Spin Resonance Spectroscopy: Principles and applications

Spin Resonance Spectroscopy

Principles and Applications

Chandran Karunakaran

Associate Professor of Chemistry, Biomedical Research Lab, VHNSN College (Autonomous), Virudhunagar, Tamilnadu, India

Table of Contents

Cover image

Title page

Copyright

List of Contributors

Preface

Abbreviations

Chapter One. Principles of Nuclear Magnetic Resonance and Pulsed Nuclear Magnetic Resonance

1.1. Introduction

1.2. Spin Magnetic Properties

1.3. Theory of Nuclear Magnetic Resonance and Electron Spin Resonance Spectroscopy

1.4. Comparison of Nuclear Magnetic Resonance and Electron Spin Resonance

1.5. Nuclear Magnetic Resonance Instrumentation

1.6. Chemical Shift

1.7. Nuclear Magnetic Resonance Reference Samples

1.8. Spin–Spin Coupling

1.9. Classification of Nuclear Magnetic Resonance System

1.10. Types of Spin–Spin Splitting

1.11. Low- and High-Resolution Nuclear Magnetic Resonance

1.12. Simplification of Complex Nuclear Magnetic Resonance Spectra

1.13. Sensitivity and Boltzmann Statistics

1.14. Pulsed/Fourier Transform Nuclear Magnetic Resonance

1.15. Relaxation Process and Relaxation Times

1.16. Measurement of T1 and T2

1.17. Bloch Equation

1.18. Lineshape and Line Width Analysis

1.19. Nuclear Magnetic Resonance and Electron Paramagnetic Resonance Problems

Chapter Two. 1H and 13C Nuclear Magnetic Resonance Spectroscopy

2.1. Interpretation of 1H Nuclear Magnetic Resonance

2.2. Assignment of 1H Nuclear Magnetic Resonance

2.3. Factors Influencing the Chemical Shift of Protons

2.4. Coupling Constant (J)

2.5. Symmetry in Nuclear Magnetic Resonance

2.6. 13C Nuclear Magnetic Resonance Spectroscopy

2.7. 13C Chemical Shifts and Their Assignment

2.8. Additivity Rule

2.9. Steric Effects

2.10. Stereoelectronic Effects on 1JCH (Perlin Effects)

2.11. Nuclear Magnetic Double Resonance

2.12. Single Frequency Off-Resonance Decoupling (SFORD)

2.13. Distortionless Enhancement by Polarization Transfer (DEPT)

2.14. Insensitive Nuclei Enhanced by Polarization Transfer (INEPT)

2.15. Chemically Induced Dynamic Nuclear Polarization (CIDNP)

2.16. Nuclear Overhauser Effect (NOE)

2.17. Multidimensional Nuclear Magnetic Resonance Spectroscopy

2.18. 2D Nuclear Magnetic Resonance Spectroscopy

2.19. 3D Nuclear Magnetic Resonance

Chapter Three. Applications of Nuclear Magnetic Resonance

3.1. Applications of (1H, 31P, 19F, and 15N) Nuclear Magnetic Resonance to Inorganic Structural Elucidation

3.2. Dynamic Nuclear Magnetic Resonance Spectroscopy

3.3. Dynamic Exchange Process in Organic Compounds

3.4. Nuclear Magnetic Resonance of Fluxional Inorganic Molecules

3.5. Solid State Nuclear Magnetic Resonance and Magic Angle Spinning

3.6. Nuclear Magnetic Resonance of Paramagnetic Molecules

3.7. Shift Reagents

3.8. Nuclear Magnetic Resonance Imaging

Chapter Four. Electron Paramagnetic Resonance Spectroscopy

4.1. Introduction

4.2. Spin Hamiltonian of Electron Paramagnetic Resonance

4.3. Presentation of Electron Paramagnetic Resonance Spectra

4.4. Electron Paramagnetic Resonance Instrumentation

4.5. Parallel and Perpendicular Modes Transitions

4.6. Choice of Experimental Conditions

4.7. Parameters Setting for Electron Paramagnetic Resonance Spectra

4.8. Determination of g-Values From Experimental Electron Paramagnetic Resonance Spectra

4.9. Electron Paramagnetic Resonance Reference Samples

4.10. Isotropic Hyperfine Interaction

4.11. Factors Affecting the Magnitude of g-Values

4.12. g-Tensor and Anisotropy

4.13. A-Tensor and Anisotropy

4.14. Measurement of Electron Paramagnetic Resonance Spectra

4.15. Zero-Field Splitting, Kramer's Theory, and Degeneracy

4.16. Exchange Coupled Electron Paramagnetic Resonance

