See also: NMR

The nuclear magnetic resonance is a technique of Spectroscopie applied to the particles or atomic whole of particles which have a nuclear Spin not no one.

It is a phenomenon by which a core of the atom considered absorbs the electromagnetic radiations of a specific Fréquence in the presence of a fort Magnetic field. Isidor Isaac Rabi discovered this phenomenon in 1938. Magnetic resonance, thereafter, was applied to the detection of the light Atome S such as the Hydrogène.

Its applications relate to the Physique, the Chimie and the Medical imagery.

Principle

In this method one uses the Spin cores of the atoms. Certain cores have a null spin (those of which the number of Proton S and Neutron S are all the two pars) but others have a spin nuclear different from zero, which implies that one can associate a to them magnetic Moment of spin which behaves like a magnetic Moment (i.e a kind of small magnet). The atoms of Carbon 12 and Oxygène 16 are very widespread but their nuclear spin is null. On the other hand the Hydrogène has only one proton and its nuclear magnetic moment is thus nonnull: the magnetic resonance of hydrogen (of the proton) is used. It is in particular important to point out that the nuclear adjective employed here has no relationship with the phenomena of Radioactivité, but referred right to the Atomic nucleus.

The quantum physics teach us that one magnetic moment of 1/2 placé spin in an external magnetic field can have 2 possible energies (2 energy levels). One can for example note it in experiments for electrons, it is the Effet Zeeman. The NMR consists in modifying the nuclear moment magnetic, in other words to make pass the core of an energy level to another, (what amounts “turning over” the spin) by absorption of a photon: when the energy of the photon (and leaving the frequency the electromagnetic wave) allows this transition there is resonance . For the usual fields (about the Tesla) the resonance of the proton takes place in the field of the Ondes radio (100  MHz approximately): 42 MHz in a field of 1,0 T and 63 MHz in a field of 1,5 T.

The mathematical relation existing between the magnetic field imposed of standard $B\_\; \{0\}$ and the frequency of resonance (reversal of spin) $\backslash \; nu$ is very simple:

$\backslash \; nu\_\; \{0\}\; =\; \backslash \; gamma\; \backslash \; times\; \{B\_\; \{0\}\; \backslash \; over\; 2\; \backslash \; pi\}$

where $\backslash \; gamma$ is the gyromagnetic Rapport characteristic of each studied core.

Thus the following table gives the values of $\backslash \; gamma$ for the most current cores:

One sees thus that the frequency of the electromagnetic wave necessary for the resonance of the proton is approximately 4 times higher than that necessary for the resonance of 13 C.

The transition from the spin towards its return to balance (relieving) involves the emission of an electromagnetic wave which can be detected by a sensor.

In Imagery by magnetic resonance (IRM), one thus measures the answer of each Voxel (basic volume of the examined sample), determining the density in protons and reconstituting an image.

History

• In 1938, Isidor Isaac Rabi discovers the phenomenon of magnetic resonance on molecular jets.

• In 1946, Felix Bloch and Edward Mills Purcell specify the concept of frequency of resonance.
• It is under the term of zeugmatomography ( zeugma being a Greek term meaning “union”) that its application in imagery, created in 1973 appeared by Paul Lauterbur, Nobel Prize of physiology and medicine in 2003 for this invention.

Uses

The nondestructive character of this analytical technique led to various developments of this method which from now on is employed in medicine to study the human body (IRM), or in Organic chemistry to carry out structural analyzes.

It is a tool of Biophysique very much used into genomic structural to obtain a “image” in 3D of the molecules of the alive one.

NMR in organic chemistry

The NMR is the tool for analysis currently more used in Organic chemistry. It makes it possible to obtain qualitative or quantitative information on the sample analyzed, according to the technique employed. The cores most often studied are the ¹H, the ¹³C, the ¹⁷O, the ³¹P and the ¹⁹F which present, for the majority, a nuclear spin not no equal to 1/2. The ¹⁴N as for him presents a nuclear spin equal to 1 while that of oxygen is of 5/2.

The sample to be analyzed is put in solution in a deutérié solvent (see ²D, a Isotope of the ¹H presenting a nuclear spin equal to one). This solvent, generally of the Chloroform deutérié, (CDCl₃) is normally invisible in NMR of the proton, since deuterium has a frequency of resonance quite different from that of hydrogen (the gyromagnetic report/ratio is worth 6,54 in the case of MHz/T Deuterium).

Let us consider the NMR with the ¹H: chemical environment of the hydrogen atoms which are connected chemically to the molecules of the influential sample on the frequency of resonance of those; thus, the hydrogen of a grouping alcohol (- OH) $\backslash \; naked\; 1$ will have a frequency of resonance higher than that of the hydrogen of a grouping carboxyl (- COOH) $\backslash \; naked\; 2$. As the difference between these two frequencies is of some Herz, the chemists defined another size: the chemical shift ( chemical shift ), and refers at a frequency of reversal of spin standard, that of the tétraméthylsilane (TMS), which one introduces into the sample.

The relation between the frequency of resonance $\backslash \; naked\; 1$ and the chemical shift corresponding $\backslash \; delta\; 1$ is given by following calculation:

$\backslash \; delta\_1\; =\; 1000000\; *\; (\backslash \; naked\; TMS\; -\; \backslash \; nu\_1)/\backslash \; naked\; TMS$

because $\backslash \; naked\; TMS$ is higher than the frequency of resonance of the majority of the types of protons.

