Preface xv
About the Editors xvii
List of Abbreviations xix
Notation xxiii
1 Introduction 1
Henner Schmidt-Traub and Reinhard Ditz
1.1 Chromatography, Development, and Future Trends 1
1.2 Focus of the Book 4
1.3 Suggestions on How to Read this Book 4
References 6
2 Fundamentals and General Terminology 9
Andreas Seidel-Morgenstern
2.1 Principles and Features of Chromatography 9
2.2 Analysis and Description of Chromatograms 13
2.2.1 Voidage and Porosity 13
2.2.2 Retention Times and Capacity Factors 16
2.2.3 Efficiency of Chromatographic Separations 17
2.2.4 Resolution 20
2.2.5 Pressure Drop 23
2.3 Mass Transfer and Fluid Dynamics 25
2.3.1 Principles of Mass Transfer 25
2.3.2 Fluid Distribution in the Column 27
2.3.3 Packing Nonidealities 28
2.3.4 Extra-Column Effects 29
2.4 Equilibrium Thermodynamics 29
2.4.1 Definition of Isotherms 29
2.4.2 Models of Isotherms 31
2.4.2.1 Single-Component Isotherms 31
2.4.2.2 Multicomponent Isotherms Based on the Langmuir Model 33
2.4.2.3 Competitive Isotherms Based on the Ideal Adsorbed Solution Theory 34
2.4.2.4 Steric Mass Action Isotherms 37
2.4.3 Relation Between Isotherms and Band Shapes 38
2.5 Column Overloading and Operating Modes 44
2.5.1 Overloading Strategies 44
2.5.2 Beyond Isocratic Batch Elution 45
References 46
3 Stationary Phases 49
Michael Schulte
3.1 Survey of Packings and Stationary Phases 49
3.2 Inorganic Sorbents 50
3.2.1 Activated Carbons 50
3.2.2 Synthetic Zeolites 54
3.2.3 Porous Oxides: Silica, Activated Alumina, Titania, Zirconia, and Magnesia 54
3.2.4 Silica 55
3.2.4.1 Surface Chemistry 57
3.2.4.2 Mass Loadability 59
3.2.5 Diatomaceous Earth 59
3.2.6 Reversed Phase Silicas 60
3.2.6.1 Silanization of the Silica Surface 60
3.2.6.2 Silanization 60
3.2.6.3 Starting Silanes 61
3.2.6.4 Parent Porous Silica 61
3.2.6.5 Reaction and Reaction Conditions 62
3.2.6.6 Endcapping 62
3.2.6.7 Chromatographic Characterization of Reversed Phase Silicas 63
3.2.6.8 Chromatographic Performance 63
3.2.6.9 Hydrophobic Properties Retention Factor (Amount of Organic Solvent for Elution), Selectivity 65
3.2.6.10 Shape Selectivity 65
3.2.6.11 Silanol Activity 67
3.2.6.12 Purity 68
3.2.6.13 Improved pH Stability Silica 68
3.2.7 Aluminum Oxide 69
3.2.8 Titanium Dioxide 70
3.2.9 Other Oxides 71
3.2.9.1 Magnesium Oxide 71
3.2.9.2 Zirconium Dioxide 71
3.2.10 Porous Glasses 72
3.3 Cross-Linked Organic Polymers 73
3.3.1 General Aspects 74
3.3.2 Hydrophobic Polymer Stationary Phases 77
3.3.3 Hydrophilic Polymer Stationary Phases 78
3.3.4 Ion Exchange (IEX) 79
3.3.4.1 Optimization of Ion-Exchange Resins 81
3.3.5 Mixed Mode 88
3.3.6 Hydroxyapatite 88
3.3.7 Designed Adsorbents 91
3.3.7.1 Protein A Affinity Sorbents 91
3.3.7.2 Other IgG Receptor Proteins: Protein G and Protein L 96
3.3.7.3 Sorbents for Derivatized/Tagged Compounds: Immobilized Metal Affinity Chromatography (IMAC) 96
3.3.7.4 Other Tag-Based Affinity Sorbents 101
3.3.8 Customized Adsorbents 102
3.3.8.1 Low Molecular Weight Ligands 105
3.3.8.2 Natural Polymers (Proteins, Polynucleotides) 108
3.3.8.3 Artificial Polymers 111
3.4 Advective Chromatographic Materials 111
3.4.1 Adsorptive Membranes and Grafted-Polymer Membranes 114
3.4.2 Adsorptive Nonwovens 115
3.4.3 Fiber/Particle Composites 117
3.4.4 Area-Enhanced Fibers 117
3.4.5 Monolith 118
3.4.6 Chromatographic Materials for Larger Molecules 121
3.5 Chiral Stationary Phases 121
