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NUCLEAR MAGNATIC RESONANCE SPECTROSCOPY
INTRODUCTION
BASICS
PRINCIPLE
INSTRUMENTATION
SHEILDING AND DESHIELDING
&APPLICATIONS
INTRODUCTION
Nuclear magnetic resonance (NMR) is a physical phenomenon based
upon the quantum mechanical magnetic properties of an atom's
nucleus.
NMR also commonly refers to a family of scientific methods that exploit
nuclear magnetic resonance to study molecules ("NMR spectroscopy").
The method of NMR was first developed by E.M. Purcell and Felix
bloch(1946)
Major application of NMR spectroscopy lies in the area of synthetic
organic chemistry, inorganic chemistry, bio-organic chemistry, bioinorganic chemistry,
NMR Historic Review
NMR Historic Review
1924
Pauli proposed the presence of nuclear magnetic moment to explain the
hyperfine structure in atomic spectral lines.
1930
Nuclear magnetic moment was detected using refined Stern-Gerlach
experiment by Estermann.
1939
Rabi et al. First detected unclear magnetic resonance phenomenon by
applying r.f. energy to a beam of hydrogen molecules in the Stern-Gerach
set up and observed measurable deflection of the beam.
1946
Purcell et al. at Harvard reported nuclear resonance absorption in paraffin
wax.
Bloch et al. at Stanford found nuclear resonance in liquid water.
1949
Chemical shift phenomenon was observed.
1952
Nobel prize in Physics was awarded to Purcell and Bloch.
1966
Ernst and Anderson first introduce the Fourier Transform technique into
NMR.
NMR.
Late in the 1960s:




Solid State NMR was revived due to the effort of Waugh.
and associates at MIT.
Biological application become possible due to the introduction
superconducting magnets.
NMR imaging was demonstrated.
1970
2D NMR was introduced.
1980s
Macromolecular structure determination in solution by NMR was
achieved.
1991
Nobel prize in Chemistry was awarded to Richard Ernst.
1990s
Continuing development of heteronuclear multi-dimensional NMR permit
the determination of protein structure up to 50 KDa.
MRI become a major radiological tool in medical diagnostic.
2002
NMR
Applications
Nobel prize in Chemistry
was awarded
to Kurt Wuthrich
(One of themost, if not the most, important analytical spectroscopic tool.)
1. Biomedical applications:
Edward M. Purcell
1912-1997
Felix Bloch
1905-1983
Richard R. Ernst
1933-
Kurt Wuthrich
1938-
CW NMR 40MHz
1960
800 MHz
NMR Spectroscopy
Where is it?
1nm
(the wave)
Frequency
(the transition)
(spectrometer)
102
10
X-ray
103
UV/VIS
electronic
X-ray
104
Infrared
Vibration
UV/VIS
Infrared/Raman
Fluorescence
105
106
Microwave
Rotation
107
Radio
Nuclear
NMR
Before using NMR
What are N, M, and R ?
Properties of the Nucleus
Nuclear spin
Nuclear magnetic moments
The Nucleus in a Magnetic Field
Precession and the Larmor frequency
Nuclear Zeeman effect & Boltzmann distribution
When the Nucleus Meet the right Magnet and radio wave
Nuclear Magnetic Resonance
Nuclear magnetic moments
Magnetic moment  is another important parameter for a nuclei
 =  I (h/2)
I: spin number; h: Plank constant;
: gyromagnetic ratio (property of a nuclei)
1H:
I=1/2 ,  = 267.512 *106 rad T-1S-1
13C:
I=1/2 ,  = 67.264*106
15N:
I=1/2 ,  = 27.107*106
PRINCIPLE
• Subatomic particles (electrons, protons and neutrons) can be
imagined as spinning on their axes.
•
In many atoms (such as 12C) these spins are paired against each
other, such that the nucleus of the atom has no overall spin.
• However, in some atoms (such as 1H and 13C) the nucleus does
possess an overall spin. The rules for determining the net spin of a
nucleus are as follows;
1. If the number of neutrons and the number of protons are both even, then the
nucleus has NO spin.
2 . If the number of neutrons plus the number of protons is odd, then the
nucleus has a half-integer spin (i.e. 1/2, 3/2, 5/2)
3.If the number of neutrons and the number of protons are both odd, then the
nucleus has an integer spin (i.e. 1, 2, 3)
Natural %
Abundanc
e
Isotope
Spin
(I)
Magnetic
Moment
(μ)*
Magnetogyri
c
Ratio (γ)†
1H
99.9844
1/2
2.7927
26.753
2H
0.0156
1
0.8574
4,107
11B
81.17
3/2
2.6880
--
13C
1.108
1/2
0.7022
6,728
17O
0.037
5/2
-1.8930
-3,628
19F
100.0
1/2
2.6273
25,179
29Si
4.700
1/2
-0.5555
-5,319
31P
100.0
1/2
1.1305
10,840
1. A spinning charge generates a magnetic field.The resulting spinmagnet has a magnetic moment (μ) proportional to the spin.
