A Complete Introduction to MODERIN NMR Spectroscopy Roger s Macomber
Dedication First, to those closest to me: my parents, Roxanne, Barbara, Dan, and Juliann. And second, to the memory of Thomas L. Jacobs of UCLA, the person who inspired my life-long interest in organic chemistry. Without all of you, this book would never have tto being
CONTENTS Preface Frequently Used Symbols and Abbreviations Chapter 1 SPECTROSCOPY: SOME PRELIMINARY CONSIDERATIONS 1. I What is NMR Spectroscopy? erties of Electromagnetic Radiation 1.3 Interaction of Radiation with Matter: The Classical Picture 1. 4 Uncertainty and the Question of Time Scale Chapter Summary Review Problems Chapter 2 MAGNETIC PROPERTIES OF NUCLEI 2.1 The structure of an atom 2.2 The Nucleus in a Magnetic Field 669 2.3 Nuclear Energy Levels and Relaxation Times 2.4 The Rotating Frame of Reference 2.5 Relaxation Mechanisms and Correlation Times Chapter Summary Additional Resou Review Problems Chapter 3 OBTAINING AN NMR SPECTRUM 3. 1 Electricity and Magnetism 3.2 The NMR Magnet 3.3 Signal Generation the Old Way: The Continuous-Wave(CW) Experiment 3.4 The Modern Pulsed Mode for Signal Acquisition 3.5 Line Widths, Lineshape, and Sampling Considerations 3.6 Measurement of Relaxation Times
vi CONTENTS Additional Resources Review Problems Chapter 4 A LITTLE BIT OF SYMMETRY 4.1 Symmetry Operations and Distinguishability 4. 2 Conformations and Their Symmetry 4.3 Homotopic, Enantiotopic, and Diastereotopic Nucle 4.4 Accidental Equivalence Chapter Summary Additional Resources 4534444 Review Problems Chapter 5 THE H ANDC NMR SPECTRA OF TOLUENE 5.1 The H NMR Spectrum of Toluene at 80 MHz 5.2 The Chemical Shift Scale 5.3 The 250-and 400-MHz 'H NMR Spectra of Toluene 690 5.4 The"C NMR Spectrum of Toluene at 20.1, 62.9, and 100.6 MHz 5.5 Data Acquisition Parameters 64 Review Problems 67 Chapter 6 CORRELATING PROTON CHEMICAL SHIFTS WITH MOLECULAR STRUCTURE 68 6. 1 Shielding and Deshielding 6.2 Chemical Shifts of Hydrogens Attached to Tetrahedral Carbon 6.3 Vinyl and Formyl Hydrogen Chemical Shifts 6.4 Magnetic Anisotropy 6.5 Aromatic Hydrogen Chemical Shift Correlations 6.6 Hydrogen Attached to Elements Other than Carbon Chapter Summary References 804788566 Additional Resources Review Problems Chapter7 CHEMICAL SHIFT CORRELATIONS FORC AND OTHER ELEMENTS 7. 1 3C Chemical Shifts Revisited 88 7.2 Tetrahedral (sp Hybridized) Carbons 88 7.3 Heterocyclic Structures 7. 4 Trigonal Carbons 92 7.5 Triply Bonded Carbons 7.6 Carbonyl Carbons 96 7.7 Miscellaneous Unsaturated Carbons 7. 8 Summary ofC Chemical Shifts 7.9 Chemical Shifts of Other Elements 10l Chapter Summary References 102 Review Problems SELF-TEST I Chapter 8 FIRST-ORDER (WEAK)SPIN-SPIN COUPLING 110 8.1 Unexpected Lines in an NMR Spectrum l10
8.2 The ' H Spectrum of Diethyl Ether l10 8.4 The Spin-Spin Coupling Checkllisrplified Picture 8.3 Homonuclear H Coupling: The Sin I 12 13 8.5 The n+I Rule l14 8.6 Heteronuclear Spin-Spin Coupling l17 8.7 Review Examples Chapter Summary Chapter 9 FACTORS THAT INFLUENCE THE SIGN AND MAGNITUDE OF J: SECOND-ORDER (STRONG) COUPLING EFFECTS 9. 1 Nuclear Spin Energy Diagrams and the Sign of 9.2 Factors that Influence J: Preliminary Considerati 9.3 One-Bond Coupling Constants 136 9.4 Two-Bond( Geminal)Coupling Constants 9.5 Three-Bond(Vicinal)Coupling Constants 138 9.6 Long-Range Coupling Constants .7 Magnetic Equivalence 143 9.8 Pople Spin System Notation 9.9 Slanting Multiplets and Second-Order(Strong Coupling) Effects 145 9.10 Calculated Spectra 9l1 The Ax→AB→A2 Continuum 9.12 More About the ABX System: Deceptive Simplicity and Virtual Coupling Chapter Summary References Review Problems Chapter 10 THE STUDY OF DYNAMIC PROCESSES BY NMR 10. 