NMR

Spectroscopy:

Introduction History of NMR

NMR Hardware and Software

Solution NMR

Sample Preparation
Presentation of Data

Important NMR Milestones

1938 - NMR in molecular beams

Rabi (Columbia University)
1946 - NMR of Liquids and Solids
Purcell, Torrey, Pound (Harvard)
Bloch, Hansen, Packard (CalTech)
1952 - First commercial NMR spectrometer
1962 - First Superconducting Magnet for
NMR
1968 - First Pulse Fourier Transform NMR
1969 - First Concept of MRI Scanners
1971 - First 2D NMR Experiment COSY
(Jean Jeener)
1985 - Protein Structures
2

NMR Nobel Prize

Winners

1944 Isador Rabi

1952 Felix Bloch
& Edwin Purcell
1991 Richard Ernst
2002 Kurt Wthrich
2003 Paul Lauterbur
& Sir Peter
Mansfield
3

From: Bruker SpinReport, Vol 153

Laukien Prize Winners

1999 Konstantin Pervushin, Roland Riek, Gerhard Wider, and Kurt Wthrich;
TROSY
2000 Lucio Frydman;
Quadrupolar MQMAS
2001 Peter Boesiger, Klaas Prmann, Markus Weiger;
Sensitivity-encoded magnetic
resonance imaging
2002 Ad Bax, Aksel Bothner-By and James Prestegard;
Residual dipolar couplings of weakly
aligned molecules in solution
2003 Jacob Schaefer;
REDOR Technique for Solid State NMR
2004 Lewis E. Kay,
NMR of Biological Macromolecules
2005 Stephan Grzesiek,
J couplings across hydrogen bonds
2006 Thomas Szyperski, Eriks Kupce, Ray Freeman, and Rafael Bruschweiler;
Acceleration of Multidimensional NMR
by novel procedures for scanning data space and efficiently processing results to obtain a
conventional spectral representation
2007 Robert G. Griffin;
High-field dynamic nuclear polarization (DNP) for sensitivity enhancement in
solid-state MAS NMR
2008 Malcom H. Levitt;
Optimized pulses and pulse sequences to enhance the power of liquid &
solid state NMR
2009 Daniel P. Weitekamp;
PASADENA and BOOMERANG
significantly improve NMR force detection by circumventing the problems of inhomogeneous
magnetic fields
2010 Paul T. Callahan;
Contributions to the study of polymeric and heterogeneous materials by advanced NMR exchange, diffusion
and relaxation techniques, and for his innovative q-space-diffusion-related developments that were relevant
in the context of the development of diffusion-tensor imaging.
2011 Daniel Rugar, John Mamin, and John Sidles;
Magnetic Resonance Force Microscopy
(MRFM).
4
2012 Klaes Golman and Jan Henrik Ardenkjaer-Larsen:
Dissolution-DNP NMR

Varian Prize Winners

2012 Ray Freeman and Weston A. Anderson

Nuclear Magnetic Double Resonance
2011 Gareth Alun Morris, The University of Manchester, UK
INEPT
2010 Martin Karplus, Harvard University, Cambridge, Massachusetts Karplus equations
2009 Albert W. Overhauser, Purdue University, West Lafayette, IN
NOE & Dynamic
Polarization
2008 Alexander Pines, UC Berkeley, and Lawrence Berkeley National Laboratory Cross
Polarization
2007 Alfred G. Redfield, Brandeis University, Waltham, Massachusetts Spin Dynamics
2006 John S. Waugh, MIT, Cambridge, Massachusetts
Average Hamiltonian Theory
(AHT)
2005 Nicolaas Bloembergen, University of Arizona, Tucson, Arizona Nuclear Magnetic
Relaxation
2004 Erwin L. Hahn, Professor Emeritus, University of California, Berkeley Spin Echoes
2002 Jean Jeener, Universite Libre de Bruxelles, Belgium
Two-dimensional NMR

NMR Spectroscopy
NUCLEAR
MAGNETIC
RESONANC
E
6

The NMR Spectrometer

Chapter 13

Schematic NMR Spectrometer

Fourier transformation and the NMR spectrum

RF Pulse

TheNMRspectrum

Fourier
transform

TheFouriertransform(FT)is
acomputationalmethodfor
analyzingthefrequencies
presentinanoscillatingsignal

Magic angle
(54.7)

NMR
Probes
Solids

Solids

Liquids

10

Liquids

NMR Signal
Generation
Spectrometer:

RF Generation:

90 180
Pulse (Sequences):
RD

Receiver:

DE

AQ

FT

11

NMR Samples
Types of NMR sample holders
Sample preparation
Spectrum quality

12

Types of NMR Sample

Holders
Solution NMR
Sample Tube
Spinners
Solid State
Sample Rotors

NMR Sample
Tubes with Caps

13

NMR Sample
Preparation
Tubes and Caps:

NMR tubes are a standard length (7 and 9 inch). When chipped

(and reduced in length) they should not be reused as an
unbalanced tube will not spin.

