Chard, Stephen John; (1998) On-line monitoring and control of fractional protein precipitation and recovery processes. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

To operate downstream processes at high yield and constant fraction, it is necessary to be able to monitor them on-line and control to preset conditions. On-line measurement eliminates manual sampling followed by retrospective analysis, the process conditions adjusted in real-time. Two examples are presented, firstly on-line particle analysis techniques are compared to those off-line which are more reliable. Solid-liquid separation is determined by properties such as size, concentration and density difference between the suspending fluid. Hence on-line sizing has been attained by light diffraction and the concentration predicted by spectrophotometry with a knowledge of the relative refractive index of the particles. This was related to data from the electrical sensing zone method. Centrifugal sedimentation may not be run on-line, but provided additional information of density difference. In the second part of the study two-cut fractional precipitation of the product enzyme alcohol dehydrogenase (ADH) from contaminating proteins according to differences in solubility with the precipitant ammonium sulphate is discussed. An online rig has been designed incorporating continuous precipitation, sample preparation using a microcentrifuge and measurement by stopped-flow analysis. Errors of the rig have been assessed by repeat measurements. The nature of the precipitation profile means the strategy was to reduce those relevant to control of the process, then tuning of the analytical times and consumption of materials. The microcentrifuge was shown to be the largest source. The fit of the ADH fraction remaining soluble by a model has been considered. Simplex and Kalman filter algorithms which update the model parameters after every measurement cycle were tested by simulation and off-line. The Kalman filter was preferred, on-line monitoring and control then being demonstrated for a pH and a concentration step change in the feed.

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