3.5.1 Cellulose- and Amylose-Based CSP 122
3.5.2 Antibiotic CSP 128
3.5.3 Cyclofructan-Based CSP 128
3.5.4 Synthetic Polymers 128
3.5.5 Targeted Selector Design 130
3.5.6 Further Developments 132
3.6 Properties of Packings and Their Relevance to Chromatographic Performance 132
3.6.1 Chemical and Physical Bulk Properties 132
3.6.2 Morphology 133
3.6.3 Particulate Adsorbents: Particle Size and Size Distribution 133
3.6.4 Pore Texture 134
3.6.5 Pore Structural Parameters 137
3.6.6 Comparative Rating of Columns 137
3.7 Sorbent Maintenance and Regeneration 138
3.7.1 Cleaning in Place (CIP) 138
3.7.2 CIP for IEX 140
3.7.3 CIP of Protein A Sorbents 140
3.7.4 Conditioning of Silica Surfaces 143
3.7.5 Sanitization in Place (SIP) 145
3.7.6 Column and Adsorbent Storage 145
References 146
4 Selection of Chromatographic Systems 159
Michael Schulte
4.1 Definition of the Task 164
4.2 Mobile Phases for Liquid Chromatography 167
4.2.1 Stability 168
4.2.2 Safety Concerns 172
4.2.3 Operating Conditions 172
4.2.4 Aqueous Buffer Systems 176
4.3 Adsorbent and Phase Systems 178
4.3.1 Choice of Phase System Dependent on Solubility 178
4.3.2 Improving Loadability for Poor Solubilities 180
4.3.3 Dependency of Solubility on Sample Purity 183
4.3.4 Generic Gradients for Fast Separations 184
4.4 Criteria for Choosing Normal Phase Systems 184
4.4.1 Retention in NP Systems 186
4.4.2 Solvent Strength in Liquidâ"Solid Chromatography 188
4.4.3 Pilot Technique Thin-Layer Chromatography Using the PRISMA Model 190
4.4.3.1 Step (1): Solvent Strength Adjustment 199
4.4.3.2 Step (2): Optimization of Selectivity 199
4.4.3.3 Step (3): Final Optimization of the Solvent Strength 200
4.4.3.4 Step (4): Determination of the Optimum Mobile Phase Composition 200
4.4.4 Strategy for an Industrial Preparative Chromatography Laboratory 202
4.4.4.1 Standard Gradient Elution Method on Silica 203
4.4.4.2 Simplified Procedure 204
4.5 Criteria for Choosing Reversed Phase Systems 206
4.5.1 Retention and Selectivity in RP Systems 208
4.5.2 Gradient Elution for Small Amounts of Product on RP Columns 212
4.5.3 Rigorous Optimization for Isocratic Runs 213
4.5.4 Rigorous Optimization for Gradient Runs 217
4.5.5 Practical Recommendations 222
4.6 Criteria for Choosing CSP Systems 223
4.6.1 Suitability of Preparative CSP 223
4.6.2 Development of Enantioselectivity 224
4.6.3 Optimization of Separation Conditions 226
4.6.3.1 Determination of Racemate Solubility 226
4.6.3.2 Selection of Elution Order 226
4.6.3.3 Optimization of Mobile/Stationary Phase Composition, Including Temperature 226
4.6.3.4 Determination of Optimum Separation Step 227
4.6.4 Practical Recommendations 227
4.7 Downstream Processing of mAbs Using Protein A and IEX 231
4.8 Size-Exclusion Chromatography (SEC) 236
4.9 Overall Chromatographic System Optimization 237
4.9.1 Conflicts During Optimization of Chromatographic Systems 237
4.9.2 Stationary Phase Gradients 241
References 246
5 Process Concepts 251
Malte Kaspereit and Henner Schmidt-Traub
5.1 Discontinuous Processes 252
5.1.1 Isocratic Operation 252
5.1.2 Gradient Chromatography 253
5.1.3 Closed-Loop Recycling Chromatography 256
5.1.4 Steady-State Recycling Chromatography (SSRC) 258
5.1.5 Flip-Flop Chromatography 259
5.1.6 Chromatographic Batch Reactors 260
5.2 Continuous Processes 261
5.2.1 Column Switching Chromatography 262
5.2.2 Annular Chromatography 262
5.2.3 Multiport Switching Valve Chromatography (ISEP/CSEP) 263
