Nuclear Magnetic Resonance

Nuclear Magnetic Resonance PRINCIPLES OF NMR SPECTROSCOPY Contents Principles of nuclear magnetic resonance The nmr spectrometer Basic principles in nmr application NMR tools used to obtain information Nuclear spin state Electric quadrupole and nuclear magnetic moment Nuclear magnetic resonance Energy consideration of the precessing nucleus NMR experimentation and data acquisition Operation the NMR machine Sample preparation Deuterated NMR solvents 1

NMR Spectroscopy Objectives To explain the principles of nmr as regard to nuclear spin To Describe the spinning of nucleus as it interacts with applied magnetic field and radiofrequency. To elaborate on how a 1 H-nmr spectrum is generated To explain nuclear spin magnetization and relaxation in the course of data acquisition To distinguish absorption (= residual peaks) of various solvents used in 1 H-nmr spectroscopy 2

Principles of nuclear magnetic resonance Spectroscopic technique which uses the longer wavelengths (radio frequency) absorption to give information about number, connectivity of each type of nuclei (e.g hydrogen/carbon) and the nature of its chemical environment. This low energy radiation affects only molecular vibration and nuclear spins. 3

Principles of NMR The energy transition in NMR: for electron spin, the change in energy ( E ) corresponds to energy in microwave region for nuclear spin, the E corresponds to the energy of radio waves. 4

The NMR spectrometer The NMR machine is designed as a strong, big gascylinder like with two gas compartments, and shim tube running from top through the centre of the cylinder to the bottom The lower part of a shim tube which is surrounded with main coil for supplying electric field and magnetic field. 5

The NMR spectrometer.. The helium dewar is surrounded by a liquid nitrogen dewar to reduce loss of the helium which is very expensive. The radio frequency are sent in and out through the probehead to the processor 6

7

The NMR spectrometer.. An NMR spectrometer showing; (a) Opening to probe, (b) Vacuum-jacketed cryostat (c) Solenoid coil of superconducting alloy, suspended in liquid Helium (d) Lines of force project several feet into Since the NMR machine is all about powerful magnetic field, the line of forces projects several feet into the room which are concentrated at the top and bottom. Thus, working with iron objects like watches can be grabbed iron objects and accelerate. Also people with medically inserted magnetic sensitive materials are advice not to work close to the NMR facility. the 11/13/2014 room. Organic Spectroscopy 8

Basic components of an NMR spectrometer Powerful magnet radio frequency generator radio frequency receiver, Sweep coils and Sample tube 9

Schematic representation of an NMR spectrometer 10

Sample NMR tubes These are special-purpose glass tubes that are manufactured to certain specifications of highquality glass that holds the sample. For evenly spinning avoid breaking allowing proper absorption of RF by the sample. 15

Basic principles in NMR applications Experiments on NMR involve absorption of radiofrequencies on electromagnetic spectrum, of which the signals are recorded as the function of frequencies. The plot of applied radiofrequency against absorption frequencies (usually in ppm) on a chemical shift scale is called an NMR spectrum. 16

Basic principles in NMR applications A signal in an NMR spectrum is referred to as resonance. 1 H and 13 CNMR spectroscopic experiment gives the number, type and connectivity of hydrogen and carbon atoms in a molecule, respectively. However, the only sensitive isotopes that are being detected are protons and carbon-13s 17

QN. Account for types and number of protons and carbon-13 of ethanol and pent-4-en-2-one, respectively H 3 C H 2 C O H H 3 C O C CH 2 CH CH 2 types of H atoms = 3 Number of H atoms= 6 types of C atoms= 5 Number of C atoms = 5 NB: The NMR spectra gives information about the nature of the chemical environment of each magnetically active nucleus in the molecule. 18

NMR tools used to obtain information There are three primary NMR tools used to obtain information (1) Chemical shift which are concerned with local nuclear environment, (2) Coupling constant (J) concerned with torsion angles and (3) Nuclear overhauser Effect (NOE) concerned with internuclear distance 19

Two variables that characterize NMR (1) an applied magnetic field,(β o ) and (2) the frequency of the radiation used for resonance (in MHz) (To be discussed later) Therefore, the NMR spectroscopy is explained under the consideration of three phenomena, the property of the nucleus: Nuclear spin state Nuclear magnetic moment Nuclear magnetic resonance 20

