nmr arises from the fact that certain atomic nuclei have a property called “ spin ”
DESCRIPTION
Nuclear Magnetic Resonance (NMR). Nucleus of NMR Active atom. Static Magnetic Field Generated by the NMR Spectrometer. NMR arises from the fact that certain atomic nuclei have a property called “ spin ” - PowerPoint PPT PresentationTRANSCRIPT
NMR arises from the fact that certain atomic nuclei have a property called “spin”
“Spin” is caused by circulating nuclear charge and can be thought of as bar magnet that adopts a preferred orientation in the presence of a static magnetic field
Static Magnetic Field Generated by the NMR Spectrometer
Nucleus of NMRActive atom
NMR spins adopt specific quantitized states; for spin ½ nuclei, the type of nuclei most commonly studied, there are two such states, and .
corresponds to preferred low energy configuration, while corresponding to the non-preferred high energy configuration.
Nuclear Magnetic Resonance (NMR)
Energy can be applied by the NMR spectrometer in the form of an oscillating magnetic field at a precisely defined frequency (and hence energy) that induces transitions from to .
Oscillating magnetic field is applied through the probe, which basically consists of a coil surrounding the NMR tube attached to a user controlled oscillating frequency source on the spectrometer console.
Nuclear Magnetic Resonance (NMR)
NMR Chemical Shifts
= physical constant for a given type of nucleus (ratio of magnetic moment and angular momentum)
h = Planck’s constant
Bo = static magnetic field strength
Prediction, based on the fact that all nuclei in the sample are placed in the same magnet, is that all nuclei of one type (1H,
13C, 15N, 31P etc) would have exactly the same NMR frequency
Predictions Do Not Match Reality
= chemical shielding tensor
Frequency
Ile in D2O
12
1
HDO
121
(Acquisition time = 30 s)
Chemical Shielding
Shielding arises from the various ways by which electrons“shield” the nuclear spin from the external magnetic field (Bo)
Physical mechanism relates to induced circulation of electrons that oppose static magnetic field (Lentz’ Law)
Shielding (tensors) can be determined through ab initio calculations. This, however, is computationally expensive, and not routinely applied to large molecules, such as proteins.
Classic Approaches to Shielding Local electronic structure; electronegativity of attached groups, bond lengths, bond angles, and conformation (dihedral angles)
Anisotropy of local groups (circulating electrons from aromatic rings for example)
Hydrogen bonds
Electric field effects that polarize bonds
Chemical Shielding Trends for Protons
Functional Groups
Proteins
Frequency
Frequency
Chemical Shifts Can Change Dramatically with Changes in Conformation
8 M Urea
No Urea
Chemical Shielding & Chemical Shifts
Recall
Bo field dependence of frequency makes comparison of spectradifficult from one instrument to another
Hence, report relative ’s, not absolute ’s
Chemical Shift (ppm) = =
peak = frequency of signal of interestref = frequency of reference signal
IUPAC-IUB Shift Standard for Proteins
Sodium-2,2-dimethyl-2-silapentane-5-sufonate (DSS)
J-coupling
Ha
Hb
J-couplings in Ile
Ile in D2O
1
2
1
1
2 1
1
NMR Active Nuclei in Biomolecules
Sensitivity of NMR
& spin states willassume a Boltzman distribution
Implications: Highest sensitivity w/ higher & higher Bo
1D 13C Natural Abundance Spectrum of Ile
Ile in D2O (1H Decoupled)
(Acquisition time = 4 hr)
1
2 1
13C ppm13C ppm
112
CO
13C ppm