NMR Spectroscopy is a powerful technique that can provide detailed information on the topology, dynamics and three-dimensional structure of molecules in solution and the solid state

1. Nuclear Magnetic Resonance
By Mehwish Nawaz

2. Contents
• Spectroscopy
• NMR
• Principle
• Steps
• Types
• Pros and Cons

3. Spectroscopy
• Spectroscopy means the dispersion of light
into component colors.
• As per analytical science, every element or
compound has unique characteristic
spectrum. Each compound absorbs and
disperses light over a certain range of
wavelength.

4. Types of spectroscopy
• Absorption spectroscopy – uses electromagnetic
spectra in which a substance absorbs: IR, NMR
• Emission spectroscopy – uses electromagnetic
spectra in which a substance emits:
Luminescence
• Scattering spectroscopy – measures the amount
of light that a substance scatter at certain
wavelengths, incident angles and polarization
angles: Raman

5. Introduction
• NMR spectroscopy is the use of NMR phenomena to
study the physical, chemical, electronic and structural
information about molecules due to either the
chemical shift, Zeeman effect, or the Knight shift effect,
or a combination of both, on the resonant frequencies
of the nuclei present in the sample.
• An NMR instrument allows the molecular structure of a
material to be analyzed by observing and measuring
the interaction of nuclear spins when placed in a
powerful magnetic field.

6. NMR
• It is a powerful technique that can provide
detailed information on the topology,
dynamics and three-dimensional structure of
molecules in solution and the solid state.

7. Why NMR?
• For the analysis of molecular structure at the
atomic level, electron microscopes and X-ray
diffraction instruments can also be used, but
the advantages of NMR are that
• Sample measurements are non-destructive
• There is less sample preparation required.

8. Principle
• Nuclear magnetic resonance (NMR) spectrometry
is based on the net absorption of energy in the
radiofrequency region of the electromagnetic
spectrum by the nuclei of those elements that
have spin angular momentum and a magnetic
moment.
• For the nuclei of a particular element,
characteristic absorption, or resonance
frequencies, and other spectral features provide
useful information on identity and molecular
structure

9. NMR principle
When a nucleus that possesses a magnetic
moment (such as a hydrogen nucleus 1H, or carbon
nucleus 13C) is placed in a strong magnetic field, it
will begin to precess, like a spinning top.

10. Electromagnetic Spectrum
Radio
Frequency

11. Radio waves  Nuclear Spin  NMR Spectroscopy
When low-energy radio waves interact with a molecule,
they can change the nuclear spins of some elements,
including 1H and 13C.

12. NMR Working

13. Steps of NMR

14. NMR

15. Resonant frequency
• It refers to the energy of the absorption, and the
intensity of the signal that is proportional to the
strength of the magnetic field. NMR active nuclei
absorb electromagnetic radiation at a frequency
characteristic of the isotope when placed in a
magnetic field.
• Acquisition of spectra
• Upon excitation of the sample with radio
frequency pulse, a nuclear magnetic resonance
response is obtained. It is a very weak signal and
requires sensitive radio receivers to pick up.

16. Chemical shift
• A chemical shift is defined as the difference in
parts per million (ppm) between the
resonance frequency of the observed proton
and tetramethylsilane (TMS) hydrogens. TMS
is the most common reference compound in
NMR, it is set at δ=0 ppm

17. Spin-spin coupling (splitting)
• The interaction between the spins of
neighboring nuclei in a molecule may cause
the splitting of NMR spectrum. This is known
as spin-spin coupling or splitting. The splitting
pattern is related to the number of equivalent
H-atom at the nearby nuclei.

18. Information given by NMR spectra
• Chemical shift
Information about the composition of atomic groups within
the molecule.
• Spin-Spin coupling constant
Information about adjacent atoms.
• Relaxation time
Information on molecular dynamics.
• Signal intensity
Quantitative information, e.g. atomic ratios within a
molecule that can be helpful in determining the molecular
structure, and proportions of different compounds in a
mixture.

19. Types
• Two common types of NMR spectroscopy are
used to characterize organic structure:
• 1H NMR:- Used to determine the type and
number of H atoms in a molecule
• 13C NMR:- Used to determine the type of
carbon atoms in the molecule

20. Proton or 1H NMR
• The most common for of NMR is based on the
hydrogen-1 (1H), nucleus or proton. It can give
information about the structure of any
molecule containing hydrogen atoms.

21. Interpretation of 1HNMR spectra
• Number of signals - Indicates how many
"different kinds" of protons are present.
• Position of signals - Indicates something about
(chemical shift) magnetic (electronic)
environment of protons
• Relative intensity of signals Proportional to
number of protons present
• Splitting of signals (spin spin coupling) Indicates
the number of nearby nuclei usually protons

22. 13Carbon NMR
• The 1D 13Carbon NMR experiment is much less sensitive than Proton (1H) but has a
much larger chemical shift range.
• Its low natural abundance (1.108%) and proton decoupling means that spin-spin
couplings are seldom observed.
• This greatly simplifies the spectrum and makes it less crowded.
• 13C is a low sensitivity nucleus that yields sharp signals and has a wide chemical
shift range.
• A typical analysis of a 13C NMR spectrum consists of matching expected chemical
shifts to the expected moieties.
• Our NMR service provides 13C NMR along with many other NMR techniques.

24. Types of 2D NMR
• Through bond
• COSY, TOCSY, heteronuclear correlation
• Through space
• NOESY, ROESY, HOESY

25. Nuclear Overhauser Effect
• When a specific nucleus is magnetically excited
and its neighbor is at equilibrium, relaxation
occurs between the two nuclei.
• The smaller the physical distance between the
equilibrium nucleus and the excited nucleus, the
bigger the expected change.
• This dependence can be used to estimate the
distance between nuclei.
• NOE is used to observe through space
correlations in NOESY and ROESY experiments.

26. • NOE Caused by dipolar coupling between
nuclei.
• The local field at one nucleus is affected by
the presence of another nucleus.
• The result is a mutual modulation of
resonance frequencies

27. Applications
• Analysis of Molecular Structure and Identification of
Unknown Chemical Substance
• By studying the peaks of nuclear magnetic resonance
spectra, chemists can determine the structure of many
compounds.
• A chemist can determine the identity of a compound by
comparing the observed nuclear precession frequencies to
known frequencies.
• Dynamics(chemical reaction speed, identification of binding
site, interaction)
• Diffusion Coefficient (molecular weight, conformation of
polymer)
• Relaxation Time (molecular mobility, interatomic distance)

28. Cont..
• Nuclear magnetic resonance imaging, better known as
magnetic resonance imaging (MRI) is an important
medical diagnostic tool used to study the function and
structure of the human body
• NMR is used to generate metabolic fingerprints from
biological fluids to obtain information about disease
states or toxic insults.
• Biochemical information can also be obtained from
living tissue (e.g. human brain tumors) with the
technique known as in vivo magnetic resonance
spectroscopy or chemical shift NMR Microscopy.

29. MRI

30. Pros
• Provides high resolution information
• Does not require a protein crystal and is not
affected by crystal contacts
• Can be used to study flexible proteins
• Reflects conformational averaging

31. Cons
• Any diamagnetic or paramagnetic ions present
in the formation can affect the tool response.
• Expensive
• Requires high concentrations of soluble
protein
• Can not be applied to large proteins (800kD
max so far)
• Can not be used with amyloid fibrils