4.17. Triplet State Electron Paramagnetic Resonance (Dipole–Dipole Interaction)

4.18. Optically Detected Magnetic Resonance (ODMR)

4.19. Nuclear Zeeman Interaction

4.20. Nuclear Quadrupole Interaction

4.21. Jahn-Teller Splitting

4.22. Jahn-Teller Distortion

4.23. Computer Simulation and Softwares

Chapter Five. Advances in Electron Paramagnetic Resonance

5.1. Multifrequency Electron Paramagnetic Resonance

5.2. Pulsed Electron Paramagnetic Resonance

5.3. Saturation Recovery Electron Paramagnetic Resonance

5.4. Time-Resolved Electron Paramagnetic Resonance

5.5. Electron Spin Echo Envelope Modulation (ESEEM)

5.6. Hyperfine Sublevel Correlation (HYSCORE)

5.7. Multiresonance Electron Paramagnetic Resonance

5.8. Electron–Nuclear Double Resonance (ENDOR)

5.9. TRIPLE Resonance

5.10. Continuous Wave Electron–Electron Double Resonance

5.11. Electron Paramagnetic Resonance Imaging

Chapter Six. Applications of Electron Paramagnetic Resonance

6.1. Applications of Electron Paramagnetic Resonance to Transition Metal Complexes

6.2. Electron Paramagnetic Resonance of Exchange Coupled Systems/Clusters

6.3. Electron Paramagnetic Resonance of Free Radicals and Spin Trapping

6.4. Electron Paramagnetic Resonance Spin Trapping—Indirect Method

6.5. Spin Labeling Versus Spin Probes

6.6. Electron Paramagnetic Resonance Dosimetry

Index

Copyright

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List of Contributors

Chandran Karunakaran,     VHNSN College (Autonomous), Virudhunagar, India

Murugesan Balamurugan,     VHNSN College (Autonomous), Virudhunagar, India

Paulraj Santharaman,     VHNSN College (Autonomous), Virudhunagar, India

Raju Rajkumar,     VHNSN College (Autonomous), Virudhunagar, India

Murugesan Karthikeyan,     VHNSN College (Autonomous), Virudhunagar, India

Dhinesh Kumar Marimuthu,     VHNSN College (Autonomous), Virudhunagar, India

Preface

This book is primarily intended to provide the fundamentals of the spin resonance spectroscopy including nuclear magnetic resonance (NMR) and electron paramagnetic resonance (EPR), methods, advances, and applications to basic science viz., biology, chemistry, physics, and also medicine. It is hoped that the reader will be able to acquire the knowledge of both the NMR and EPR in a single-source book and the state-of-the-art developments in this field as it stands today. This book covers the syllabus of leading universities and colleges. Thus, this book caters to the needs of students of basic sciences, researchers, and R&D scientists in industries, who wish to understand, enhance, and broaden their knowledge and expertise on the fundamentals, technologies, and applications of spin resonance spectroscopy. The book begins with the introduction of NMR and ESR, theory, instrumentation, and comparison of them. Chapter 1 discusses the parameters obtained from NMR, mechanism of spin–spin splitting, classification of NMR systems, pulse/Fourier transform NMR, relaxation processes and times, Bloch equations, and line width/shape. In Chapter 2, the advances in ¹H and ¹³C NMR viz., double-resonance NMR including nuclear Overhauser effect, single frequency off-resonance decoupling, distortionless enhancement by polarization transfer, insensitive nuclei enhanced by polarization transfer, chemically induced dynamic nuclear polarization, and multidimensional viz., 2D NMR including homonuclear and heteronuclear and 3D NMR are described. In Chapter 3, the applications of NMR to structural elucidation of inorganic compounds especially ³¹P and ¹⁹F NMR are highlighted. Dynamic NMR, solid-state NMR (magic-angle spinning), NMR of paramagnetic molecules, shift and contrast reagents, and magnetic resonance imaging are also discussed.

