Download NMR Nuclear Magnetic Resonance Spectroscopy

NMR
Nuclear Magnetic
Resonance Spectroscopy
Evdoxia Coutouli-Argyropoulou
Professor of Organic Chemistry
Chemistry Department
Aristotle University of Thessaloniki
NMR is a spectroscopic technique, thus relies on the interaction
between material and electromagnetic radiation
The nuclear magnetic moment
Any motion of a charged particle has an associated magnetic field. This means,
that a magnetic dipole is created, just like an electrical current in a loop creates a
magnetic dipole,which corresponds to a magnetic moment µ.
According to the classical picture the atomic nucleus, assumed to be spherical,
rotates about an axis and thus, posses a nuclear or intrinsic angular momentum
P and because its charge shows also magnetic moment µ which is connected
with P with a proportionality constant γ known as gyromagnetic ratio
µ = γP
Quantum mechanical description
Quantum mechanical considerations show that, like many other atomic properties,
this angular momentum is quantized:
where ħ = h/2π, (h is Planck’s constant).
I is the nuclear quantum number that can have values I = 0, 1/2, 1, 3/2, 2,up
to 7.
If a nucleus with angular momentum P and magnetic moment µ is placed in a
static strong magnetic field B0, the angular momentum takes up an
orientation such that its component Pz along the direction of the field is an
integral or half-integral multiple of ħ:
where mI is the magnetic or directional quantum number with values mI
= I, I - 1,., -I, (2I + 1) different values of mI, and equal number of possible
orientations of the angular momentum and magnetic moment in an external
magnetic field B0.
For 1H and 13C nuclei, which have I = ½, there are two mI-values(+½ and .½)
Thus, if these nuclei are immersed in an external magnetic field, they can only
be regarded as being effectively lined up with the field (mI = +½) or against the
field (mI = -½). The energy difference ∆E between the states is:
∆E = γħB and since ∆E = hν
ν = |γ|Β/2π (Larmor equation)
To observe a nuclear magnetic absorption, we have to adjust either the
frequency ν of the radiation or the strength of the magnetic field at the
nucleus, B until Larmor equation holds, until the point where resonance
(energy absorption) occurs.
Larmor precession
The movement of a magnetic moment, µ, in a uniform externally applied strong
magnetic field Β0 under the condition of constant total energy traces out a cone
about Β0, which is analogous to the motion of a gyroscope running infrictionfree bearings under the influence of the Earth.s field Such motion isreferred to
in general as Larmor precession.
The precession frequency, ν0, is given
by the Larmor equation:
ν0 = |γ|Β0/2π
Effect of a second field B1. Resonance condition
Suppose there is an additional externally applied, but
weak magnetic field, B1,perpendicular to B0. Such a
field will also exert a torque on µ, tending to change
the angle θ between µ and B0. If B1 is fixed in
direction and magnitude, it will alternately try to
increase and decrease θ as µ precesses. Since B1 is
stated to be weak, the net effect will be a slight
wobbling in the precession of µ. If B1 is not fixed, but
is rotating about B0 with the same frequency as the
precession of µ, its orientation with respect to µ will
be constant and then the torque exerted on µ by B1
will always be away from B0. Consequently, an
accumulated effect on µ is possible. Since changing
θ corresponds to changing the energy of µ in B0,
this condition is described as resonance.The
frequency, ν, of the field B1 required must equal the
Larmor precession frequency. The energy for the
change of θ is derived from the rotating field B1,
which is supplied by radio frequency electromagnetic
radiation.
The nuclear quantum number
A particular nucleus is composed of p protons and n neutrons, its total mass is p
+ n, its total charge is +p and its total spin will be a vector combination of p + n
spins, each of magnitude 1/2. The atomic mass is usually specified for each
nucleus by writing it as a prefix to the nuclear symbol, e.g.12C indicates the
nucleus of carbon having an atomic mass of 12. This nucleus contains six
protons and six neutrons to make up a mass of 12. The nucleus 13C (an isotope
of carbon) has six protons and seven neutrons. Each nuclear isotope, being
composed of a different number of protons and neutrons will have its own total
spin value. Although the spin of a particular nucleus cannot be exactly predicted
the following empirical rules have been formulated:
(i) Nuclei with both p and n even have zero spin (e.g. 4He, 12C, 16O).
