Principles and Selected Applications of Diffusion-Ordered NMR Spectros

Principles and selected applications of
Diffusion-Ordered NMR Spectroscopy
Stphane Viel, Ph. D.

Assistant Professor
Aix-Marseille University
Molecular Sciences Institute II (UMR-6263)
Chemometrics and Spectroscopy Laboratory
Marseilles (France)
DOSY ?
Diffusion Ordered NMR Spectroscopy
Number of publications
60
50
40
30
20
10
0
1992
Web of Science, 12 / 2007
1996
2000
Year
2004
2008
DOSY ?
Diffusion Ordered NMR Spectroscopy
Number of publications
60
50
40
30
20
10
0
1992
Web of Science, 12 / 2007
1996
2000
Year
2004
2008
NMR and Diffusion

1950
1954
1965
PGSE
Pulsed Gradient Spin Echo
NMR and Diffusion

1981
1987
1992
DOSY
Diffusion Ordered SpectroscopY
NMR and Diffusion
PGSE
1965
Pulsed Gradient Spin Echo
1992
DOSY
Diffusion Ordered SpectroscopY
General outline
Part 1: Theory about molecular mobility
Self-diffusion
Study of self-diffusion by NMR
Principles of Pulsed Gradient Spin Echo (PGSE)
Diffusion ordered NMR spectroscopy (DOSY)
Part 2: Selected applications of DOSY
Self-diffusion
Random translational motion of molecules
or ions that arises from the thermal energy
under conditions of thermodynamic
equilibrium
No thermal gradient (convection)
No concentration gradient (mutual diffusion)
Self-diffusion by Brown, 1828

Random jostling of molecules which leads
to their net displacement over time
s 2n Dt
time = t
s
time = 0
root mean square displacement
self-diffusion coefficient
9
Self-diffusion coefficient D
D is related to the hydrodynamic volume of
the diffusing particle through
D self-diffusion coefficient
kT
D
f
k Boltzmanns constant
T absolute temperature
f friction factor
10
Self-diffusion coefficient D
D is related to the hydrodynamic volume of
the diffusing particle through
D self-diffusion coefficient
kT
D
f
Sphere
k Boltzmanns constant
T absolute temperature
f friction factor
f 6a
11
Stokes Einstein equation

For a sphere diffusing in an isotropic and
continuous medium of viscosity :
Diffusion
kT
D
6a
Molecular Size 12

Pulsed Gradient Spin Echo (PGSE)
PGSE
Stejskal and Tanner, 1965
Gradients of magnetic field (Pulsed)
Gradient
Pulses
ON
OFF
ON
OFF
ON
OFF
OFF
Time
13

Principle: 2 steps
1. Spatially label the nuclear spins using
gradients of magnetic field.
2. Monitor their displacement by measuring
their spatial positions at 2 distinct times.
14
Larmor frequency
In NMR, each nuclear spin is identified by
its Larmor precession frequency 0
B0
0 B0
Nuclear magnetogyric ratio
15
Magnetic field gradient

Magnetic field
gradient
Spatially dependent
magnetic field
For a single and constant gradient

oriented along the z direction
g g ez
16
Magnetic field gradient

Magnetic field
gradient
Spatially dependent
magnetic field
For a single and constant gradient

oriented along the z direction
g g e ez
Notion of effective gradient
Coherence
order
ge p g
17
Phase shift of nuclear spins

Assume that the magnetic field
gradient is active during a time
A nuclear spin acquires a phase shift
( z ) ( B0 g e z )
Static Field
18
Phase shift of nuclear spins

( z ) ( B0 g e z )
Gradient
19
Phase shift
nuclear
spins
Nuclear
spinofspatial
labelling
( z ) ( B0 g e z )
The spatial position of the nuclear
spins is encoded into a phase shift
20
Rotating frame
In NMR, a common simplification
consists in describing the evolution of
the magnetization in a frame rotating at
the Larmor frequency 0
For nuclear spins on resonance, the
phase shift reduces to
( z ) g e z
21
Spin Echo or Hahn Echo (SE)

