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In addition, this textbook teaches the fundamentals of all the major analytical techniques used in the pharmaceutical laboratory, and teaches the international pharmacopoeias and guidelines of importance for the field. It is primarily intended for the pharmacy student, to teach the requirements in “analytical chemistry” for the 5 years pharmacy curriculum, but the textbook is also intended for analytical chemists moving into the field of pharmaceutical analysis.
"It is teaches the fundamentals of all the major analytical techniques and their application in pharmaceutical laboratories, and it is valuable resource for pharmacy students and analytical chemists working in pharmaceutical analysis." (Analytical and Bioanalytical Chemistry, 1 August 2012)
1 Introduction to Pharmaceutical Analysis 1
1.1 Applications and Definitions 1
1.2 The Life of Medicines 4
1.3 The Quality of Medical Products 8
1.4 Summary 11
2 International Pharmacopoeias, Regulations and Guidelines 13
2.1 Overview of Legislation 13
2.2 Legislation and Regulations for Industrial Production 14
2.3 Life Time of Drugs and Drug Substances 17
2.4 Pharmacopoeias 18
2.5 International Harmonization 19
2.5.1 International Conference on Harmonization 20
2.5.2 Pharmacopoeial Discussion Group 20
2.6 Legislation and Regulations for Pharmacy Production 20
2.7 Summary 21
3 Fundamental Chemical Properties, Buffers and pH 23
3.1 pH and pKa 23
3.2 Partition 25
3.3 Stereochemistry 28
3.4 Stability Testing 29
3.5 Summary 30
4 Fundamentals of Pharmaceutical Analysis 33
4.1 What is a Pharmaceutical (Chemical) Analysis? 33
4.2 How to Specify Quantities and Concentrations? 35
4.3 Basic Laboratory Equipment 37
4.3.1 The Analytical Balance 37
4.3.2 Pipettes 41
4.3.3 Volumetric Flasks 44
4.3.4 Burettes 47
4.4 How to Make Solutions and Dilutions 47
4.5 Calibration of Analytical Methods 49
4.6 Errors, Accuracy, and Precision 50
4.6.1 Systematic and Random Errors 50
4.6.2 Accuracy and Precision 51
4.7 Statistics 52
4.7.1 Mean Value and Standard Deviation 52
4.7.2 Confidence Intervals 54
4.7.3 Comparison of Means with a t-Test 55
4.7.4 Q-Test to Reject Outliers 56
4.7.5 Linear Regression with the Method of Least Squares 57
4.7.6 How to Present an Analytical Result 58
4.8 Some Words and Concepts 62
4.8.1 Analysis and Determination 62
4.8.2 Sample Replicates and Measuring Replicates 62
4.8.3 Interference 62
4.8.4 Blind Samples 62
5 Titrimetric Methods 65
5.1 Introduction 65
5.2 Acid–Base Titrations 72
5.3 Acid–Base Titrations in Non-Aqueous Media 75
5.4 Redox Titrations 78
5.5 Other Principles of Titration 81
5.6 Summary 82
6 Introduction to Spectroscopic Methods 83
6.1 Electromagnetic Radiation 83
6.2 Molecules and Electromagnetic Radiation 85
6.3 Atoms and Electromagnetic Radiation 86
6.4 Summary 88
7 UV Spectrophotometry 89
7.1 Principle of Quantitative Determination 89
7.2 Principle of Identification 94
7.3 Which Substances Have Strong UV Absorbance? 95
7.4 Instrumentation 95
7.5 Practical Work and Method Development 99
7.6 Areas of Usage and Performance 101
7.7 System Testing 101
7.8 Summary 102
8 IR Spectrophotometry 103
8.1 IR Spectrophotometry 103
8.2 Instrumentation 106
8.3 Scope 109
8.4 Instrument Calibration 109
8.5 NIR Spectrophotometry 110
8.6 Applications 112
8.7 Summary 114
9 Atomic Spectrometry 115
