Applied NMR spectroscopy for chemists and life scientists

Content: Preface XV 1 Introduction to NMR Spectroscopy 1 1.1 Our First 1D Spectrum 1 1.2 Some Nomenclature: Chemical Shifts, LineWidths, and Scalar Couplings 2 1.3 Interpretation of Spectra: A Simple Example 5 1.4 Two-Dimensional NMR Spectroscopy: An Introduction 9 Part One Basics of Solution NMR 11 2 Basics of 1DNMR Spectroscopy 13 2.1 The Principles of NMR Spectroscopy 13 2.2 The Chemical Shift 16 2.3 Scalar Couplings 17 2.4 Relaxation and the Nuclear Overhauser Effect 20 2.5 Practical Aspects 23 2.5.1 Sample Preparation 23 2.5.2 Referencing 25 2.5.3 Sensitivity and Accumulation of Spectra 27 2.5.4 Temperature Calibration 29 2.6 Problems 30 Further Reading 31 3 1H NMR 33 3.1 General Aspects 33 3.2 Chemical Shifts 34 3.2.1 Influence of Electronegativity of Substituents 35 3.2.2 Anisotropy Effects 35 3.2.3 Other Factors Affecting Chemical Shifts: Solvent, Temperature, pH, and Hydrogen Bonding 37 3.2.4 Shift Reagents 37 3.3 Spin Systems, Symmetry, and Chemical or Magnetic Equivalence 39 3.3.1 Homotopic, Enantiotopic, and Diastereotopic Protons 42 3.3.2 Determination of Enantiomeric Purity 43 3.4 Scalar Coupling 44 3.4.1 First-Order Spectra 45 3.4.2 Higher-Order Spectra and Chemical Shift Separation 47 3.4.3 Higher-Order Spectra and Magnetic Equivalence 49 3.5 1H 1H Coupling Constants 50 3.5.1 Geminal Couplings 50 3.5.2 Vicinal Couplings 50 3.5.3 Long-Range Couplings 52 3.5.4 1HCouplings to Other Nuclei 52 3.6 Problems 54 Further Reading 55 4 NMRof13C and Heteronuclei 57 4.1 Properties of Heteronuclei 57 4.2 Indirect Detection of Spin-1/2 Nuclei 59 4.3 13C NMR Spectroscopy 59 4.3.1 The 13C Chemical Shift 60 4.3.2 X,13C Scalar Couplings 64 4.3.3 Longitudinal Relaxation of 13C Nuclei 68 4.3.4 Recording 13C NMR Spectra 68 4.4 NMR of Other Main Group Elements 70 4.4.1 Main Group Nuclei with I D 1/2 71 4.4.2 Main Group Nuclei with I >
1/2 75 4.5 NMR Experiments with Transition Metal Nuclei 78 4.5.1 Technical Aspects of Inverse Experiments with I D 1/2 Metal Nuclei 79 4.5.2 Experiments with Spin I >
1/2 Transition Metal Nuclei 81 4.6 Problems 82 Further Reading 84 Part Two Theory of NMR Spectroscopy 85 5 Nuclear Magnetism A Microscopic View 87 5.1 The Origin of Magnetism 87 5.2 Spin An Intrinsic Property of Many Particles 88 5.3 Experimental Evidence for the Quantization of the Dipole Moment: The Stern Gerlach Experiment 93 5.4 The Nuclear Spin and Its Magnetic Dipole Moment 94 5.5 Nuclear Dipole Moments in a Homogeneous Magnetic Field: The Zeeman Effect 96 5.5.1 Spin Precession 98 5.6 Problems 103 6 Magnetization A Macroscopic View 105 6.1 The Macroscopic Magnetization 105 6.2 Magnetization at Thermal Equilibrium 106 6.3 Transverse Magnetization and Coherences 108 6.4 Time Evolution of Magnetization 109 6.4.1 The Bloch Equations 110 6.4.2 Longitudinal and Transverse Relaxation 112 6.5 The Rotating Frame of Reference 115 6.6 RF Pulses 117 6.6.1 Decomposition of the RF Field 118 6.6.2 Magnetic Fields in the Rotating Frame 119 6.6.3 The Bloch Equations in the Rotating Frame 120 6.6.4 Rotation of On-Resonant and Off-Resonant Magnetization under the Influence of Pulses 121 6.7 Problems 122 7 Chemical Shift and Scalar and Dipolar Couplings 125 7.1 Chemical Shielding 125 7.1.1 The Contributions to Shielding 127 7.1.2 The Chemical Shifts of Paramagnetic Compounds 131 7.1.3 The Shielding Tensor 132 7.2 The Spin Spin Coupling 133 7.2.1 Scalar Coupling 