2. In the presence of an external magnetic field (B0), two spin states
exist, +1/2 and -1/2.
3.The magnetic moment of the lower energy +1/2 state is aligned with
the external field, but that of the higher energy -1/2 spin state is
opposed to the external field. Note that the arrow representing the
external field points North.
In an Applied Magnetic Field
•Nuclei with 2 allowed spin states
can align either with or against the
field, with slight excess of nuclei
aligned with the field
•The nuclei precess about an axis
parallel to the applied magnetic
field, with a frequency called the
Larmor Frequency (w)
Larmor Frequency is Proportional
to the Applied Magnetic Field
Slow precession in small
magnetic field
Faster precession in larger
magnetic field
Nuclear Zeeman effect
• Zeeman effect: when an atom is placed in an external magnetic field, the
energy levels of the atom are split into several states.
• The energy of a give spin sate (Ei) is directly proportional to the value of mI
and the magnetic field strength B0
Spin State Energy EI=- . B0 =-mIB0 r(h/2p)
•For a nucleus with I=1/2, the energy difference between two states is
ΔE=E-1/2-E+1/2 = B0 r(h/2p)
m=1/2
m=-1/2
The Zeeman splitting is proportional to the strength of the magnetic
field
Boltzmann distribution
 Quantum mechanics tells us that, for net absorption of radiation to
occur, there must be more particles in the lower-energy state than in the
higher one.
 If no net absorption is possible, a condition called saturation.
 When it’s saturated, Boltzmann distribution comes to rescue:
Pm=-1/2 / Pm=+1/2 = e -DE/kT
where P is the fraction of the particle population in each state,
T is the absolute temperature,
k is Boltzmann constant 1.381*10-28 JK-1
Anything that increases the population difference will give rise to a
more intense NMR signal.
Nuclear Magnetic Resonance Spectrometer
How to generate signals?
B0: magnet
B1: applied small energy
Rf Energy Can Be Absorbed
•Precessing nuclei
generates an oscillating
electric field of the same
frequency
•Rf energy with the
same frequency as
the Larmor frequency
can be applied to the
system and absorbed
by the nuclei
 The Nucleus in a Magnetic Field
Precession and the Larmor frequency
• The magnetic moment of a spinning nucleus processes with a
characteristic angular frequency called the Larmor frequency w, which is a
function of r and B0
Remember  =  I (h/2) ?
Angular momentum dJ/dt=  x B0
Larmor frequency w=rB0
Linear precession frequency v=w/2p= rB0/2p
J
 When the Nucleus Meet the Magnet
Nuclear Magnetic Resonance
•For a particle to absorb a photon of electromagnetic radiation, the particle
must first be in some sort of uniform periodic motion
• If the particle “uniformly periodic moves” (i.e. precession)
at precession, and absorb erengy. The energy is E=hvprecession
v
•For I=1/2 nuclei in B0 field, the energy gap between two spin states:
DE=rhB0/2p
DE =hvphoton
• The radiation frequency must exactly match the precession frequency
Ephoton=hvprecession=hvphoton=DE=rhB0/2p
This is the so called “ Nuclear Magnetic RESONANCE”!!!!!!!!!
 Magnet B0 and irradiation energy B1
B0 ( the magnet of machine)
(1) Provide energy for the nuclei to spin
Ei=-miB0 (rh/2p)
Larmor frequency w=rB0
(2) Induce energy level separation (Boltzmann distribution)
The stronger the magnetic field B0, the greater separation
between
different
nuclei in the spectra
Dv =v1-v2=(r1-r2)B0/2p
(3) The nuclei in both spin states are randomly oriented around the
z axis.
M z=M, Mxy=0
( where M is the net
nuclear magnetization)
What happen before irradiation
• Before irradiation, the nuclei in both spin states are processing with
characteristic frequency, but they are completely out of phase, i.e.,
randomly oriented around the z axis. The net nuclear magnetization M is
aligned statically along the z axis (M=Mz, Mxy=0)
What happen during irradiation
When irradiation begins, all of the individual nuclear magnetic moments become
phase coherent, and this phase coherence forces the net magnetization vector M
to process around the z axis. As such, M has a component in the x, y plan,
Mxy=Msina. a is the tip angle which is determined by the power and duration of
the electromagnetic irradiation.
z
Mo
a
x
x
B1
wo
y
y
a deg pulse
Mxy
90 deg pulse
B1(the irradiation magnet, current induced)
(1) Induce energy for nuclei to absorb, but still spin at w or
vprecession
Ephoton=hvphoton=DE=rhB0/2p=hvprecession
And now, the spin jump to the higher energy ( from
m= –1/2
m=1/2m= – 1/2)
m= 1/2
(2) All of the individual nuclear magnetic moments become phase
coherent, and the net M process around the z axis at a angel
M z=Mcosa
Mxy=Msina.
