Chem-806 Identification of organic and inorganic compounds by advance NMR techniques Tool box 2D-NMR: Homonuclear 2D-NMR: Heteronuclear 3D-NMR
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Chem-806Identification of organic and inorganic
compounds by advance NMR techniques
Tool box
2D-NMR: Homonuclear
2D-NMR: Heteronuclear
3D-NMR
Parameter Consideration• Receptivity
– Spin Quantum Number– Resonance Frequency– Sensitivity– Natural abundance
• Frequency Shift– Absolute Frequency– Relative Chemical Shift Scale (Referencing)
• Relaxation – T1 and T2
– Definition– Mechanisms– Measurement– NOE
Multinuclear NMR
Z Nuclei Spin I Frequency % natural abundance
1 1H ½ 100.00 99.98
6 13C ½ 25.15 1.108
7 14N 1 7.23 99.935
15N ½ 10.136 0. 365
9 19F ½ 94.103 100
15 31P ½ 40.43 100
At Bo=2.35 Tesla
When sampling a nuclei, following parameters shoud be considered:
• Sensitivity• Natural abundance• Relaxation time : T1 recycle time, T2 acquisition Time• Influence of H-decoupler: {NOE and J}
n0 =gB0
2p
Sensitivity and Receptivity
The sensitivity of a nuclei depends on:
1. Magnetic Field ( g)2. Population excess ( g)3. Magnetic field induced in receiver coil ( g)
Sensitivity = k * gx3 * Ix(Ix +1)
e.g. g(13C)/g(1H) = 1/4 13C less sensitive than proton (64 less)
Receptivity Rx = ax * Sensitivity
Where ax = natural abundance
e.g. Relative receptivity of 1H and 13C
R =CH aC * gC
3
aH * gH3 = 0.01 * (25)3
1 * (100)3 = 1.6 * 10-4 R = 0.83FH
T1 considerationsSpin lattice relaxation time T1 “lifetime” of First Order Rate process
z
yx
90x
z
yx
Mo
My
z
yx
z
yx
z
yx
Mo5 T1t ..t
Magnitude of T1 is highly dependant on : 1. the type of nuclei2. State of the sample
T1 governs the efficiency of the NMR experiment : recycle time
For 1H in solution T1 can be 0.01 to 100 sec.For low g nuclei – spin ½ - relaxation can be much longer!
Recovery of the magnetization along the Z axis
Relaxation (T1)
Dinitrobenzene: T1
NO2
NO2
HH
H
H
H2H4/H6 H5
180 90
t
Inversion recovery : 13C
180 90
tD1= 5T1
t = 0.03 s
t = 1.5 s
t = 3 s
t = 6 s
t = 50 s
3
2
4
1
5
6
7CH39
CH38
OH
CH310
T1 = tnull / ln2 = 1.443 * tnull
e.g. C2 => T1 = 4.3 s (tnull=3 )
Helping relaxationOne approach of reducing relaxation time is by the addition of paramagnetic relaxation reagent (Chromium III acetylacetonate => Cr(acac)3)
1s delay, 30o pulseWithout Cr(acac)3
With Cr(acac)3
Intensity vs Pulse Interval
Mz
t
M(t) = Mo(1-e-t/T1)
PW=90o
D1 AQ
NSz
yx
90x
z
yx
Mo
My
t = pulse interval = D1 + AQ
t1 * T1
M(t)0.63 M0
2 * T1 0.86 M0
3 * T1 0.95 M0
4 * T1 0.98 M0
5 * T1 0.99 M0
10 * T1 0.99995 M0
Optimum recycle delay (pulse interval) with 90` pulse
Total experiment time fixed
During the experiment, t (D1+AQ) is repeated for NS
PW=90o
D1 AQ
NS
t = pulse interval = D1 + AQ
t Sensitivity
.1 T1 .3
.2 T1 .41
.5 T1 .56
.75 T1 .61
1 T1 .63
1.26 T1 .64
1.5 T1 .63
2 T1 .55
Optimum delay
Optimum angle with D1 < T1
PW<90o
D1 AQ
NS
D1 = 0
t = pulse interval = AQ
z
yx
PWx
Mo M
z
yx
Optimum angle Ernst angle
a = cos-1 et/T1
t aE T1 (t=1)
100 T1 90o .01
10 T1 90o .1
2.5 T1 86.3o .4
1.5 T1 77.1o .67
1. T1 68.4o 1
0.5 T1 52.7o 2
0.25 T1 38.8o 4
0.1 T1 25.2o 10
0.01 T1 8.1o 100
Short T1
Long T1
Sensitivity curves for different pulse and different delays and relaxation time
Steady State
T2 Consideration
T2
Refocusing of field inhomogeneity
Carr-Purcell-Meiboom-Gill
CPMG used to get rid of broad signals
Polystyrene (50,000) + camphor
90 180
t
t = 1.5 ms
t
SW and Memory sizeO1
SW
The spectral window (SW) and the carrier offset (O1) are chosen to match entire spectra (to avoid Fold-over and aliasing)
For a given SW, the time (Dwell time) between 2 data point is defined by the Nyquist theorem (1/2SW).