134 7.2.2 Quadrupolar Coupling 140 7.2.3 Dipolar Coupling 141 7.3 Problems 144 Further Reading 145 8 A Formal Description of NMR Experiments: The Product Operator Formalism 147 8.1 Description of Events by Product Operators 148 8.2 Classification of Spin Terms Used in the POF 149 8.3 Coherence Transfer Steps 151 8.4 An Example Calculation for a Simple 1D Experiment 152 Further Reading 153 9 A Brief Introduction into the Quantum-Mechanical Concepts of NMR 155 9.1 Wave Functions, Operators, and Probabilities 155 9.1.1 Eigenstates and Superposition States 156 9.1.2 Observables of Quantum-Mechanical Systems and Their Measured Quantities 157 9.2 Mathematical Tools in the Quantum Description of NMR 158 9.2.1 Vector Spaces, Bra s, Ket s, and Matrices 158 9.2.2 Dirac s Bra Ket Notation 159 9.2.3 Matrix Representation of State Vectors 160 9.2.4 Rotations between State Vectors can be Accomplished with Tensors 161 9.2.5 Projection Operators 162 9.2.6 Operators in the Bra Ket Notation 163 9.2.7 Matrix Representations of Operators 165 9.3 The Spin Space of Single Noninteracting Spins 166 9.3.1 Expectation Values of the Spin-Components 168 9.4 Hamiltonian and Time Evolution 169 9.5 Free Precession 169 9.6 Representation of Spin Ensembles The Density Matrix Formalism 171 9.6.1 Density Matrix at Thermal Equilibrium 173 9.6.2 Time Evolution of the Density Operator 173 9.7 Spin Systems 175 9.7.1 Scalar Coupling 176 Part Three Technical Aspects of NMR 179 10 The Components of an NMR Spectrometer 181 10.1 The Magnet 181 10.1.1 Field Homogeneity 182 10.1.2 Safety Notes 183 10.2 Shim System and Shimming 184 10.2.1 The Shims 184 10.2.2 Manual Shimming 185 10.2.3 Automatic Shimming 186 10.2.4 Using Shim Files 187 10.2.5 Sample Spinning 187 10.3 The Electronics 187 10.3.1 The RF Section 188 10.3.2 The Receiver Section 189 10.3.3 Other Electronics 189 10.4 The Probehead 189 10.4.1 Tuning and Matching 190 10.4.2 Inner and Outer Coils 191 10.4.3 Cryogenically Cooled Probes 191 10.5 The Lock System 192 10.5.1 The 2H Lock 192 10.5.2 Activating the Lock 193 10.5.3 Lock Parameters 194 10.6 Problems 194 Further Reading 194 11 Acquisition and Processing 195 11.1 The Time Domain Signal 197 11.2 Fourier Transform 199 11.2.1 Fourier Transform of Damped Oscillations 199 11.2.2 Intensity, Integral, and Line Width 200 11.2.3 Phases of Signals 201 11.2.4 Truncation 202 11.2.5 Handling Multiple Frequencies 202 11.2.6 Discrete Fourier Transform 203 11.2.7 Sampling Rate and Aliasing 204 11.2.8 How Fourier Transformation Works 205 11.3 Technical Details of Data Acquisition 209 11.3.1 Detection of the FID 209 11.3.2 Simultaneous and Sequential Sampling 210 11.3.3 Digitizer Resolution 213 11.3.4 Receiver Gain 214 11.3.5 Analog and Digital Filters 215 11.3.6 Spectral Resolution 216 11.4 Data Processing 217 11.4.1 Digital Resolution and Zero Filling 217 11.4.2 Linear Prediction 219 11.4.3 Pretreatment of the FID: Window Multiplication 220 11.4.4 Phase Correction 227 11.4.5 Magnitude Mode and Power Spectra 229 11.4.6 Baseline Correction 230 11.5 Problems 231 Further Reading 232 12 Experimental Techniques 233 12.1 RF Pulses 233 12.1.1 General Considerations 234 12.1.2 Hard Pulses 235 12.1.3 Soft Pulses 236 12.1.4 Band-Selective RF Pulses 237 12.1.5 Adiabatic RF Pulses 238 12.1.6 Composite Pulses 240 12.1.7 Technical Considerations 241 12.1.8 Sources and Consequences of Pulse Imperfections 243 12.1.9 RF Pulse Calibration 244 12.1.10 Transmitter