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EFFECT OF WETTING/DRYING ON THE
CONFORMATIONAL ARRANGEMENT OF A
HETEROGENEOUS ORGANIC MIXTURE AS ASSESSED
BY SOLID STATE 13C NMR SPECTROSCOPY
Università degliStudi di Palermo
P. Conte, A.E. Berns, H. Philipp, P. Burauel, H.-D. Narres, H. Vereecken
Forschungszentrum Jülich
Agrosphere
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1. Solid state NMR spectroscopy is a very powerful tool
to analyze insoluble natural organic matter (NOM)2. It is the only way to obtain quantitative information
on the chemical nature of NOM
3. It provides information on the chemical nature of
organic matter directly in bulk soils4. It can be used to study the interactions between
NOM and the inorganic moieties of soils as well asthe interactions with environmental pollutants
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Structural unit proposed by Flaig (1960)
Models of humic substances: the macromolecular structures
Structural unit proposed by Stevenson (1982)
Structural unit proposed by Schulten(1993)
Structural unit proposed by Stein(1997)
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Models of humic substances: the supramolecular structure
Weak dispersive forces are mainly involved in the conformational stabilization of NOM
Saccharides
Alkyl chains
Aromatic systems
Peptides
Cations
Are the molecular
properties of naturalorganic matter affected bythe experimental handling?
• proposed for the first time by Tschapek, Wasowski and Torres Sanchez, Plant and Soil 63,261-271 (1981)• resumed in Conte and Piccolo, Environmental Science & Technology 33, 1682-1690 (1999)• fully reviewed in Sutton and Sposito, Environmental Science & Technology 39, 9009-9015(2005)
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Analyses of the effect of sample preparation on
conformational behavior of some standard organic systems
CH3
CH3
CH3
CH3
CH3
CH3
O H
O
C H3
O HO
CMC
HMB
FERAC
SDS
CH3 O
S
O
O
O Na
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Analyses of the effect of sample preparation on
conformational behaviour of some standard organic systems
1. Solid state NMR of pure materials
2. Solid state NMR on dry mixture
3. Solid state NMR on mixture added with water and dried in desiccator (P2O5)
4. Solid state NMR on mixture added with water and freeze dried
Weight ratio 1:1:1:1
Molar ratio 1:52:77:93 for CMC, SDS, FERAC, and HMB, respectively
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EXPERIMENTAL SETUP: VCT EXPERIMENTS
-50050100150200250300
∗
−
−−∗
−∗
−∗=
−
CH
CH
CH
T
CT H T
T
H T
CT
H T
T I t I
)(1
exp1)(
exp)(
1)(1
1
1
1
0
ρ
ρ ρ
CT
I(t)
0 1000 2000 3000 4000 5000 6000 7000 8000
4
6
8
1012
14
16
18
20
22
24
26
28
30
32
34
36
38
40
I n t e n s i t y ( a . u . )
Contact time (µµµµs)
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EXPERIMENTAL SETUP: VCT EXPERIMENTS
-50050100150200250300
T CH : 1. fast local motions → high T CH values; 2. high amount of protons → low T CH values
T 1 ρ ρρ ρ (H) : proton spin lattice in the rotating frame; fast local motions →
shorterT 1 ρ (H) values. High proton concentration → faster spin diffusion with shorter T 1 ρ (H) values.
