NMR SPECTROSCOPY

PART I BASIC PRINCIPLES OF NMR SAMPLE PREPARATION SCHEME OF NMR INSTRUMENT

CHEM 127.1 Maam Fatima

Argamino, C.; Buenaseda, A.; Gajigan, A.

Basic Principles of NMR

Introduction

NMR - identify the carbonhydrogen framework of an organic compound

In many atoms (such as 12C) spins are paired such that the nucleus of the atom has no overall spin.

atoms (such as 1H,13C 15N, 19F, 31P.) the nucleus does possess an overall spin 12C = no spin = no magnetic dipole

NMR phenomenon

absence of an applied magnetic field

the

nuclear spins are randomly

applied magnetic field

the

nuclei twist and turn to align themselves with or against the field

NMR phenomenon

The energy difference (E) between the and states depends on the strength of the applied magnetic field (Bo)

Bo E

Radiation required is in the radiofrequency (rf) region of the electromagnetic spectrum and is called rf radiation

NMR phenomenon

Sample - subjected to a pulse of radiation (equal to E)

Nuclei promoted

Nuclei undergo relaxation

emit electromagnetic signals whose frequency depends on

The NMR spectrometer detects these signals

displays a plot of signal frequency versus intensityan NMR spectrum

NMR phenomenon

Resonance flipping back and forth of nuclei between spin states] the magnetic field is proportional to the operating frequency (MHz).

if the spectrometer has a more powerful magnet, it must have a higher operating frequency
v = frequency = gyromagnetic ratio (depends on the magnetic moment of the particular kind of nucleus)

Fourier Transform (FT) spectrometer

Dispersive IR
viewing

each component frequency sequentially

Fourier Transform IR
all

frequencies are examined simultaneously Radiation source interferometer detector

Fourier Transform (FT) spectrometer

Individual protons absorb frequency Protons relax

produce a complex signal free induction decay (FID) E intensity of the signal decays as the nuclei lose the energy

computer converts the intensity-versus-time data into intensity- versusfrequency information in a mathematical operation known as a Fourier transform

Fourier Transform (FT) spectrometer

Michelson interferometer
allows

some wavelengths to pass through but blocks others due to wave interference is modified for each new data point by moving one of the mirrors

Typical Layout of FTIR spectrometer

Shielding

protons do not experience the same applied magnetic field cloud of electrons partly shields the nucleus

Protons in electron-rich environments

shielded and appear at lower frequencies less shielded and appear at higher frequencies

Protons in electron-poor environments

Number of Signals

Chemical Shift

position at which a signal occurs in an NMR spectrum

a

measure of how far the signal is from the reference signal Does not depend on operating frequency

Relative positions of signals

The closer the protons are to the electronwithdrawing group, the less they are shielded from the applied magnetic field, so the higher the frequency (i.e., the farther downfield)

Characteristic Values of Chemical Shift

Integration of NMR Signals

the area under each signal is proportional to the number of protons that gives rise to the signal

1.6 : 7.0 = 1 : 4:4 = 2 : 8.8 =2:9

Splitting of Signals

Splitting is caused by protons bonded to adjacent (i.e., directly attached) carbons.

splitting
where

of a signal is described by N + 1 rule

N is the number of equivalent protons bonded to adjacent carbons

Splitting of Signals

Hb split into a quartet by Ha, spilt into triplet by Hc Peaks depend on J (overlap is possible)

Splitting of Signals

diff coupling constants = use (N + 1) separately similar J = use (N + 1) as if the adjacent H are equivalent

Coupling constant

The distance (Hz) between two adjacent peaks of a split NMR signal

useful in analyzing complex NMR spectra because protons on adjacent carbons can be identified by identical coupling constants

Resolution of HNMR Spectra

To observe separate signals with clean splitting patterns, the difference in the chemical shifts of two adjacent protons (in Hz) must be at least 10 times the value of the coupling constant (J).

13C
13

NMR

C NMR requires Fourier transform technique

because the signals obtained from a single scan are weak to be distinguished from background electronic noise.

