核磁共振基础
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第2章 核磁共振基础
NMR Spectroscopy
Chapter 2 Basic Principles of NMR
核磁共振波谱学
Nuclear Magnetic Resonance (NMR) Spectroscopy
第2章 核磁共振基础
2.1 核自旋和共振 2.2 核磁共振基本概念 2.2.1 驰豫 2.2.2 化学位移 2.2.3 自旋偶合 2.3 核磁共振实验 2.3.1 核磁共振仪 2.3.2 测定方法 2.3.3 核磁共振谱图
H1 H1 H3 H2 H2
d1
t1
Spin-lock
H
1
H
X X
3
H
1
H
3
X O X
X X
X
R1
R2 X H
2
H
4
O
H
2
H
4
HSQC - Heteronuclear Single Quantum Coherence
(correlates X-1H chemical shifts via one-bond 1J(XH) couplings)
R. R. Ernst
1991, Chemistry HR & 2D NMR
K. Wuethrich 2002, Chemistry NMR for Bio-Macromol.
P. Lauterbur
P. Mansfield
2003 Medicine MRI
2003年度诺贝尔生理学和医学奖获得者
核磁共振成像技术的发现，医学诊断和生物细胞研究领域的突破性成就。
NMR, An Evergreen
• • • • • • • • • • 1939 Rabi, Magnetic resonance for atomic nuclei (1944 Noble Prize) 1945 Bloch & Purcell, NMR phenomenon, (1952 Noble Prize) 1953 Varian Company, The first commercial NMR spectrometer 1957 13C NMR 1959 Solid state NMR 1964 High resolution NMR (Superconducting magnet) 1965 Pulse Fourier Transform NMR (FT-NMR) 1967 Chemically Induced Dynamic Nuclear Polarization (CIDNP) 1976 Ernst, Methodology in high resolution NMR (1991 Noble Prize) 1973,1982 Lauterbur & Mansfield, Hospital NMR Imaging (2003 Nobel Prize) • 1983, 1990 Wuethrich, NMR for bio-macromolecules (2002 Nobel Prize) • 1995 LC-NMR, LC-MS-NMR
Applications of NMR
Time-Resolved CIDNP
Yang, Gomberg 2000, 2000
Applications of NMR
NMR Imaging (MRI)
P. Lauterbur P. Mansfield 2003 Nobel Prize in Medicine
H
1
H
3
H4
X
X O X
X X
R1
H4
R2 X H
2
H
4
O
H
2
H
4
H3
TOCSY - TOtal Correlation SpectroscopY
(identifies protons belonging to the same scalar coupling network)
H3
90 Trim
H4 H4
13C
203.61 172.72 171.22 170.35 167.04 167.01 141.95 137.99 133.71 133.65 133.23 131.97 130.21 129.18 129.03 128.72 128.70 128.37 127.04 127.04
shifts (int)
84.41 81.19 79.05 76.52 75.57 74.98 73.20 72.41 72.18 58.65 55.04 45.64 43.19 35.72 35.63 26.88 22.63 21.83 20.84 14.83 9.57
(2) (2) (1) (2) (1) (2) (2) (2) (1) (1) (1) (1) (1) (1) (1) (1) (1) (1) (1) (1)
NMR Microscopy
Substructural elements
X-Ray UV IR MS NMR (+) + + + +
full connectivity
+ +? +
3D structure
+ +
The NMR strategy and the NMR tools:
-
number of protons, 1H chemical shifts, 1H-1H couplings ---> 1H spectrum number of magnetically nonequivalent carbons ---> 13C spectrum number of NMR-detectable other nuclei (19F, 31P, …, 15N?) ---> X spectrum 1H-13C direct correlation (one-bond) ---> HSQC, (C,H COSY, HMQC) identification if 1H spin systems ---> TOCSY 1H coupling network ---> COSY 1H-X (X=13C, 15N, 19F, 31P, etc) long-range correlation ---> HMBC( COLOC ) stereochemistry, 3D structure ---> NOESY, ROESY
COSY - COrrelation SpectroscopY
(correlates scalarly coupled protons)
The gradient DQF-COSY sequence:
1H
H3
H4H4
H1 H1 H3 H2 H2
H2
d1
t1
H2 H3
Grad.
H1 H1
H
1
H
X X
3
NMR, An Evergreen
Phys.
Biol.
NMR
Chem.
Med.
