Magnetic Resonance

Principal Forms of Magnetic Resonance Spectrometry

• NUCLEAR MAGNETIC RESONANCE (NMR)
  - spectrometry based on energy transitions of spinning NUCLEI in applied magnetic field. Radiofrequency radiation. The more important method
• ELECTRON PARAMAGNETIC RESONANCE (EPR)
  - spectrometry based on energy transitions of spinning ELECTRONS in applied magnetic field. Microwave radiation. Lesser importance

Nuclear Magnetic Resonance (NMR)
THEORY

(a) Magnetic moments of spinning nuclei - no external magnetic field

- - - - - - - - -> r.f absorbed <- - - - - - - - - r.f. emitted

(b) Possible energy transitions if magnetic field applied

NMR Spectrometer

Continuous wave NMR Spectrometer

8.3.1  How NMR Spectrometer Works

Absorption of r.f when energy gap corresponds to transmitter frequency

Resonant Condition.  NMR Signal received

Applicability of NMR

Common nuclei capable of giving NMR

Chemical Shift

An effect due to shielding by orbital electrons.
----> resonance field affected by chemical environment.

eg p-xylene

NMR Spectrum

Spin Coupling

Also called SPIN-SPIN SPLITTING

An effect due to SHIELDING BY NEIGHBOURING NUCLEI with magnetic moments.

eg. CH2ClCHCl2, or in full:	H Cl | | Cl-C-C-H     | | H Cl
Chemical shift gives --->
Then, spin coupling Triplet splitting of CH peak by combined spin of 2H's of CH2Cl	Doublet splitting of CH2 peak by spin of H of CHCl2
Overall spectrum

Basis of Peak Splitting by Spin Coupling
Equivalent protons normally don't split each other's resonance
Non-equivalent protons on adjacent C atoms cause splitting in accordance with the number of possible spin states of the nucleus that produces the splitting

CHCl2 2 spin states of H atom + 1/2 & - 1/2 ( & ) Thus the CH2Cl peak is split into a doublet CH2Cl 3 combined spin states of 2 H atoms + 1/2 + 1/2 net +1 ( & ) + 1/2 - 1/2 net 0 ( & ) - 1/2 + 1/2 net 0 ( & ) - 1/2 - 1/2 net -1 ( & ) Thus the CHCl2 peak is split into a triplet

8.7 Pulsed NMR with Fourier transform

Continuous wave NMR lacks resolution to analyse complex biological macromolecules.
For high resolution use Pulsed - FT NMR
Magnetic field strength is fixed, sample is subjected to pulse of r.f. containing wide band of frequencies. Protons are excited out of alignment with the magnetic field, then relax back to the aligned state.
Measure time decay of signal as nuclei relax

Apply Fourier transform maths -->

Advantages of pulsed-FT method:

• Higher resolution
• Opportunity for different types of NMR experiment by varying...
  duration & form of r.f. pulse, and
  interval between pulse & signal measurement.

    COSY Correlation Spectroscopy
    NOESY Nuclear Overhauser Effect Spectroscopy
  (information about through space interactions where atoms are near neighbours - elicit tertiary protein structure)

Applications of NMR in Biology & Biochemistry

• Spatial arrangements of atoms in macromolecules - tertiary folding of proteins.
  Complementary to X-ray crystallography, but gives results on molecules in solution.

• Identification analysis of small molecules -
  distinguish similar structural isomers (& sometimes stereoisomers)
  Note use of D₂O as solvent - exchangeable H atoms (bonded to O or N) replaced by D, which does not give NMR signal.

• Phosphorus NMR
  Follow reactions of important metabolites e.g. ATP, sugar phosphates in perfused living tissue. (abundant ³¹P gives NMR)

• Magnetic Resonance Imaging (MRI)

Magnetic Resonance Imaging (MRI)

Medical diagnostic technique - imaging of soft tissue (contrast X-ray for imaging of hard tissue). Requires strong homogeneous magnetic field over large space.

Locates protons, typically in body H₂O & in lipids, can give information on physical state (e.g whether blood flowing normally) and chemical environment (ability to distinguish tumour from normal tissue).