NUCLEAR MAGNETIC RESONANCE Nuclear Magnetic Resonance is the study of the properties of the molecules containing magnetic nuclei

by applying an external magnetic field. Electrons exhibit two types of motion ...the orbital motion around the nuclei and spinning motion. certain nuclei are considered to be spinning along an axis. Since the nuclei is charged particle, this results in development of magnetic field. The individual nucleons have a spin of but their orbital moments are quenched by the interactions present in the nuclei. A spinning charge generates a magnetic field along the spinning axis. Spin of each nucleon couple together leading to total spin angular momentum of I. where I = spin quantum number. The spin quantum number I: 1. fixed characteristic property of the nucleus 2. may be or multiple of . Orbital and spin angular momentum give rise to magnetic moments. If I>0, the nuclei can possess magnetic moment with a constant magnitude and orientation/ state that is determined by the value of mI where mI = -I to I leading to (2I+1) orientations. The following table can help to predict empirically the value of I : Mass number A even even odd odd Atomic number Z even odd even odd Number Of neutrons even odd odd even Spin I 0 Integral value Half- Integral value Half- Integral value
1 2

Examples
12

C,16O

H, 11B 14N,17O

H, 13C, 15N, 19F, 31P

The nuclear magnetic moment: mI. Hence = mI where = magnetogyric ratio (experimentally determined). Also for a spinning particle of charge e and mass m : = e/2m c. where c = velocity of light This model fails to predict the magnetic moment of proton which is found to be 2.79 times greater than the predicted value (Nuclear magneton). The modified expression for magnetic moment is thus, = g e/2mp g= nuclear g factor equivalent to Lande splitting factor. In an absence of an external magnetic field the (2I +1) magnetic states are at the same energy level. When the nucleus is subjected to an external magnetic field, the various magnetic states split according to their potential energy. Ez = - zB = - mI B where B = external magnetic field in the z direction.

m=-1/2

m=-1

E= B

m=0

E m=1/2 m=1 Ext B No Ext B Ext B

No Ext B

E = B= h (for transition to next higher mI ) Hence = B(1/2) where = Larmor frequency (lie in the radiofrequency region 106-109 Hz) . The distribution of the population will be given by Boltzmann distribution. N/ N = e (-E/ kT) Resonance is said to occur when the frequency applied equals the Larmor frequency and a strong absorption occurs as the spins make transition from to state. Thus Nuclear Magnetic Resonance is the resonant absorption of the radiofrequency by magnetic nuclei exposed to a magnetic field. When viewed classically, the nuclei when subjected to an external field, rather than aligning in the direction of the external magnetic field as a bar magnet, the nuclei precesses about the field direction with the preccessional frequency equal to Larmor frequency.

Simply increasing magnetic field increase the precessional frequency. To align the nuclei in the field direction second magnetic field is applied. The magnetic vector of the radiofrequency field oscillates perpendicular to the applied magnetic field and flips the nuclear spins at resonance. A voltage is induced which is amplified and recorded.

Block Diagram of NMR:

MAGNET COMPUTER

A stable, homogenous magnetic field is a prerequisite for NMR. The magnets used are given below:

PROBE PREAMPLIFIER RECEIVER DETECTOR TRANSMITTER

Features Field strength

Permanent 1.4T max 1. rugged and inexpensive 2.Temperature induced field drifts and hence not suitable for long spectral accumulations

Electromagnet 1.4-2.4T 1.Considerable space req, 2. resistive magnets and the current used causes considerable heating: hence stabilization req

Superconductor 12T or even greater 1. Nb-Sn-Tl alloys used at liquid helium temp ( 4.2K) in the form of solenoids 2. provides stable fields, economical, small size.

Comments

The sample along with the spinner is inserted from the top of the spectrometer and is taken down to the probe using the compressed air cushion. The probe consists of a radiofrequency circuit. The coil or inductor is wound and positioned so that the magnetic field generated by it is perpendicular to the magnets field. Helmholtz coil and solenoid coil are widely used. The probe is tuned by adjusting the current in the capacitors for maximum absorption of frequency which

influences the sensitivity. The signal from the probe needs to be amplified using a preamplifier. A broadband preamplifier amplifies signal over a very large frequency range whereas a tuned preamplifier detects a narrow band of intensity. The signal is digitized using analog to digital (Ato-D) converter.. There are two general types of NMR instrument; (a) continuous wave (b) Fourier transform Early experiments were conducted with continuous wave (C.W.) instruments, and in 1970 the first Fourier transform (F.T.) instruments became available and are now used widely. Continuous wave NMR instruments In the Continuous wave NMR spectrometers sample is held in a strong magnetic field, and the frequency of the source is slowly scanned (in some instruments, the source frequency is held constant, and the field is scanned). The signal producing precessing magnetization is a response to a weak oscillating transverse magnetic field. Fourier transform NMR instruments In FT-NMR, 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 major advantage of the FT-NMR is increased sensitivity. Recording multiple spectras and adding them together increases the sensitivity as noise is random phenomenon , and adds as the square root of the number of spectra recorded. For example, if one hundred spectra of a compound were recorded and summed, then the noise would increase by a factor of ten, but the signal would increase in magnitude by a factor of one hundred - giving a large increase in sensitivity. However, if this is done using a continuous wave instrument, the time needed to collect the spectra is very large (one scan takes two to eight minutes). Chemical Shift: Keeping in view the energy splitting in presence of an external magnetic field protons of a compound should resonate at the same frequency as an isolated proton. However this is not the case. Nuclear magnetic moments interact with the local magnetic field B loc. The circulating electrons lead to electronic orbital angular momentum which gives rise to a small additional magnetic field at the nuclei and is proportional to the applied field. Since nuclei of different chemical groups will have different electronic structure near the nucleus, hence these will experience different net magnetic field and will have different Larmor frequency. B exp= B appl+ Bloc Bloc B appl Bloc = - B appl Where = shielding contant and is different for different nuclei depending on electronic distribution.. Hence, B exp = B appl (1-) Since, = Bloc (1/2 )

Therefore, = B appl (1-) (1/2 ) Thus the resonance frequency of a nuclei varies with the chemical environment.