5.2.4 Isocratic Simulated Moving Bed (SMB) Chromatography 264
5.2.5 SMB Chromatography with Variable Process Conditions 268
5.2.5.1 Varicol 269
5.2.5.2 PowerFeed 270
5.2.5.3 Partial-Feed, Partial-Discard, and Fractionation-Feedback Concepts 271
5.2.5.4 Improved/Intermittent SMB (iSMB) 271
5.2.5.5 Modicon 273
5.2.5.6 FF-SMB 273
5.2.6 Gradient SMB Chromatography 274
5.2.7 Supercritical Fluid Chromatography (SFC) 275
5.2.7.1 Supercritical Batch Chromatography 276
5.2.7.2 Supercritical SMB processes 277
5.2.8 Multicomponent Separations 277
5.2.9 Multicolumn Systems for Bioseparations 278
5.2.9.1 Multicolumn Capture Chromatography (MCC) 279
5.2.9.2 Multicolumn Countercurrent Solvent Gradient Purification (MCSGP) 286
5.2.10 Countercurrent Chromatographic Reactors 288
5.2.10.1 SMB Reactor 288
5.2.10.2 SMB Reactors with Distributed Functionalities 290
5.3 Choice of Process Concepts 292
5.3.1 Scale 292
5.3.2 Range of kâ 292
5.3.3 Number of Fractions 293
5.3.4 Example 1: Lab Scale; Two Fractions 293
5.3.5 Example 2: Lab Scale; Three or More Fractions 294
5.3.6 Example 3: Production Scale; Wide Range of kâ 296
5.3.7 Example 4: Production Scale; Two Main Fractions 297
5.3.8 Example 5: Production Scale; Three Fractions 298
5.3.9 Example 6: Production Scale; Multistage Process 300
References 302
6 Modeling of Chromatographic Processes 311
Andreas Seidel-Morgenstern
6.1 Introduction 311
6.2 Models for Single Chromatographic Columns 311
6.2.1 Equilibrium Stage Models 312
6.2.1.1 Discontinuous Model According to Craig 313
6.2.1.2 Continuous Model According to Martin and Synge 315
6.2.2 Derivation of Continuous Mass Balance Equations 316
6.2.2.1 Mass Balance Equations 318
6.2.2.2 Convective Transport 320
6.2.2.3 Axial Dispersion 320
6.2.2.4 Intraparticle Diffusion 321
6.2.2.5 Mass Transfer Between Phases 321
6.2.2.6 Finite Rates of Adsorption and Desorption 322
6.2.2.7 Adsorption Equilibria 323
6.2.3 Equilibrium Model of Chromatography 323
6.2.4 Models with One Band Broadening Effect 329
6.2.4.1 Equilibrium Dispersion Model 329
6.2.4.2 Finite Adsorption Rate Model 331
6.2.5 Continuous Lumped Rate Models 331
6.2.5.1 Transport Dispersion Models 332
6.2.5.2 Lumped Finite Adsorption Rate Model 333
6.2.6 General Rate Models 333
6.2.7 Initial and Boundary Conditions of the Column 335
6.2.8 Dimensionless Model Equations 336
6.2.9 Comparison of Different Model Approaches 338
6.3 Including Effects Outside the Columns 343
6.3.1 Experimental Setup and Simulation Flow Sheet 343
6.3.2 Modeling Extra-Column Equipment 345
6.3.2.1 Injection System 345
6.3.2.2 Piping 345
6.3.2.3 Detector 345
6.4 Calculation Methods and Software 346
6.4.1 Analytical Solutions 346
6.4.2 Numerical Solution Methods 346
6.4.2.1 Discretization 346
6.4.2.2 General Solution Procedure and Software 349
References 350
7 Determination of Model Parameters 355
Andreas Seidel-Morgenstern, Andreas Jupke, and Henner Schmidt-Traub
7.1 Parameter Classes for Chromatographic Separations 355
7.1.1 Design Parameters 355
7.1.2 Operating Parameters 356
7.1.3 Model Parameters 356
7.2 Concept to Determine Model Parameters 357
7.3 Detectors and Parameter Estimation 359
7.3.1 Calibration of Detectors 359
7.3.2 Parameter Estimation 360
7.3.3 Evaluation of Chromatograms 362
7.4 Determination of Packing Parameters 363
7.4.1 Void Fraction and Porosity of the Packing 363
7.4.2 Axial Dispersion 363