Notes : the proton concentration is more important than local molecular mobility in protonated organic systems
VCT experiments overestimate T 1 ρ ( H) values
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EXPERIMENTAL SETUP: VSL EXPERIMENTS
Pulse sequence for T 1 ρ ρρ ρ ( (( ( H) measurements
-50050100150200250300
I(t)
t (= delay)
−=
)(
exp)(1
0
H T
t I t I
ρ
0.000 0.005 0.010 0.015 0.020 0.025 0.030 0.035
-200
0
200
400
600
800
1000
1200
1400
I n t e n s i t y
( a . u
. )
delay (s)
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50100150
1 7 7
1 0 3 8
3
7 4
6 1
-COOH C1 C4
C2,3,5
C6
Carboxymethylcellulose sodium salt (CMC)
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-50050100150200250300
7 4
C2,3,5
Dry mixture
Water suspensionand dried indesiccator
Water suspensionand freeze Dried
Carboxymethylcellulose sodium salt (CMC)
Only the signal of
carbons 2, 3 and 5 isvisible in the spectra ofthe mixtures
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CMC
0
20
40
60
80
100
120
pure dry mix dried in desiccator freeze dried
T C H
( u s )
74 ppm
CMC – Results from VCT
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CMC
0
20
40
60
80
100
120
pure dry mix dried in desiccator freeze dried
T C H
( u s )
74 ppm
CMC – Results from VCT
CMC
2.4
2.5
2.6
2.7
2.8
2.9
3.0
3.1
3.2
pure dry mix dried in desiccator freeze dried
T 1 ρ ρρ ρ ( H ) (
m s )
74 ppm
CMC – Results from VSL for T 1ρρρρ(H)
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Possible arrangement of CMC in the different conditions
Pure CMC (water amount 11.5%)
CMC
2.4
2.5
2.6
2.7
2.8
2.9
3.0
3.1
3.2
pur e dri ed m ix aft er es sic at or aft er freez e dry ing
T 1 ∠ ∠∠ ∠ ( H ) ( m s )
74ppm
= components = water
Mixing of the dry components(total amount of water 3.3%)
Addition of H2Odesiccator
Hydrogens from residualwater (5.0%) and fromother close molecules
Freeze drying
Hydrogens from residualwater (2.4%) and fromother close molecules
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050100150
1 3 1
1 7
CH3
CH3
CH3
CH3
CH3
CH3
Aromatic C
Methyl C
1st order SSB2nd order SSB
Hexamethylbenzene (HMB)
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-50050100150200250300
CH3
CH3
CH3
CH3
CH3
CH3
HMB – NOT PRESENT
Aromatic C
Methyl C
Confirmation byHPLC measurements
Hexamethylbenzene (HMB)
Dry mixture
Water suspension
and dried indesiccator
Water suspensionand freeze Dried
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HMB
0
500
1000
1500
2000
2500
3000
131 ppm 17 ppm
T C H
( u s )
puredry
mix
dried
in des.
puredry
mix
dried
in des.
HMB – Results from VCT
HMB
0.0
20.0
40.0
60.0
80.0
100.0
120.0
140.0
160.0
131 ppm 17 ppm
T 1
( H )
( m s )
puredry
mix
dried
in des. pure drymix
driedin des.
HMB – Results from VSL for T 1ρρρρ(H)
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Possible arrangement of HMB in the different conditions
π−π interactions
HMB =Addition of other componentsto obtain mixture
HMB
H2O
Addition of water
HMB
desiccatorHMB
Entropy driven compactionof the HMB molecules
Freeze-dried
The π−π interactionsbetween the HMBmolecules are too weak
to prevent removing ofHMB from the mixture
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Ferulic Acid
OH
O
CH3
OHO
12
3
4
6
7
89
10 1=126 ppm
2=126 ppm
3=148 ppm
4=149 ppm
5=114 ppm
6=112 ppm
7=56 ppm
8=145 ppm
9=109 ppm
10=173 ppm
50100150
1 7 3
1 4 9
1 4 8
1 4 5
1 2 6
1 1 4
1 1 2
1 0 9
5 6
C10
C4 C3
C8C5
C1, C2 C6C7
C9
Peak assignment done with the aid of dipolar dephasing experiments
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Ferulic Acid