H DESHIELDED Eneg H <E neg X

H Eneg H >E neg X SHIELDED

The electrons can either shield or deshield the proton depending on the orientation. The resonance frequencies are expressed in terms of an empirical quantity called chemical shift = {(- 0)/ 0opp} * 106 where 0 = resonance frequency for tetramethylsilane (TMS)

The advantage of the scale is that shifts are independent of the applied field . Tetramethylsilane (TMS) is an internal standard used for calibrating chemical shift for 1H, 13C and 29Si NMR spectroscopy. The twelve hydrogen atoms in tetramethylsilane molecule are equivalent,a single peak appears which is assigned 0.0. Similarly, all four carbon atoms in a tetramethylsilane molecule are equivalent. In a fully decoupled 13C NMR spectrum, the carbon in the tetramethylsilane appears as a singlet, allowing for easy identification. The chemical shift of this singlet is also set to be 0.0 in the 13C spectrum, and all other chemical shifts are determined relative to it. TMS is highly volatile which allows easy recovery of sample and inert.it is soluble in organic solvents. Since TMS is insoluble in water other soluble salts are used. TSP (3- ( trimethylsilyl) propionic acid) is used as a reference material for aqueous solutions.

Hence the chemical shift values that are the peak positions help to determine the type of chemical group present. The intensity of these peaks helps to determine the number of protons of that group.

The Fine structure: The splitting of resonances into individual lines is called the fine structure of the spectrum. The resonance frequencies are modified by the influence of other nuclei on the local magnetic field of the proton. Measure of these interaction are the scalar coupling constant J (Hz) and these lead to the splitting of the peaks in the intensity ratio as given by the Pascals Triangle. 1 1 1 1 2 1 1 3 3 1 1 4 6 4 1 and so on.... In general n equivalent nuclei of spin split the peak of nearby nuclei into n +1 lines. This splitting helps to determine the number of neighbouring protons.
CH3CH2OH CH3 (t)

OH

CH2 (q)

The 13C NMR is similar to the proton NMR. This magnetically active isotope of carbon is only 1.1% of naturally occurring carbon. A similar set of information as in the number of the signals which indicate the different types of carbon , splitting of signals which indicate the H attached to the carbon and chemical shifts values indicate the electronic environment and hybridization of carbon can be obtained,. Normally a proton decoupled spectrum of carbon which shows no splitting is used.

2D spectroscopy: As the number and type of protons in a compound increases, the complexity of the spectra increases. Two-dimensional NMR spectra provide more information about a molecule than one-dimensional NMR spectra and are especially useful in determining the structure of a molecule, particularly for molecules that are too complicated to work with using onedimensional NMR. Here a second frequency dimension is introduced and a second Fourier transformation is applied on the FID obtained in this frequency. The resulting peaks are in the form of surface in three dimensional space which are sliced from the top and contour plot is obtained. Correlation spectroscopy COSY is one of several types of two-dimensional nuclear magnetic resonance (NMR) spectroscopy. If COSY is done using the same nuclei we have Homonuclear COSY spectroscopy. When done on different nuclei we have Heteronuclear COSY spectroscopy. Applications of NMR: The versatile NMR spectroscopy is an important tool in chemistry for the study of chemical structure. NMR is widely used for structural determination of compounds of both synthesised and natural products in additional to the one-dimensional NMR spectroscopy, two dimensional

approaches have been developed for the determination of the structure of complex molecules like polymers, proteins and nucleic acids. Time domain NMR spectroscopy is used to study molecular dynamics in solutions. Relaxation studies viz T1 and T2 studies are done in-vitro for evaluating MRI contrast agents. NMR of solid samples can help determine molecular structures. There are NMR methods for measuring diffusion coefficients. Nowdays, metabolic profiling is the emerging area in which serum , urine , saliva and other biological fluids are used. Proton NMR spectroscopy can provide details on the concentration and mobility of many low-molecularmass (Mr) metabolites in tissue and in the blood and plasma of several animal species,including humans. It can also be applied to the study of metabolite concentrations in plasma in disease states such as diabetes mellitus, renal failure, gingivitis etc. The advantage of the technique is that is non invasive, rapid, economical, requires only a small sample, potentially nondestructive to the samples and involves no separation or derivatization steps. The analytes studied can be detected down to about 100 micromol/L.

Ref: CLN. CHEM. 40/7, 1245-1250 (1994)

For extra reference please refer to: 1. 0rganic Chemistry, Morrison and Boyd. 2. Physical Chemistry, PW Atkins. 3. Nuclear Magnetic Resonance Spectroscopy, Frank A Bovey.

Experimental details: 1. Demonstration and Discussion of proton NMR of ethyl benzene. 2. Demonstration and discussion of C-NMR. 3. Demonstration and discussion of NMR of serum/urine ( biological fluid). 4. Demonstration and discussion of 2D NMR