INFORMATION TO USERS While the most advanced technology has been used to photograph and reproduce this manuscript, the quality of the reproduction is heavily dependent upon the quality of the material submitted. For example: # Manuscript pages may have indistinct print. In such cases, the best available copy has been filmed. ® Manuscripts may not always be complete. In such cases, a note will indicate that it is not possible to obtain missing pages. ® Copyrighted material may have been removed from the manuscript. In such cases, a note will indicate the deletion. Oversize materials (e.g., maps, drawings, and charts) are photographed by sectioning the original, beginning at the upper left-hand corner and continuing from left to right in equal sections with small overlaps. Each oversize page is also filmed as one exposure and is available, for an additional charge, as a standard 35mm slide or as a 17”x 23" black and white photographic print. Most photographs reproduce acceptably on positive microfilm or microfiche but lack the clarity on xerographic copies made from the microfilm. For an additional charge, 35mm slides of 6”x 9” black and white photographic prints are available for any photographs or illustrations that cannot be reproduced satisfactorily by xerography. Order Number 8717718 On-line, adaptive, optimal control of a high-density, fed-batch fermentation of streptomyces C5 Schlasner, Steven Mark, Ph.D. The Ohio State University, 1987 UMI 300N.ZeebRd. Ann Arbor, MI 48106 PLEASE NOTE: In all cases this material has been filmed in the best possible way from the available copy. Problems encountered with this document have been identified here with a check mark V 1. Glossy photographs or p a g e s. 2. Colored illustrations, paper or print______ 3. Photographs with dark background_____ 4. Illustrations are poor copy_______ 5. Pages with black marks, not original copy 6. Print shows through as there is text on both sides of page_ 7. Indistinct, broken or small print on several pages 8. Print exceeds margin requirements______ 9. Tightly bound copy with print lost in spine_______ 10. Computer printout pages with indistinct print. 11. Page(s)____________ lacking when material received, and not available from school or author. 12. Page(s)____________ seem to be missing in numbering only as text follows. 13. Two pages numbered . Text follows. 14. Curling and wrinkled pages______ 15. Dissertation contains pages with print at a slant, filmed as received__________ _ 16. O t h e r ____________ _________ ___ _______ University Microfilms International ON-LINE, ADAPTIVE, OPTIMAL œNTROL OF A HIGH-DENSITY, FED-BATCH FERMENTATION OF STREPTOMYCES C5 DISSERTATION Presented In Partial Fulfillment of the Requirements for the Degree Doctor of Philosophy in the Graduate School of The Ohio State University By Steven Mark Schlasner, B.A., B.S., M.B.A., M.S. ***** The Ohio State University 1987 Dissertation Committee: Approved by W-K Lee — Q o_ D.R. Skidmore Adviser W.R. Strohl Department of Chemical Engineering Copyright by Steven Mark Schlasner 1987 To My Wife, Betsy 11 ACKNOWLEDGEMENTS I express heartfelt appreciation to Drs. Won-Kyoo Lee and William R. Strohl for their guidance and insight throughout the research. Thanks, too, to Dr. Duane R. Skidmore, the remaining member of my dissertation committee, for his suggestions and comments. Gratitude is expressed to Donald E. Ordaz for his invaluable assistance during the experimental portion of the research. Appreciation is also expressed to Michael L. Dekleva and Michael B. Kukla for their technical assistance. To my wife, Betsy, daughter, Jacqueline, and other members of my family, I offer thanks for their patience, support and encouragement. iii VITA May 23, 1952 .................... B o m - Glendive, Montana 1974 ........................... B.A., St. Olaf College, Northfield, Minnesota 1974 - 1978..................... United States Air Force Officer Ellsworth Air Force Base, South Dakota 1977 ......................... M.B.A., University of South Dakota, Vermillion, South Dakota 1980 ........................... B.S., South Dakota School of , Mines and Technology, Rapid City, South Dakota 1980 - Present ................... United States Air Force Reserve Officer, United States Air Force Wright Aeronautical Laboratories, Wright-Patterson Air Force Base, Ohio 1983 ........... ............... M.S., The Ohio State University Columbus, Ohio FIELDS OF STUDY Major Field: Chemical Engineering Studies in chemical process control with Dr. WonHCyoo Lee. Minor Field: Microbiology Studies in industrial microbiology with Dr. William R. Strohl. iv TABLE OF CONTENTS DEDICATION ................................................. 