1 Reversible and Irreversible Dynamic Processes 10.2 Reversible Intramolecular Processes Involving Rotation Around Bonds 159 10.3 Simple Two-Site Intramolecular Exchange 10.4 Reversible Intramolecular Chemical Processes 10.5 Reversible Intermolecular Chemical Processes 10.6 Reversible Intermolecular Complexation 10.7 Other Examples of Reversible Complexation: Chemical Shift Reagents Chapter Summary 171 References Review Problems 172 Chapter 11 ELECTRON PARAMAGNETIC RESONANCE SPECTROSCOPY AND CHEMICALLY INDUCED DYNAMIC NUCLEAR POLARIZATION 176 11.1 Electron Paramagnetic Resonance 176 11.2 Free Radicals 176 1 1.3 The g Factor 11.4 Sensitivity Considerations 1 1.5 Hyperfine Coupling and the a value 11.6 A Typical EPR Spectrum 181 11.7 CIDNP: Mysterious Behavior of NMR Spectrometers 182 11. 8 The Net Effect 11.9 The Multiplet Effect 11.10 The Radical-Pair Theory of The Net Effect I1.11 The Radical-Pair Theory of the Multiplet Effect 187
viIi CONTENTS 11 12 A Few Final Words about CIDNP Chapter Summary References 189 Review Problems 189 Chapter 12 DOUBLE-RESONANCE TECHNIQUES AND COMPLEX PULSE SEQUENCES 12. 1 What is Double Resonance? 191 12.2 Heteronuclear Spin Decoupling 192 12. 3 Polarization Transfer and the Nuclear Overhauser Effect 193 12.4 Gated and Inverse Gated Decoupling 196 12.5 Off-Resonance Decouplin 196 12.6 Homonuclear Spin Decoupling 198 12.7 Homonuclear Difference NOE: The Test for Proximity 198 12.8 Other Homonuclear Double-Resonance Techniques 12.9 Complex Pulse Sequences 20l 12. 10 The J-Modulated Spin Echo and the APT Experiment 203 12. 11 More About Polarization Transfer 12 12 Distortionless Enhancement by Polarization Transfer 210 210 References 2l1 Additional Resources Review Problems 211 Chapter 13 TWO- DIMENSIONAL NUCLEAR MAGNETIC RESONANCE 215 13. 1 What is 2D NMR Spectroscopy? 215 13.2 2D Heteroscalar Shift-Correlated Spectra 218 13.3 2D Homonuclear Shift-Correlated Spectra 222 13.4 NOE Spectroscopy(NOESY) 13.5 Hetero- and Homonuclear 2D J-Resolved Spectra 13.6 ID and 2D INADEQUATE 230 13.7 2D NMR Spectra of Systems Undergoing Exchange 236 Chapter Summary 236 References Additional Resources Review problems 236 SELF-TEST II 244 Chapter 14 NMR STUDIES OF BIOLOGICALLY IMPORTANT MOLECULES 252 14.1 Introduction 14.2 NMR Line Widths of Biopolymers 14.3 Exchangeable and Nonexchangeable Protons 253 4. 4 Chemical Exchange 254 14.5 The Effects of pH on the NMR Spectra of Biomolecules 256 14.6 NMR Studies of Proteins 256 14.7 NMR Studies of Nucleic Acids 263 14.8 Lipids and Biological Membranes 270 14.9 Carbohydrat Chapter Summary 277 References
Additional Resources 277 277 Chapter 15 SOLID-STATE NMR SPECTROSCOPY 15.1 Why Study Materials in the Solid State? 283 15.2 Why is NMR of Solids Different from NMR of Fluids? 15.3 Chemical Shifts in Solids 15.4 Spin-Spin Coupling 15.5 Quadrupole Coupling 297 15.6 Overcoming Long T]: Cross Polarization Chapter Summary Additional Resources Review Problems Chapter 16 NMR IN MEDICINE AND BIOLOGY: NMR IMAGING 16.1 A window into Anatomy and Physiology 306 16.2 Biomedical NMR 16.3 Pictures with NMR: Magnetic Resonance Imaging 16.4 Image Contrast 16.5 Higher Dimensional Imaging 32 16.6 Chemical Shift Imaging 16.7 NMR Movies: Echo Planar Imaging 16.8 NMR Microscopy 16.9 In Vivo NMR Spectroscopy 326 16.10 Nonmedical Applications of MRI Chapter Summary Additional Resources 33 Appendix 1 ANSWERS TO REVIEW PROBLEMS 334 Appendix 2 PERIODIC TABLE SUBJECT INDEX 370 CHEMICAL COMPOUNDS INDEX 379
PREFACE In the decade since the first version of this book was written, will be well prepared to solve any molecular structure prob the field of nuclear magnetic resonance(NMR)spectroscopy lem given a complete set of NMr data. And you will be able has made, quite literally, quantum leaps. Computer-controlled to proceed confidently to any advanced treatise on NMR NMR spectrometers with high-field superconducting mag Above all, I have tried to make the text clear, logical, and nets, once available only to the most well-funded institutions, interesting to read. There are hundreds of figures and actual are now commonplace. Two-dimensional NMR techniques, spectra to illustrate the many topics. The vast majority of in their infancy a decade ago, have blossomed into an indis- spectra were obtained on modern high-field spectrometers pensable array of tools to elucidate the molecular