Nobel Prize Laureates related to NMR
I. I. Rabi 1944, Physics Resonance of nuclei
F. Bloch E. M. Purcell 1952, Physics Discovery of NMR
S. P. Mansfield (彼得－曼斯菲尔德), University of Nottingham, 英国
P. C. Lauterbur(保罗－劳特布尔), University of Illinois, 美国
Increase of Magnetic Field
21.3T 18.8T 900 17.6T 800 750 14.1T 11.74T 600 500 9.4T 400 4.7T 2.35T 200 0.71T 100 30
140
120
100
80
60
40
20
ppm
1H
8.06 7.67 7.54 7.44 7.43 7.41 7.35 7.33 7.28 6.91 6.21 6.16 5.72 5.61 4.87 4.72 4.33 4.23 4.13 3.73
shifts (int)
3.48 2.47 2.38 2.31 2.29 2.22 2.17 2.10 1.81 1.73 1.72 1.62 1.53 1.17 1.08 (1) (1) (1) (3) (1) (1) (3) (imp.) (1) (1) (3) (3) (H2O) (3) (3)
1960 1970 1980 1990 2000
800
H Frequency (MHz)
600
400
1
200
0
Year
核磁共振仪
1953年，美国varian公司研制成世界上第 一台商品化的核磁共振仪（30MHz）
核磁共振
经历了磁场超导化和脉冲傅立叶变换技 术两次重大革命 大大提高了仪器的分辨率和灵敏度 使核磁共振的研究对象从液体扩展到固 体 实验技术从一维扩展到多维 应用范围从有机小分子扩展到生物大分 子
Applications of NMR
Chemically Induced Dynamic Nuclear Polarization (CIDNP)
(Ph C O O2) 80 C
o
PhCOO HS
.
Ph H
Ph
.
- CO2
Fischer, 1967
Yang, et al., JCS Perkin 2, 1994, 585 ChemComm., 1996, 451
HPLC-NMR / MS
Applications of NMR
Dynamic NMR (DNMR)
Inversion Barrier ~ 10 kcal / mol
Inversion Barrier ~ 21 kcal / mol Yang, Chin. J. Chem., 1990, 428 Yang, Chin. J. Chem., 1990, 430
Proton-Carbon Pairs
8.06 7.67 7.54 7.44 7.42 7.41 7.35 7.33 7.28 6.91 6.21 6.16 5.72 5.61 4.87 (2) (2) (1) (2) (1) (2) (2) (2) (1) (1) (1) (1) (1) (1) (1) 130.21 127.04 133.71 128.70 131.97 127.04 129.03 128.72 128.37 (d*) (d*) (d) (d*) (d) (d*) (d*) (d*) (d) 4.72 4.33 4.23 (1); 4.13 3.73 3.48 2.47 (1); 1.81 2.38 2.31 2.29 (1); 2.22 2.17 1.73 1.72 1.62 1.17 1.08 (1) (1) (1) (1) (1) (1) (1) (3) (1) (3) (1) (3) (3) (3) (3) 73.20 72.18 76.52 45.64 (d) (d) (t) (d)
1H
d1
t1/2
t1/2
13C
Dec.
Grad H3 C1 C6
H C
1
H2
H4
H5a H5b
H1
H6
H
6
H C H C
5
Cl Cl C
4
C C
3 2
C5 C4
H H
H H
H H
C3
C2
1H chemical shifts
C13 chemical shifts
Results from the HSQC experiment:
核磁共振
已经成为鉴定有机化合物结构以及研究 分子结构、构型、构象和化学反应动力 学等方面的重要的方法 在有机化学、生物化学、药物化学、物 理化学、无机化学及多种工业部门研究 中成为最活跃和不可缺少的一部分
Nuclei can be studied by NMR
X X
X X
X X X X X X
O H CH3 HN H OH O O H H H HO H O O O H O CH3 O CH3 H3C H3C O O H O CH3 OH H H H H
紫杉醇
Asym. Carbon (11)
211 = 2048 stereoisomers !!!
?
1H
spectrum of Taxol
(2)
(2) (2) (2) (2) (2)
51 protons
47 carbons
Applications of NMR
3D Structure of Bio-macromolecules
K. Wuethrich
2002 Nobel Prize in Chemistry
Applications of NMR
Fra Baidu bibliotek
Applications of NMR
Molecular Structure in Solution
High resolution 1D & 2D NMR
?
R. R. Ernst 1991 Nobel Prize in Chemistry
Taxol: anticancer drug (MW: 853.906) (MS) C47H51NO14 ( the number of constitutional isomers >> 1030 !!!!!)
