provided. The characteristics of nearand mid-infrared spectra of several microbial, mammalian and plant cell cultures are documented. Some of the factors critical to quantifying chemical species in fermentation samples, from the perspective of on-line implementation for control, are considered.
072 On-Line Monitoring of Volatile Compounds in Alcoholic Fermentations
069 On-Line Monitoring of the Physiological State of the Cell Population in a Bioreactor T. Yoshida, K.B. Konstantinov, W. Ruenglertpanyakul, R.M. Matanguihan, pp 15-20
M.N. Pons, T. Cacjot, A. Groboillot,
C. Mathis, M. Miiller, S. Schutze, pp 35-40 The use of a gas membrane sensor connected to a gas chromatograph or mass spectrometer is described to monitor the concentration of volatile compounds produced during alcoholic fermentations in which the type and concentration of natural substrates, the inoculation rates and the temperature were varied.
The concept of the physiological state control of bioreactor systems using a knowledge-based approach is discussed from the viewpoint of plant structure. The structure of the bioreactor system is elucidated, and the various state variables, especially physiological state variables, are classified. As an example of work on sensor development, an application of a di-electric measurement method is investigated. This will possibly provide a powerful tool for detecting the physiological state of cells such as a viability index. A multi-mode cascade of neural networks with a shape-analysis module has been designed for the estimation of unmeasurable variables and identification of the physiological state of the cell population.
073 Image Analysis for Quantification of Fungal Biomass and Differentiation C.R. Thomas, G.C. Paul, C.A. Kent, H.L. Packer, E. Keshavarz-Moore, M.D. Lilly, pp 41-44 Fully-automatic image analysis methods are described for the measurement of vacuolation in filamentous fungi
733
734
Abstracts
growing in dispersed form in submerged cultures. For chrysogenum biomass, volumes of cytoplasmic and vacuolised or degenerated regions can be converted successfully into dry weight estimations. The vacuolation can be characterised in detail for physiological studies. Although off-line, the method is rapid enough to be considered for use as a process control tool. Links between cytoplasmic content and antibiotic production are being sought. The method might be used to provide data for structured models of fungal fermentations, and for control of product formation in those fermentations.
PeniciUium
074 Modelling of Filamentous Microorganisms J. Nielsen, pp 45-52 Based on reported mechanisms for filamentous growth, a simple morphologically structured growth model is set up. The model is based on a structuring of the biomass into three types of cells: apical cells, subapical cells, and hyphal cells. The model can describe the morphology of filamentous microorganisms, e.g. the hyphal growth unit, and it is compared with experimental data for the morphology of Geotrichum candidum grown in a submerged culture.
075 Structured Segregated Model for Optimal Profit Control of Baker's Yeast Production J.Q. Yuan, K.H. Bellgardt, pp 53-58 In fed-batch production of baker's yeast the sugar flow profile must regard the Crabtree-Effect and the oxygen transfer capacity of the reactor to avoid loss of yield. This paper describes optimal fed-batch control strategies which also consider the final product quality. Optimization is based on a three-part mathematical process model: the structured growth model, the agedistribution model and the cost model. New parts of the model are presented and simulation results are compared with experimental data. The optimum operation is then investigated. Unlike usual control trajectories, consideration of the quality criterion leads to a pulsed control of substrate flow. The model was verified in laboratory-scale experiments.
076 Modelling the Cyathus Striatus Fermentation Process: Comparison of Three Methods N.A. Jalel, V. Hags, A.R. Mirzal, R. Boeker, A. Munack, J.R. Leigh, pp 59-64 In controlling a fermentation process, the most difficult aspect is that of finding a suitable model. This paper describes three different approaches to modelling the Cyathus Striatus fermentation process. First, conventional biotechnological wisdom has been used to develop a set of equations whose parameters have been determined numerically. Second, a linear multivariable model has been fitted to experimentally obtained data. The equations have been integrated to generate a set of time-varying equations. Thirdly, neural nets are trained on experimental data. The approaches are compared in regard to their utility in state estimation and in trajectoryfollowing control.
077 An Analysis of the Impact of Controlled Variables Selection on the Operation of Anaerobic Digestion Processes M. Perrier, D. Dochain pp 65-70 Different evaluated process. Chemical
adaptive nonlinear controller designs are for the operation of the anaerobic digestion The selected controlled variables are the Oxygen Demand, propionate concentration,
and dissolved hydrogen concentration. The manipulated variable is the dilution rate. Although the responses are model-specific, the methodology can be applied to an actual lab scale or industrial scale bioreactor. The study reveals the intrinsic behaviour of the balances between the assumed bacterial species. The methodology demonstrates the usefulness of advanced control •techniques to get a better understanding of a process, even if the controller designs are based on incomplete knowledge.
078 Issues in the Optimization, Estimation and Control of Fed-Batch Bioreactors Using Tendency Models P. Tsobanakls, S.H. Lee, J.A. Phillips, C. Georgakls, pp 71-76 The tendency modeling approach for optimization, estimation, and control is applied to bioreaetors. This approach uses a "tendency" or "grey" model based on the available fundamental knowledge of the process and emphasizes the incorporation of new process data such as the power, the oxygen transfer capacity, and the rheological characteristics of the broth. The techniques is demonstrated on simulation studies and preliminary experimental results of penicillin fermentation. The model can be updated periodically, utilizing data collected both on-line and off-line. The updated model can be used off-line to develop optimal feeding policies. On-line, a Kalman filter can utihze the tendency model to estimate poorly, infrequently, or unmeasured states.
079 Substrate Control In Fedbatch Cultivations using a Model-Based Modification of a PI-Regulator P. Hagander, O. Hoist, pp 77-82 A fedbatch process shows exponential growth under ideal conditions. Good substrate concentration control requires that the regulator can track an exponentially growing feed demand. This paper proposes replacing an I-term in standard PI-control with a model-basexl observer for an exponentially growing demand. The regulator can now track the demand without error. Pseudomonas cepacia was grown on salicylate as sole carbon and energy source. On-line measurement of salicylate concentration was carried out using a filtration system. Introducing more instability into the controller requires attention to the anti-windup features. No such problems were found during the cultivations or in simulations of the effect of conceivable disturbances.
080 Can we Identify Biotechnical Processes? G. Bastin, L. Chen, V. Chotteau, pp 83-88 The identification problem of biotechnological processes is threefold: (i) determination of the number of biological reactions, (ii) identification of the underlying reaction network, (iii) identification of the kinetics. In most practical cases, these three parts of the identification problem can be completely deeoupled from one another.
081 Some Improvements in the Identification of Bioprocesses A. Munack, pp 89-94 In identification of dynamic models for bioteelmical processes, one is sometimes confronted with the situation, that models with different structure may be equally well fitted to the measured data. Some possibilities for experiment design in order to detect the most suitable model structure are presented and applied to a growth model of Cyathus striatus.