Nuclear spin states NMR spectroscopy technique is only sensitive to any atomic nucleus which possesses either odd mass number or odd atomic number or both and thus has a quantized spin angular momentum and a magnetic moment ( ). Proton and carbon-13 have the spin quantum number l = ½ and has two allowed spin states, +½ and ½. 21

Nuclear spin states Spin quantum number Recall that l is a physical constant for each nucleus, and there are 2 l + 1 allowed spin states which range with integral differences from + l to - l that is - l (-l + 1),, (l 1), l. For the proton, allowed spin state = 2 l + 1 then, 2(½) + 1 = 2 allowed spin states 22

Electric quadrupole moment Electric Quadrupole Moment (eq) is a parameter which describes the effective shape of the ellipsoid (non-spherical) of nuclear charge distribution. Usually, particular nuclei may have spherical charge distribution or non-spherical (i.e ellipsoid: either prolate or oblate) charge 23

Illustration of spherical charge and no-spherical nuclear charge distribution Spherical charge distribution Prolate charge distribution 24

eq moment A non-zero quadrupole moment indicates that the charge distribution is not spherically symmetric. That is, all nucleus with l = 0 or l = ½ have approximately spherical charge distribution within their nuclei hence small electric quadrupole moment. Whereas those with l > ½ have non-spherical (ellipsoidal) charge distribution within their nucleus hence large electric quadrupole moment (eq). 25

eq moment Thus, all nuclei with non-zero spin (l >0) have magnetic moment, μ. Therefore, conventionally the value of eq is taken to be positive if the ellipsoid is prolate and negative if it is oblate 1 H 12 C 16 O 19 F eq negligible 2 H < 14 N << 35 Cl, 37 Cl << 79 Br, 81 Br 127 I Increasing eq (absolute value) 26

Nuclear magnetic moment The intrinsic magnitude of the generated dipole is expressed in nuclear magnetic moment, μ. The magnetic moment is generated by the charge and spin of a charged particle. The nuclear magnetic moment varies from isotope to isotope of an element and it can align with an externally applied magnetic field of strength B o in only 2 l +1 ways, either re-enforcing or opposing B o. So, nuclear magnetic moment can only be zero if the numbers of protons and of neutrons are both even. 27

Nuclear magnetic moment.. A characteristic of the collection of protons and neutrons (which are fermions) is that a nucleus of odd mass number, A will have a half-integer spin and a nucleus of even A will have integer spin. If the number of neutrons and the number of protons are both even, then the nucleus has NO spin. e. g If the number of neutrons + protons is odd, then the nucleus has a half-integer spin of, for, and, respectively. If the number of neutrons and the number of protons their sum is odd, then the nucleus has an integer spin (i.e. = 1 28

Spin state of some isotopic elements with respect to atomic number and atomic masses Nuclear spin (l ) Half integer l = 1/2, 3/2, 5/2, Integer l = 1 For zero I = 0 Have Have Have Odd Atomic mass Odd or Even Atomic number Even Atomic mass Odd Atomic number Even Atomic mass Even Atomic number l = : 1 H, 13 C, 19 F l =1: 2 H, 14 N l = 0: 12 C, 16 O l = : 11 B l = : 17 O 29

NMR phenomenon Applied magnetic field, circulation of electrons and induced field Local diamagnetic current & Diamagnetic anisotropy Precessing nucleus 30

Applied magnetic field, circulation of electrons and induced field Many moving charge (like proton nucleus) generates a magnetic field of its own and a weak secondary field (B sec ) due to movement of electron around it that opposes applied field and therefore shields the proton nucleus. Hydrogen nucleus may have +½ (α-spin state) or ½ spin (β-spin state) with magnetic moment, μ. B o B i e - e - B o 31

Precessing nucleus To understand this phenomenon, we need to consider hydrogen nuclei as a tiny bar magnet having North pole (N) and South pole (S) when placed in magnetic field. Thus, like-poles do repel each other and opposite poles attract each other. In this look, like-poles repel each other, thus in the spin states it s the - spin state which is raised at higher energy. 32

Illustration of proton like a small magnetic bar placed in a magnetic field. 33

Precessing nucleus... When the frequency of the oscillating electric field component of the incoming radiation just matches the frequency of the electric field generated by the precessing nucleus the two fields can couple, and energy can be transformed from the incoming radiation to the nucleus thus causing spin change. This situation is called Resonance. Spinning proton has electron which also do spin and both generate magnetic field. Therefore, the effective field (B eff ) is the difference between B o and B sec 34