Without magnetic field gradients
Echo
Signal
22

With magnetic field gradients
coding
decoding
23

Echo
( z1 )
( z2 )
24

p=1
p ( g ) z1
p=-1
p ( g ) z2
25

Echo
g ( z1 26z2 )

Attenuation factor
g ( z1 27z2 )
Attenuation factor
I echo
exp D q
I0
3
Iecho: Intensity at the echo with gradients

I0: Intensity at the echo without gradients
D: Self-diffusion coefficient
: gradient pulse duration
: Diffusion time
q g
q: gradient pulse area
28
How do we actually obtain D?
exp D g
3
Normalizedfactor
intensity
Attenuation

1,0
0,8
0,6
0,4
0,2
0,0
0
10
20
Gradient strength g (G.cm-1)
30
29
How do we actually obtain D?
exp D g
3
Normalizedfactor
intensity
Attenuation

1,0
0,8
F
I
T
0,6
0,4
0,2
0,0
0
10
20
Gradient strength g (G.cm-1)
30
30
Stimulated Echo (STE)

coding
decoding
31
BPP-STE-LED sequence
Stimulated Echo (STE) with Bipolar gradient (BPP)
pulses and longitudinal eddy current delay (LED)
32
The BPP-STE-LED sequence

Stimulated Echo (STE):
T1 relaxation vs. T2 relaxation
No artefacts due to J modulation
Bipolar gradient pulses (BPP):

Reduced eddy currents
Longitudinal Eddy currents Delay (LED):

Less spectral distortions due to eddy currents
33

34



35

36



37

Echo Signal
38
Squence
BPP-STE-LED
39
How can we use PGSE data?

A
SIZE
DA > DC > DB
NMR spectrum (frequency scale, ppm)

DA DB
DA
DC
DC
ppm
40
SIZE
NMR spectrum (ppm scale)

DA DB
DA
DA > DC > DB
James & McDonald, 1978

Stilbs & Moseley, 1978-80
DC
DC
ppm
S
I
Z
E
41
Size Resolved Spectrometry

B
SIZE
NMR spectrum (ppm scale)
ppm
DA > DC > DB
Stilbs, 1981
42
ppm
Low
DB
DC
DA
43
High
ppm A
DOSY
Low
DB
DC
DA
44
High
DOSY
Diffusion Ordered NMR SpectroscopY
Morris & Johnson, 1992
Antalek, B. Concepts in Magn. Reson 2002, 14, 225-258

45
DOSY
Signal processing
Many processings
available:
- MaxEnt (Delsuc, M. A.)
- DECRA (Antalek, B.)
- CORE (Stilbs, P.)
- MCR (van Gorkom, L. C. M.)
- MULVADO (Huo, R.)
46
DOSY
Signal processing
Many processings
available:
- MaxEnt (Delsuc, M. A.)
- DECRA (Antalek, B.)
- CORE (Stilbs, P.)
- MCR (van Gorkom, L. C. M.)
- MULVADO (Huo, R.)
47
DOSY map
48
Adapted from Nilsson et al.
Distortions due to spectral overlap
49
Adapted from Nilsson et al.
iRRT
inverse
Regularized
Resolvent
Transform
V. Mandelshtam
A. J. Shaka
Mixture of 2
isomers
Armstrong, G. S.; Loening, N. M.; Curtis, J. E.; Shaka, A. J.; Mandelshtam, V. A., J. Magn. Reson. 2003, 163, 139
50
Thureau, P.; Thvand, A.; Ancian, B.; Escavabaja, P.; Armstrong, G. S.; Mandelshtam, V. A., ChemPhysChem
2005, 6, 1
General outline
Part 1: Theory about molecular mobility
Self-diffusion
Principles of Pulsed Gradient Spin Echo (PGSE)
Diffusion ordered NMR spectroscopy (DOSY)
Part 2: Selected applications of DOSY