Chapter 4 presents the basics of EPR, spin Hamiltonian to describe the observed spectra, determination of g-value and factors affecting it, Kramer's degeneracy, zero-field splitting, exchange-coupled EPR, triplet state, free radical EPR, optically detected magnetic resonance, Jahn Teller effect, and spectral simulation software. In Chapter 5, the recent development and advances in EPR viz., multifrequency, high-field EPR, and pulse EPR including electron spin echo envelope modulation, hyperfine sublevel correlation, and multifrequency EPR viz., electron nuclear double resonance, (continous wave and pulsed), TRIPLE, and double electron–electron resonance are discussed. Chapter 6 presents the applications of EPR to various transition metal ions, explore the active site of enzymes, fast flow, rapid freeze-quench, spin trapping, and spin label EPR to detect and monitor the dynamics of unstable free radicals; in vivo EPR and EPR imaging using spin probes are also highlighted. The above chapters thus attempt to provide the essential background knowledge and up-to-date advances in this field. The book should thus serve as introductory text for those who intend to specialize in either the theoretical or practical applications. This textbook is also hoped to be a fruitful launch pad for many careers in spin resonance spectroscopy.

Thanks are due to the contributing authors of this book for their active support and continuous cooperation throughout the course of preparation and all those great and eminent researchers in this field in and around the globe whose valuable contributions made this work possible. It is a real pleasure to acknowledge my mentors Prof. R. Murugesan, Prof. B. Kalyanaraman, Prof. William Antholine, Prof. Brian Benett, and all the teachers for their inspiration and guidance. Many thanks are also due to my collaborative researchers viz., Prof. Brian Volkmann of Medical College of Wisconsin, Prof. John McCraken, Michigan State University, Prof. Anumantha Kanthasamy, Iowa State University, Prof. Narayan Avadhani University of Pennsylvania, Prof. S. Subramanian, IIT-Chennai, Dr. M. Velayutham, University of Pittsburgh, Dr. K.R. Justin Thomas, IIT, Roorkee, Dr. Srigiridhar Kotamraju and Dr. Shasi Kalivendi, IICT, Hyderabad, Dr. B. Kalpana, DIPAS-DRDO, Dr. Sushil Kumar Singh, SSPL-DRDO, Delhi, Prof. Anuradha Dhanasekeran, Anna University, Chennai, Prof. Mainak Das, IIT-Kanpur, Dr. V. Shanmugaiah, Madurai Kamaraj University, Prof. Jonathan Claussen, Iowa State University, Dr. A. Rajendran, Dr. D. Christopher Durairaj, Dr. S. Elango and all colleagues for their input and sharing knowledge, the Principal and Managing Board of our institution for their encouragement and cooperation, Prof. C.S. Gopinath, National Chemical Lab, Pune, and Dr. Sujatha Venkataraman, University of Colorado for giving their suggestions and critical comments to improve the book, and my wife and kids for their perseverance and support during the preparation of the book. I am indebted to my present and past research scholars Dr. S. Rajesh, Dr. U.S.E. Arivudainambi, Dr. S. Rajasingh, Dr. S. Prakash, Dr. P. Gurusamy, Dr. V. Ramesh, Dr. M. Pandiaraj, Dr. T. Madasamy, Dr. M. Balamurugan, Dr. G. Muneeswaran, Dr. P. Santharaman, Mr. R. Rajkumar, Mr. M. Karthikeyan, and Mr. M. Dhinesh Kumar for their contributions enabling me to undertake this task and complete it. Finally, it has been a great experience to work with Elsevier publisher of this book and special thanks to the entire them for their effort in making this book possible.

Abbreviations

Chapter One

Principles of Nuclear Magnetic Resonance and Pulsed Nuclear Magnetic Resonance

Chandran Karunakaran, Raju Rajkumar, and Murugesan Balamurugan     VHNSN College (Autonomous), Virudhunagar, India

Abstract

Spin resonance spectroscopy deals with the nuclear and electron spin resonance absorptions in the radio and microwave frequencies used for investigating diamagnetic (I  ≥  1/2) and paramagnetic (S  ≥  1/2) systems, respectively. This chapter focuses on the theory of nuclear magnetic resonance (NMR), its instrumentation, and comparison with electron paramagnetic resonance spectroscopy. The parameters obtained from NMR viz., chemical shift and spin–spin splitting including their types (first and second orders), characteristics, and mechanisms (dipolar and indirect scalar coupling) are discussed. Furthermore, the classification of NMR systems including chemically and magnetically equivalent nuclei is presented. The simplification techniques for complex NMR spectra are also highlighted. It also includes the principles of pulse and Fourier transform NMR and different methods viz., free induction decay, saturation recovery, spin echo (SE), inversion recovery, and stimulated SE to investigate the relaxation process and times. The time evolution of magnetization is described by Bloch equation. The different lineshapes and factors affecting line width are also discussed.

