Contents

1 Solid-State Fermentation Bioreactor Fundamentals: Introduction and

Overview 1

David A. Mitchell, Marin Berovic, and Nadia Krieger

1.1 What Is "Solid-state Fermentation"? 1

1.2 Why Should We Be Interested in SSF? 3

1.3 What Are the Current and Potential Applications of SSF? 5

1.4 Why Do We Need a Book on the Fundamentals of SSF Bioreactors? 6

1.5 How Is this Book Organized? 8

1.5.1 Introduction to Solid-State Fermentation and Bioreactors 9

1.5.2 Introduction to the Various Classes of SSF Bioreactors 9

1.5.3 Fundamentals of Modeling of SSF Bioreactors 10

1.5.4 Modeling Case Studies of SSF Bioreactors 11

1.5.5 Key Issues Associated with SSF Bioreactors 11

1.5.6 A Final Word 12

Further Reading 12

2 The Bioreactor Step of SSF: A Complex Interaction of Phenomena 13

David A. Mitchell, Marin Berovic, Montira Nopharatana, and Nadia Krieger

2.1 The Need for a Qualitative Understanding of SSF 13

2.2 The General Steps of an SSF Process 14

2.3 The Bioreactor Step of an SSF Process 16

2.4 The Physical Structure of SSF Bioreactor Systems 17

2.4.1 A Macroscale View of the Phases in an SSF Bioreactor 17

2.4.2 A Microscale Snapshot of the Substrate Bed 20

2.5 A Dynamic View of the Processes Occurring 22

2.5.1 A Dynamic View with a Time Scale of Seconds to Minutes 22

2.5.2 A Dynamic View with a Time Scale of Hours to Days 24

2.6 Where Has this Description Led Us? 31

Further Reading 32

3 Introduction to Solid-State Fermentation Bioreactors 33

David A. Mitchell, Marin Berovic, and Nadia Krieger

3.1 Introduction 33

3.2 Bioreactor Selection and Design: General Questions 34

3.2.1 The Crucial Questions 35

3.2.2 Other Questions to Consider 36

3.3 Overview of Bioreactor Types 38

3.3.1 Basic Design Features of the Various Bioreactor Types 38

3.3.2 Overview of Operating Variables 40

3.4 A Guide for Bioreactor Selection 41

Further Reading 43

4 Heat and Mass Transfer in Solid-State Fermentation Bioreactors: Basic

Principles 45

David A. Mitchell, Marin Berovic, Oscar F. von Meien, and Luiz F.L. Luz Jr

4.1 Introduction 45

4.2 An Overall Balance Over the Bioreactor 45

4.3 Looking Within the Bioreactor in More Detail 47

4.3.1 Phenomena Within Subsystems Within the Bioreactor 47

4.3.2 Transfer Between Subsystems When the Substrate Bed Is Treated as a Single Pseudo-Homogeneous Phase 50

4.3.3 Transfer Between Subsystems When the Substrate Bed Is Treated as Two Separate Phases 51

4.3.4 Bulk Gas Flow Patterns and Pressure Drops 53

4.3.5 Mixing Patterns in Agitated Beds of Solids 56

Further Reading 56

5 The Scale-up Challenge for SSF Bioreactors 57

David A. Mitchell, Oscar F. von Meien, Luiz F.L. Luz Jr, and Marin Berovic

5.1 Introduction 57

5.2 The Challenges Faced at Large Scale in SLF and SSF 57

5.3 The Reason Why Scale-up Is not Simple 58

5.4 Approaches to Scale-up of SSF Bioreactors 63

Further Reading 64

6 Group I Bioreactors: Unaerated and Unmixed 65

David A. Mitchell, Nadia Krieger, and Marin Berovic

6.1 Basic Features, Design, and Operating Variables for Tray-type Bioreactors 65

6.2 Use of Bag Systems in Modern Processes 66

6.3 Heat and Mass Transfer in Tray Bioreactors 67

6.3.1 Oxygen Profiles Within Trays 67

6.3.2 Temperature Profiles Within Trays 69

6.3.3 Insights from Dynamic Modeling of Trays 71

6.4 Conclusions 75

Further Reading 76

7 Group II Bioreactors: Forcefully-Aerated Bioreactors Without Mixing 77

David A. Mitchell, Penjit Srinophakun, Nadia Krieger, and Oscar F. von Meien

7.1 Introduction 77

7.2 Basic Features, Design, and Operating Variables for Packed-Bed Bioreactors 77

7.3 Experimental Insights into Packed-Bed Operation 81

7.3.1 Large-Scale Packed-Beds 82

7.3.2 Pilot-Scale Packed-Beds 83

7.3.3 Laboratory-scale Packed-beds 84

7.4 Conclusions on Packed-Bed Bioreactors 93

Further Reading 94

8 Group III: Rotating-Drum and Stirred-Drum Bioreactors 95

David A. Mitchell, Deidre M. Stuart, Matthew T. Hardin, and Nadia Krieger

8.1 Introduction 95

8.2 Basic Features, Design, and Operating Variables for Group III Bioreactors 95

8.3 Experimental Insights into the Operation of Group III Bioreactors 98

8.3.1 Large-Scale Applications 98

8.3.2 Pilot-Scale Applications 100

8.3.3 Small-Scale Applications 101

8.4 Insights into Mixing and Transport Phenomena in Group III Bioreactors 104

8.4.1 Solids Flow Regimes in Rotating Drums 105

8.4.2 Gas Flow Regimes in the Headspaces of Rotating Drums 110

8.5 Conclusions on Rotating-Drum and Stirred-Drum Bioreactors 112

Further Reading 114

9 Group IVa: Continuously-Mixed, Forcefully-Aerated Bioreactors 115

David A. Mitchell, Nadia Krieger, Marin Berovic, and Luiz F.L. Luz Jr

9.1 Introduction 115

9.2 Basic Features, Design, and Operating Variables of Group IVa Bioreactors 115

9.3 Where Continuously-Agitated, Forcefully-Aerated Bioreactors Have Been Used 117

9.3.1 Stirred Beds with Mechanical Agitators 117

9.3.2 Gas-Solid Fluidized Beds 121

9.3.3 Bioreactors Mixed by the Motion of the Bioreactor Body 123

9.4 Insights into Mixing and Transport Phenomena in Group IVa Bioreactors 125

9.5 Conclusions on Group IVa Bioreactors 128

Further Reading 128

10 Group IVb: Intermittently-Mixed Forcefully-Aerated Bioreactors 129

David A. Mitchell, Oscar F. von Meien, Luiz F.L. Luz Jr, Nadia Krieger, J. Ricardo Pérez-Correa, and Eduardo Agosin