51
Chiral recognition
Chiral recognition of dipeptides in a
biomembrane model
C. Bombelli, S. Borocci, F. Lupi, G. Mancini, L. Mannina,

A. L. Segre, S. Viel
52
J. Am. Chem. Soc. 2004, 126, 13354-13362
Introduction
The organization of biomembranes is based
on molecular recognition phenomena (chiral
recognition)
To investigate the non covalent interactions
involved in such systems, models are used
-
CO2 Na
Here
O
C11H23
CO2 Na
H
C11H23
O
we used Sodium N-doceanoyl-L-prolinate (SDP)

C. Bombelli, S. Borocci, F. Lupi, G. Mancini, L. Mannina, A. L. Segre, S. Viel
J. Am. Chem. Soc. 2004, 126, 13354-13362
53
Introduction (2)
We studied by NMR the chiral recognition in
SDP micelles of 2 dipeptides
5
Ditryptophan (1)
4
3a
7a
H
CO2
+
NH3
N '
H
'
2'
Diphenylalanine (2)
7'a 7'
3'a
4'
6'
5'
CO2
+
NH3
N
H
NMR techniques: 1H, PGSE, ROESY

+Molecular mechanic calculations
J. Am. Chem. Soc. 2004, 126, 13354-13362
54
H experiments: LL/DD couple
Ditryptophan (1)
+SDP micelles
Diphenylalanine (2)
+SDP micelles
J. Am. Chem. Soc. 2004, 126, 13354-13362
55
H experiments: LD/DL couple
Ditryptophan (1)
+SDP micelles
Diphenylalanine (2)
+SDP micelles
J. Am. Chem. Soc. 2004, 126, 13354-13362
56
PGSE experiments
Monitor the D values of the dipeptides by
PGSE experiments
2-site model: dipeptide in equilibrium
between the bound (b) and free (f) phase
D
Free
State
SDP
Bound
State

J. Am. Chem. Soc. 2004, 126, 13354-13362
57
PGSE experiments
Monitor the D values of the dipeptides by
PGSE experiments
2-site model: dipeptide in equilibrium
between the bound (b) and free (f) phase
D
SDP
Dobs xb Db (1 xb ) D f
J. Am. Chem. Soc. 2004, 126, 13354-13362
58
PGSE experiments
Determine the partition coefficient of the
dipeptides in the 2 phases
DP micellar
p
DP aqueous
Vaqueous
xb
p
1 xb Vmicellar
J. Am. Chem. Soc. 2004, 126, 13354-13362
59
PGSE experiments
Bound molar fractions xb and partition coefficients p
Dipeptide:
Xb
LD-1
0.981 0.004
1900 407
DL-1
0.981 0.004
1900 407
DD-1
0.959 0.004
860 87
LL-1
0.962 0.004
931 100
LD-2
0.79 0.03
138 25
DL-2
0.79 0.03
138 25
DD-2
0.69 0.03
82 12
LL-2
0.69 0.03
82 12

J. Am. Chem. Soc. 2004, 126, 13354-13362
60
PGSE experiments
Dipeptide:
Xb
LD-1
0.981 0.004
1900 407
DL-1
0.981 0.004
1900 407
DD-1
0.959 0.004
860 87
LL-1
0.962 0.004
931 100
LD-2
0.79 0.03
138 25
DL-2
0.79 0.03
138 25
DD-2
0.69 0.03
82 12
LL-2
0.69 0.03
82 12

J. Am. Chem. Soc. 2004, 126, 13354-13362
61
PGSE experiments
Dipeptide:
Xb
LD-1
0.981 0.004
1900 407
DL-1
0.981 0.004
1900 407
DD-1
0.959 0.004
860 87
LL-1
0.962 0.004
931 100
LD-2
0.79 0.03
138 25
DL-2
0.79 0.03
138 25
DD-2
0.69 0.03
82 12
LL-2
0.69 0.03
82 12

