High-Field NMR Experiments in the Upper-Level Laboratory Courses at Furman University

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High-Field NMR Experiments in the Upper-Level
Laboratory Courses at Furman University

• Tim Hanks, Moses Lee and
• Larry Trzupek
• Dept. of Chemistry, Furman University

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Current NMR Instrumentation

• Varian EM-360A (1981 31,000) CW H-1 only
• Varian VXR-300S (1988 329,000) FT H-1/C-13
  and multinuclear VT computer system updated to
  Sun SPARCstation 5 in 1996
• Varian Inova 500 (1996 454,000) FT
  indirect detection probe, H-1 N-15/P-31 VT
  Sun SPARC station 5 computer system.

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Calendar/Chemistry Curriculum at Furman
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Enantioselective Epoxidation (Hanks Chem 23)
NMR Techniques - H-1 NMR analysis of
fructose-based intermediates, ?-methylstyrene,
and epoxide product - C-13 NMR analysis of the
chiral catalyst and epoxide product - use of
chiral lanthanide shift reagent for determination
of enantiomeric purity
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Enantioselective Epoxidation (Hanks Chem 23)
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Enantioselective Epoxidation (Hanks Chem 23)
(33)
(86)
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Enantioselective Epoxidation (Hanks Chem 23)
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Enantioselective Epoxidation (Hanks Chem 23)
H-1 NMR analysis, trans-?-methylstyrene, vinyl
region
6.6 6.4
6.2
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Enantioselective Epoxidation (Hanks Chem 23)
C-13 NMR analysis, chiral oxirane precursor
180 140
100 60
20 ppm
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Enantioselective Epoxidation (Hanks Chem 23)
NMR determination of enantiomeric purity using
Eu(hfc)3
4.8 4.4 4.0
3.6 3.2 ppm
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Enantioselective Epoxidation (Hanks Chem 23)
NMR determination of enantiomeric purity results
Typical yield of epoxide product 60 Typical
enantiomeric excess 84ee Reference
Catalytic Asymmetric Epoxidation Using a
Fructose-Derived Catalyst Andy
Burke, Patrick Dillon, Kyle
Martin and Tim Hanks, J.
Chem. Ed., accepted for publication, 1999.
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Structure of a Tricyclic Compound (Lee Chem 23)
Features - Microscale preparation -
Multi-step reaction sequence - Use of basic 2-D
NMR to establish structure Initial Reactants
cyclopentadiene, maleic anhydride Target
Compound endo-9-methoxycarbonyl-3-oxatricyclo
4,2,1,0-2-nonane
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Structure of a Tricyclic Compound (Lee Chem 23)
(yield 40 - 70)
References W. J. Shepard, J. Chem. Ed., 40,
40-41 (1963) L. F. Fieser
and K. L. Williamson, Organic Experiments, 7th
ed., D. C. Heath, pp. 283-294 (1992)
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Structure of a Tricyclic Compound (Lee Chem 23)
COSY spectrum of tricyclic product
Reference The Microscale Synthesis and the
Structure Determination of Endo-9-methoxycar-
bonyl-3-oxatricyclo4,2,1,0-2-n
onane M. Lee, J. Chem. Ed., 69, A172-A173 (1992)
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Detailed NMR Characterization (Trzupek Chem 34)
Goals to develop - a basic understanding
of 2D NMR methods - the ability to carry out 2D
experiments independently - the ability to
process 2D data productively - a facility with
the interpretation of such data Requirements
assignment of - all H-1 resonances (chemical
shift, mulitplet pattern) - all C-13 resonances
(chemical shift) - all H-H coupling constant
values - all C-H coupling constant values
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Detailed NMR Characterization (Trzupek Chem 34)
NMR Techniques Available - simple H-1 spectrum
- resolution-enhanced H-1 spectrum - proton
decoupled H-1 spectrum - use of lanthanide shift
reagents - relay COSY - multiple quantum
filtered COSY - homonuclear 2D-J - simple
C-13 spectrum - heteronuclear 2D-J -
HETCOR -spectral simulation (H-1)
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Detailed NMR Characterization (Trzupek Chem 34)
Sample requirements - ready availability
(commercial or easily prepared) - good purity,
solubility - overlapping proton resonances -
complex splitting patterns - manageable
molecular size (5 to 8 types of Hs) Typical
candidates
(5-hexen-2-one)
(3,4-pentadien-1-ol)
(8-hydroxyquinoline)
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Detailed NMR Characterization (Trzupek Chem 34)
Results, 3,4-pentadien-1-ol COSY spectrum
HE HD HC
HB HA
E
D
C
B
D
A