(ii) Nuclei with both p and n odd and mass even, have integral spin [e.g. 2H,
14N (I = 1), 10B (I = 3)].
(iii) Nuclei with odd mass have half-integral spins [e.g. 1H, 13C, 15N (I = 1/2),
17O (I = 5/2)].
Nuclei with I ≠ 0 are characterized magnetic and it is possible to give NMR
signals.
Properties of some magnetic nuclei
a magnetic
dipole moment in units of the nuclear magneton, eh/(4πΜp), where Mp is
the mass of the proton.
Frequencies for B=9.4 tesla
The macroscopic nature of NMR spectroscopy
Population of energy levels
Whenever we measure the spectrum of a molecule, we actually obtain the response
from a very large number of similar molecules. In a magnetic field the spin magnetic
moments orientate themselves in two directions with different populations N
according to the Boltzmann distribution.
Where Nα and Nβ are the numbers of particles in the lower and upper levels,
respectively, and T is the absolute temperature, k is the Boltzmann constant (k =
1.38066 × 10-23 JK-1) and ∆Ε the energy gap between the two levels.
For protons in a field B of 9.4 Tesla (1T =1 kg s-2 A-2), at a temperature of 300 K,
∆E/kT = 6.4 × 10-5 and Nα / Nβ = 3.2x10-5 or surplus population one part in
about 31000.
The net result of this population difference is an overall magnetization, M0, of the
sample in the direction of the z axis.
Proton the most popular NMR nucleus
In NMR spectroscopy where the upward transitions outnumber the
downward transitions by only one in 104-106, it is as if one detects only
one nucleus in every 104-106. It is therefore of crucial importance to
optimize signal strengths, for example by using strong external magnetic
fields B0 to maximize ∆Ε.
Similarly, nuclei with large gyromagnetic ratio and high natural
abundance are favoured, hence the popularity of 1H as an NMR nucleus.
The vast majority of molecules of interest to chemists contain hydrogen
atoms and, as this nucleus has one of the strongest resonances, it is not
surprising that 1H NMR has found the widest application.
Fourier Transform NMR
The screening factor- Chemical shift
The signal frequency that is detected in (NMR) spectroscopy is proportional to
the magnetic field applied to the nucleus. This would be a precisely
determined frequency if the only magnetic field acting on the nucleus was the
externally applied field. But the response of the atomic electrons to that
externally applied magnetic field is such that their motions produce a small
magnetic field at the nucleus which usually acts in opposition to the externally
applied field. The effective magnetic field at the nucleus can be expressed in
terms of the externally applied field B0 by the expression
where  is called the shielding factor or screening factor. The factor  is
small - typically 10-5 for protons and <10-3 for other nuclei.
This change in the effective field on the nuclear spin causes the NMR
signal frequency to shift. The magnitude of the shift depends upon the
type of nucleus and the details of the electron motion in the nearby
atoms and molecules. It is called a "chemical shift". The precision of
NMR spectroscopy allows this chemical shift to be measured, and the study
of chemical shifts has produced a large store of information about the
chemical bonds and the structure of molecules.
Chemical Shift ranges.
In practice the chemical shift is usually indicated by a symbol  which
is defined in terms of a standard reference.
1H
NMR spectra
The 1H-NMR spectrum of ethanol.
i)Identical nuclei, i.e. 1H nuclei, give rise to different absorption positions
when in different chemical surroundings.
(ii) The area (step-like curve is the integrated signal)of an absorption peak
is proportional to the number of equivalent nuclei (i.e.nuclei with the same
chemical shift).
(iii) Protons of CH3 and CH2 give rise to a triplet and a quartet, each with a
rather distinct intensity distribution. The splitting of resonances into
individual lines is called the fine structure of the spectrum.
The spin-spin coupling
The splitting of resonances into individual lines is called the fine structure of the
spectrum. It arises because each magnetic nucleus may contribute to the local field
experienced by the other nuclei and so modify their resonance frequencies. The strength
of the interaction is expressed in terms of the scalar coupling constant, J. Spin coupling
constants are independent of the strength of the applied field and they are expressed in
hertz (Hz). Spin-spin coupling is transmitted through chemical bonds and the coupling
constant, J, is a sensitive parameter for the types of bonds involved and for their spatial
orientation in the molecule.