In the quoted cases, the chemical shift is worth approximately 4 for RO- H , and 11 for R-COO- H .

The NMR of the ¹H can be relatively fast (order of idea: 2 min) and allows an easy quantitative analysis. Thanks to the interpretation of nature of the solid masses obtained (bytes) and with the empirical knowledge of the chemical shifts of the protons present in each functional grouping, it is possible to determine the developed structure of all the organic molecules per application of a simple logical reasoning.

The NMR of the ¹³C makes it possible to find all carbons of the molecule, grace there too to the empirical knowledge of the chemical shifts of carbons belonging to various functional groupings. The recent apparatuses make it possible to obtain quickly the spectra NMR ¹H and ¹³C, those with or without decoupling of the proton.

NMR in medical imagery and biophysics

The imagery by nuclear magnetic resonance (IRM) is a technique of Medical imagery making it possible to have a sight 2D or 3D of part of the body. This technique is very useful for the observation of the Cerveau. Thanks to the various sequences (Sequence IRM), one can observe various fabrics with very high contrasts because the resolution in contrast is definitely better than that of the scanner.

See the article Imagery by magnetic resonance

NMR in structural biology

Beside X-ray crystallography, the NMR became a method of study of the biological macromolecules in solution. It does not require obtaining monocrystals and makes it possible to study proteins, nucleic acids with concentrations millimolaires. The multidimensional techniques of NMR result in correlating the frequencies of several spins and to solve ambiguities related to the spectral superpositions. Proteins of molecular mass from 10 to 30 kDa can be analyzed as well as oligonucléotides several tens of pairs of bases.

NMR homonucléaire without isotopic marking

Historically, the proteins were studied by the NMR of the proton (isotope ¹H) present in abundance. A first stage consists in allotting resonances, i.e. to establish a correlation between the signals of the spectrum and the hydrogen atoms of the molecule. Two key experiments are used, the experiment of correlation by the scalar couplings ( HOHAHA or TOCSY ) and the experiment of correlation through space by effect Overhauser ( NOESY ). This attribution is known as sequential, because it operates by relative positioning of a core compared to its neighbors by using the information of the peptide sequence. This process of attribution (similar to a Puzzle) becomes increasingly complex as the size of protein increases; moreover, the Overhauser effect is transmitted through space and does not allow to distinguish from the close cores in the peptide sequence and those close in space. Errors are thus possible which are detected only at the end of the process, when certain parts of the puzzle remain on the carpet.

Once the allotted spectra, information are then used in a quantitative way: the scalar couplings inform about the plane angles and the Overhauser effects on interatomic distances (up to 4-6 Angströms). This information is introduced into programs of molecular modeling to seek one or more conformations of the molecules compatible with the data. The strategy is comparable with that of the geometrician who measures distances and angles between buildings and recomputes a plan of city. Except that the range of the distances measured in NMR is weak compared to the evaluated objects; the accumulation of the experimental errors and/or the low number of data led to badly definite structures locally.

Two difficulties limit the size of the macromolecules étudiables: the complexity of the spectra and the individual width of each signal. If the size of double protein, the number of resonances in the spectrum will double without dispersion (i.e. the spectral width) not increasing. A solution consists in resorting to a spectrometer NMR with higher field… and thus much more expensive. If the double size, the molecular mass increases and the protein rotates more slowly by diffusive movement. That led to broader signals, because transverse relieving becomes more powerful. The increase in the magnetic field is without effect… and it is necessary to find another parade with the problem (reduction of the Viscosité, reduction of the number of close protons…).

NMR hétéronucléaire with isotopic marking

In order to solve the spectral superpositions in the large molecules, it proved to be necessary to pass from the NMR 2D to the NMR 3D. Not very profitable tests were carried out with the beginning of the year 1990 to combine sequences HOHAHA and NOESY in a three-dimensional experiment. If one has a protein entirely enriched in ¹⁵N and ¹³C, one can conceive experiments of correlation between spins, only based on couplings scalar and allowing to connect all the peptide skeleton as well as the side chains. The naturally abundant Isotopes are respectively the ¹⁴N ( quadrupolar core ) and the ¹²C ( invisible core in NMR ), but the majority of proteins being obtained by bacterial surexpression, it are possible to make cultures on mediums enriched isotopiquement.

This new strategy calls upon a series of experiments 3D triples resonance: 3D because a three-dimensional spectrum is obtained, triple resonance because the frequencies of three different cores are detected. For reasons of sensitivity, all these sequences leave the ¹H (core with the high gyromagnetic report/ratio) and end in the detection of the same core. In the experiment HNCO , one thus establishes a correlation between the proton amide (^HN), his nitrogen (NR) and the carbonyl (CO) of the preceding amino-acid. The scalar couplings used for the transfers of coherence are couplings ¹J (with a connection) and thus relatively important (¹JNH = 90 Hz and ¹JNCO=15Hz). That thus ensures a great effectiveness of transfer even in the case of high proteins of mass (width of line). To note that this experiment HNCO makes it possible to connect by scalar coupling of the amino-acids, which the strategy homonucléaire did not allow ( absence of scalar coupling 3J through the peptide connection )

See too

Internal bonds

External bonds

• Abbreviations and Acronymes used in NMR

• French Grouping of study of magnetic resonance (GERM)
• course in line
• CARA Computer Aided Assigment Resonance, free software.
• http://www.cis.rit.edu/htbooks/mri/inside.htm