7.4.3 Pressure Drop 364
7.5 Adsorption Isotherms 365
7.5.1 Determination of Adsorption Isotherms 365
7.5.2 Estimation of Henry Coefficients 365
7.5.3 Static Isotherm Determination Methods 370
7.5.3.1 Batch Method 370
7.5.3.2 Adsorptionâ"Desorption Method 370
7.5.3.3 Circulation Method 371
7.5.4 Dynamic Methods 371
7.5.5 Frontal Analysis 371
7.5.6 Analysis of Dispersed Fronts 378
7.5.7 Peak Maximum Method 380
7.5.8 Minor Disturbance/Perturbation Method 380
7.5.9 Curve Fitting of the Chromatogram 383
7.5.10 Data Analysis and Accuracy 384
7.6 Mass Transfer Kinetics 386
7.6.1 Correlations 386
7.6.2 Application of Method of Moments 388
7.7 Plant Parameters 389
7.8 Experimental Validation of Column Models and Model Parameters 391
7.8.1 Batch Chromatography 391
7.8.2 Simulated Moving Bed Chromatography 394
7.8.2.1 Model Formulation and Parameters 394
7.8.2.2 Experimental Validation 400
References 404
8 Process Design and Optimization 409
Andreas Jupke, Andreas Biselli, Malte Kaspereit,Martin Leipnitz, and Henner Schmidt-Traub
8.1 Basic Principles and Definitions 409
8.1.1 Performance, Costs, and Objective Functions 409
8.1.1.1 Performance Criteria 410
8.1.1.2 Economic Criteria 411
8.1.1.3 Objective Functions 412
8.1.2 Degrees of Freedom 413
8.1.2.1 Categories of Parameters 413
8.1.2.2 Dimensionless Operating and Design Parameters 414
8.1.3 Scaling by Dimensionless Parameters 418
8.1.3.1 Influence of Different HETP Coefficients for Every Component 419
8.1.3.2 Influence of Feed Concentration 420
8.1.3.3 Examples for a Single-Column Batch Chromatography 421
8.1.3.4 Examples for SMB Processes 424
8.2 Batch Chromatography 426
8.2.1 Fractionation Mode (Cut Strategy) 426
8.2.2 Design and Optimization of Batch Chromatographic Columns 427
8.2.2.1 Process Performance Depending on Number of Stages and Loading Factor 427
8.2.2.2 Design and Optimization Strategy 432
8.2.2.3 Other Strategies 436
8.3 Recycling Chromatography 437
8.3.1 Design of Steady-State Recycling Chromatography 437
8.3.2 Scale-Up of Closed-Loop Recycling Chromatography 440
8.4 Conventional Isocratic SMB Chromatography 445
8.4.1 Considerations to Optimal Concentration Profiles in SMB Process 445
8.4.2 Process Design Based on TMB Models (Shortcut Methods) 446
8.4.2.1 Triangle Theory for an Ideal Model with Linear Isotherms 447
8.4.2.2 Triangle Theory for an Ideal Model with Nonlinear Isotherms 449
8.4.2.3 Shortcut to Apply the Triangle Theory on a System with Unknown Isotherms Assuming Langmuir Character 452
8.4.3 Process Design and Optimization Based on Rigorous SMB Models 455
8.4.3.1 Estimation of Operating Parameter 456
8.4.3.2 Optimization of Operating Parameters for Linear Isotherms Based on Process Understanding 457
8.4.3.3 Optimization of Operating Parameters for Nonlinear Isotherms Based on Process Understanding 458
8.4.3.4 Optimization of Design Parameters 460
8.5 Isocratic SMB Chromatography Under Variable Operating Conditions 465
8.5.1 Performance Comparison of Varicol and Conventional SMB 466
8.5.2 Performance Comparison of Varicol, PowerFeed, and Modicon with Conventional SMB 470
8.5.3 Performance Trends Applying SMB Concepts Under Variable Operating Conditions 475
8.6 Gradient SMB Chromatography 476
8.6.1 Step Gradient 476
8.6.2 Multicolumn Solvent Gradient Purification Process 482
8.7 Multicolumn Systems for Bioseparations 487
8.7.1 Design of Twin-Column Capture SMB 488