Always clean the tubes thoroughly after use.

First use the solvent you were using to recover your previous
sample,
then rinse several times with acetone and finally dry the sample
tube laying flat on a layer of kimwipes or placed upside-down on
a kimwipe in a beaker or Erlenmeyer flask. Choose the container
so the tubes stand vertically. Dont heat the tubes above 50 C, as
the glass might warp.
Always store unused, clean tubes uncapped and laying on a flat
surface.

Tube caps are disposable and replacements can be easily

obtained in bags of 100 ($5) or 1000 ($40 at www.wilmad.com).

Degassing Samples:

NMR spectra recorded using degassed solvents usually benefit

from reduced half-height line-width and thus better S/N. (O2 gas
is paramagnetic!)
14

There are several ways of degassing your sample:

NMR Sample
Preparation

Quantity:

For proton NMR spectra of small organic compounds (up to

MW=500) anything between 1 and 20 mg of sample will be fine.
Concentrated solutions can be viscous and may result in broad
signals.
Very dilute samples could be masked by impurities and solvent
peaks.

Carbon-13 is present at approximately 1.1 % natural

abundance.
It is intrinsically less sensitive than protons (approx. six
thousand times).
Please provide as much sample as possible, 50 - 100 mg (or
more) is fine.
Preparing two samples - one dilute sample for proton NMR and
one concentrated sample for carbon NMR is a useful, but
unnecessary practice.

Solvent height (volume) should be uniform, 5 cm or 2 inches

equal 0.5 ml.
The ends of the sample distort the field homogeneity, shimming
on each sample corrects this effect and takes just a minute or
so. However, vastly different solvent heights (volumes) prevent
complete correction and require many minutes shimming to
achieve acceptable homogeneity.
15

NMR Sample
Preparation
5 mm

Use clean + dry NMR tubes and caps

(tubes can be re-used, caps should not!)
0.5 ml deuterated solvent
(i.e. CDCl
3 ,C
6 6D , acetone-d
6 ,etc.)
substrate requirements for routine spectra:
10 mg for proton NMR
100 mg for carbon-13 NMR
min. filling height of tube: 2 inches (5 cm)
Cleaning of tubes:
1. rinse with solvent you were using
2. rinse with acetone
3. dry in (vacuum-)oven at low temperature

16

Clean
clear
solution

GOOD!

NMR Sample
Preparation
Suspension
or opaque
solution

Precipitate

Concentration
gradient

Two
phases

Bad Samples!

Not
enough
solvent

17

NMR Sample
Preparation

Shimming
improves the
magnetic field
homogeneity
If the magnetic field is not uniform within the sample, molecules in
different positions will experience different field strengths.
This will produce broad, distorted, or additional signals.18

Good and bad NMR

Spectra
are the result of:
Homogeneity of magnetic field
Sample preparation
Choice of solvent
Data acquisition parameters
Processing procedures

19

Good spectrum
ppm

20

ppm

Good spectrum
ppm

Peak picking

Integrals

ppm scale
21

ppm

Good spectrum
ppm

22

ppm

Bad spectrum ?

23

Bad spectrum !

No units specified for

axis and peak picking

Signal/Noise
ratio bad

24

Bad spectrum ?

25

Bad spectrum !

Tall signals
are cut off

26

Bad spectrum ?

27

Bad spectrum !

Signals too small

(only allowed when trying to
compare signal intensities between
different spectra)

28

Bad spectrum ?

29

Bad spectrum !

Broad signals

Possible reasons:
poor shimming
viscous sample
sample too concentrated
suspended particles in sample
excessive line broadening may
have been used during processing

30

Bad spectrum ?

31

Bad spectrum ?

32

Bad spectrum ?

Areas without signals should be excluded.

(If you want to print all your spectra with a
default range, i.e. 0-10 ppm, dont forget to
print detailed expansions.)

33

34

The nmr spectra included in this presentation have been

taken from the SDBS database with permission.
National Institute of Advanced Industrial Science and Te
chnology
(http://www.aist.go.jp/RIODB/SDBS/menu-e.html)

Nuclear Magnetic Resonance (nmr)

- the nuclei of some atoms spin:

H,

C,

13

- the nuclei of many atoms do not spin:

F,

19

H,

C,

12

O,

16

- moving charged particles generate a magnetic field ()

- when placed between the poles of a powerful magnet, spinning
nuclei will align with or against the applied field creating an
energy difference. Using a fixed radio frequency, the magnetic
field is changed until the E = EEM. When the energies match,
the nuclei can change spin states (resonate) and give off a
magnetic signal.
E

magnetic field = 14,092 gauss

for 1H v = 60,000,000 Hz (60 MHz)
nmr spectrum

intensity

magnetic field

10
0

chemical shift (ppm)

H nuclei are shielded by the magnetic field produced by the

surrounding electrons. The higher the electron density around
the nucleus, the higher the magnetic field required to cause
resonance.
1

CH3Cl
versus
CH4
lower electron
higher electron
density
density
resonate at lower
resonate at higher
applied field
applied field

Information from 1H-nmr spectra:

1. Number of signals: How many different types of
hydrogens in the molecule.
2. Position of signals (chemical shift): What types of
hydrogens.
3. Relative areas under signals (integration): How
many hydrogens of each type.
4. Splitting pattern: How many neighboring
hydrogens.