J. Am. Chem. Soc. 2004, 126, 13354-13362
62
Conformations of 1 isomers by NMR and

Molecular mechanic calculations (1)
Buffer
LL-1 + Buffer
DL-1 + Buffer

J. Am. Chem. Soc. 2004, 126, 13354-13362
63

SDP micelles (LL/DD couple)
LL-1 + SDP micelles
DD-1 + SDP micelles

J. Am. Chem. Soc. 2004, 126, 13354-13362
64

SDP micelles (DL/LD couple)
DL-1 + SDP micelles
LD-1 + SDP micelles

J. Am. Chem. Soc. 2004, 126, 13354-13362
65
Binding modes of 1 isomers

to SDP micelles
LL/DD
couple

J. Am. Chem. Soc. 2004, 126, 13354-13362
66
Binding modes of 1 isomers

to SDP micelles
LD/DL
couple

J. Am. Chem. Soc. 2004, 126, 13354-13362
67
Chemical exchange
Determining chemical exchange rates in
nucleobases
P. Thureau, B. Ancian, S. Viel, A. Thvand

Chem. Comm. 2006, 200-202
P. Thureau, B. Ancian, S. Viel, A. Thvand
Chem. Comm. 2006, 1884-1886
68
Hydrogen bonding in nucleic acids

Thymine Adenine
DNA
Uracil Adenine
RNA
P. Thureau, B. Ancian, S. Viel, A. Thvand Chem. Comm. 2006, 200-202

69
Effect of chemical exchange in DOSY

Uridine
HO
HO
O
O
N
OH
N
O
H2O
70
Model
Simple 2-site exchange N-H +H O
2
T = 50 ms
T = 200 ms

HOH+N-H
T= 900 ms
71
Model
2
T = 50 ms
T = 200 ms

HOH+N-H
T= 900 ms
72
Model
2
T = 50 ms
T = 200 ms

HOH+N-H
T= 900 ms
73
Uracil exchange constants Ka

2
HOH+N-H
H1 ka= 8 s-1
H3 ka= 18 s-1

74
Thymine exchange constants Ka

2
HOH+N-H
H1 ka= 5 s-1
H3 ka= 7 s-1

75
Self-aggregation
Investigations of complexes in
solution
S. Viel, L. Mannina, A. L. Segre

Tetrahedron Lett. 2002, 43, 2515-2519
C. Sanna, C. La Mesa, L. Mannina, P. Stano, S. Viel, A. L. Segre
Langmuir 2006, 22, 6021-6031
76
Introduction
stacking interactions are important in
organic chemistry and for biological systems
Here we consider 2 types of organic molecules
bearing an aromatic ring and characterized by
a: - low molecular weight (< 400 Da)
- low H2O solubility
Studied by:
- NMR (1H, PGSE, NOESY)

- DLS
- Physicochemical measurements
S. Viel et al. Tetrahedron Lett. 2002, 43, 2515-2519

C. Sanna et al. Langmuir 2006, 22, 6021-6031
77
Molecules under study

Name
X
METO
CH3
ACET
CH3
PRET CH2CH3
Y
CH(CH3)CH2OCH3
CH2OCH2CH3
CH2CH2OCH2CH2CH3
Class B
Name
X
1A
H
1B
CH3
1C
OCH3
1D
H
Class A
Y
Y
H
H
H
CH3
Z
H
H
H
CH3
O
O
NO2
X
z

78
H experiments
H spectra of dilute aqueous solutions of

METO,
METO ACET and PRET,
PRET (Conc < sol)
1
Monomeric resonances
7.4
7.3
7.2
7.1
7.0
6.9
ppm 1.2

1.1
1.0
0.9
ppm
79
H experiments

METO,
METO ACET and PRET,
PRET (Conc > sol)
1
Monomeric resonances
Extra resonances
7.4
7.3
7.2
7.1
7.0
6.9
ppm 1.2