What happen after irradiation ceases
•After irradiation ceases, not only do the population of the states revert to a
Boltzmann distribution, but also the individual nuclear magnetic moments
begin to lose their phase coherence and return to a random arrangement
around the z axis.
(NMR spectroscopy record this process!!)
•This process is called “relaxation process”
•There are two types of relaxation process : T1(spin-lattice relaxation) &
T2(spin-spin relaxation)
• Relaxation processes
•
How do nuclei in the higher energy state return to the lower state?
•
Emission of radiation is insignificant because the probability of reemission of photons varies with the cube of the frequency.
At radio frequencies, re-emission is negligible.
•
Ideally, the NMR spectroscopist would like relaxation rates to be fast - but not too
fast.
•
If the relaxation rate is fast, then saturation is reduced. If the relaxation rate is too
fast, line-broadening in the resultant NMR spectrum is observed.
•
There are two major relaxation processes;
•
Spin - lattice (longitudinal) relaxation
•
Spin - spin (transverse) relaxation
T1 (the spin lattice relaxation)
• How long after immersion in a external field does it take for a collection of
nuclei to reach Boltzmann distribution is controlled by T1, the spin lattice
relaxation time.
(major Boltzmann distribution effect)
•Lost of energy in system to surrounding (lattice) as heat
( release extra energy)
•It’s a time dependence exponential decay process of Mz components
dMz/dt=-(Mz-Mz,eq)/T1
T2 (the spin –spin relaxation)
•This process for nuclei begin to lose their phase coherence and return to
a random arrangement around the z axis is called spin-spin relaxation.
•The decay of Mxy is at a rate controlled by the spin-spin relaxation time T2.
dMx/dt=-Mx/T2
dMy/dt=-My/T2
dephasing
NMR Parameters
 Chemical Shift
•
The chemical shift of a nucleus is the difference between the resonance
frequency of the nucleus and a standard, relative to the standard. This
quantity is reported in ppm and given the symbol delta,

= (n - nREF) x106 / nREF
•
In NMR spectroscopy, this standard is often tetramethylsilane, Si(CH3)4,
abbreviated TMS, or 2,2-dimethyl-2-silapentane-5-sulfonate, DSS, in
biomolecular NMR.
•
The good thing is that since it is a relative scale, the d for a sample in a
100 MHz magnet (2.35 T) is the same as that obtained in a 600 MHz
magnet (14.1 T).
Alcohols, protons a
Deshielded
(low field)
Acids
Aldehydes
Aromatics
Amides
to ketones
Olefins
Aliphatic
Shielded
(up field)
ppm
15
10
7
5
2
0
TMS
The NMR scale (, ppm)
• We can use the frequency scale as it is. The problem is that
since Bloc is a lot smaller than Bo, the range is very small
(hundreds of Hz) and the absolute value is very big (MHz).
• We use a relative scale, and refer all signals in the spectrum
to the signal of a particular compound.
w - wref
=
ppm (parts per million)
wref
The good thing is that since it is a relative scale, the  in a
100 MHz magnet (2.35 T) is the same as that obtained for
the same sample in a 600 MHz magnet (14.1 T).
•
CH3
H 3C
Si
CH3
CH3
Tetramethyl silane (TMS) is used as
reference because it is soluble in most
organic solvents, inert, volatile, and has
12 equivalent 1Hs and 4 equivalent 13Cs:
Other references can be used, such as the residual solvent
peak, dioxane for 13C, etc. What reference we use is not
critical, because the instrument (software/hardware) is
calibrated internaly. Don’t use them if you don’t need to...
HO-CH2-CH3
w0=rBeffect
low
field
wo
high
field
•
Notice that the intensity of peak is proportional to the number of H
 J-coupling
•Nuclei which are close to one another could cause an influence on each
other's effective magnetic field. If the distance between non-equivalent
nuclei is less than or equal to three bond lengths, this effect is observable.
This is called spin-spin coupling or J coupling.
1
H
13
1
1
H
H
three-bond
C
one-bond
•Each spin now seems to has two energy ‘sub-levels’ depending on the state
of the spin it is coupled to:
J (Hz)
ab
I
S
bb
S
I
aa
ba
I
S
The magnitude of the separation is called coupling constant (J) and has units
of Hz.