-50050100150200250300
OH
O
CH3
OHO
12
3
4
6
7
89
10 1=126 ppm
2=126 ppm3=148 ppm
4=149 ppm
5=114 ppm
6=112 ppm
7=56 ppm
8=145 ppm
9=109 ppm
10=173 ppm
Dry mixture
Water suspensionand dried indesiccator
Water suspensionand freeze Dried
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FERAC
0
100
200
300
400
500
600
700
173 ppm 149 ppm 148 ppm 145 ppm 126 ppm 114 ppm 112 ppm 109 ppm 56 ppm
T
C H
( u s )
pure
dried mix
after essicator
after freeze drying
Ferulic Acid – VCT results (T CH)
FERAC
250.0
300.0
350.0
400.0
450.0
500.0
550.0
600.0
650.0
173 ppm 149 ppm 148 ppm 145 ppm 126 ppm 114 ppm 112 ppm 109 ppm 56 ppm
T 1 r ( H )
pure
dried mix
after essicator
after freeze drying
FERAC – Results from VSL for T 1ρρρρ(H)
dry mix
after desiccator
pure
after freeze drying
dry mix
after desiccator
pure
after freeze drying T 1 ρ
( H ) ( m s )
H R2
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Possible arrangement ofFERAC in the differentconditions
OHO
OH
O
CH3
OHO
OH
O
CH3
H R1
H R2
H R3
H R1
H R2
H R3
HO
H
HO
H
HO
H
HO
H
HO
H
HO
H
HO
H
H R1
H R2
H R3
H
O
H
HO
H
OO
OH
O
CH3
HO
H
H
OH
HO
H
HO
H
OO
O
O
CH3
HO
H
H
OH
H
OH
H R1
H R2
H R3
HO
H
OO
OH
O
CH3
HO
H
H
OH
H
OH
addition of components
addition of water
drying in essicator
freeze-drying
slowly relaxing centers
quickly relaxing centers
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SDS
102030405060
5 2
3 3
2 5
1 5
C 1C 11 C 12
C 2-10
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SDS
-50050100150200250300
Dry mixture
Water suspensionand dried indesiccator
Water suspensionand freeze Dried
CH3 O
S
O
O
O Na
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SDS
0
50
100
150
200
250
300
52 ppm 33 ppm 25 ppm 15 ppm
T C H ( u s ) pure
dried mix
after essicator
after freeze drying
SDS – Results from VCT (T CH)
dry mix
after desiccator
pure
after freeze drying
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SDS
0
50
100
150
200
250
300
52 ppm 33 ppm 25 ppm 15 ppm
T C H ( u s ) pure
dried mix
after essicator
after freeze drying
SDS – Results from VCT (T CH)
dry mix
after desiccator
pure
after freeze dryingSDS
0.0
100.0
200.0
300.0
400.0
500.0
600.0
52 ppm 33 ppm 25 ppm 15 ppm
T 1
r ( H ) ( m s ) pure
dried mix
after essicator
after freeze drying
SDS – Results from VSL for T 1ρρρρ(H)
dry mix
after desiccator
pure
after freeze drying T
1 ρ ρρ ρ
( H ) ( m s )
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Possible arrangement of SDS in the different conditions
Water amount 1%
OS
O
O
ONa
HH
H
HH
HH
HH
HH
HH
HH
HH
HH
HH
HH
HH Dry mixing
= SDS = other components of the mixture = water
Increase of rigidity
Addition of waterdesiccator
Freeze drying
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CONCLUSIONS
Sample handling (wetting and drying) affects molecular properties ofthe organic components of a mixture
Molecular dynamics changes according to the closest proximity of themolecular systems following dissolution and drying at different conditions
Residual water also plays an important role in the relaxationmechanisms
The ππππ−−−−ππππ interactions in the aromatic HMB are not strong enough toprevent removal of such system from the mixture following the freezedrying under forced vacuum conditions
CARE MUST BE USED IN INTERPRETING MOLECULAR DYNAMICSRESULTS FROM NMR EXPERIMENTS ON COMPLEX ORGANICMIXTURES SUCH AS THOSE BELONGING TO EXTRACTED NATURALORGANIC MATTER
THANK YOU FOR YOUR
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THANK YOU FOR YOUR
ATTENTION
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