Keywords

Chemical shift; Coupling constant; FID; Lineshape and line width; NMR spin system; Relaxation times; Spin echo

Contents

1.1 Introduction

1.2 Spin Magnetic Properties

1.3 Theory of Nuclear Magnetic Resonance and Electron Spin Resonance Spectroscopy

1.3.1 Larmor Frequency

1.3.2 Resonance Condition

1.4 Comparison of Nuclear Magnetic Resonance and Electron Spin Resonance

1.5 Nuclear Magnetic Resonance Instrumentation

1.6 Chemical Shift

1.7 Nuclear Magnetic Resonance Reference Samples

1.7.1 Tetramethylsilane

1.7.2 4,4-Dimethyl-4-silapentane-1-sulfonic Acid

1.7.3 4,4-Dimethyl-4-silapentane-1-ammonium Trifluoroacetate

1.7.4 Nuclear Magnetic Resonance Solvents

1.8 Spin–Spin Coupling

1.8.1 Characteristics of Spin–Spin Coupling

1.8.2 Mechanism of Nuclear Spin–Spin Interaction

1.8.2.1 Direct Interaction (Dipole–Dipole Coupling D)

1.8.2.2 Indirect or Scalar Coupling (Spin–Spin Splitting J)

1.9 Classification of Nuclear Magnetic Resonance System

1.9.1 Chemically and Magnetically Equivalent Nuclei

1.10 Types of Spin–Spin Splitting

1.10.1 First-Order Splitting

1.10.2 Second-Order Splitting

1.10.3 Effect of J on Energy Levels of AB System

1.11 Low- and High-Resolution Nuclear Magnetic Resonance

1.11.1 Low-Resolution Nuclear Magnetic Resonance

1.11.2 High-Resolution Nuclear Magnetic Resonance

1.12 Simplification of Complex Nuclear Magnetic Resonance Spectra

1.12.1 Using Shift Reagents

1.12.2 High-Field Nuclear Magnetic Resonance

1.12.3 Deuterium Exchange and Labeling

1.12.4 Spin Decoupling/Double Resonance Method

1.12.5 Multidimensional Nuclear Magnetic Resonance

1.12.6 Magic Angle Spinning

1.13 Sensitivity and Boltzmann Statistics

1.14 Pulsed/Fourier Transform Nuclear Magnetic Resonance

1.14.1 Nuclear Magnetic Resonance Pulse

1.14.2 Quadrature Detection

1.15 Relaxation Process and Relaxation Times

1.15.1 Spin-Lattice Relaxation Time (T1)

1.15.2 Spin–Spin Relaxation Time (T2)

1.15.3 Mechanism of T1 and T2

1.16 Measurement of T1 and T2

1.16.1 Free Induction Decay Method and Saturation Recovery

1.16.2 Spin Echo Method (or) Hahn Echo Method

1.16.2.1 Significance of Spin Echo Method

1.16.3 Stimulated Echo

1.16.4 Inversion Recovery Method

1.17 Bloch Equation

1.18 Lineshape and Line Width Analysis

1.18.1 Lineshape

1.18.1.1 Lorentzian Lineshape

1.18.1.2 Gaussian Lineshape

1.18.1.3 Voigt Lineshape

1.18.2 Line Width

1.19 Nuclear Magnetic Resonance and Electron Paramagnetic Resonance Problems

References

1.1. Introduction

Spin magnetic resonance spectroscopy is broadly classified into two types, viz., (1) nuclear magnetic resonance (NMR) spectroscopy and (2) electron spin resonance spectroscopy (ESR)/electron paramagnetic resonance (EPR)/electron magnetic resonance based on the nuclear spin and electron spin, respectively. NMR was independently discovered in the year 1946 by the American physicists F. Bloch (1905–83) of Stanford University and E.M. Purcell (1912–97) of Harvard University. They shared the 1952 Physics Nobel prize. ESR was first observed by the Russian physicist E. Zavoisky [1a]. It is interesting to note here that the famous Dutch physicist C.J. Gorter [1b] was the first person to look for NMR in bulk matter. Because of his bad luck, he failed to observe resonance as the sample was easily saturated because of its large spin-lattice relaxation time. These two branches viz., NMR and ESR are essentially similar, based on the same principles. However, NMR spectra are observed in the radiofrequency (RF) region and ESR spectra in the microwave region of electromagnetic spectrum. The application of ESR is limited to species containing unpaired electrons such as organic and inorganic free radicals, triplet states, and the ions of the d-block and f-block elements. The first application of NMR to chemistry was made in 1951 when the NMR spectrum of ethanol was recorded. Since then, NMR technique has almost completely been monopolized by chemists, biochemists, and molecular biologists for structural elucidation, physicists being primarily involved in its theoretical and instrumentation aspects.