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Detailed NMR Characterization (Trzupek Chem 34)
Results, 8-hydroxyquinoline
9
8
7 ppm
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Detailed NMR Characterization (Trzupek Chem 34)
Results, 8-hydroxyquinoline homonuclear 2D-J
HF
HE
HB
HD
HC
HA
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Detailed NMR Characterization (Trzupek Chem 34)
Results, 5-hexene-2-one heteronuclear 2D-J
Hd
Hf
He
c
a
e
d
g
f
(CDCl3)
Ha
Hc
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Detailed NMR Characterization (Trzupek Chem 34)
Results, 3,4-pentadien-1-ol H-1 simulation
HC
HA
D
E
C
B
D
A
HD
HE
HB
(simulated)
(actual)
5 4
3 2 ppm
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Detailed NMR Characterization (Trzupek Chem 34)
Results, 5-hexene-2-one HETCOR
Ce
Cf Cd
Cc
B
C
Ca
c
a
e
E
A
D
d
F
g
f
HC
HA,B
HE
HF
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Detailed NMR Characterization (Trzupek Chem 34)
Results, 5-hexene-2-one student report sheet
H chem shift multiplet J(x,y)
J(x,y) J(x,y) A 5.04
tdd AB, 2.2 AC, 17.0 AD,
1.1 B 4.97 tdd AB,
2.2 BC, 10.4 BD, 1.4 C 5.82
ddt AC, 17.0 BC, 10.4
CD, 6.6 D 2.33 tddd
BD, 1.4 CD, 6.6 DE, 7.4 E 2.55
t DE, 7.4 F
2.16 s C chem shift
J(H) a 114.9 160 (A,B) c
136.7 156 (C) d 27.5
122 (D,D) e 42.5 124
(E,E) f 29.7 128 (F,F,F) g 207.8
--- (all chemical shifts in ppm all J values in
Hz)
B
C
c
a
e
E
A
d
D
F
g
f
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3D structure of AZTMP by NMR (Lee Chem 44)
Background - bioactivation of AZT AZT
---1---gt AZTMP ---2----gt AZTDP ---3---gt AZTTP -
reaction rate of step 2 (thymidylate kinase) - v.
slow - consequence build-up of AZTMP
imbalance in the
nucleoside pool (the basis of AZT
toxicity) Goal - to determine if the solution
conformation of AZTMP is significantly
different from that of AMP and if that dif-
ference might be the basis for the sluggish
kinase reaction
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3D structure of AZTMP by NMR (Lee Chem 44)
NMR techniques employed - H-1 spectrum - P-31
spectrum - COSY analysis - homonuclear
decoupling (use of the above to assign proton
chemical shifts and ob- tain H-H coupling
constants throughout the molecule) -
determination of T1 relaxation time values for
each H - acquisition of NOE difference spectra
(use of the above to obtain non-bonded
distances between selected protons in the
molecule)
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3D structure of AZTMP by NMR (Lee Chem 44)
AZTMP H-1 spectrum in buffer (201 D2O/DMSO-D6)
(DMSO-d5)
5
1
4
3
2
(HOD)
T-H6
H1
T-CH3
8
6
4
2 ppm
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3D structure of AZTMP by NMR (Lee Chem 44)
Peak assignments COSY spectrum of AZTMP
H-1
5
1
4
3
2
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3D structure of AZTMP by NMR (Lee Chem 44)
J-values by homonuclear decoupling
5
1
H-2
4
3
H-2
2
H-2
H-2
Decoouple At H-3
gt
2.40 2.36 // 2.24
2.20 ppm
2.40 2.36 // 2.24 2.20 ppm
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3D structure of AZTMP by NMR (Lee Chem 44)
Dihedral angles from the Karplus relationship
H-H coupling J (Hz) ???degrees? 1-2
7.0 136 1-2 7.0 20
2-3 5.5 30 2-3 5.5
130 3-4 3.7 128
4-5 2.2 57 4-5 2.9
53 5-5 14.0 ---
5-P 6.1 --- 5-P 4.6
--
5
1
4
3
2
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3D structure of AZTMP by NMR (Lee Chem 44)
Additional conformational features from the
J-values
5
1
4
3
2
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3D structure of AZTMP by NMR (Lee Chem 44)
Inversion-recovery method for the determination
of T1s
5
1
4
3

2
0.9
( H-1)
0.7

0.5

0.3

0.1

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3D structure of AZTMP by NMR (Lee Chem 44)
Graphical use of inversion-recovery data to get
T1 values

0.9
( H-1)
0.7

0.5

0.3

0.1

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3D structure of AZTMP by NMR (Lee Chem 44)
Determination of the glycosidic torsional angle, ?
- obtain NOEs for irradiation at thymine H-6
- use known H-6/CH3 distance, NOE, and T1 to
obtain molecular correla- tion time, ?c - use
?c thus determined, other NOEs, and other T1s
to get other distances
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3D structure of AZTMP by NMR (Lee Chem 44)
Results solution-phase conformational structure
of AZTMP
Comparison to structure of TMP very similar
conclusion some other factor (steric bulk of
azido group) responsible for poor interaction
with the thymidylate kinase. M. Lee, J. Chem.
Ed., 73, 184-187 (1996)
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Acknowledgments
- National Science Foundation - Keck
Foundation - Milliken Foundation - Furman
Chemistry Alumni - Furman University