Thematic use of ribavirin as an example to illustrate
NMR principles and techniques
Brant L. Kedrowski and William F. Wacholtz, University of Wisconsin Oshkosh, Oshkosh, Wisconsin 54901
Abstract
Conformations and 1D NOE
HO
a
OH
b/c
e/f/g
e/f/g
*
a
HDO
e/f/g
*
*
+ D2O
*
b c
d e
h i j
g
f
k l H2O
a
ppm
k l H2O
Rotation about ribavirin’s amide bond can be studied at experimentally
accessible temperatures.
The effect of temperature on
chemical shifts of NH and OH
protons is also clearly visible (only
80 °C
NH and OH signals shift upfield
with increasing temps)
74 °C
Coalescence
Temperature = 74 °C
70 °C
O
N
H
N
60 °C
H
b/c
H
H
H
N
HO
N
O
40 °C
H
H
H
H
22 °C
HO
OH
b
c
H-Oxygen
.
h i j
Students can assign protons b, c, and e-g as OH or NH through exchange
with D2O.
H-Carbon
H-Nitrogen
g
f
Variable temperature NMR
The aromatic ring and multiple heteroatoms also make ribavirin a good
example for discussing deshielding effects and chemical shifts.
inequivalent
methylene protons
d e
Deuterium-proton exchange
Unique protons & chemical shift
inequivalent
amide protons
b c
b c
d e
g
f
5.9
5.8
5.7
5.6
5.5
5.4
5.3
5.2
5.1
5.0
4.9
4.8
4.7
4.6
4.5
4.4
4.3
4.2
4.1
4.0
3.9
3.8
3.7
3.6
3.5
3.4
Ribavirin serves as a good example to illustrate spin-spin splitting,
including inverted splitting tree analysis.
13C
Students can assign protons d-l definitively by selective decoupling.
(Millions)
FG
D E
C
AB
H
l
k
j
i
h
g
HETCOR example
f
e
c
11.9 Hz
11.9 Hz
5.4 Hz
5.4 Hz
4.3 Hz
5.4 Hz
g
Splitting in coupled
protons k and l
affected
j
H
Hl
ddd
dt
O
i h
N
150.0
140.0
130.0
120.0
110.0
100.0
90.0
80.0
70.0
60.0
50.0
H
N
H
HO
C
H
HO
triplet
e
f
g
h
i
j
N
H
N
O
H
N
k
l
d
e
f
g
h
i
j
k
l
160.0
150.0
160
150.0
X : parts per Million : 13C
140.0
130.0
F G
D E
OH
140
140.0
130.0
120.0
110.0
110.0
8.0
(Millions)
1) Unique protons
6) NOE & conformations
2) Chemical shifts
7) Variable temperature NMR
3) Spin-spin splitting
8) 13C NMR, DEPT
4) H-D exchange
9) Homonuclear 2D NMR
5) Selective decoupling
10) Heteronuclear 2D NMR
H
Acknowledgements
100.0
120 ppm 100
120.0
6.0
1. Williams, K. R.; King, R. W J. Chem. Educ. 1990, 67, A125-A137.
Proton decoupled
13C NMR
X : parts per Million : 13C
160.0
CH2
CH
A
B
4.0
Reference
H
H
HO
2.0
H
H
H
C
quartets
(approximate)
A/B
CH2
O
H
H
CH
N
0
Ribavirin is useful in teaching the following NMR topics:
CH3
CH
DEPT 135
C
doublets
d
H
d
H
e
OH
H
f
Hk
H
k/l
H
H
HO
N
b
Summary
O
Example:
Irradiate proton g
d
a
NMR, DEPT
Carbon NMR and DEPT techniques can be introduced with ribavirin
N
10.0
Heteronuclear 2D techniques introduced with ribavirin
160.0
Selective 1H,1H decoupling
0
X : parts per Million : 1H
X : parts per Million : 13C
Illustrating splitting and n+1 rule
3.3
2D Heteronuclear NMR
k l H2O
h i j
6.0
0
*
*
d
4.0
• Commercially available (~$40/100 mg, TCI America, Aldrich, Acros, etc.)
H
e
5.0
H
*
H
f
6.0
* H
g
20.0 40.0
N
O
• Interesting backstory (used clinically to treat serious viral infections)
N
j
i
h
7.0
• All multiplets are first order, making splitting of analysis straightforward
H
l
k
8.0
HO
H
COSY example
9.0
• All protons are unique and give well-resolved 1H NMR signals
N
H
Y : parts per Million : 1H
H
H
(Thousands)
• Asymmetric structure with four contiguous stereogenic centers
N
Y : parts per Million : 1H
Ribavirin
• Multiple functional groups with protons in diverse chemical environments
Ribavirin illustrates how asymmetry and restricted rotation can make
protons inequivalent.
O
Homonuclear 2D techniques introduced with ribavirin.
g
e
f
d
k l
i
j
h
6.0 5.9 5.8 5.7 5.6 5.5 5.4 5.3 5.2 5.1 5.0 4.9 4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 0
Ribavirin’s Properties Make it Useful in NMR Education
10.0
Conformations of ribavirin can be studied by NOE analysis.
(Millions)
The molecule ribavirin serves as an excellent example for illustrating a variety of NMR principles and instrumental
techniques. Ribavirin was used in a thematic fashion as a recurring example throughout a semester-long interpretative
spectroscopy course. The goal was to provide a familiar molecular framework in which to introduce new concepts and techniques
of increasing complexity. Proton NMR spectra were acquired at 270 MHz (carbon at 67.5 MHz) in DMSO-d6
2D Homonuclear
1
NMR
100.0
90.0
80.0
70.0
80
90.0
80.0
60.0
60
70.0
60.0
•
UW Oshkosh Chemistry Department
•
NSF-ILI Program (USE-9153034) for funds to purchase the JEOL
GSX-270 NMR spectrometer