1.1
1.0
0.9
ppm
80
H experiments

METO,
METO ACET and PRET,
PRET (Conc > sol)
1
Well resolved
Upfield shifted
7.4
7.3
7.2
7.1
7.0
6.9
ppm 1.2

1.1
1.0
0.9
ppm
81
PGSE experiments (DOSY display)

PGSE on a dilute
aqueous solution
of ACET
Log D
ppm
2 -1
(m s )
Aggregate
-10.5
Much lower
diffusion
coefficient
-10.0
-9.5
Monomer
-9.0
7

ppm
82
PGSE experiments
Hydrodynamic radii
(Stokes Einstein,
Sphere)
Conc
DNMR
(mM) (10-11 m2 s-1)
RH
(nm)
Monomer
ACET
METO
PRET
50
46
43
53
51
50
0.4
0.4
0.4
Aggregate
ACET
METO
PRET
50
46
43
1.7
1.3
1.9
11
15
10

83
NOESY experiments
NOESY spectrum
of a dilute aqueous
solution of ACET
400 ms
Color of cross peaks:
Blue : Negative
Green/Yellow : Positive
84
NOESY experiments
NOESY spectrum
of a dilute aqueous
solution of ACET
400 ms
Blue : Negative cross-peak
Green/Yellow : Positive crosspeak
Spin
Diffusion
85
NOESY experiments
NOESY spectrum
of a dilute aqueous
solution of ACET
10 ms
Blue : Negative
Green/Yellow : Positive
86
DLS experiments
Hydrodynamic radii
of the aggregates
were also estimated
by DLS
Conc
D
RH
(mM) (10-13 m2 s-1) (nm)
Aggregate
METO
13
PRET
3
ACET
2
2.8
7.8
6.5
700
250
300

ACET
PRET
METO
87
Physico-chemical properties
Surface
Tension
Osmotic
Coeff
Activity
Coeff
Rel. viscosity

88
Molecular weight
Diffusion-Ordered NMR Spectroscopy: a
versatile tool for the molecular weight
determination of uncharged
polysaccharides
S. Viel, D. Capitani, L. Mannina, A. L. Segre

Biomacromolecules 2003, 4, 1843-1847
89
Introduction
Polysaccharides constitute a major class of
biomacromolecules and play key roles in
biological recognition processes.
Their structural elucidation relies mainly on
NMR, but a complete characterization may also
require the molecular weight (MW).
Available techniques: Photonic Correlation
Spectroscopy, Gel Permeation Chromatography
Drawbacks: sample manipulation
90
Diffusion and Mass

Strictly, diffusion relates to molecular size. A
calibration is hence required to establish the
relationship between diffusion coefficient and
molecular weight
Pullulan (linear polysaccharide)
6 fractions (kDa): 5.8; 12; 28.3; 100; 180 and 853
Studied by PGSE experiments

91
Diffusion and Mass

5.8 kDa

100 kDa
853 kDa
92
Determination of Molecular Weight:

Pullulan as a Model Sample
1E-9
Pullulan
(m2/s)
1E-10
1E-11
100
1000
10000

100000
1000000
MW
(Da)
93

Calibration curve
1E-9
Pullulan
Calibration Curve
(m2/s)
1E-10
1E-11
100
1000
10000

100000
1000000
MW
(Da)
94

Check
with another polysaccharide
1E-9
Calibration Curve
Dextran
(m2/s)
1E-10
1E-11
100
1000
10000

100000
1000000
MW
(Da)
95

Check with oligosaccharides
1E-9
Calibration Curve
Dextran
Cyclodextrins - Oligosaccharide
(m2/s)
1E-10
1E-11
100
1000
10000

100000
1000000
MW
(Da)
96

Check with saccharides
1E-9
Calibration Curve
Dextran
Cyclodextrins - Oligosaccharide
Saccharides
(m2/s)
1E-10
1E-11
100
1000
10000