8.7.2 Modeling of Multicolumn Capture processes 490
References 493
9 Process Control 503
Sebastian Engell and Achim Kienle
9.1 Standard Process Control 504
9.2 Advanced Process Control 504
9.2.1 Online Optimization of Batch Chromatography 505
9.2.2 Advanced Control of SMB Chromatography 507
9.2.2.1 Purity Control for SMB Processes 508
9.2.2.2 Direct Optimizing Control of SMB Processes 510
9.2.3 Advanced Parameter and State Estimation for SMB Processes 515
9.2.4 Adaptive Cycle-to-Cycle Control 517
9.2.5 Control of Coupled Simulated Moving Bed Processes for the Production of Pure Enantiomers 519
References 521
10 Chromatography Equipment: Engineering and Operation 525
Henner Schmidt-Traub and Arthur Susanto
10.1 Challenges for Conceptual Process Design 525
10.1.1 Main Cost Factors for a Chromatographic System 527
10.1.2 Conceptual Process Design 528
10.1.2.1 A Case Study: Large-Scale Biotechnology Project 529
10.2 Engineering Challenges 533
10.2.1 Challenges Regarding Sanitary Design 535
10.2.2 Challenges During Acceptance Tests and Qualifications 539
10.3 Commercial Chromatography Columns 540
10.3.1 General Design 541
10.3.1.1 Manually Moved Piston 542
10.3.1.2 Electrically or Hydraulically Moved Piston 542
10.3.2 High- and Low-Pressure Columns 543
10.3.2.1 Chemical Compatibility 544
10.3.2.2 Frit Design 546
10.3.2.3 Special Aspects of Bioseparation 549
10.4 Commercial Chromatographic Systems 551
10.4.1 General Design Aspects: High-Pressure and Low-Pressure Systems 551
10.4.2 Material 553
10.4.3 Batch Low-Pressure Liquid Chromatographic (LPLC) Systems 553
10.4.3.1 Inlets 553
10.4.3.2 Valves to Control Flow Direction 555
10.4.3.3 Pumps 556
10.4.3.4 Pump- and Valve-Based and Gradient Formation 556
10.4.4 Batch High-Pressure Liquid Chromatography 558
10.4.4.1 General Layout 558
10.4.4.2 Inlets and Outlets 559
10.4.4.3 Pumps 559
10.4.4.4 Valves and Pipes 562
10.4.5 Continuous Systems: Simulated Moving Bed 563
10.4.5.1 General Layout 563
10.4.5.2 A Key Choice: The Recycling Strategy 565
10.4.5.3 Pumps, Inlets, and Outlets 566
10.4.5.4 Valves and Piping 566
10.4.6 Auxiliary Systems 567
10.4.6.1 Slurry Preparation Tank 567
10.4.6.2 Slurry Pumps and Packing Stations 568
10.4.6.3 Cranes and Transport Units 568
10.4.6.4 Filter Integrity Test 568
10.4.7 Detectors 569
10.5 Packing Methods 571
10.5.1 Column and Packing Methodology Selection 571
10.5.2 Slurry Preparation 572
10.5.3 Column Preparation 574
10.5.4 Flow Packing 575
10.5.5 Dynamic Axial Compression (DAC) Packing 577
10.5.6 Stall Packing 577
10.5.7 Combined Method (Stall+DAC) 578
10.5.8 Vacuum Packing 580
10.5.9 Vibration Packing 581
10.5.10 Column Equilibration 582
10.5.11 Column Testing and Storage 583
10.5.11.1 Test Systems 583
10.5.11.2 Hydrodynamic Properties and Column Efficiency 584
10.5.11.3 Column and Adsorbent Storage 585
10.6 Process Troubleshooting 585
10.6.1 Technical Failures 586
10.6.2 Loss of Performance 587
10.6.2.1 Pressure Increase 587
10.6.2.2 Loss of Column Efficiency 590
10.6.2.3 Variation of Elution Profile 591
10.6.2.4 Loss of Purity/Yield 592
10.6.3 Column Stability 592
10.7 Disposable Technology for Bioseparations 593
10.7.1 Prepacked Columns 596
10.7.2 Membrane Chromatography 597
References 599
Appendix A Data of Test Systems 601
A.1 EMD53986 601
A.2 Tr¶gerâs Base 602
A.3 Glucose and Fructose 604
A.4 β-Phenethyl Acetate 606
References 607
Index 609