C47H51NO14
5.2 mg (10 mmol) in CDCl3
4.9
4.8
4.7
4.6
4.5
4.4
4.3
4.2
ppm
8
7
6
5
4
3
2
ppm
13C
spectrum of Taxol
Experimental time: 14h
C47H51NO14
130.0
129.0
128.0
ppm
200
180
160
NMR Spectroscopy
Chapter 2 Basic Principles of NMR
核磁共振波谱学
Nuclear Magnetic Resonance (NMR) Spectroscopy
第2章 核磁共振基础
2.1 核自旋和共振 2.2 核磁共振基本概念 2.2.1 驰豫 2.2.2 化学位移 2.2.3 自旋偶合 2.3 核磁共振实验 2.3.1 核磁共振仪 2.3.2 测定方法 2.3.3 核磁共振谱图
H1 H1 H3 H2 H2
d1
t1
Spin-lock
H
1
H
X X
3
H
1
H
3
X O X
X X
X
R1
R2 X H
2
H
4
O
H
2
H
4
HSQC - Heteronuclear Single Quantum Coherence
(correlates X-1H chemical shifts via one-bond 1J(XH) couplings)
R. R. Ernst
1991, Chemistry HR & 2D NMR
K. Wuethrich 2002, Chemistry NMR for Bio-Macromol.
P. Lauterbur
P. Mansfield
2003 Medicine MRI
2003年度诺贝尔生理学和医学奖获得者
核磁共振成像技术的发现，医学诊断和生物细胞研究领域的突破性成就。
NMR, An Evergreen
• • • • • • • • • • 1939 Rabi, Magnetic resonance for atomic nuclei (1944 Noble Prize) 1945 Bloch & Purcell, NMR phenomenon, (1952 Noble Prize) 1953 Varian Company, The first commercial NMR spectrometer 1957 13C NMR 1959 Solid state NMR 1964 High resolution NMR (Superconducting magnet) 1965 Pulse Fourier Transform NMR (FT-NMR) 1967 Chemically Induced Dynamic Nuclear Polarization (CIDNP) 1976 Ernst, Methodology in high resolution NMR (1991 Noble Prize) 1973,1982 Lauterbur & Mansfield, Hospital NMR Imaging (2003 Nobel Prize) • 1983, 1990 Wuethrich, NMR for bio-macromolecules (2002 Nobel Prize) • 1995 LC-NMR, LC-MS-NMR
Applications of NMR
Time-Resolved CIDNP
Yang, Gomberg 2000, 2000
Applications of NMR
NMR Imaging (MRI)
P. Lauterbur P. Mansfield 2003 Nobel Prize in Medicine
H
1
H
3
H4
X
X O X
X X
R1
H4
R2 X H
2
H
4
O
H
2
H
4
H3
TOCSY - TOtal Correlation SpectroscopY
(identifies protons belonging to the same scalar coupling network)
H3
90 Trim
H4 H4
13C
203.61 172.72 171.22 170.35 167.04 167.01 141.95 137.99 133.71 133.65 133.23 131.97 130.21 129.18 129.03 128.72 128.70 128.37 127.04 127.04
shifts (int)
84.41 81.19 79.05 76.52 75.57 74.98 73.20 72.41 72.18 58.65 55.04 45.64 43.19 35.72 35.63 26.88 22.63 21.83 20.84 14.83 9.57
(2) (2) (1) (2) (1) (2) (2) (2) (1) (1) (1) (1) (1) (1) (1) (1) (1) (1) (1) (1)
NMR Microscopy
Substructural elements
X-Ray UV IR MS NMR (+) + + + +
full connectivity
+ +? +
3D structure
+ +
The NMR strategy and the NMR tools:
-
number of protons, 1H chemical shifts, 1H-1H couplings ---> 1H spectrum number of magnetically nonequivalent carbons ---> 13C spectrum number of NMR-detectable other nuclei (19F, 31P, …, 15N?) ---> X spectrum 1H-13C direct correlation (one-bond) ---> HSQC, (C,H COSY, HMQC) identification if 1H spin systems ---> TOCSY 1H coupling network ---> COSY 1H-X (X=13C, 15N, 19F, 31P, etc) long-range correlation ---> HMBC( COLOC ) stereochemistry, 3D structure ---> NOESY, ROESY
COSY - COrrelation SpectroscopY
(correlates scalarly coupled protons)
The gradient DQF-COSY sequence:
1H
H3
H4H4
H1 H1 H3 H2 H2
H2
d1
t1
H2 H3
Grad.
H1 H1
H
1
H
X X
3
NMR, An Evergreen
Phys.
Biol.
NMR
Chem.
Med.