100000
1000000
MW
(Da)
97
Molecular Weight
Use of Pulsed Field Gradient Spin-Echo
NMR as a tool in MALDI method
development for polymer Mw
determination
M. Mazarin, S. Viel, B. Allard-Breton, A. Thvand, L. Charles

Anal. Chem. 2006, 78, 2758-2764
98
Polymers
M. Mazarin, S. Viel, B. Allard-Breton, A. Thvand, L. Charles

Anal. Chem. 2006, 78, 2758-2764
pMAM
99
Polymers PS
D0PS=f(Mw)
D=f[PS]
1e-8
CDCl3
1.4e-9
0.5412
2.721448e-8
1.2e-9
1e-9
D0 (m2.s-1)
2 -1
D (m .s )
1.0e-9
8.0e-10
6.0e-10
1e-10
4.0e-10
2.0e-10
0.0
0.0
1.0
2.0
3.0
4.0
-1
Concentration (mg.mL )
Mw 309
(D0 = 1.172e-9)
Mw 972
(D0 = 6.596e-10)
Mw 3460
(D0 = 3.405e-10)
Mw 9830
(D0 = 1.973e-10)
Mw 23800 (D0 = 1.189e-10)

Mw 74500 (D0 = 5.903e-11)
1e-11
10
100
1000
10000
Molecular weight Mw (Da)
D = k Mw -a
100000
1000000
PS : Comparison Mw : SEC, NMR and MS

PS standards
SEC (provider)
NMR
MALDI-TOF-MS
PS 400
309
334
434 10
-8.1
40.5
966
918 6
-0.6
-5.6
3278
3470 4
-5.2
0.3
8986
9375 10
-8.6
-4.6
22907
22890 49
-3.8
-3.8
83539
83416 203
12.1
12.0
(Sigma-Aldrich)
PS 1000
972
(Fluka)
PS 3000
3460
(Fluka)
PS 10000
9830
(Fluka)
PS 20000
23800
(Fluka)
PS 70000
(Fluka)
74500
101
Analysis of mixtures (part I)

Improved 3D DOSY-TOCSY experiment
for mixture analysis
S. Viel, S. Caldarelli
Chem. Comm. 2008, in press
102
Introduction
Overlapping signals severely complicate
DOSY analysis
A typical solution is the addition of another
frequency dimension to spread the signals out
Drawback: time consuming experiments due
to the requirement of sampling the indirect
frequency dimension
103
Speeding up 3D NMR
experiments
Various methodologies have been proposed
to speed up 3D NMR experiments (FDM)
104
Speeding up 3D NMR
experiments
to speed up 3D NMR experiments (FDM)
One possibility is Hadamard (there are other
ones ... ..3D iRRT would be great!)
105
Speeding up 3D NMR
experiments
to speed up 3D NMR experiments
One possibility is Hadamard (there are other
ones ... ..3D iRRT would be great!)
In Hadamard NMR spectroscopy, the
evolution time in the indirect dimension of the
2D block is replaced by phase-encoded
multisite selective excitation
106
Hadamard encoding
A B C D
Pulse 1
Hadamard family
matrices H
Matrix dimension N:
N = 2k (k = 1, 2, 3)
+
+
H
H
H
H
+
+
+
+
+
+
Pulse 2
Pulse 3
Pulse 4
M chemical sites M N
107
Hadamard encoding
A B C D
Pulse 1
Hadamard family
matrices H
Matrix dimension N:
N = 2k (k = 1, 2, 3)
+
+
H
H
H
H
+
+
+
+
+
+
Pulse 2
Pulse 3
Pulse 4
108
Hadamard encoding
A B C D
Pulse 1
Hadamard family
matrices H
Matrix dimension N:
N = 2k (k = 1, 2, 3)
+
+
H
H
H
H
+
+
+
+
+
+
Pulse 2
Pulse 3
Pulse 4
109
Hadamard encoding
A B C D
Pulse 1
Hadamard family
matrices H
Matrix dimension N:
N = 2k (k = 1, 2, 3)
+
+
H
H
H
H
+
+
+
+
+
+
Pulse 2
Pulse 3
Pulse 4
Signal B = + 1 2 + 3 4
110
Proposed pulse sequence
ZQC filters
Thrippleton, M. J.; Keeler, J., Angew. Chem. Int. Ed. 2003, 42, 3938-3941.
Cano, K. E.; Thrippleton, M.; Keeler, J.; Shaka, A. J., J. Magn. Reson. 2004, 167, 291-297.
111
Proof of principle (1)