Patterns of coupling
Pascal’s triangle.The intensity distribution of the A resonance of an AXn system can
be constructed by considering the splitting caused by 1, 2, . n nuclei.
13C
NMR spectra
Two-Dimensional NMR Spectra
2D 1H,13C-correlation
spectrum of a
neuraminic acid derivative.
Nuclear Relaxation
Application of external radiation at the correct frequency disturb the Bollzmann
equilibrium. At resonance, a net absorption will occur because there are more
nuclei in the lower energy state. After the application of the radiofrequency the
system tends to restore the Boltzmann equilibrium These restoring processes
are known as relaxation, and they effectively provide a continuous supply of
(excess) nuclei in the lower level of excitation.
It is convenient to consider two different relaxation mechanisms, each of which is
effectively a first-order rate process characterized by its own time constant: spinlattice relaxation, of time constant T1, occurs because there is exchange of energy
between the spin states and the surrounding medium; and spin-spin relaxation, of
time constant T2, occurs with exchange of energy between different nuclear spins.
The relaxation times scale is connected with the molecular mobility in materials
such as segmental motion in polymers and translational motion of small molecules in
liquids embedded in the pores of wood and stones. Slow motion correlates with
short and fast motion with high T2. Distributions of these parameters are obtained
by NMR signals acquired with suitable sequences of RF impulses (spin echo).
The NOE phemomenon
Nuclear Overhauser Effect
NOE: change in intensity of one resonance when the spin transitions of another
are perturbed from their equilibrium populations
NOE is observed for spin I when spin S is perturbed by a second radiofrquency.
The two spins should “communicate” through dipole-dipole interaction. Thus,
NOE is related to molecular motion. Small molecules exhibit positive NOEs.
Large molecules exhibit negative NOEs
Since the dipolar mechanism is a direct through-space interaction with an
inverse sixth-power dependence on the internuclear distance of the dipoledipole coupled nuclei, therefore, ΝΟΕ∝ 1/r6. Hence, the observation for of a
NOE for a certain pair of protons is a good indication that they are located close
to one another (within 5 Å) in the same or in different molecules.
Solid state NMR
Besides the indirect coupling through bonds, couplings can also be experienced
directly between the magnetic dipole moments of nuclei through space, which is
known as direct dipolar coupling, D. This interaction is characterized by the angle
between the internuclear vector and the external magnetic field (θ), and by the
coupling constant, bIS
1
DIS bIS (3cos2  1)
2
 0 I  S
bIS 
4 r 3 IS
In solutions the direct dipolar coupling as well as and the chemical shift anisotropy
are averaged to zero by the rapid motion of the molecules. In solids a similar
averaging effect can be produced by spinning the sample very rapidly about an
axis inclined with an angle of 54.74° with respect to the external magnetic field.
This is called the magic angle.
  arccos
1
 54.74
3
MAS (Magic Angle Spinning) technique
Strong homonuclear couplings that give rise to homogeneous line
broadening can only be effectively suppressed using spinning speeds
exceeding the magnitude of the coupling interactions. Otherwise the
spectrum will exhibit a pattern of side-bands, where the isotropic line is
surrounded by lines on both sides separated in frequency by the spinning
speed. Consequently, 1H is not commonly used directly. In contrast, 13C is
only 1% abundant and has a small magnetogyric ratio which results in weak
rarely occurring homonuclear interactions. Hence, no homogeneous line
broadening occurs and high resolution is possible even at modest spinning
rates.
MAS technique a routine method especially for 13C spectra.
NMR instrumentation
Unilateral Mobile NMR Sensors
A breakthrough for the NMR application to Cultural
Heritage
These sensors allow one to study arbitrarily sized objects non-invasively
by combining open magnets and surface RF coils to generate a sensitive
volume external to the sensor and inside the object under investigation,
but the price to be paid is the inhomogenity of the magnetic field. The
availability of this instrumentation makes it nowadays possible to
measure NMR parameters such as proton density, relaxation times,
diffusion coefficients. The most common data recorded are relaxation
measurements of 1H because the proton is the most sensitive NMR
nucleus, and relaxation can be measured despite the inhomogeneous
magnetic field that typically accompanies a simple magnet design.