10.1 Introduction 129

10.2 Basic Features of Group IVb Bioreactors 129

10.3 Experimental Insights into the Performance of Group IVb Bioreactors 131

10.3.1 Large-Scale Intermittently-Mixed Bioreactors 131

10.3.2 Pilot-Scale Intermittently-Mixed Bioreactors 135

10.3.3 Laboratory-Scale Intermittently-Mixed Bioreactors 138

10.4 Insights into Mixing and Transport Phenomena in Group IVb Bioreactors 138

10.5 Conclusions on Group IVb Bioreactors 140

Further Reading 140

11 Continuous Solid-State Fermentation Bioreactors 141

Luis B. R. Sánchez, Morteza Khanahmadi, and David A. Mitchell

11.1 Introduction 141

11.2 Basic Features of Continuous SSF Bioreactors 141

11.2.1 Equipment 141

11.2.2 Flow Patterns: Real-Flow Models 146

11.3 Continuous Versus Batch Mode of Operation 148

11.3.1 Reduction of Upstream and Downstream Investment 148

11.3.2 Uniformity of the Product from Batch and Continuous

Bioreactors 149

11.3.3 Enhanced Production Rates 150

11.3.4 Contamination 150

11.4 Comparison by Simulation of the Three CSSFBs 152

11.4.1 Continuous Tubular Flow Bioreactors (CTFBs) with Recycling... 152

11.4.2 Continuous Rotating Drum Bioreactor (CRDB) 154

11.4.3 Continuous Stirred Tank Bioreactor (CSTB) 155

11.4.4 Evaluation of the Various CSSFB Configurations 156

11.5 Scientific and Technical Challenges for CSSFBs 158

Further Reading 158

12 Approaches to Modeling SSF Bioreactors 159

David A. Mitchell, Luiz F.L. Luz Jr, Marin Berovic, and Nadia Krieger

12.1 What Are Models and Why Model SSF Bioreactors? 159

12.2 Using Models to Design and Optimize an SSF Bioreactor 161

12.2.1 Initial Studies in the Laboratory 161

12.2.2 Current Bioreactor Models as Tools in Scale-up 163

12.2.3 Use of the Model in Control Schemes 164

12.3 The Anatomy of a Model 164

12.4 The Seven Steps of Developing a Bioreactor Model 167

12.4.1 Step 1: Know What You Want to Achieve and the Effort You

Are Willing to Put into Achieving It 170

12.4.2 Step 2: Draw the System at the Appropriate Level of Detail and Explicitly State Assumptions 170

12.4.3 Step 3: Write the Equations 171

12.4.4 Step 4: Estimate the Parameters and Decide on Values for the

Operating Variables 173

12.4.5 Step 5: Solve the Model 174

12.4.6 Step 6: Validate the Model 175

12.4.7 Step 7: Use the Model 177

Further Reading 177

13 Appropriate Levels of Complexity for Modeling SSF Bioreactors 179

David A. Mitchell, Luiz F.L. Luz Jr, Marin Berovic, and Nadia Krieger

13.1 What Level of Complexity Should We Aim for in an SSF

Bioreactor Model? 179

13.2 What Level of Detail Should Be Used to Describe the Growth Kinetics? 179

13.2.1 Growth Should Be Treated as Depending on Which Factors? 180

13.2.2 Is It Worthwhile to Describe the Spatial Distribution of the

Biomass at the Microscale? 182

13.2.3 Typical Features of the Kinetic Sub-models 183

13.3 What Level of Detail Should Be Used to Describe Transport Processes? 183

13.4 At the Moment Fast-Solving Models Are Useful 185

13.5 Having Decided on Fast-Solving Models, How to Solve Them? 188

13.6 Conclusions 188

Further Reading 189