NMR spectroscopy takes advantage of the magnetic properties of certain nuclei and records the absorption of energy due to transition between quantized nuclear spin energy levels. In an NMR experiment, the spectrometer is tuned to the frequency of a particular magnetic nucleus, viz., ¹H, ¹³C, ¹⁹F, etc. It is thus a very powerful technique, the closest analogy being a powerful microscope that allows the chemist to see the structure of molecules in solution. Actually, the NMR experiment does not directly show how all the atoms are connected. However, it is up to the chemist to take the information provided by NMR spectra to build a model of the molecule [1–12].

1.2. Spin Magnetic Properties

NMR is possible owing to the magnetic spin properties of certain nuclei. In addition to charge and mass, several nuclei also possess a property called nuclear spin. Because nuclei have a charge, they generate a magnetic field due to its spinning motion and hence magnetic moment.

There are useful empirical rules relating mass number, atomic number (Z), and nuclear spin quantum number (I):

In other words, the rules for determining the net spin of a nucleus are expressed as follows:

1. If the number of neutrons and protons are both even, then the nucleus has no spin (i.e., I  =  0).

2. If the number of neutrons plus the number of protons is odd, then the nucleus has a half-integer spin (i.e., I  =  1/2, 3/2, 5/2, etc.).

3. If the number of neutrons and the number of protons are both odd, then the nucleus has an integer spin (i.e., I  =  1, 2, 3, etc.).

Because NMR depends on the existence of a nuclear spin, nuclei with I  =  0 have no NMR spectrum (e.g., ⁴He, ¹²C, ¹⁶O, ¹⁸O, ³²S). The most important class of nuclei, which are NMR active those with I  =  1/2 (e.g., ¹H, ¹³C, ¹⁵N, ¹⁹F) and nuclei with I  >  1/2 (e.g., ¹¹B, I  =  3/2; ²H, ¹⁰C, ¹⁴N, I  =  1), have quadrupole moments, a nonspherical distribution of nuclear charge, which results in broad absorption lines and makes observation of NMR spectra more difficult. The quadrupole moment can even affect the lineshape of neighboring nuclei. For example, the resonances of protons bonded to nitrogen or boron atoms are generally broad in ¹H NMR spectra. We shall thus be primarily concerned with nuclei with I  =  1/2. A list of some common isotopes with I  =  1/2 is provided in Table 1.1.

Table 1.1

Natural abundances of some common nuclear magnetic resonance–active isotopes with I  =  1/2

a μ in units of nuclear magnetons  =  5.05  ×  10−²⁷  JT−¹.

1.3. Theory of Nuclear Magnetic Resonance and Electron Spin Resonance Spectroscopy

Consider a nucleus of bare proton with nuclear spin I  =  1/2, so that mI  =  ±1/2, associated with it a magnetic dipole moment μ that is proportional to the angular momentum. Similarly, an isolated electron with S  =  1/2 and ms  =  ±1/2 also possess magnetic moment.

When the nucleus of magnetic moment μ is placed in a magnetic field B0, the interaction energy (E) is given by

The splitting of the nuclear and electron spin energy levels in an external magnetic field (B0) is shown above. The diagram is not drawn to the scale. The nuclear splitting should be even smaller than it is shown.

1.3.1. Larmor Frequency

In the presence of applied magnetic field (B0), the interaction between μ and B0 produces a force called torque, which causes the nuclear magnetic moment (μ) to precess about z-axis as shown in Fig. 1.1 (similar to gyroscope). This process is called Larmor precession. The angular frequency of precession is known as Larmor frequency (ω) given by

Figure 1.1  Larmor frequency.