Nobel Prize Laureates related to NMR
I. I. Rabi 1944, Physics Resonance of nuclei
F. Bloch E. M. Purcell 1952, Physics Discovery of NMR
S. P. Mansfield (彼得－曼斯菲尔德), University of Nottingham, 英国
P. C. Lauterbur(保罗－劳特布尔), University of Illinois, 美国
Increase of Magnetic Field
21.3T 18.8T 900 17.6T 800 750 14.1T 11.74T 600 500 9.4T 400 4.7T 2.35T 200 0.71T 100 30
140
120
100
80
60
40
20
ppm
1H
8.06 7.67 7.54 7.44 7.43 7.41 7.35 7.33 7.28 6.91 6.21 6.16 5.72 5.61 4.87 4.72 4.33 4.23 4.13 3.73
shifts (int)
3.48 2.47 2.38 2.31 2.29 2.22 2.17 2.10 1.81 1.73 1.72 1.62 1.53 1.17 1.08 (1) (1) (1) (3) (1) (1) (3) (imp.) (1) (1) (3) (3) (H2O) (3) (3)
1960 1970 1980 1990 2000
800
H Frequency (MHz)
600
400
1
200
0
Year
核磁共振仪
1953年，美国varian公司研制成世界上第 一台商品化的核磁共振仪（30MHz）
核磁共振
经历了磁场超导化和脉冲傅立叶变换技 术两次重大革命 大大提高了仪器的分辨率和灵敏度 使核磁共振的研究对象从液体扩展到固 体 实验技术从一维扩展到多维 应用范围从有机小分子扩展到生物大分 子
Applications of NMR
Chemically Induced Dynamic Nuclear Polarization (CIDNP)
(Ph C O O2) 80 C
o
PhCOO HS
.
Ph H
Ph
.
- CO2
Fischer, 1967
Yang, et al., JCS Perkin 2, 1994, 585 ChemComm., 1996, 451
HPLC-NMR / MS
Applications of NMR
Dynamic NMR (DNMR)
Inversion Barrier ~ 10 kcal / mol
Inversion Barrier ~ 21 kcal / mol Yang, Chin. J. Chem., 1990, 428 Yang, Chin. J. Chem., 1990, 430
Proton-Carbon Pairs
8.06 7.67 7.54 7.44 7.42 7.41 7.35 7.33 7.28 6.91 6.21 6.16 5.72 5.61 4.87 (2) (2) (1) (2) (1) (2) (2) (2) (1) (1) (1) (1) (1) (1) (1) 130.21 127.04 133.71 128.70 131.97 127.04 129.03 128.72 128.37 (d*) (d*) (d) (d*) (d) (d*) (d*) (d*) (d) 4.72 4.33 4.23 (1); 4.13 3.73 3.48 2.47 (1); 1.81 2.38 2.31 2.29 (1); 2.22 2.17 1.73 1.72 1.62 1.17 1.08 (1) (1) (1) (1) (1) (1) (1) (3) (1) (3) (1) (3) (3) (3) (3) 73.20 72.18 76.52 45.64 (d) (d) (t) (d)
1H
d1
t1/2
t1/2
13C
Dec.
Grad H3 C1 C6
H C
1
H2
H4
H5a H5b
H1
H6
H
6
H C H C
5
Cl Cl C
4
C C
3 2
C5 C4
H H
H H
H H
C3
C2
1H chemical shifts
C13 chemical shifts
Results from the HSQC experiment:
核磁共振
已经成为鉴定有机化合物结构以及研究 分子结构、构型、构象和化学反应动力 学等方面的重要的方法 在有机化学、生物化学、药物化学、物 理化学、无机化学及多种工业部门研究 中成为最活跃和不可缺少的一部分
Nuclei can be studied by NMR
X X
X X
X X X X X X
O H CH3 HN H OH O O H H H HO H O O O H O CH3 O CH3 H3C H3C O O H O CH3 OH H H H H
紫杉醇
Asym. Carbon (11)
211 = 2048 stereoisomers !!!
?
1H
spectrum of Taxol
(2)
(2) (2) (2) (2) (2)
51 protons
47 carbons
Applications of NMR
3D Structure of Bio-macromolecules
K. Wuethrich
2002 Nobel Prize in Chemistry
Applications of NMR
Fra Baidu bibliotek
Applications of NMR
Molecular Structure in Solution
High resolution 1D & 2D NMR
?
R. R. Ernst 1991 Nobel Prize in Chemistry
Taxol: anticancer drug (MW: 853.906) (MS) C47H51NO14 ( the number of constitutional isomers >> 1030 !!!!!)
C47H51NO14
5.2 mg (10 mmol) in CDCl3
4.9
4.8
4.7
4.6
4.5
4.4
4.3
4.2
ppm
8
7
6
5
4
3
2
ppm
13C
spectrum of Taxol
Experimental time: 14h
C47H51NO14
130.0
129.0
128.0
ppm
200
180
160