14 The Kinetic Sub-model of SSF Bioreactor Models: General Considerations 191

David A. Mitchell and Nadia Krieger

14.1 What Is the Aim of the Kinetic Analysis? 191

14.2 How Will Growth Be Measured Experimentally? 194

14.2.1. The Problem of Measuring Biomass in SSF 194

14.2.2 Indirect Approaches to Monitoring Growth 196

14.3 What Units Should Be Used for the Biomass? 197

14.3.1 Grams of Biomass per Gram of Fresh Sample 199

14.3.2 Grams of Biomass per Gram of Dry Sample 199

14.3.3 Grams of Biomass per Gram of Initial Fresh or Dry Sample 200

14.3.4 Which Set of Units Is Best to Use for Expressing the Biomass? 201

14.4 Kinetic Profiles and Appropriate Equations 201

14.5 Conclusions 204

Further Reading 205

15 Growth Kinetics in SSF Systems: Experimental Approaches 207

David A. Mitchell and Nadia Krieger

15.1 Experimental Systems for Studying Kinetics 207

15.1.1. Flasks in an Incubator 208

15.1.2. Columns in a Waterbath 210

15.1.3. Comparison of the Two Systems 211

15.2 Experimental Planning 211

15.3 Estimation of Biomass from Measurements of Biomass

Components 214

15.3.1 Suitable Systems for Undertaking Calibration Studies 214

15.3.2 Conversion of Measurements of Components of the Biomass 216

15.3.3 Limitations of these Calibration Methods 217

15.4 Conclusion 217

Further Reading 217

16 Basic Features of the Kinetic Sub-model 219

David A. Mitchell, Graciele Viccini, Lilik Ikasari, and Nadia Krieger

16.1 The Kinetic Sub-model Is Based on a Differential Growth Equation 219

16.2 The Basic Kinetic Expression 220

16.3 Incorporating the Effect of the Environment on Growth 222

16.3.1 Incorporating the Effect of Temperature on Growth 225

16.3.2 Incorporating the Effect of Water Activity on Growth 228

16.3.3 Combining the Effects of Several Variables 230

16.4 Modeling Death Kinetics 231

16.4.1 General Considerations in Modeling of Death Kinetics 231

16.4.2 Approaches to Modeling Death Kinetics that Have Been Used 232

16.5 Conclusion 234

Further Reading 234

17 Modeling of the Effects of Growth on the Local Environment 235

David A. Mitchell and Nadia Krieger

17.1 Introduction 235

17.2 Terms for Heat, Water, Nutrients, and Gases 237

17.2.1 Metabolic Heat Production 237

17.2.2 Water Production 238

17.2.3 Substrate and Nutrient Consumption 238

17.2.4 Oxygen Consumption and Carbon Dioxide Production 239

17.2.5 General Considerations with Respect to Equations for the

Effects of Growth on the Environment 243

17.3 Modeling Particle Size Changes 244

17.3.1 An Empirical Equation for Particle Size Reduction 244

17.3.2 How to Model Particle Size Changes in Bioreactor Models? 245

17.4 Product Formation - Empirical Approaches 246

17.5 Conclusions 247

Further Reading 247

18 Modeling of Heat and Mass Transfer in SSF Bioreactors 249

David A. Mitchell, Oscar F. von Meien, Luiz F.L. Luz Jr, and Marin Berovic

18.1 Introduction 249

18.2 General Forms of Balance Equations 249

18.3 Conduction 252

18.3.1 Conduction Across the Bioreactor Wall 252

18.3.2 Conduction Within a Phase 253

18.4 Convection 255

18.4.1 Convection at the Bioreactor Wall 255

