NMR Spectroscopy

Part II. Signals of NMR

Free Induction Decay (FID)•FID represents the time-domain

response of the spin system following application of an radio-frequency pulse.

•With one magnetization at , receiver coil would see exponentially decaying signal. This decay is due to relaxation.

Fourier TransformThe Fourier transform relates the time-domain f(t) data with the frequency-domain f(w) data.

Fourier Transform

Fourier Transform

NMR line shape

Lorentzian line

A amplitude

W half-line width

202

2

4 xxW

AWy

Resolution

Definition

For signals in frequency domain it is the deviation of the peak line-shape from standard Lorentzian peak. For time domain signal, it is the deviation of FID from exponential decay. Resolution of NMR peaks is represented by the half-height width in Hz.

Resolution

Resolution-digital resolution

Resolution Measurement

half-height width:10~15% solution of 0-dichlorobenzene

(ODCB) in acetone

Line-shape:Chloroform in acetone

Resolution Factors affect resolution

Relaxation process of the observed nucleus

Stability of B0 (shimming and deuterium locking)

Probe (sample coil should be very close to the sample)

Sample properties and its conditions

Sensitivity Definition

signal to noise-ratio

A : height of the chosen peakNpp : peak to peak noise

ppN

Ans 5.2/

Sensitivity Measurement1H 0.1% ethyl benzene in deuterochloroform13C ASTM, mixture of 60% by volume deuterobenzen

e and dioxan or 10% ethyl benzene in chloroform

31P 1% trimehylphosphite in deuterobenzene15N 90% dimethylformamide in deutero-dimethyl-

sulphoxide19F 0.1% trifluoroethanol in deuteroacetone2H, 17O tap water

Sensitivity

Factors affect sensitivity

Probe: tuning, matching, size

Dynamic range and ADC resolution

Solubility of the sample in the chosen solvent

Spectral Parameters Chemical Shift

Caused by the magnetic shielding of the nuclei by their surroundings. d-values give the position of the signal relative to a reference compound signal.

Spin-spin CouplingThe interaction between neighboring nuclear dipoles leads to a fine structure. The strength of this interaction is defined as spin-spin coupling constant J.

Intensity of the signal

Chemical Shift Origin of chemical shift

shielding constant

Chemically non-equivalent nuclei are shielded to different extents and give separate resonance signals in the spectrum

000 1 BBBBeff

0' 1

22BBeff

Chemical Shift

Chemical Shift – scale or abscissa scale

612

121

12

120

12

20

2

10

1

10parameter shift Chemical

2

12

12

B

B

B

Chemical Shift

Shielding CH3Br < CH2Br2 < CH3Br < TMS

610frequency observing

(ppm) 82.6101090

614CHBr 6

63

90 MHz spectrum

Abscissa Scale

Chemical Shift

is dimensionless expressed as the relative shift in parts per million ( ppm ).

is independent of the magnetic field of proton 0 ~ 13 ppm

of carbon-13 0 ~ 220 ppm

of F-19 0 ~ 800 ppm

of P-31 0 ~ 300 ppm

Chemical Shift

Charge density Neighboring group

AnisotropyRing currentElectric field effectIntermolecular interaction (H-bonding &

solvent)

ieRNlocalpara

localdia

2

//3cos1

43

1

r

N

Chemical Shift – anisotropy of neighboring group

Differential shielding of HA and HB in the dipolar field of a magnetically anisotropic neighboring group

susceptibility

r distance to the dipole’s center

~2.88

~9-10

Chemical Shift – anisotropy of neighboring group

• Electronegative groups are "deshielding" and tend to move NMR signals from neighboring protons further "downfield" (to higher ppm values).

• Protons on oxygen or nitrogen have highly variable chemical shifts which are sensitive to concentration, solvent, temperature, etc.

• The -system of alkenes, aromatic compounds and carbonyls strongly deshield attached protons and move them "downfield" to higher ppm values.

•Electronegative groups are "deshielding" and tend to move NMR signals from attached carbons further "downfield" (to higher ppm values). •The -system of alkenes, aromatic compounds and carbonyls strongly deshield C nuclei and move them "downfield" to higher ppm values. •Carbonyl carbons are strongly deshielded and occur at very high ppm values. Within this group, carboxylic acids and esters tend to have the smaller values, while ketones and aldehydes have values 200.

Ring Current The ring current is induced form the delocalized ele

ctron in a magnetic field and generates an additional magnetic field. In the center of the arene ring this induced field in in the opposite direction t the external magnetic field.

Ring Current -- example

Spin-spin coupling

Spin-spin coupling

AX system

AX2 system

Spin-spin coupling

AX3 system

Multiplicity RuleMultiplicity M (number of lines in a multiplet)

M = 2n I +1

n equivalent neighbor nuclei

I spin numberFor I= ½

M = n + 1

AX4I=1; n=3

Example AX4 system

Order of Spectrum

Zero order spectrum

only singlet

First order spectrum

>> J

Higher order spectrum

~ J

AMX system

Spin-spin coupling Hybridization of the atoms Bond angles and torsional angles Bond lengths Neighboring -bond Effects of neighboring electron lone-pairs Substituent effect

JH-H and Chemical Structure

Geminal couplings 2J (usually <0)

H-C-H bond anglehybridization of the carbon atomsubstituents

Geminal couplings 2J bond angle

Geminal couplings 2J

Substituent Effects Effect of Neighboring -

electrons

Vicinal couplings 3JH-H

Torsional or dihedral angles Substituents HC-CH distance H-C-C bond angle

Vicinal couplings 3JH-H dihedral

angles

Karplus curves

tg JJJ 333

3

12

3

1

Chemical Shift of amino acid

http://bouman.chem.georgetown.edu/nmr/interaction/chemshf.htm

Automated Protein Chemical Shift Predictionhttp://www.bmrb.wisc.edu:8999/shifty.html

BMRB NMR-STAR Atom Table Generator for Amino Acid Chemical Shift Assignments

http://www.bmrb.wisc.edu/elec_dep/gen_aa.html

Chemical Shift Prediction

http://bouman.chem.georgetown.edu/nmr/interaction/chemshf.htm

Example 1