Through NMR relaxation, the state of matter can be analyzed locally, and
the signal amplitude gives the proton density.
Usually, with these devices, to reach different depth of measurements
inside the investigated object, the tuning frequency must be electronically
switched. For instance, different probeheads each one tuned at the
proper frequency, are used to measure at selected depths
NMR-MOUSE (MObile Universal Surface Explorer
A well-known stray field sensor is the commercially available NMR-MOUSE
(mobile universal surface explorer), which is small and can readily be carried to an
object to be studied.
(a) Superconducting high-field NMR magnet for high-resolution NMR spectroscopy in the laboratory. The
sample is centered inside the magnet. (b) NMR-MOUSE (black) mounted on a computer-operated lift
(blue plate), compact spectrometer, and computer for measurement. The object to be measured is
placed on the black plate above the NMR-MOUSE. (c) Large version of the NMR MOUSE mounted on a
support for lateral displacement to measure profiles up to 25 mm depth into a wall. (d) Principle
components of the NMR-MOUSE and measurement arrangement.
NMR applications
Food chemistry
1H-
NMR spectra of the ethanolic extract of the plant Origanum vulgare
(Greek oregano).The arrows denote the presence of the antioxidant
compound rosmarinic acid.
Gerothanassis, I.P., Exarchou, V., Lagouri, V., Troganis, A., Tsimidou, M., & Boskou, D.
Journal of Agricultural and Food Chemistry, 46 (1998), 4185-4192.
Clinical applications
1H-NMR
spectra (400 MHz) of urine from a healthy subject and from
one patient suffering from Paraquat intoxication
Bairaktari, E., Katopodis, K., Siamopoulos, K. C., & Tsolas, O.,Clinical Chemistry, 44
(1998), 1256-1261.
Study of Macromolecular biomolecules
Fragment of a protein polypeptide chain. NOE is used as a tool for identifying the
aminoacid sequence as well as for determining internuclear distances in Å.
Wüthrich, K., Accounts in Chemical Research, 22 (1989), 36-44.
Wüthrich, K., Science, 243 (1989), 45-50.
NMR as a microscope
NMR microscopy of a fresh fruit (on the left) and of a frozen one (on
the right).Note that the two images were taken without the crosscut of
the fruit.
Gerothanassis, I.P., Troganis, A., Exarchou, V., Barbarossou, K. Chemistry Education:
Research and Practice in Europe, 3 (2002), 229-252.
Magnetic tomography
Magnetic Resonance Imaging (MRI): Anatomical view of the brain
Nuclear Magnetic Resonance to characterize
and monitor Cultural Heritage
•Liquid phase NMR
•Solid state NMR
•Mobile NMR
Characterization of oil paints
Linolenic acid
Linoleic acid
1H
Y = methyl of linolenic fatty chains; X = part of
the triplet of linoleic fatty chain; Z = methyl of
saturated and oleic fatty chains; S =
methylenic protons of saturated chains; E =
methylenic protons bonded to C3 of all fatty
chains; A = allylic protons bonded to C8 of
linolenic fatty chain and allylic protons of
linoleic chain; B = allylic protons of oleic fatty
chain; D = diallylic proton of linolenic fatty
chain;H = diallylic proton of linoleic fatty chain;
F = methylenic protons bonded to C2 of all
fatty chains; G = glycerol moiety; O = double
bonds.
By comparing the spectra, it was deduced
that there is a net reduction of
polyunsaturated fatty chains in the oil painting
extract.
NMR spectra at 600.13 MHz of (a)
linseed oil and (b) an oil painting extract
D.Capitani , V.Di Tullio, N., ProiettiProgress in Nuclear Magnetic Resonance
Spectroscopy 64 (2012) 29–69.
Application of NMR to the study of solid resins
Resins are produced by woody plants
on a worldwide basis. Resin fossilizes
over millions of years into a robust
material sometimes called amber.
Fossil resin can be characterized and
classified in groups by solid-state 13C
NMR spectroscopy.