TOCSY spectrum
of a mixture of:
- Methanol (M)
- Ethanol (E)
- Propanol (P)
- Valine (V)
112
Proof of principle (2)
113
Effect of signal overlapping

Propanol
OH
2-Butanol
OH
114
Effect of signal overlapping (2)

OH
OH
Time saving factor: 64
115
Analysis of mixtures (part II)

Enhanced diffusion-edited NMR
spectroscopy of mixtures using
chromatographic stationary phases
S. Viel, F. Ziarelli, S. Caldarelli

Proc. Natl. Acad. Sci. U. S. A. 2003, 100, 9696-9698
116
Introduction
PGSE experiments allow compounds to be
discriminated according to differences in their
effective size (mixture analysis)
Corollary: similar sized compounds CANNOT
be resolved by PGSE
Can we selectively
slow down the diffusion
of some components of
the mixture?
Chromatographic
phases

Proceedings of the National Academy of Sciences of the United States of America 2003, 100, 9696-
117
Principle
A chromatographic phase interacts selectively
with some of the mixture components (for
instance: polarity/apolarity)
Discrimination is achieved according to
apparent diffusion rates
(instead of free self-diffusion coefficients)

118
Problem: spectral resolution!

H of Sol. + Stationary phase
High Resolution
Conventional NMR
Magic Angle Spinning:
solid state technique
1

119
Problem: spectral resolution!

H of Sol. + Stationary phase
High Resolution
Conventional NMR
Magic Angle Spinning:
solid state technique
1
HRMAS NMR

120
HRMAS
HRMAS
probe
HRMAS rotor

121
Example 1
Mixture 1:
- Dichlorophenol
- Ethanol
- Heptane

122
Example 1
Mixture 1:
- Dichlorophenol
- Ethanol
- Heptane
+
SiO2
123
Example 2
Mixture 2:
- Naphtalene
- Dec-1-ene
- Ethanol

124
Example 2
Mixture 2:
- Naphtalene
- Dec-1-ene
- Ethanol
+
C18
125
Research directions
Improve resolution of complex mixtures
Characterize new chromatographic phases
Investigate chromatographic phenomenon
Discriminate stereoisomers

126
PFG MAS diffusion

measurements
Pulsed field gradient magic angle
spinning NMR self-diffusion
measurements in liquids
S. Viel, F. Ziarelli, G. Pags, C. Carrara, S. Caldarelli

J. Magn. Reson. 2008, 190, 113-123
127
Gradients and MAS probes
Courtesy of Bruker Instruments

J. Magn. Reson. 2008, 190, 113-123
128
Magic gradient

J. Magn. Reson. 2008, 190, 113-123
129
Magic gradient
Stator
Gradients

J. Magn. Reson. 2008, 190, 113-123
130
Gradient calibration: Profile

Hahn echo on a H2O/D2O sample with gradient
during acquisition
Adapted from Hurd et al.