•N neighboring spins: split into N + 1 lines
Single spin:
One neighboring spins: - CH – CH -
Two neighboring spins:
- CH2 – CH -
• From coupling constant (J) information, dihedral angles can be derived ( Karplus
equation)
3
J NHa  6.4 cos 2 (  60)  1.4 cos(  60)  1.9
3
J ab 1  9.5 cos 2 ( 1  120)  1.6 cos( 1  120)  1.8
3
J ab 2  9.5 cos 1  1.6 cos 1  1.8
χ2
χ1
2
N
Cγ
Cβ
Cα
ψΨ
N
ω
C’
 Nuclear Over Hauser Effect (NOE)
•The NOE is one of the ways in which the system (a certain spin) can release
energy. Therefore, it is profoundly related to relaxation processes. In
particular, the NOE is related to exchange of energy between two spins that
are not scalarly coupled (JIS = 0), but have dipolar coupling.
• The NOE is evidenced by enhancement of certain signals in the spectrum
when the equilibrium (or populations) of other nearby are altered. For a two
spin system, the energy diagram is as following:
bb
W1I
ab
W2I
W0IS
W1S
W1S
ba
S
W1I
aa
•W represents a transition probability, or the rate at which certain transition
can take place. For example, the system in equilibrium, there would be W1I
and W1S transitions, which represents single quantum transitions.
INSTRUMENTATION
1. MAGNET
Permanent magnets
Conventional electromagnets and
Super conducting magnets
2. SAMPLE PROBE
3. FIELD SWEEP GENARETOR
4. THE RADIO FREQUENCY SOURCE
5. THE SIGNAL DETECTOR&
RECORDER SYSTEM
The spectrometer
Preparation for NMR Experiment
1.
Sample preparation
Which buffer to choose? Isotopic labeling?
Best temperature?
Sample Position ?
N
2.
S
What’s the nucleus or prohead?
A nucleus with an even mass A and even charge Z  nuclear spin I is zero
Example: 12C, 16O, 32S  No NMR signal
A nucleus with an even mass A and odd charge Z  integer value I
Example: 2H, 10B, 14N  NMR detectable
A nucleus with odd mass A  I=n/2, where n is an odd integer
Example: 1H, 13C, 15N, 31P  NMR detectable
3. The best condition for NMR Spectrometer?
 Wobble : Tune & Match & Shimming
Tune
Match
RCVR
0%
Absorption
100%
4.
Frequency
What’s the goal?  Which type of experiment you need?
Different experiments will result in different useful information
5. NMR Data Processing
 The FID (free induction decay) is then Fourier transform to frequency domain
to obtain vpression ( chemical shift) for each different nuclei.
Time (sec)
frequency (Hz)
roperties of Some Deuterated NMR Solvents
Solvent
B.P. °C
Residual
1H signal (δ)
Residual
13C signal (δ)
acetone-d6
55.5
2.05 ppm
206 & 29.8
ppm
acetonitrile-d3
80.7
1.95 ppm
118 & 1.3 ppm
benzene-d6
79.1
7.16 ppm
128 ppm
chloroform-d
60.9
7.27 ppm
26.4 ppm
cyclohexane-d12
78.0
1.38 ppm
26.4 ppm
dichloromethane-d2
40.0
5.32 ppm
53.8 ppm
dimethylsulfoxide-d6
190
2.50 ppm
39.5 ppm
nitromethane-d3
100
4.33 ppm
62.8 ppm
pyridine-d5
114
7.19, 7.55 &
8.71 ppm
150, 135.5 &
123.5 ppm
tetrahydrofuran-d8
65.0
1.73 & 3.58
ppm
67.4 & 25.2
ppm
Limitations of nmr spectroscopy
1.Its lack of sensitivity. fairly large numbers are requried.minimum sample size
is about0.1ml having minimum concentrations of about on1%
2.Limited number of nuclei which may be usefully studied with this technique.
3.Inmost of the cases ,the technique is limited to liquid samples or to a liquid
capable of solutions in a suitable solvents or of melting at a temperature
below 260oc
4.In some compounds two different types of hydrogen atoms resonance at
similar resonance frequencies .this results in an overlap of spectra .hence
the interpretation of spectra becomes difficult.
APPLICATIONS
1.determination of optical purity
2.study of molecular interactions
3.quantative analysis: assay components, surfactant chain length
Determination, hydrogen analysis, iodine value, moisture
analysis
4.elemental analysis
5.Multicomponentmixture analysis
6.magnetic resonance imaging
7. NMR has also been used in various special fields that includes
industrial quality control, biology, engineering and medicine
8.Structure elucidation
•
•
other applications
Molecular conformation in solution
Quantitative analysis of mixtures containing known compounds
Determining the content and purity of a sample
Through space connectivity (over Hauser effect)
Chemical dynamics (Line shapes, relaxation phenomena)
Solid State NMR is widely popular for the characterization of polymers,
rubbers, ceramics, and molecular sieves.
Thank you