9.1 Atomic Absorption Spectrometry 115
9.2 Instrumentation 118
9.3 Applications and Performance 121
9.4 Practical Work and Method Development 122
9.5 Atomic Emission Spectrometry 123
9.6 Instrumentation 124
9.7 Summary 124
10 Chromatography 127
10.1 General Principles 127
10.2 Retention 131
10.3 Column Efficiency 133
10.4 Selectivity 135
10.5 Peak Symmetry 136
10.6 Resolution 138
10.7 Chromatographic Techniques 140
10.8 Summary 140
11 Chromatographic Separation Principles 141
11.1 General Introduction 141
11.2 Normal Phase Chromatography 142
11.2.1 Silica 142
11.2.2 Interactions 143
11.2.3 Order of Elution 144
11.2.4 Other Stationary Phases 145
11.2.5 Mobile Phases 146
11.2.6 Summary of Normal Phase Chromatography 147
11.3 Reversed Phase Chromatography 148
11.3.1 Stationary Phases 148
11.3.2 Retention Mechanisms 150
11.3.3 Mobile Phases 152
11.3.4 Ion-Pair Chromatography 155
11.3.5 Summary of Reversed Phase Chromatography 155
11.4 Hydrophilic Interaction Chromatography 156
11.5 Chiral Separations 156
11.6 Size Exclusion Chromatography 158
11.6.1 Principle 158
11.6.2 Summary of SEC 160
11.7 Ion Exchange Chromatography 160
12 Thin-Layer Chromatography 163
12.1 Introduction 163
12.2 Apparatus 164
12.3 TLC Plates 166
12.4 Stationary Phases 166
12.5 Mobile Phases 167
12.6 Chromatographic Development 168
12.7 Detection 169
12.8 Applications of TLC 169
12.9 Quantitative Analysis and Instrumentation 170
12.10 Summary 171
13 High Performance Liquid Chromatography 173
13.1 Introduction 173
13.2 The Chromatographic Separation Process 175
13.3 The Column 177
13.4 Pumps 180
13.5 Detectors 182
13.5.1 UV detector 182
13.5.2 Fluorescence Detector 184
13.5.3 Electrochemical Detector 186
13.5.4 Refractive Index, Evaporative Light Scattering and Corona Discharge Detectors 186
13.5.5 Combination of Detectors 187
13.6 Injectors 187
13.7 Mobile Phases 188
13.8 Solvents for Sample Preparation 189
13.9 Reporting the Results 189
13.10 Summary 190
14 Gas Chromatography 191
14.1 Introduction 191
14.2 Apparatus 192
14.3 Temperature 193
14.4 Carrier Gas 195
14.5 Stationary Phases 196
14.6 Selectivity in GC 197
14.7 Columns 198
14.7.1 Capillary Columns 198
14.7.2 Packed Columns 199
14.8 Injection Systems 200
14.8.1 Injection Systems for Capillary Columns 200
14.8.2 Injection Systems for Packed Columns 202
14.9 Detectors 203
14.9.1 Flame Ionization Detector 203
14.9.2 Nitrogen–Phosphorus Detector 203
14.9.3 Thermal Conductivity Detector 204
14.9.4 Electron Capture Detector 204
14.9.5 Mass Spectrometry Detector 206
14.10 Derivatization 206
14.10.1 Silylation 206
14.10.2 Alkylation 207
14.10.3 Acylation 207
14.11 The Uses of GC 208
14.12 More Advanced GC techniques 209
14.13 Summary 209
15 Capillary Electrophoresis 211
15.1 Principle and Theory 211
15.2 Electroosmotic Flow 213
15.3 Instrumentation 214
15.4 The Capillary 217
15.5 Sample Introduction 218
15.6 Capillary Zone Electrophoresis; an Example 221
15.7 Micellar Electrokinetic Chromatography 222
15.8 Chiral Separations 224
15.9 Coated Capillaries 225
15.10 Non-Aqueous CE 229
15.11 Summary 229
16 Mass Spectrometry 231
16.1 Introduction 231
16.2 Basic Theory 233
16.3 Electron Ionization 236
16.4 Identification using Electron Ionization Spectra 237
16.5 Characterization of Totally Unknowns using Electron Ionization Spectra 239
16.6 Chemical Ionization 244
16.7 Electrospray Ionization 246
