dr tim claridge - modern nmr techniques for chemical structure elucidation
DESCRIPTION
NMR TechniquesTRANSCRIPT
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Modern NMR techniques for chemical structure
elucidation
-ethanol and beyond...
Dr. Tim Claridge
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Introduction
Historical overview
NMR Instrumentation
Introducing NMR spectroscopy
NMR techniques for structure elucidation
Undergraduate NMR course content
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What is NMR spectroscopy?
Nuclear- dealing with the property of nuclear spin Magnetic- interaction of nuclear spins with applied magnetic fields
Resonance Spectroscopy- excitation of these nuclear spins
Why do we use it?
Molecular structure
conformation
dynamics
interactions
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Nuclear Spin
+
spin
magnetic moment
precession B0
External magnetic
field
N
S
-
and Resonance
DE
a
b S
N External magnetic
field
Electromagnetic pulse of energy DE DE = hn
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History: the beginning
1946: Nuclear Magnetic Resonance
(Nuclear induction)
Purcell, Torrey and Pound- 1 kg Paraffin wax
Bloch, Hansen and Packard- 850 ml water
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Organic NMR spectroscopy
1951: First published high-resolution NMR spectrum:
Neat ethanol @ 30 MHz
HO-CH2-CH3
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And now
10 mg incubation product from antibiotic biosynthesis
pathway (700 MHz, cryogenic probe)
NH
HO2C
CO2H
Me
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NMR instrumentation
Magnet 400 MHz
robot
console
anti-vibration platform
preamplifier
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0
200
400
600
800
1000
1950 1960 1970 1980 1990 2000 2010
1H
Fre
qu
en
cy (
MH
z)
Year
20 MHz/year
Magnet development
11.7
14.1
16.4
18.8
21.1
8.5
7.0
B0 /Tesla
23.5
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Superconducting magnet
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NMR probeheads
Radiofrequency transmit & receive
coils
Tuning circuitry
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Cryogenic probeheads
Probe detection coils @ ~25K
NMR signal preamplifier @ ~ 70K
Sample @ ~ 300K
Thermal noise reduced significantly
Cold head
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Introducing NMR Spectroscopy
The electromagnetic spectrum
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8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 ppm
Hydrogen NMR spectrum (1H)
H2C
CH3
1) Chemical shift
2) Spin-coupling fine structure
3) Absorption intensities: Peak integration
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Introducing NMR Spectroscopy
Features of the NMR Spectrum
1. Chemical shifts
Tell us about the local chemical environments of each nucleus
2. Spin-spin couplings
Tell us about the interactions of each nucleus with its neighbouring nuclei
3. Peak intensities (integrals)
Tell us the relative number of nuclei in each chemical environment
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Chemical shifts reflect chemical environments: nuclei act as
undercover spies reporting on their surroundings
Feature I: Chemical Shifts (d)
9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 ppm
1H Spectrum
frequency
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The origin of chemical shifts
+
N External magnetic
field
Electron spins surround nucleus their own magnetic fields shield nucleus from external field ..differences in electronic shielding produce differences in chemical shifts
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Feature II: Spin Spin couplings (J)
A nucleus can sense the presence of its neighbours across bonds: nuclear spies again
Strength of interaction reflected in the spin-spin coupling constant, J (Hz)
1.21.31.41.51.61.71.81.92.02.12.22.32.42.52.62.72.82.9 ppm
H2C
CH3
Bond to HQ. Come in HQ Im next to three hydrogensout!
and Im next to two hydrogensout!
J
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Spin coupling patterns and multiplicities
doublet (d) double-
doublet (dd)
triplet (t)
triple doublet (td)
double triplet (dt)
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1.52.02.53.03.54.04.55.05.56.06.57.07.5 ppm
Feature III: Resonance intensities
Total resonance intensity reflects the number of nuclei in that environment: the number of spies present
Calculated by integration of peak areas
H2C
CH3
5 2 3
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NMR techniques
for chemical structure elucidation
NMR
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NMR in chemical research Hydrogen (1H) NMR spectra
O
H
BnO
H
BnO
H
H
OBnH
CH2
BnO
O
H
O
H
BnO
H
H
OBnH
CH2
STol
BnO
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Multi-pulse NMR- spin gymnastics
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Carbon NMR and editing 12C is NMR inactive, 13C is NMR active but only 1% abundant
Standard carbon spectra hide multiplicity information (eg CH vs CH2 vs
CH3)- simple stick appearance
Chemical shifts indicate chemical environments
Carbon editing experiments indicate protonation state
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Two-dimensional (2D) NMR Spectroscopy
Maps or correlates nuclear interactions within molecules:
Through-bond coupling (J)
Indicates presence of bonding connectivity
Through-space coupling (nOe)
Indicates close spatial proximity -> 3-dimensional shapes of molecules & stereochemistry
2D as two frequency axes
8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 ppm
1D
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2D contour presentation
Top-down view: Contour plot
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ppm
0.51.01.52.02.53.03.54.04.5 ppm
5
4
3
2
1
H2N CH C
CH
OH
O
OH
CH3
Threonine
coupled hydrogen
spin system
2D 1H-1H Correlation
Spectroscopy (COSY)
Maps spin-spin coupled partners
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2D COSY
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2D 1H-1H Total Correlation Spectroscopy (TOCSY)
Maps all spin-spin interactions within molecular fragments or even complete molecules
A B C D
F2
F1
COSY
A B C D
F2
F1
TOCSY
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2D TOCSY
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HH
H
H
A
B
C
D
12
3
Heteronuclear correlation methods: 1H-13C
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2D Heteronuclear correlation
13C
1H
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Biological chemistry: Peptides of 19 amino-acids
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Chemical biology: Protein of 110 amino acids
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1H-15N correlation: 110 amino acids Isotopically 15N labelled protein
1H
15N
Structure Drug binding Protein-protein interactions
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3D NMR methods
for protein structures: triple-resonance NMR 1H/13C/15N
HNCA
1H
15N
1H
15N
13C
N
C
C
N
H
H
C
C
O
O
N
C
C
N
H
H
C
C
O
O
N
C
C
N
H
H
C
C
O
O
HNCACO HNCO
N
C
C
N
H
H
C
C
O
O
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13C/15N labelled
Protein
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Medicine:
Magnetic Resonance Imaging
Functional MRI Unit, Oxford
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Undergraduate NMR spectroscopy
Often part of Organic Spectroscopy courses (+UV, IR, MS)
YEAR Material
2nd 1H & 13C chemical shifts, spin-spin couplings, resonance intensities
Origins of chemical shifts and couplings Correlating chemical shifts with structural environments Correlating spin-spin couplings with structural features
Influences on NMR spectra: stereochemistry, conformation, dynamics
Properties of other NMR active nuclei: 19F, 31P, 11B Instrumentation & Fourier transform NMR
3rd Through-space correlations: nuclear Overhauser effects
defining stereochemistry and 3D shapes of molecules
2D NMR methods
1H-1H through-bond correlations 1H-13C through-bond correlations 1H-1H through-space correlations
Physical behaviour of nuclear spins: relaxation
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Thank you for your attention