1. Number of signals: How many different types of

hydrogens in the molecule.
Magnetically equivalent hydrogens resonate at the same
applied field.
Magnetically equivalent hydrogens are also chemically
equivalent.
# of signals?

CH4

CH3CH3

number of signals?

H3C
H3C
one

C
C

CH3
CH3

one
CH3

one

CH3
two

CH3
H3C C CH3
Br
one

CH3CH2CH2-Br
three

CH3CH2-Br
two

CH3CHCH3
Cl
two

CH3CHCH2CH3
Br

Cl-CH2CH2CH2-Cl

two

four
CH3
CH2Cl

three

2. Position of signals (chemical shift): what types of

hydrogens.
primary
0.9 ppm
secondary 1.3
tertiary 1.5
aromatic
6-8.5
Note: combinations
allyl
1.7
may greatly influence
benzyl 2.2-3
chemical shifts. For
chlorides 3-4 H-C-Cl
example, the benzyl
bromides 2.5-4 H-C-Br
hydrogens in benzyl
iodides 2-4 H-C-I
alcohols
3.4-4 H-C-O
chloride are shifted to
alcohols
1-5.5 H-O(variable)lower field by the

chlorine and resonate

at 4.5 ppm.

reference compound = tetramethylsilane (CH 3)4Si @ 0.0 ppm

magnetic field

remember:

chemical shift
convention: let most upfield signal = a, next most upfield = b,
etc.

tms

toluene
CH3
b

chemical shifts

H3C
a
H3C

C
C

CH3
a
CH3

CH3 a
a

b
CH3

CH3
H3C C CH3
a
a
Br

CH3CH2CH2-Br

CH3CH2-Br

a b a
CH3CHCH3
Cl

b
d c a
CH3CHCH2CH3
Br

b a b
Cl-CH2CH2CH2-Cl

CH3

CH2Cl
b
c

3. Integration (relative areas under each signal): how

many hydrogens of each type.
a
b
c
CH3CH2CH2Br a
b
c

3H
2H
2H

a:b:c=3:2:2

a b a
CH3CHCH3
Cl

6H
1H

a:b=6:1

a
b

integration

H3C
a
H3C
a 12 H

C
C

CH3
a
CH3

a 12 H
CH3 a
a

a 6H

b
CH3
a 6H
b 4H

CH3
H3C C CH3
a
a
Br
a 9H
a

CH3CH2CH2-Br
a 3H
b 2H
c 2H

CH3CH2-Br
a 3H
b 2H
a b a
CH3CHCH3
Cl
a 6H
b 1H

b
d c a
CH3CHCH2CH3
Br
a
b
c
d

3H
3H
2H
1H

b a b
Cl-CH2CH2CH2-Cl
a 2H
b 4H

CH3

CH2Cl
b
c

a 3H
b 2H
c 4H

Integration: measure the height of each step in the

integration and then calculate the lowest whole number
ratio: a:b:c = 24 mm : 16 mm : 32 mm = 1.5 : 1.0 : 2.0
3H : 2H : 4H

If the formula is known ( C8H9OF ), add up all of the

steps and divide by the number of hydrogens = (24 + 16
+ 32 mm) / 9H = 8.0 mm / Hydrogen. a = 24 mm / 8.0
mm/H 3 H; b = 16 mm/8.0 mm/H 2H;
c
= 32 mm/8.0 mm/H 4H.

4. Splitting pattern: how many neighboring hydrogens.

In general, n-equivalent neighboring hydrogens will split a 1H
signal into an ( n + 1 ) Pascal pattern.
neighboring no more than three bonds away
n

n+1

Pascal pattern:

singlet

2
3
4

doublet
1

3
6

triplet
1

quartet

quintet

note: n must be equivalent neighboring hydrogens to

give rise to a Pascal splitting pattern. If the neighbors
are not equivalent, then you will see a complex pattern
(aka complex multiplet).
note: the alcohol hydrogen OH usually does not split
neighboring hydrogen signals nor is it split. Normally a
singlet of integration 1 between 1 5.5 ppm (variable).

splitting pattern?