16.8 Atmospheric Pressure Chemical Ionization 247
16.9 High-Resolution Mass Spectrometry 248
16.10 Instrumentation 250
16.11 Chromatography Coupled with Mass Spectrometry 253
16.12 Quantitative GC-MS and LC-MS 256
16.13 Areas of Usage and Performance 257
16.14 Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry 257
16.15 Inductively Coupled Plasma Mass Spectrometry 258
16.16 Summary 259
17 Miscellaneous Chemical Techniques 261
17.1 Potentiometric Determination of Ions using Ion-Selective Electrodes 261
17.2 Paper Chromatography 263
17.3 Supercritical Fluid Chromatography 264
17.4 Gel Electrophoresis 265
17.5 Iso-Electric Focusing 267
17.6 Nuclear Magnetic Resonance Spectrometry 268
17.7 Raman Spectrometry 270
18 Sample Preparation 273
18.1 Why is Sample Preparation Required? 273
18.2 Main Strategies 274
18.3 Recovery and Enrichment 276
18.4 Protein Precipitation 278
18.5 Liquid–Liquid Extraction 279
18.5.1 Fundamentals 279
18.5.2 A Closer Look at the Theory 279
18.5.3 Extraction Solvents 282
18.5.4 Calculation of Recovery 283
18.5.5 Multiple Extractions 285
18.5.6 LLE with Back-Extraction 286
18.6 Solid–Liquid Extraction 287
18.7 Solid Phase Extraction 287
18.7.1 Fundamentals 287
18.7.2 The SPE Column 288
18.7.3 Conditioning 289
18.7.4 Equipment 290
18.7.5 Reversed-Phase SPE 290
18.7.6 Secondary Interactions 292
18.7.7 Ion Exchange SPE 293
18.7.8 Mixed-Mode SPE 295
18.7.9 Normal-Phase SPE 297
18.8 Summary 298
19 Analytical Chemical Characteristics of Selected Drug Substances 299
19.1 Amitriptyline and Mianserin 299
19.2 Morphine and Codeine 301
19.3 Ibuprofen and Naproxen 302
19.4 Furosemide 304
19.5 Paracetamol (Acetaminophen) 306
19.6 Neutral Drugs 307
20 Quantification and Quality of Analytical Data 309
20.1 Peak Height and Peak Area 309
20.2 Calibration Methods 310
20.2.1 External Standard Method 310
20.2.2 Internal Standard Method 313
20.2.3 Standard Addition 314
20.2.4 Normalization 314
20.3 Validation 314
20.3.1 Analytical Procedure 317
20.3.2 Accuracy 317
20.3.3 Precision 318
20.3.4 Specificity 320
20.3.5 Detection Limit 320
20.3.6 Quantification Limit 321
20.3.7 Linearity and Range 321
20.3.8 Robustness 323
20.3.9 Test Methods in the European Pharmacopeia 325
20.4 System Suitability 325
20.4.1 Adjustment of Chromatographic Conditions 326
21 Chemical Analysis of Drug Substances 327
21.1 What is a Pharmaceutical Raw Material, how is it Produced and why must it be Controlled? 327
21.2 The Pharmacopoeias – the Basis for Control of Pharmaceutical Raw Materials 330
21.3 Which Contaminants are Found in Raw Materials, What are the Requirements in a Maximum Content and Why? 337
21.3.1 Well Defined Chemical Compounds 339
21.3.2 Mixtures of Organic Compounds 343
21.4 How to Check the Identity of Pharmaceutical Raw Materials 344
21.4.1 Overview of the Identification Procedures 344
21.4.2 Techniques used for the Identification of Well Defined Chemical Compounds 344
21.4.2.1 Infrared Absorption Spectrophotometry 344
21.4.2.2 Ultraviolet and Visible Absorption Spectrophotometry 347
21.4.2.3 Thin-Layer Chromatography 351
21.4.2.4 Melting Point 352
21.4.2.5 Polarimetry 353
21.4.2.6 High Performance Liquid Chromatography 356
21.4.2.7 Chloride and Sulfate Identification 359
21.5 How to Test for Impurities in Pharmaceutical Raw Materials 359
21.5.1 Main Purity Tests for Well Defined Chemical Compounds 359