The total number of data point (TD) acquired is related to the Acquisition time AQ (DW*TD)
The digital resolution depends on the window and on the number of points placed in that window
Digital resolution = 2 * SW/ TD = 1/AQ
TD = 2 * SW * AQ
Sharp lines have long FID (long T2*),
broad peaks have short FID (short T2*),
AQ ~ 3 * T2
Nyquist Theorem
DIGITALLIZATION
• Accuracy• Speed
Digitallization: Convert FID (Volt/Time) in Digital formDigitallization process is limited by:
Carrier Offset or Transmitter Offset or “O1” is the frequency of the irradiating field. It is also the “Reference” or “Rotating Frame” frequency
The “Window” or “Spectral Width” also called “SW” define the range of frequencies that can be measured
r.f.
O1SW
The Sampling Rate => 2 Points/Cycle Dwell Time = DW= _1__2*SW
If Maximum Frequency to be sampled is fmax = SW
We must sample at a rate no less than 2 * SWsec.
Digital Resolution
The amount of memory limit the accuracy of the signal to be recorded
For a given # of memory (# Points -> TD (time domain)), one obtain:
NP (real) and NP (Imaginary) 2 2
Digital Resolution = D.R. = Df (Separation between 2 points)
D.R. = 2 * SW NP
Digitallization : resolution and Acquisition time
Example
At 200 MHz If: SW=2000 Hz (10 ppm)
TD = 16,000 points (16K)
What is the Digital Resolution:
D.R. = 2*SW/TD = 4000 / 16,000 = 1 / 4 = 0.25 Hz
What is the Acquisition Time AQ:
AQ = TD * DW = TD / (2 * SW) = 4 seconds
D.R. = 1 / AQ = 2 * SW / TD
C13-NMR : proton decoupling
C
O
OH CH2
CHCH3
OH
Heteronuclear nOe
For nuclei having positive g : (e.g. 13C)Decoupling proton can produce higher signals due to nOe.Enhancement is dependant on motion and distance between interacting nuclei
4a
8a
5
8
6
7
3
2
4
O O
CH39
AcO
AcO
As a consequence, quaternary carbons are much smaller than protonated carbons
Heteronuclear nOe
• Can yield higher positive signal for nuclei with positive g (e.g. 13C)
• For nuclei with negative g (e.g. 29Si, 15N) can yield larger (negative) signals or for partial T1DD can null the signal!!!
e.g. 15N {1H}
Without NOES = A0 =1
NOEmax gH
2 gN
~ -5
S =A0+NOE= -4
100% NOE50% NOE
NOE=-2.5
S =A0+NOE= -1.5
20% NOE
S =A0+NOE= 0
NOE=-1
Refocusing : echo formation
Delay as a sequence building block
JAX
dA
J2
J2
-
AX Spin system A=1H, X=13C, JAX
For each isochromats the distance they run in the xy frame during a delay t is:
Distance = Frequency (cycles/sec) * delay (sec)
Distance = 2p * (+/-) J/2 * t
x x x x
J= 10 Hz t (1/2J) = 0.05 s
J=100 Hz t (1/2J) = 0.005 s
J=140 Hz t (1/2J) = 0.00357 s
DelayDist.
t=00
t=1/4Jp/4 (45o)
t=1/2Jp/2 (90o)
t=1/Jp (180o)
APT experiment
APT Pulse Sequence
Multiplet Modulation
APT : 8 msec delay
APT : 6 msec delay
INEPT experiment
INEPT
Comparing NOE Enhancement and
INEPT Enhancement
INEPT sequence
Multiplet distortion in INEPT: 29Si
Multiplet distortion in coupled INEPT: 13C
Multiplet distortion in coupled INEPT: 13C
29Si – INEPT coupled
Refocused INEPT
Multiplicity Modulation : HX
Multiplicity Modulation : XH2
Multiplicity Modulation : XH3
Decoupled INEPT: Menthol
Optimum delay in decoupled INEPT
Decoupled INEPT: compare normal and INEPT 29Si
DEPT
Intensity vs
pulse angle
Normal 13C
DEPT-45
DEPT-90
DEPT-90
DEPT-135Menthol in Acetone-d6
DEPT Ipsenol
DEPT Editing
ADEPT Editing
The BIRD pulse
Refocusing : echo formation
Field Gradient in NMR
Gradient Field: A Way to Speed up 2D & 3D NMR =>
replace phase Cycling
If gradient gz is applied during tg
Each isochromats accumulate phase:
F = 2p * Df * tg
Coherences precess at n * Df Where n is the coherence order
GE-COSY: Gradient Enhance COSY
Homonuclear 2DJ-NMR
Different format in 2D-NMR
2DJ stack plot
2DJ- contour
2DJ- expansion
2DJ-ROTATE
2DJ-rotate
NMR processing: apodization Window
• Line Broadning (LB) : Exponential Multiplication improves the signal to noise ratio (at the expense of resolution)
Processing : line broadening
Processing : line broadening
NMR processing: apodization Window
• Line broadning : Exponential multiplication improves the signal to noise ratio (at the expense of resolution)
• Resolution Enhancement reverse exponential + Gaussian function also traf function, also sine function (for 2D – magnitude mode)
Processing : resolution enhancement
Processing : sine bell (phased mode)
Processing : sine bell (magnitude mode)
Processing : sine bell (Power mode)
Processing : Qsine (phased mode)
Processing : Qsine (magnitude mode)
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