H3C
a
H3C
a 12 H singlet

C
C

CH3
a
CH3

a 12 H singlet
CH3 a

a
a 6 H singlet

b
CH3

a 6 H singlet
b 4 H singlet

CH3
H3C C CH3
a
a
Br
a 9 H singlet
a

CH3CH2CH2-Br
a 3 H triplet
b 2 H complex
c 2 H triplet

CH3CH2-Br
a 3 H triplet
b 2 H quartet
a b a
CH3CHCH3
Cl
a 6 H doublet
b 1 H septet

b
d c a
CH3CHCH2CH3
Br
a
b
c
d

3H
3H
2H
1H

a 2 H quintet
b 4 H triplet

triplet
doublet
complex
complex

CH3

CH2Cl
b
c

b a b
Cl-CH2CH2CH2-Cl

a 3 H singlet
b 2 H singlet
c 4 H ~singlet

CH3CH2-OH
a 3 H triplet
b 2 H quartet
c 1 H singlet

Information from 1H-nmr spectra:

1. Number of signals: How many different types of
hydrogens in the molecule.
2. Position of signals (chemical shift): What types of
hydrogens.
3. Relative areas under signals (integration): How
many hydrogens of each type.
4. Splitting pattern: How many neighboring hydrogens.

cyclohexane

singlet

12H

2,3-dimethyl-2-butene
H3C
CH3
C C
H3C
CH3

a singlet
12H

benzene

a singlet
6H

p-xylene

H3C

CH3

a
b

a singlet
6H
b singlet
4H

tert-butyl bromide

CH3
H3C C CH3
Br

a singlet 9H

ethyl bromide
a
b
CH3CH2-Br
a
b

triplet 3H
quartet 2H

1-bromopropane

a
b
c
CH3CH2CH2-Br
a
b
c

triplet
3H
complex 2H
triplet
2H

isopropyl chloride

a
b a
CH3CHCH3
Cl
a
b

doublet 6H
septet
1H

2-bromobutane
b
d c
a
CH3CHCH2CH3
Br
a
b
c
d

triplet
doublet
complex
complex

3H
3H
2H
1H

o-methylbenzyl chloride
a
CH3

b
CH2Cl

a singlet 3H
b singlet 2H
c ~ singlet 4H

ethanol

a
c
b
CH3CH2-OH
a
b
c

triplet 3H
singlet 1H
quartet 2H

c
ethylbenzene

b a
CH2CH3

a triplet 3H
b quartet
2H
c ~singlet 5H

p-diethylbenzene
a

CH3CH2
a triplet 6H
b quartet 4H
c singlet 4H

CH2CH3

m-diethylbenzene

o-diethylbenzene

2-bromo-2-methylbutane

CH3
b CH3CCH2CH3 a
Br c
a triplet 3H
b singlet 6H
c quartet 2H
overlap

b&c

di-n-propylether

a
CH3CH2CH2-OCH2CH2CH3
a
b
c

triplet
6H
complex 4H
triplet
4H

1-propanol
a
b
d
c
CH3CH2CH2-OH
a
b
c
d

triplet
complex
singlet
triplet

3H
2H
1H
2H

C11H16

a 9H = 3CH3, no neighbors

9H

c 5H = monosubstituted
benzene
b 2H, no neighbors

CH3
CH2 C CH3
CH3
neopentylbenzene

5H

2H

C4H8Br2
a = 6H, two CH3 with no neighbors
6H

(CH3)2C
b = CH2, no neighbors & shifted
downfield due to Br

CH3
H3C C CH2
Br Br

2H

C7H8O
c = monosubst. benzene
5H

H2C

OH

b = CH2
c = OH

2H

1H

C4H9Br
a doublet 1.04 ppm
b complex 1.95 ppm
c doublet 3.33 ppm

6H
1H
2H

a = two equivalent CH3s with one neighboring H (b?)

c = CH2 with one neighbor H (also b)

CH3
CH3CHCH2Br
a
b c

6H doublet
b 1H complex
c 2H doublet

C10H13Cl
a singlet 1.57 ppm 6H
b singlet 3.07 ppm 2H
c singlet 7.27 ppm 5H

a = two-equilalent CH3s with no neighbors

c = monosubstituted benzene ring
b = CH2

b CH3
CH2 C CH3
Cl

a singlet 6H
b singlet 2H
c singlet 5H

13

C nmr

C ~ 1.1% of carbons

13

1) number of signals: how many

different types of carbons
2) splitting: number of hydrogens on the
carbon
3) chemical shift: hybridization of
carbon sp, sp2, sp3
4) chemical shift: evironment

13

C-nmr

2-bromobutane
a
c
d b
CH3CH2CHCH3
Br

mri
magnetic
resonance
imaging

This image is
copyrighted,
and used by
kind permission
of Joseph
Hornak for use
in Chem-312.

http://www.cis.rit.edu/htbooks/mri/