Pulse Calibration 245 12.1.11 Decoupler Pulse Calibration (13C and 15N) 246 12.2 Pulsed Field Gradients 247 12.2.1 Field Gradients 247 12.2.2 Using Gradient Pulses 248 12.2.3 Technical Aspects 250 12.3 Phase Cycling 251 12.3.1 The Meaning of Phase Cycling 251 12.4 Decoupling 255 12.4.1 How Decoupling Works 255 12.4.2 Composite Pulse Decoupling 256 12.5 Isotropic Mixing 257 12.6 Solvent Suppression 257 12.6.1 Presaturation 258 12.6.2 Water Suppression through Gradient-Tailored Excitation 259 12.6.3 Excitation Sculpting 260 12.6.4 WET 260 12.6.5 One-Dimensional NOESY with Presaturation 260 12.6.6 Other Methods 261 12.7 Basic 1D Experiments 262 12.8 Measuring Relaxation Times 262 12.8.1 Measuring T1 Relaxation The Inversion-Recovery Experiment 262 12.8.2 Measuring T2 Relaxation The Spin Echo 263 12.9 The INEPT Experiment 266 12.10 The DEPT Experiment 268 12.11 Problems 270 13 The Art of Pulse Experiments 271 13.1 Introduction 271 13.2 Our Toolbox: Pulses, Delays, and Pulsed Field Gradients 272 13.3 The Excitation Block 273 13.3.1 A Simple 90y Pulse Experiment 273 13.3.2 The Effects of 180y Pulses 273 13.3.3 Handling of Solvent Signals 274 13.3.4 A Polarization Transfer Sequence 275 13.4 The Mixing Period 277 13.5 Simple Homonuclear 2D Sequences 278 13.6 Heteronuclear 2D Correlation Experiments 279 13.7 Experiments for Measuring Relaxation Times 281 13.8 Triple-Resonance NMR Experiments 283 13.9 Experimental Details 284 13.9.1 Selecting the Proper Coherence Pathway: Phase Cycles 284 13.9.2 Pulsed Field Gradients 286 13.9.3 N-Dimensional NMR and Sensitivity Enhancement Schemes 288 13.10 Problems 289 Further Reading 289 Part Four Important Phenomena and Methods in Modern NMR 291 14 Relaxation 293 14.1 Introduction 293 14.2 Relaxation: The Macroscopic Picture 293 14.3 The Microscopic Picture: Relaxation Mechanisms 294 14.3.1 Dipole Dipole Relaxation 295 14.3.2 Chemical Shift Anisotropy 297 14.3.3 Scalar Relaxation 298 14.3.4 Quadrupolar Relaxation 298 14.3.5 Spin Spin Rotation Relaxation 299 14.3.6 Paramagnetic Relaxation 299 14.4 Relaxation and Motion 299 14.4.1 A Mathematical Description of Motion: The Spectral Density Function 300 14.4.2 NMR Transitions That Can Be Used for Relaxation 302 14.4.3 The Mechanisms of T1 and T2 Relaxation 303 14.4.4 Transition Probabilities 304 14.4.5 Measuring Relaxation Rates 306 14.5 Measuring 15N Relaxation to Determine Protein Dynamics 306 14.5.1 The Lipari Szabo Formalism 307 14.6 Measurement of Relaxation Dispersion 310 14.7 Problems 313 15 The Nuclear Overhauser Effect 315 15.1 Introduction 315 15.1.1 Steady-State and Transient NOEs 318 15.2 The Formal Description of the NOE: The Solomon Equations 318 15.2.1 Different Regimes and the Sign of the NOE: Extreme Narrowing and Spin Diffusion 320 15.2.2 The Steady-State NOE 321 15.2.3 The Transient NOE 324 15.2.4 The Kinetics of the NOE 324 15.2.5 The 2D NOESY Experiment 325 15.2.6 The Rotating-Frame NOE 327 15.2.7 The Heteronuclear NOE and the HOESY Experiment 329 15.3 Applications of the NOE in Stereochemical Analysis 330 15.4 Practical Tips for Measuring NOEs 332 15.5 Problems 333 Further Reading 334 16 Chemical and Conformational Exchange 335 16.1 Two-Site Exchange 335 16.1.1 Fast Exchange 338 16.1.2 Slow Exchange 340 16.1.3 Intermediate Exchange 340 16.1.4 Examples 342 16.2 Experimental Determination of the Rate Constants 344 16.3 Determination of the Activation Energy by Variable-Temperature NMR Experiments 