structure of Because of the way I structured the book, I recommend that compounds as complex as proteins. And who is unaware of you proceed through it chapter by chapter, rather than skip- the growing importance of NMR to medical diagnosis ping around. Nonetheless, I have tried to help those readers through the technique of magnetic resonance imaging(MRI)? who skip around by adding liberal references to earlier sec Nuclear magnetic resonance has become ubiquitous in tions that have supporting information on each topic such divergent fields as chemistry, physics, material science, Soon after starting Chapter I you will note that I have biology, medicine, and forensic science, among others. In adopted a semiprogrammed approach. That is, there are fre- writing this book it has been my goal to provide a monograph quent example problems(with solutions)to test your mastery aimed at anyone, not just chemists, with an interest in learning of the topic at hand. To get the most from your reading, try to about NMR. Medical students, whose interest lie not so much work each problem as you encounter it. They often contain in molecular structure as it does in the three-dimensional important additional information about the material just cov distribution of magnetic nuclei such as hydrogen, will find ered. Then, at the end of each chapter there is a chapter what they are looking for in the beginning three chapters, summary and several review problems to see if you have which discuss the physics of NMR signal generation, and mastered the concepts in that chapter. There are also two Chapter 16, which describes MRI. If your interests are in self-tests(after Chapters 7 and 13)that will help you assess biochemistry, Chapter 14 will be very useful to you. The your overall mastery of the subject. The answers to these majority of you will use NMR to elucidate molecular struc- review and self-test problems appear in Appendix I tures, so in Chapters 4-13 I have tried to give you everything I hope you enjoy this book and that it inspires you to learn you will need to get that job done efficiently all you can about modern NMR methods. I would also appre A first-year college chemistry course is the only scientific ciate greatly any feedback you would like to offer background I have assumed the reader has. All the necessary details are developed from the most basic level. The approach Malibu, California is relatively nonmathematical, with only a few simple equa Roger S Macomber tions. But do not let this fool you. By the end of the book you
ACKNOWLEDGMENTS Many individuals have contributed mightily to this book, and on the slopes, so I expected to get a lot of writing done Before without their help I could not have gone forward with the I left Utah, over 120 inches of fresh powder had fallen. Need I say more? About two years ago Jeff Holtmeier, a consulting editor at While at Utah a number of individuals offered to contribute ohn Wiley, approached me to see if I had any interest in example spectra for inclusion in the book. I have mentioned writing an updated edition of my earlier book on NMr the source of each contributed spectrum in the text or figure spectroscopy, published in early 1988. When I said yes, he set legend, but let me introduce them here, as well: Bobby L. about getting the original edition reviewed by a half-dozen Williamson(with Peter Stangs group), Alan Sopchik( with experts in the field. With these in hand, he single-handedly Wes Bentrude's group), Dhileepkumar Krishnamurthy and promoted the project through the appropriate Wiley channels, Stan McHardy(with Gary Keck's group), and John Bender resulting in a contract in late 1995 and Soren Giese(with Fred Wests group). I also wish to thank As the prospectus for the new edition was being developed, Steve Fetherston and Rosemary Laufer, able staff members several of my colleagues agreed to contribute chapters in their who made my visit even more productive and enjoyable particular areas of expertise, George Kreishman and Elwood Back again in