J. Magn. Reson. 2008, 190, 113-123
131
Gradient calibration: Profile

6%
95%

J. Magn. Reson. 2008, 190, 113-123
132
Gradient calibration: strength
Rotor:

J. Magn. Reson. 2008, 190, 113-123
133
Gradient calibration: strength
Rotor:
V = 12 L
V = 50 L
G = 6.0 G cm-1 A-1

J. Magn. Reson. 2008, 190, 113-123
134
Effect of spinning
12 L
Water
ACN
Water
ACN

J. Magn. Reson. 2008, 190, 113-123
135
Effect of spinning
Water
ACN
Water
ACN
50 L

J. Magn. Reson. 2008, 190, 113-123
136
Results: ACN 4 kHz

12 L

J. Magn. Reson. 2008, 190, 113-123
50 L
137
Results

J. Magn. Reson. 2008, 190, 113-123
138
Results
PEO 116kDa D2O 4 kHz

J. Magn. Reson. 2008, 190, 113-123
PEO 116kDa CDCl3 3 kHz
139
Research directions
Improve resolution of complex mixtures
Characterize new chromatographic phases
Investigate chromatographic phenomenon
Discriminate stereoisomers

140
Mixture of:
- Benzene
- Naphthalene
- Anthracene
ODS phase
Silica gel
(ACN/H2O, 90/10)
HPLC
PFG MAS
G. Pags et al. Anal. Chem. 2006, 78, 561-566
G. Pags et al. Angew. Chem. Int. Ed. 2006, 45, 5950-5953
141
Merci
142
Grazie
143
Thank you !
A
144
A
145

Principles and Selected Applications of Diffusion-Ordered NMR Spectros

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Principles and Selected Applications of Diffusion-Ordered NMR Spectros

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Principles and selected applications of

Diffusion-Ordered NMR Spectroscopy

Stphane Viel, Ph. D.

Web of Science, 12 / 2007

Web of Science, 12 / 2007

NMR and Diffusion

NMR and Diffusion

NMR and Diffusion

Pulsed Gradient Spin Echo

Part 2: Selected applications of DOSY

Self-diffusion by Brown, 1828

root mean square displacement

Stokes Einstein equation

Study of self-diffusion by NMR

Study of self-diffusion by NMR

Magnetic field gradient

For a single and constant gradient

Magnetic field gradient

For a single and constant gradient

Notion of effective gradient

Phase shift of nuclear spins

Phase shift of nuclear spins

Spin Echo or Hahn Echo (SE)

Spin Echo or Hahn Echo (SE)

Spin Echo or Hahn Echo (SE)

Spin Echo or Hahn Echo (SE)

Spin Echo or Hahn Echo (SE)

Spin Echo or Hahn Echo (SE)

Iecho: Intensity at the echo with gradients

How do we actually obtain D?

Gradient strength g (G.cm-1)

How do we actually obtain D?

Gradient strength g (G.cm-1)

Stimulated Echo (STE)

The BPP-STE-LED sequence

Bipolar gradient pulses (BPP):

Longitudinal Eddy currents Delay (LED):

The BPP-STE-LED sequence

The BPP-STE-LED sequence

Bipolar gradient pulses (BPP):

Longitudinal Eddy currents Delay (LED):

The BPP-STE-LED sequence

The BPP-STE-LED sequence

Bipolar gradient pulses (BPP):

Longitudinal Eddy currents Delay (LED):

The BPP-STE-LED sequence

How can we use PGSE data?

NMR spectrum (frequency scale, ppm)

NMR spectrum (ppm scale)

James & McDonald, 1978

Size Resolved Spectrometry

NMR spectrum (ppm scale)

Antalek, B. Concepts in Magn. Reson 2002, 14, 225-258

Distortions due to spectral overlap

Part 2: Selected applications of DOSY

C. Bombelli, S. Borocci, F. Lupi, G. Mancini, L. Mannina,

we used Sodium N-doceanoyl-L-prolinate (SDP)

NMR techniques: 1H, PGSE, ROESY

H experiments: LL/DD couple

H experiments: LD/DL couple

C. Bombelli, S. Borocci, F. Lupi, G. Mancini, L. Mannina, A. L. Segre, S. Viel

C. Bombelli, S. Borocci, F. Lupi, G. Mancini, L. Mannina, A. L. Segre, S. Viel

C. Bombelli, S. Borocci, F. Lupi, G. Mancini, L. Mannina, A. L. Segre, S. Viel