346 16.4 Problems 348 Further Reading 349 17 Two-Dimensional NMR Spectroscopy 351 17.1 Introduction 351 17.2 The Appearance of 2D Spectra 352 17.3 Two-Dimensional NMR Spectroscopy: How Does It Work? 354 17.4 Types of 2D NMR Experiments 357 17.4.1 The COSY Experiment 358 17.4.2 The TOCSY Experiment 359 17.4.3 The NOESY Experiment 362 17.4.4 HSQC and HMQC Experiments 364 17.4.5 The HMBC Experiment 365 17.4.6 The HSQC-TOCSY Experiment 366 17.4.7 The INADEQUATE Experiment 367 17.4.8 J-Resolved NMR Experiments 368 17.5 Three-Dimensional NMR Spectroscopy 370 17.6 Practical Aspects of Measuring 2D Spectra 370 17.6.1 Frequency Discrimination in the Indirect Dimension: Quadrature Detection 370 17.6.2 Folding in 2D Spectra 376 17.6.3 Resolution in the Two Frequency Domains 377 17.6.4 Sensitivity of 2D NMR Experiments 378 17.6.5 Setting Up 2D Experiments 379 17.6.6 Processing 2D Spectra 380 17.7 Problems 381 18 Solid-State NMR Experiments 383 18.1 Introduction 383 18.2 The Chemical Shift in the Solid State 384 18.3 Dipolar Couplings in the Solid State 386 18.4 Removing CSA and Dipolar Couplings: Magic-Angle Spinning 387 18.5 Reintroducing Dipolar Couplings under MAS Conditions 388 18.5.1 An Alternative to Rotor-Synchronized RF Pulses: Rotational Resonance 390 18.6 Polarization Transfer in the Solid State: Cross-Polarization 391 18.7 Technical Aspects of Solid-State NMR Experiments 393 18.8 Problems 394 Further Reading 394 19 Detection of Intermolecular Interactions 395 19.1 Introduction 395 19.2 Chemical Shift Perturbation 397 19.3 Methods Based on Changes in Transverse Relaxation (Ligand-Observe Methods) 398 19.4 Methods Based on Changes in Cross-Relaxation (NOEs) (Ligand-Observe or Target-Observe Methods) 400 19.5 Methods Based on Changes in Diffusion Rates (Ligand-Observe Methods) 403 19.6 Comparison of Methods 404 19.7 Problems 405 Further Reading 406 Part Five Structure Determination of Natural Products by NMR 407 20 Carbohydrates 419 20.1 The Chemical Nature of Carbohydrates 419 20.1.1 Conformations of Monosaccharides 422 20.2 NMR Spectroscopy of Carbohydrates 423 20.2.1 Chemical Shift Ranges 423 20.2.2 Systematic Identification by NMR Spectroscopy 424 20.2.3 Practical Tips: The Choice of Solvent 429 20.3 Quick Identification 430 20.4 A Worked Example: Sucrose 430 Further Reading 437 21 Steroids 439 21.1 Introduction 439 21.1.1 The Chemical Nature 440 21.1.2 Proton NMR Spectra of Steroids 441 21.1.3 Carbon Chemical Shifts 443 21.1.4 Assignment Strategies 444 21.1.5 Identification of the Stereochemistry 447 21.2 A Worked Example: Prednisone 449 Further Reading 456 22 Peptides and Proteins 457 22.1 Introduction 457 22.2 The Structure of Peptides and Proteins 458 22.3 NMR of Peptides and Proteins 461 22.3.1 1HNMR 461 22.3.2 13C NMR 464 22.3.3 15N NMR 467 22.4 Assignment of Peptide and Protein Resonances 469 22.4.1 Peptides 470 22.4.2 Proteins 473 22.5 A Worked Example: The Pentapeptide TP5 476 Further Reading 480 23 Nucleic Acids 481 23.1 Introduction 481 23.2 The Structure of DNA and RNA 482 23.3 NMR of DNA and RNA 486 23.3.1 1HNMR 486 23.3.2 13C NMR 489 23.3.3 15NNMR 490 23.3.4 31P NMR 490 23.4 Assignment of DNA and RNA Resonances 492 23.4.1 Unlabeled DNA/RNA 492 23.4.2 Labeled DNA/RNA 496 Further Reading 498 Appendix 499 A.1 The Magnetic H and B Fields 499 A.2 Magnetic Dipole Moment and Magnetization 500 A.3 Scalars, Vectors, and Tensors 501 A.3.1 Properties of Matrices 504 Solutions 507 Index 525