Based on spectral distinctions, fossil
resin found in an archaeological
context sometimes can be assigned to
a specific geographical origin on the
basis of its 13C NMR spectrum
Amber: the Organic Gemstone
J.B. Lambert,J.O. Poinar Jr, Acc. Chem. Res., 35 (2002), 628-636.
13C
CPMAS NMR spectrum at 100.13 MHz of a sample of Baltic amber.
J.B. Lambert, J.S. Frye, Science 217 (1982), 55–57.
Characterization of Textiles
A study has been carried out
13C
by
CPMAS
NMR
spectroscopy on three silk
pieces from the 12th century
sampled from three coffinsin
north-eastern Japan The
analysis focused on the
carbonyl carbon resonances
in the region 170–174 ppm
assigned to C=O of the
peptide bonds in fibroin
molecules and ascribed to
the differences in constituent
amino acid residues, such as
alanine and glycine.
13C
CPMAS NMR spectrum at 100.13 MHz
of Bombyx Mori silk
R. Chϋiτ, A. Shimaoka, K. Nagaoka, A. Kurata, M. Inoue,, Polymer 37 (1996) 3693–3696.
Characterization of Paper
The 13C CPMAS spectrum may be
considered as the ‘‘fingerprint’’ of
the solid component of the paper. In,
the spectrum of Linters paper the
sharp resonances are due to the long
fibres of cellulose in crystalline
domains, while the broad ones are due
to cellulose in amorphous domains.
The Free Induction Decay (FID) of
paper shows always two components:
a fast decaying component from the
polymeric matrix, and a slow decaying
one from the confined water. After a
Fourier transformation, the cellulose
component appears as a broad hump
on top of which a rather sharp
resonance due to water is observed. A
best fit procedure may be applied to the
FID of a piece of paper to obtain the
Linters paper. (a) 13C CPMAS NMR spectrum at 50.13 MHz
along with resonance assignments. (b) Free Induction Decay,
the fast decaying component is from cellulose the slow
decaying one is due to water. (c) 1H proton wide line
spectrum, the broad line is from cellulose (line width at half
height about 64 kHz), and the sharp line is due to bound water
molar ratio between the water
component and the polymeric
component
D. Capitani, N. Proietti, F. Ziarelli, A.L. Segre, Macromolecules 35 (2002) 5536–5543.
Biodegradation of Paper
13C
CPMAS NMR spectra
at 50.13 MHz of, on the left,
uncoated paper before (0
days) and after 2, 5, and 10
days of enzymatic attack,
and on the right, spectra of
paper coated with poly
urethane before (0 days)
and after 2, 5, 10, 15, and
30 days of enzymatic
attack.
C. Boileau, S. Pessanha, C. Tardif, K. Castro, N. Proietti, D. Capitani, S. Vicini,
J.M. Madariaga, M.L. Carvahlo, E. Princi, J. Appl. Polym. Sci., 113 (2009) 2030–2040.
Depth profiling of paintings
Photo of “Adoration of the Magi”(1470) by
Perugino and positions of the measured depth
profiles.
Depth profiles at the two marked positions
revealing differences in the thickness of
the textile layer.
NMR-MOUSE was used to obtain two depth profiles in different regions of an
ancient oil painting, the ‘‘Adorazione dei Magi’’, by Pietro Vannucci ‘‘Il
Perugino’’, dated about 1470. The profiles revealed four different layers
ascribed to the wood, the incamottatura, the primer and the paint layer. It
was also possible to measure the thickness of each layer with an accuracy of
10–15 μm.
F. Presciutti, J. Perlo, F. Casanova, S. Gloggler, C. Miliani, B. Blόmich, B.G.
Brunetti, A. Sgamellotti, Appl. Phys. Lett. 93 (2008) 033505-1–033505-3.
Αging of paint layers
NMR can detect aging of paint layers over five centuries
Longitudinal relaxation weights of paint layers with oil and tempera
binders from several centuries. T1 shortens with increasing age
indicating indicating a trend toward a more a brittle texture of older
binder
B. Blumich, F. Casanova, J. Perlo, F. Presciutti, C. Anselmi, B. Doherty, Accounts of
Chemical Research, 43 (2010) 761-770.