Embed Size (px)
Citation preview
PPM7.67.26.86.46.05.65.24.84.44.03.63.22.82.42.01.61.2
N
O
O
N
H
H
H
H
1
4
17
18
1011
12
13
14
20
21
23
227
8
9
1516
19
4 aromatic protons
Can we use 2D to assign this spectrum?
Structure first determined by Woodward in 1948 (without NMR!)
PPM (F2)5.0 4.0 3.0 2.0 PPM (F1)12011010090807060504030
N
O
O
N
H
H
H
H
1
4
17
18
1011
12
13
14
20
21
23
227
8
9
1516
19
CH2
Identify CH, CH2 and CH3 peaks on HSQC (use DEPT if necessary)
HSQC
PPM (F2)5.04.03.02.0 PPM (F1)5.65.24.84.44.03.63.22.82.42.01.61.2
N
O
O
N
H
H
H
H
1
4
17
18
1011
12
13
14
20
21
23
227
8
9
1516
19
22 23
H-22 is a good place to start
COSY
PPM (F2)5.0 4.0 3.0 2.0 PPM (F1)12011010090807060504030
N
O
O
N
H
H
H
H
1
4
17
18
1011
12
13
14
20
21
23
227
8
9
1516
19
Transfer this information to the HSQC spectrum
22
23
12 ?
H-12 can be tentatively identified from C and H shifts
HSQC
PPM (F2)3.63.22.82.42.01.6 PPM (F1)4.24.03.83.63.43.23.02.82.62.42.22.01.81.61.41.2
12 11 13
N
O
O
N
H
H
H
H
1
4
17
18
1011
12
13
14
20
21
23
227
8
9
1516
19
We can identify 11 as a CH2 and 13 as a CH from the HSQC
H-8 can distinguished from H-14 by the doublet multiplicity and shift
13-14 correlation is weak, but is seen on one side of the diagonal
138
13, 14
COSY
PPM (F2)5.0 4.0 3.0 2.0 PPM (F1)12011010090807060504030
N
O
O
N
H
H
H
H
1
4
17
18
1011
12
13
14
20
21
23
227
8
9
1516
19
22
2312
1113
8
Note that have a number of “dead ends” as far as COSY is concerned
14
HSQC
PPM (F2)2.82.42.01.6 PPM (F1)3.283.243.203.163.123.083.043.002.962.922.882.842.802.762.722.682.642.602.56
N
O
O
N
H
H
H
H
1
4
17
18
1011
12
13
14
20
21
23
227
8
9
1516
19
14
15 15 13
The protons at C-15 can be identified by their correlation to H-14
COSY
N
O
O
N
H
H
H
H
1
4
17
18
1011
12
13
14
20
21
23
227
8
9
1516
19
PPM (F2)3.63.22.82.42.01.61.2 PPM (F1)2.402.302.202.102.001.901.801.701.601.501.401.301.201.101.00
15
15
16
16
Having identified the C-15 protons, we can now see the correlation to H-16
gem
COSY
PPM (F2)5.0 4.0 3.0 2.0 PPM (F1)12011010090807060504030
N
O
O
N
H
H
H
H
1
4
17
18
1011
12
13
14
20
21
23
227
8
9
1516
19
22
2312
1113
8
14
15
This leaves 3 CH2s, at C-17, C-18 and C-20
16
HSQC
PPM (F2)3.6 3.4 3.2 3.0 2.8 PPM (F1)53.253.052.852.652.452.252.051.851.651.4
N
O
O
N
H
H
H
H
1
4
17
18
1011
12
13
14
20
21
23
227
8
9
1516
19
The C-20 protons can be identified as a pair of coupled (from COSY) doublets, not coupled to anything else
HSQC
N
O
O
N
H
H
H
H
1
4
17
18
1011
12
13
14
20
21
23
227
8
9
1516
19
PPM (F2)3.23.02.82.62.42.22.01.8 PPM (F1)2.202.162.122.082.042.001.961.921.881.841.801.761.721.681.641.60
The CH2s at C-17 and C-18 are left. The HSQC shows an overlapping CH2 at 1.94 ppm.
The chemical shift indicates that this must be the C-17 CH2.
The CH2 at 2.92 and 3.35 ppm must be C-18.
17(diagonal)
1818
COSY
PPM (F2)5.0 4.0 3.0 2.0 PPM (F1)12011010090807060504030
N
O
O
N
H
H
H
H
1
4
17
18
1011
12
13
14
20
21
23
227
8
9
1516
19
22
2312
1113
8
14
15
20
1718
16
All protonated carbons have now been assigned
HSQC
PPM (F2)4.03.83.63.43.23.02.82.6 PPM (F1)173.0172.0171.0170.0169.0168.0167.0166.0165.0164.0163.0
N
O
O
N
H
H
H
H
1
4
17
18
1011
12
13
14
20
21
23
227
8
9
1516
19
C-10
H-12H-11ax H-11eq
We can now use HMBC to assign the non-protonated carbons (and confirm other assignments, if necessary)
HMBC
PPM (F2)6.0 5.0 4.0 3.0 2.0 PPM (F1)
54.053.653.252.852.452.051.651.250.850.450.049.6N
O
O
N
H
H
H
H
1
4
17
18
1011
12
13
14
20
21
23
227
8
9
1516
19
C-7
H-1CH2-17
H-15
H-8
C-18
C-20
H-18
H-20 H-20
2 bond C-7 to H-16 correlation not observed?
HMBC
PPM (F2)3.63.22.82.42.01.61.2 PPM (F1)1.901.801.701.601.501.401.301.20
PPM (F2)3.63.22.82.42.01.61.2 PPM (F1)1.901.801.701.601.501.401.301.20
COSY TOCSY
17
13
15
12 118
16 15(diagonal) (diagonal)
14
N
O
O
N
H
H
H
H
1
4
17
18
1011
12
13
14
20
21
23
227
8
9
1516
19
PPM (F2)3.63.22.82.42.01.6 PPM (F1)4.24.03.83.63.43.23.02.82.62.42.22.01.81.61.41.2
1315
16
NOESY Avance 300Degassed sample
Degassing removes oxygen (paramagnetic) from the sample which would otherwise provide alternate relaxation pathways.
N
O
O
N
H
H
H
H
1
4
17
18
1011
12
13
14
20
21
23
227
8
9
1516
19
gem
1512
8
1315
15
8
12
NOESY
PPM (F2)3.63.22.82.42.01.61.2 PPM (F1)152.0151.0150.0149.0148.0147.0146.0145.0144.0143.0142.0141.0140.0
N
O
O
N
H
H
H
H
1
4
17
18
1011
12
13
14
20
21
23
227
8
9
1516
19
CH2-17 H-15
H-13H-11H-8
H-18
N-9
N-19
H-18H-16
15N-HMBC
15N spectroscopy is very difficult!
However, we can measure 15N shifts and determine the location of the nitrogens with HMBC (AMX 500, 10 mg sample, overnight)