13

C are weak since they exist in low amount (isotope) area under a 13C NMR signal is not proportional to the number of atoms Carbon splitting - rare ; carbon-proton coupling possible

13C

NMR

Proton coupled 13C NMR splitting is observed

SaMPLE PREPARATION

SAMPLE PREPARATION

Sample tubes
accurate dimensions keep tubes free from dust, grease etc never be cleaned with chromic acid tubes can be rinsed with distilled water and then acetone

blow nitrogen or air through the tube, with a pipette, while warming it gently for a few minutes or to leave under vacuum for some hours

SAMPLE PREPARATION

To obtain high resolution NMR spectra it is necessary that your NMR sample is free of suspended material

will increase the line width of the Spectrum broad spectral lines reduce spectral resolution and no amount of shimming can correct for this

Suspended material can easily be removed from an NMR sample by constructing a filter using cotton wool as a filtering agent

SAMPLE PREPARATION

The solute of interest can then be dissolved in a separate glass vial using deuterated solvent (e.g. CDCl3, DMSOD8, CD3OD, etc.) required to make your NMR sample After the solute has been dissolved it can be transferred directly to a NMR tube by passing the solution through the cotton filter

SAMPLE PREPARATION

the sample volume can be adjusted by adding the remaining solvent to the NMR tube so that a final sample volume of ~700 L or a sample height of ~55 mm is reached followed by vigorous shaking of the sample to effectively mix its content

SAMPLE PREPARATION

Sample Volume
best

results will be obtained using a sample volume of 0.6 - 0.8 ml for a 5mm NMR tube Smaller volumes can be used but will require much more shimming too long are also more difficult to shim and are waste costly solvent

SAMPLE PREPARATION

Sample Quality

Solid particles distort the magnetic field homogeneity because the magnetic susceptibility of a particle is different from that of a solution A sample containing suspended particles thus has a field homogeneity distortion around every single particle.

causes broad lines and indistinct spectra that cannot be corrected.

SAMPLE PREPARATION

Sample Solvent
contain

deuterium instead of hydrogen The NMR signal from the deuterium nuclei is called the NMR lock and is used by the spectrometer for stabilization. Solvents should be kept dry and free from impurities.

SAMPLE PREPARATION

Sample Solvent
Deuterium

have the same properties of the "light" solvents with the added bonus of having a substantially reduced proton signal (some solvents are > 99.99% deuterated). Deuterium is a spin 1 nucleus and hence has a reasonably short T1 compared to 1H permitting the use of rapid pulse-acquire measurement.

SAMPLE PREPARATION

Air sensitive
An

inert atmosphere or vacuum Special NMR tubes are commercially available for sealing samples under vacuum or an inert atmosphere alternative
to

attach a glass tube to the top of a regular NMR tube with a constriction Care must be taken to maintain concentricity.

SAMPLE PREPARATION

High Pressure Sample

can

be sealed in thick walled NMR tubes alternative

to

use shortsealed 4 mm tubes placed in a 5 mm tube A tiny piece of tissue paper in the bottom of the outer tube makes it less likely to break

SAMPLE PREPARATION

Solid and SemiSolid

Rotor Special
Teflon

Inserts

(white) and Kel-F (transparent)

Caps

SAMPLE PREPARATION

Solid and Semi-Solid

Red pepper washing with D2O, cutting to leaf cutting to size, size and rolling up and inserting into inserting into the the rotor rotor

INSTRUMENTATION

Continuous Wave NMR instruments

Similar in principle to optical spectrometers. The sample is held in a strong magnetic field, while frequency of the source is slowly scanned (in some instruments, the reverse is done).

Due to the lower maintenance and operating cost of cw instruments, they are still commonly used for routine 1H NMR spectroscopy at 60 MHz.

Fourier Transform NMR instruments

All frequencies in a spectrum are irradiated simultaneously with a radio frequency pulse. Following the pulse, the nuclei return to thermal equilibrium.

A time domain emission signal is recorded by the instrument as the nuclei relax.

A frequency domain spectrum is obtained by Fourier transformation.

The Pulse
Range of radiation: F 1/t Where: F-signal frequencey 1/t-bandwidth t-pulse duration