r m s b nmr spectroscopy jm franconi. r m s b mr spectrocopy in a spectrum, each signal represent...

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NMR SPECTROSCOPYNMR SPECTROSCOPY

JM FranconiJM Franconi

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In a spectrum, each signal represent the resonance frequencyof a nucleus in a specific environment

Principle of FT NMR spectroscopy

Bo is fixed

a RF pulse irradiate a broad range of frequency

t

RFirradiation

FID spectrum

sampling

f

FT

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Non selective RF pulse

In spectroscopy the RF pulse used is < 100 microsecond

RF rectangular sinc

B1 > 2F

F frequency range

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RF pulse Magnetisation evolution

Bo

MMz

MxyO

B1

During RF pulse M rotateAround x direction with an angle:

x

y

z

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spectrum parameter chemical shift

f1 f2 f3 f4

A same nucleus can have different resonance frequency

f

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chemical shift phenomenon

Bo

H1

O

Bo1 Bo2

H2Bo1

Bo

Bo2

C

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chemical shift measurement

f

reference

6

10ref

refi

i

ref i

DiamagneticHigh fieldLow electronégativity

paramagneticlow fieldhigh electronégativity

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J coupling

J coupling is an interaction between close nuclei

This interaction is independant of Bo field

It does not exit on one isolated molecule

f fA fB

JAB

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J scalar coupling

H1

C C

H2

50%(+1/2)

50%(-1/2)

H1 is influenced by the 2 H2 sites

Jcoupling is transported by chemical bond

f1 f1+dBH2 f1- dBH2

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J dipolar coupling

Coupling is also possible directly through space

H1 H2

1/r6

r

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J coupling

The Nuclei magneticaly equivalent have the same chemicalshift. They do not show any coupling effect

C

CHa

HbHc

fHa = fHb = fHc

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J coupling

Rules for multiplicity determination

A I nucleus coupled to n spin S give a signal with:

2.n.S + 1 componants

The componants relative intensity are given by a binomialDistribution.

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Signal area

NMR spectroscopy is a quantitative method

f

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spectrum quality

Concept of digital resolution

Digital resolution

True NMR resolution

At

DW

FT

Time domain Frequency domain

R = 1/AT

t

T2*

f

TR=1/T2*

CONDITION: TR > DR

DR

ft

AT = N.DW

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Sampling rules

DW

F=1/Dw

f2

f2

f1

f1f2

Application of Nquist theorem

2. F. Aq = N

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Sensitivity

3

ofy Sensitivit ofy Sensitivit

H

XaHX

-The gyromagnetic ratio and the natural isotopic abundancedetermine the NMR signal available

- the nucleus sensitivity is proportional to Bo2 and

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concept of population and coherence

E1

E2

Mz

population

E2

Cohérence=transition

Mz

E1

MR signal

RF

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Polarisation transfer (NOE) C

H

+1/2

-1/2

Niveau H1

+1/2

+1/2

Niveau C13

2 independant level of energy

C

H

NO COUPLING

+1/2+1/2

-1/2-1/2

-1/2+1/2

+1/2-1/2

Transition H1

Transition C13

COUPLING

system of 4 levels of energy

Longdistance

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Polarisation transfer (NOE)

H1 transition

C13 transition

POLARISATION

dH+dC

dH-dC

-dH-dC

H1 TRANSITION SATURATION

-dH+dC

-dH+dC

+dH-dC

-dH-dC

+5

-3

ff

dH = 4.dC

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SEQUENCESEQUENCE

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SEQUENCESEQUENCESPIN ECHO SEQUENCE

90°180°

te/2

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SEQUENCESEQUENCE

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SEQUENCESEQUENCE

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DEMI TOUR!

SEQUENCESEQUENCE

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SEQUENCESEQUENCE

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ECHO = coherence recovery

SEQUENCESEQUENCE

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SEQUENCE SPIN ECHOSEQUENCE SPIN ECHO

FIDFIDECHO DE SPINECHO DE SPIN

T2T2T2*T2*

Mxy

t

90 o 180o

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RF

GS

GP

GR

DAC on

TE TR

CONCEPT OF PULSE SEQUENCE

SPIN ECHO SEQUENCE

90°180°

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NMR SPECTROSCOPY APPLICATION

-Chemical structure determination (protein, ADN,…)

-Quantitative measurement(isotopic enhancement determination)

food and beverage quality control

-Spectroscopy in vivo (metabolism caracterisation)