18.4.2 Convective Heat Removal from Solids to Air 256

18.4.3 Convective Heat Removal Due to Air Flow Through the Bed 258

18.5 Evaporation 259

18.5.1 Evaporation from the Solids to the Air Phase 260

18.5.2 Water Removal Due to Air Flow Through the Bed 261

18.6 Conclusions 263

Further Reading 263

19 Substrate, Air, and Thermodynamic Parameters for SSF Bioreactor Models 265

David A. Mitchell, Oscar F. von Meien, Luiz F.L. Luz Jr, and Marin Berovic

19.1 Introduction 265

19.2 Substrate Properties 265

19.2.1 Particle Size and Shape 266

19.2.2 Particle Density 267

19.2.3 Bed Packing Density 268

19.2.4 Porosity (Void Fraction) 270

19.2.5 Water Activity of the Solids 271

19.3 Air Density 273

19.4 Thermodynamic Properties 274

19.4.1 Saturation Humidity 275

19.4.2 Heat Capacity of the Substrate Bed 276

19.4.3 Enthalpy of Vaporization of Water 277

Further Reading 278

20 Estimation of Transfer Coefficients for SSF Bioreactors 279

David A. Mitchell, Oscar F. von Meien, Luiz F.L. Luz Jr, and Marin Berovic

20.1 Introduction 279

20.2 Thermal Conductivities of Substrate Beds 279

20.3 Heat Transfer Coefficients Involving the Wall 280

20.3.1. Bed-to-Wall Heat Transfer Coefficients 281

20.3.2 Wall-to-Headspace Heat Transfer Coefficients 281

20.3.3 Wall-to-Surroundings Heat Transfer Coefficients 282

20.3.4 Overall Heat Transfer Coefficients 282

20.4 Solids-to-Air Heat and Mass Transfer Coefficients Within Beds 283

20.5 Bed-to-Headspace Transfer Coefficients 284

20.6 Conclusions 289

Further Reading 289

21 Bioreactor Modeling Case Studies: Overview 291

David A. Mitchell

21.1 What Can the Models Be Used to Do? 291

21.2 Limitations of the Models 292

21.3 The Amount of Detail Provided about Model Development 293

21.4 The Order of the Case Studies 294

22 A Model of a Well-mixed SSF Bioreactor 295

David A Mitchell and Nadia Krieger

22.1 Introduction 295

22.2 Synopsis of the Model 295

22.2.1 The System, Equations, and Assumptions 295

22.2.2 Values of Parameters and Variables 301

22.3 Insights the Model Gives into the Operation of Well-Mixed Bioreactors 303

22.3.1 Insights into Operation at Laboratory Scale 303

22.3.2 Insights into Operation at Large Scale 307

22.3.3 Effect of Scale and Operation on Contributions to Cooling of the Solids 310

22.4 Conclusions on the Operation of Well-Mixed Bioreactors 312

Further Reading 314

23 A Model of a Rotating-Drum Bioreactor 315

David A. Mitchell, Deidre M. Stuart, and Nadia Krieger

23.1 Introduction 315

23.2 A Model of a Well-Mixed Rotating-Drum Bioreactor 315

23.2.1 Synopsis of the Mathematical Model and its Solution 315

23.2.2 Predictions about Operation at Laboratory Scale 320

23.2.3 Scale-up of Well-Mixed Rotating-Drum Bioreactors 325

23.3 What Modeling Work Says about Rotating-Drum Bioreactors

Without Axial Mixing 328

23.4 Conclusions on the Design and Operation of Rotating-Drum Bioreactors 329

Further reading 330

24 Models of Packed-Bed Bioreactors 331

David A. Mitchell, Penjit Srinophakun, Oscar F. von Meien, Luiz F.L. Luz Jr, and Nadia Krieger