Cincinnati, Elwood Brooks, Marshall wil Brooks(UCs staff NMR spectroscopist)collaborated to write son, and graduate students Pat Hutchins, Mark Guttaduaro Chapter 14, dealing with the applications of NMR to bio- and Sheela Venkitachalam contributed additional spectra for chemistry. Jerry Ackerman, a one-time colleague of mine at the book, as did David Watt of the University of Kentucky the University of Cincinnati and now director of NMR spec- But there are two major contributors of example spectra who troscopy at Massachusetts General Hospital and assistant deserve special thanks. Sadtler Research Laboratories( Divi- professor of radiology at the Harvard Medical School, offered sion of Bio-Rad Laboratories, Inc ) through staff scientist to contribute Chapters 15 and 16 on solid-state NMR and Marie Scandone, provided many of the one-dimensional H etic resonance imaging. and C spectra. And David Lankin, a graduate of this depart Fortuitously, at that time I signed the contract, I was just ment and currently a research scientist and NMR expert at beginning a previously arranged two-quarter sabbatical leave, Searle, provided the bulk of the special 2D and related spectra during which I planned to make major progress on the new in Chapters 12 and 13, with the help of Geoffrey Cordell of edition. My first stop was the Chemistry Department at the the University of Illinois School of Pharmacy University of Hawaii, hosted by my friend and colleague, Karl Finally, three more of my faculty colleagues at UC deserve Seff. Here I spent a most enjoyable month writing drafts of special thanks. Albert Bobst advised me on the EPR section several chapters, delivering and attending seminars, surfing, of Chapter Il, Frank Meeks provided mathematical advice and biking. Then, after six weeks back in Cincinnati, I headed and Allan Pinhas helped edit the entire set of proofs. To all to the University of Utah, where I was hosted by my friends these individuals, I give a sincere thank you! Pete Stang and Wes Bentrude. Before I arrived, the winter in Utah had been exceptionally mild, with absolutely no snow Roger S Macomber
FREQUENTLY USED SYMBOLS AND ABBREVIATIONS ., an unpaired electron β Bohr magneton ↑or↓, possible orientations(with respect to Bo) of the agnetic moment of an electron or an 1=, nucleus [] concentration in moles per liter m,n-fold axis of C (I, indicates the nucleus being irradiated in a double- C(A)T, computed(axial)tomography COSY, 2D correlation spectroscopy 2D, NMR spectrum where signal intensity is a function of CSR, chemical shift reagent d, doublet(signal multiplicity) A, (1)mass number of a nucleus; (2)net CIDNP absorption D, deuterium (H) △, a change(eg△E, a change in energy) a, hyperfine coupling constant in an EPR spectrum 8. chemical shift signal position(pp a,(1)one possible orientation(with respect to Bo) of the 811.822, 833. chemical shift tensors magnetic moment of an electron or an /=7 nucleus; (2)the flip angle in a pulsed NMR experiment Aox, substituent shielding parameter Av, frequency difference(Hz) between the signal of inter- AA'BB, Pople designation for a coupled four-spin system est and the operating frequency of the spectrometer: consisting of two chemically but not magnetically also used for electric quadrupole interaction equivalent nuclei in each of two sets Avo, frequency difference(Hz) between two sites at the A/E, absorption-emission CIDNP net effect APT, attached proton test 8v, frequency difference(Hz) between the signal of inter- est and the frequency of the internal standard signal also a measure of spectral resolution Bo, external(applied)magnetic field vector 8r, spatial resolution along x axis B. oscillating magnetic field vector of the observing chan An, field of view along x axis DEPT, distortionless enhancement by polarization transfer B2, oscillating magnetic field vector of the"irradiating DMF, dimethylformamide B, possible orientation( with respect to Bo)of the magnetic moment of an electron or an /=i nucleus E, (I)energy:(2)net CIDNP emission