21.5.1.1 Appearance of Solution 361
21.5.1.2 Absorbance 364
21.5.1.3 Acidity/Alkalinity 365
21.5.1.4 Optical Rotation 365
21.5.1.5 Related Substances 366
21.5.1.6 Solvent Residues 372
21.5.1.7 Foreign Anions 372
21.5.1.8 Cationic Impurities 376
21.5.1.9 Loss on Drying 378
21.5.1.10 Determination of Water 379
21.5.2 Purity Tests for Raw Materials of the Type of Mixtures of Organic Compounds 382
21.5.2.1 Oxidizing Substances 383
21.5.2.2 Acid Value 383
21.5.2.3 Hydroxyl Value 384
21.5.2.4 Iodine Value 384
21.5.2.5 Peroxide Value 385
21.5.2.6 Saponification Value 385
21.5.2.7 Unsaponifiable Matter 386
21.5.2.8 Other Tests 386
21.5.3 Identification of the Raw Materials of the Type of Mixtures of Organic Compounds 388
21.6 How to Determine the Purity of Pharmaceutical Raw Materials 389
21.6.1 Acid–Base Titration in Aqueous Environment 389
21.6.2 Acid–Base Titration in a Non-Aqueous Environment 393
21.6.3 Redox Titrations 396
21.6.4 High Performance Liquid Chromatography 396
21.6.5 UV spectrophotometry 401
21.7 How to Control Compounds for Which no Pharmacopoeia Monograph Exists 402
21.8 How are Ph.Eur. and USP Updated? 402
22 Chemical Analysis of Final Pharmaceutical Products 405
22.1 Quality Control of Final Pharmaceutical Products 405
22.2 Monographs and Chemical Testing 406
22.3 Identification of the Active Pharmaceutical Ingredient 412
22.4 Assay of the Active Pharmaceutical Ingredient 427
22.5 Chemical Tests for Final Pharmaceutical Products 446
22.5.1 Test for Related Substances 446
22.5.2 Uniformity of Content 449
22.5.3 Dissolution 451
23 Analysis of Drugs in Biological Fluids 453
23.1 Introduction 453
23.1.1 Drug Development 453
23.1.2 Therapeutic Drug Monitoring 455
23.1.3 Forensic and Toxicological Analysis 456
23.1.4 Doping Control Analysis 457
23.2 The Biological Matrix 458
23.3 Bioanalytical Methods 460
23.3.1 Sampling 460
23.3.2 Sample Preparation 461
23.3.3 Protein Precipitation 462
23.3.4 Liquid–Liquid Extraction 463
23.3.5 Solid-Phase Extraction 463
23.3.6 Separation 464
23.3.7 Detection 464
23.3.8 Calibration and Quantification 465
23.4 Examples 466
23.4.1 Sample Preparation 466
23.4.1.1 Sample Preparation Procedure by LLE 466
23.4.1.2 Comments to the Procedure 466
23.4.1.3 Sample Preparation Procedure by LLE and Back Extraction 467
23.4.1.4 Comments to the Procedure 467
23.4.1.5 Sample Preparation Procedure by SPE 467
23.4.1.6 Comments to the Procedure 468
23.4.1.7 Sample Preparation Procedure by Protein Precipitation 468
23.4.1.8 Comments to the Procedure 468
23.4.2 Quantitative Determination 468
23.4.2.1 Quantitative Determination of Amitriptyline in Serum by LC-MS 468
23.4.2.2 Comments to the Procedure 469
23.4.2.3 Determination of Valproic Acid in Serum by GC-MS 471
23.4.2.4 Comments to the Procedure 471
23.4.3 Identification 472
23.4.3.1 Sample Preparation Procedure for Unknown Screening by Mixed Mode Cation Exchange 472
23.4.3.2 Comments to the Procedure 472
23.4.3.3 GC-MS Procedure for Unknown Screening 473
23.4.3.4 Comments to the Procedure 473
23.4.3.5 LC-MS-MS Procedure for Unknown Screening 475
23.4.3.6 Comments to the Procedure 475
Index 477
Overview
In addition, this textbook teaches the fundamentals of all the major analytical techniques used in the pharmaceutical laboratory, and teaches the international pharmacopoeias and guidelines of importance for the field. It is ...