(1.2)

(1.3)

ω0 is the precession rate called the Larmor frequency. It is defined as the frequency with which the magnetic dipolar moment of a spin precesses around an external magnetic field (B0). The constant γ is called the magnetogyric (or) gyromagnetic ratio. It relates the magnetic moment μ and the spin number I for any specific nucleus as follows

(1.4)

1.3.2. Resonance Condition

If we irradiate the sample with radio waves (MHz), the proton can absorb the energy and get promoted to the higher energy state. This absorption is called nuclear spin resonance because the frequency of the applied RF radiation and the precession coincide or resonate. Thus, the resonance condition, i.e., (hv  =  gNμNB0), is achieved by applying an RF oscillating field perpendicular to the external static field. The proton resonance frequencies for different applied field (B0) strengths (in Gauss) are shown in Table 1.2.

Each nucleus has a characteristic value of γ as shown in Table 1.1. For proton, γ  =  2.674  ×  10⁴  G−¹  s−¹. Therefore, for different nuclei with different gyromagnetic ratios, different frequencies must be applied to achieve the resonance condition. The frequencies fall in the RF region of the electromagnetic spectrum. In ESR, the resonant frequencies tend to be much higher, by virtue of the 659-times higher gyromagnetic ratio of an unpaired electron relative to a proton. For example, a typical magnetic field strength used in ESR spectrometers is 0.32  T, with a corresponding resonant frequency of about 9000  MHz (9.8  GHz), whereas in NMR, the resonant frequency is 14  MHz.

Table 1.2

Proton resonance frequency at different magnetic field

1.4. Comparison of Nuclear Magnetic Resonance and Electron Spin Resonance

1.5. Nuclear Magnetic Resonance Instrumentation

The basis of NMR experiment is to induce a transition between two consecutive nuclear spin energy levels that result when a nucleus is placed in a magnetic field. The resonance condition is given by the following equation, hv  =  gNμNB0. So, the NMR spectrum can be observed experimentally either

1. by varying the external magnetic field, keeping the frequency fixed (or)

2. by varying the frequency, keeping the magnetic field fixed.

The first approach is often preferred because it is difficult to vary the frequency at a very high level of stability. The basic components of a typical NMR spectrometer (Fig. 1.2) are as follows:

1. Electromagnet: It gives a powerful, stable, and homogeneous magnetic field. The field must be constant over the entire area of the sample and over the period of the experiment.

2 Sweep coil generator: It supplies a variable current to a secondary magnet. Then, the total applied magnetic field can be varied over a small range.

3. Sample holder: The sample container is usually a glass tube (5  mm OD) spun by an air-driven turbine to average the magnetic field over the sample container. The process is often referred to as the spinning of the sample.

Figure 1.2  (A) Block diagram of nuclear magnetic resonance (NMR) spectrometer and (B) photograph of 700   MHz Brucker spectrometer.

4. RF transmitter: It is an oscillator connected to a coil, called the transmitter coil. It generates the RF radiation and transmits the RF energy to the sample. The axis of the coil has to be perpendicular to the magnetic field, i.e., the RF is applied perpendicular to B.

5. RF receiver: It is connected to a coil, called the receiver coil, and encircles the sample (Fig. 1.2). Its axis has to be perpendicular to both the magnetic field and the axis of the transmitter coil. The transmitter and receiver coils and the sample holder are constructed into a single unit called probe. Often, the necessary small magnetic sweep field is obtained by the use of sweep generator coils built into the NMR probe.

6. Read out system: It consists of an RF amplifier, recorder/computer, and other accessories to increase the sensitivity, resolution, and accuracy.

A photograph of NMR spectrometer and schematic representation of these components are shown in Fig. 1.2. At a fixed frequency, the magnetic field at which the resonance occurs for a given nucleus is constant. Therefore, each instrument normally works only for a particular kind of nucleus. Several nuclei can be studied by changing the oscillator, and obviously, the field has also to be changed accordingly. In the case of solids, because the width of lines is expected to be large, this type of NMR setup for investigation cannot be used (see Section 3.5).

1.6. Chemical Shift

For bare proton, the field experienced by the ¹H nuclei is the same as the applied field B0. However, when proton is present in a molecule, the circulating electrons and bonded electrons or other π electrons change (increase or decrease) the magnetic field experienced by the ¹H nuclei; i.e.,

it may also be represented as

(1.5)

σ is called shielding constant.

The Larmor frequency of the precession spins in the presence of applied magnetic field.

We know that

The NMR resonance frequencies obtained from the respective fields of reference (R) and unknown compound (C) are as follows.

(1.6)

σ is small typically 10−⁵ for protons and <10−³ for other nuclei. δ is defined in terms of