11 ACKNOWLEDGEMENTS ........................................... Ill VITA ...................................................... Iv LIST OF TABLES ............................................. LIST OF F I G U R E S ............................................. ^ LIST OF S Y M B O L S ............................................. CHAPTER PAGE I . INTRODUCTION ........................................ Introduction ...................................... The Importance and Evolution of Fermentation Processes ........................................ Potential Improvements In Fermentation Processes ........................................ Goal and System Definition .......................... Obstacles to Achievement of Desired Goals ........... 9 Objectives ...................................... 11 II. LITERATURE REVIEW .................................... 15 Introduction ...................................... 15 High (Cell) Density Fermentation .................... 15 Bloreactor Configuration ........................... 19 B a t c h ......................................... 19 Continuous ....................... 20 Semi-batch .................................... 22 Plug-flow .................................... 25 Microorganisms .................................... 25 Y e a s t ......................................... 26 Escherichia coll .............................. 27 Penicllium spp................................... 27 Bacillus subtilis .............................. 28 Bacillus thuringiensis ......................... 28 Streptomyces spp................................. 28 Optimal Control Methods ........................... 29 Static optimization ............................. 29 Dynamic optimization ........................... 30 State and Parameter Estimation...................... 43 Indirect measurement ........................... 44 Filtering...................................... 44 Estimation .................................... 45 Measurement error detection ...................... 69 III. EXPERIMENTAL APPARATUS ................ 70 Fermentor ........................................ 70 Vessel ....................................... 70 Agitation...................................... 71 Gas flow ...................................... 71 Temperature control ............................. 72 Microcomputer .................................... 72 Microcomputer hardware ......................... 73 Microcomputer software ......................... 74 Peripherals ...................................... 77 Sensors and Actuators ............................. 82 Oxygen off-gas analysis ......................... 83 Carbon dioxide off-gas analysis .................. 84 Fermentor pressure measurement .................... 84 Gas flow rate measurement....................... 85 pH measurement ................................ 86 Dissolved oxygen measurement .................... 87 Dissolved carbon dioxide measurement ............. 87 Foam control .................................. 89 Glucose analyzer .............................. 90 Turbidity measurement ........................... 93 Glucose feed rate measurement.................... 94 Temperature measurement ....................... 94 Agitation r a t e ................................ 95 Stepping motor actuators .................... 96 Gas flow controllers ........................... 97 Relay box ................................ 99 Auxiliary devices ................................ 101 Air conditioning subsystem ...................... 102 Miscellaneous components ....................... 103 Continuous culture apparatus .................... 104 vi IV. ORGANISM, MATERIALS, AND METHODS ...................... 107 Microorganism ..................................... 108 Cuture Maintenance ................................. 109 Media ............................................ 110 Fermentation Preparation and Performance ....... Ill Shake-flask culture ............................. 112 Continuous culture ............................. 112 Batch and fed-batch fermentations ................ 114 Off-Line Analyses and Assays ........................ 117 Dry weight ..................................... 117 Optical density ................................. 117 Protein assay ................................... 118 Elemental biomass analyses ...................... 118 Glucose analysis ............................... 119 Nitrate analysis ........................... 119 Phosphate analysis ............................. 120 Organic acid a n a l y s i s ............................ 121 Anthracycline analysis .......................... 121 Ammmonia analysis ............................... 121 V. ESTIMATOR AND CONTROL SCHEME DEVELOPMENT ............... 123 Estimator Development ............................. 124 Optimal controller Development ...................... 133 VI.