24.1 Introduction 331

24.2 A Model of a Traditional Packed-Bed Bioreactor 331

24.2.1 Synopsis of the Mathematical Model and its Solution 333

24.2.2 Base-Case Predictions 334

24.2.3 Insights that Modeling Has Given into Optimal Design and

Operation of Traditional Packed-Beds 336

24.3 A model of the Zymotis Packed-Bed Bioreactor 341

24.3.1 The Model 341

24.3.2 Insights into Optimal Design and Operation of Zymotis

Packed-Beds 342

24.4 Conclusions on Packed-Bed Bioreactors 347

Further Reading 347

25 A Model of an Intermittently-Mixed Forcefully-Aerated Bioreactor 349

David A. Mitchell, Oscar F. von Meien, Luiz F.L. Luz Jr, and Nadia Krieger

25.1 Introduction 349

25.2 Synopsis of the Model 349

25.3 Insights the Model Gives into Operation of Intermittently-Mixed Bioreactors 353

25.3.1 Predictions about Operation at Laboratory Scale 353

25.3.2 Investigation of the Design and Operation of Intermittently-

Mixed Forcefully-Aerated Bioreactors at Large Scale 357

25.4 Conclusions on Intermittently-Mixed Forcefully-Aerated Bioreactors 360

Further Reading 362

26 Instrumentation for Monitoring SSF Bioreactors 363

Mario Fernández and J. Ricardo Pérez-Correa

26.1 Why Is It Important to Monitor SSF Bioreactors? 363

26.2 Which Variables Would We Like to Measure? 363

26.3 Available Instrumentation for On-line Measurements 365

26.4 Data Filtering 369

26.5 How to Measure the Other Variables? 371

Further Reading 374

27 Fundamentals of Process Control 375

J. Ricardo Pérez-Correa and Mario Fernández

27.1 Main Ideas Underlying Process Control 375

27.1.1 Feedback 375

27.1.2 Control Loop 376

27.1.3 Computer Control Loop 376

27.2 Conventional Control Algorithms 377

27.2.1 On/Off Control 377

27.2.2 PID Control 380

27.2.3 Model Predictive Control 385

Further Reading 386

28 Application of Automatic Control Strategies to SSF Bioreactors 387

J. Ricardo Pérez-Correa, Mario Fernández, Oscar F. von Meien, Luiz F.L. Luz Jr, and David A. Mitchell

28.1 Why Do We Need Automatic Control in SSF Bioreactors? 387

28.2 How to Control SSF Bioreactors? 388

28.3 Case Studies of Control in SSF Bioreactors 390

28.3.1 Control of the Bioreactors at PUC Chile 390

28.3.2 Model-Based Evaluation of Control Strategies 395

28.4 Future Challenges in the Control of SSF Bioreactors 400

Further Reading 401

29 Design of the Air Preparation System for SSF Bioreactors 403

Oscar F. von Meien, Luiz F.L. Luz Jr, J. Ricardo Pérez-Correa, and David A. Mitchell

29.1 Introduction 403

29.2 An Overview of the Options Available 404

29.3 Blower/Compressor Selection and Flow Rate Control 407

29.4 Piping and Connections 408

29.5 Air Sterilization 408

29.6 Humidification Columns 409

29.7 Case Study: An Air Preparation System for a Pilot-Scale Bioreactor ...410 Further Reading 412

30 Future Prospects for SSF Bioreactors 413

David A. Mitchell, Marin Berovic, and Nadia Krieger

30.1 The Increasing Importance of SSF 413

30.2 Present State and Future Prospects 414

References 417

Appendix: Guide to the Bioreactor Programs 429

A.1 Disclaimer 429

A.2 General Information and Advice 429

A.3 Use of the Well-Mixed Bioreactor Model 431

A.4 Use of the Rotating-Drum Bioreactor Model 433

A.5 Use of the Traditional Packed-Bed Bioreactor Model 435

A.6 Use of the Zymotis Packed-Bed Bioreactor Model 436

A.7 Use of the Model of an Intermittently-Mixed Forcefully-Aerated

Bioreactor 439

Index 443

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