Spin flip Alignment of proton spin states when an external magnetic field, B o is applied. Aplied magnetic field, B o spin + ½ aligned to B o spin - ½ oppose B o 35

Energy consideration of a precessing nucleus 36

Energy consideration... spin state Rel. energy E 0 E spin state B x Increasing strength of applied magnetic field, B o 37

spin state Rel. energy E 0 E B x Increasing strength of applied magnetic field, B o spin state Energy consideration... From the illustration of the precessing nucleus with an angular velocity is given by the below expression; o = o Where o is the precessional frequency also called Larmor frequency and is the magnetogyric ratio (a nuclear constant which is the ratio between magnetic moment and angular momentum). So the magnetogyric ratio relates the magnetic moment and the spin number l for any specific nucleus 38

spin state Rel. energy E 0 E B x Increasing strength of applied magnetic field, B o spin state Energy consideration... o = o = 2 ν ν = ( / 2 ) o E = hν E = (μ/l) o?? μ l μ = (h /2 ). l But ν = (μ /h. l) o so, ν = ((h /2 ). l /h. l) o ν = ( / 2 ) o Search for various plausible derivations 39

spin state Rel. energy E 0 E B x Increasing strength of applied magnetic field, B o spin state Energy consideration... The difference in energy, E between the two states is dependent on strength of the applied field, B o. The charge distribution which is slightly excess in state (a lower energy level) is explain by Boltzmann charge distribution. 40

spin state Rel. energy E 0 E spin state Energy consideration... Note: B x Increasing strength of applied magnetic field, B o When the rate of precession equals the frequency of the rf radiation applied, the absorption of rf radiation take place and the nucleus spins either with +1/2 or -1/2 Firstly, sample is placed in the magnetic field then irradiated with radiofrequency radiation. When the frequency the frequency of radiation satisfies the equation of angular velocity then magnetic component of the radiant energy becomes absorbed. Also RF may be kept constant and vary B o absorption vs frequency of can be applied The term resonance is due to excitation to the nucleus to the high energy then decayed to the ground state 41

spin state Rel. energy E 0 E B x Increasing strength of applied magnetic field, B o spin state Energy consideration Note: This equation ( E = (hγ/2π)β o ) have to be modified due to secondary magnetic field by the nucleus i.e ν = β o (1- d)/2π; d is shielding factor 42

Chemical shift Chemical shift is the frequency of signal on an NMR spectrum where the peak occurs Atoms in different chemicals environment will resonate at different frequency and will appear at different chemical shift The chemical shift scale is calibrated such that the frequency point is tetramethylsaline (TMS) 43

Why TMS? CH 3 Si H 3 C CH 3 CH 3 Since silicon is less electronegative than carbon, TMS protons are highly shielded and its signal defined as zero. Organic protons absorb downfield (to the left) of the TMS signal It s is inert to most organic samples (in old technique) 44

Chemical shift scale Chemical shift scale can be expressed in two ways frequency in Hz Frequency in ppm (expressed as d) Chemical shift in Hz is dependent upon magnetic strength, B o Whereas chemical shift in ppm (d) is independent of magnetic strength, B o There are different of NMR machines of different strength, say 60, 90, 100,.900 MHz) 45

Chemical shift scale Protons with high electron density are said to be shielded (signal appear upfield) Whereas those with low electron density are said to be deshielded (signal appear downfield) 46

Illustration of 1 H NMR spectrum of methanol 7 6 Atoms in different chemicals environment will resonate at different frequency and will appear at different chemical shift 5 H 4 3 2 less shielded lower field (downfield) H O C CH 3 H more shielded high field (upfield) 1 0 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 Increasing magnetic field strength (B o ) 47

Chemical shift expressed in Hz Chemical shift can be expressed in terms of Hz by setting the TMS peak at 0 Hz. When chemical shifts are given in Hz the applied magnetic frequency must be specified i.e 60, 90,... MHz because the chemical shift in Hz is directly proportional to B o and therefore to the applied radiofrequency 48

Chemical shift in Hz... The value of chemical shift, n in Hz is; n (in Hz)= n s n r The number of Hz shift from TMS for a given proton (or nuclei) will depend on the strength of B o In 100 MHz the shift in Hz from TMS is 5/3 larger than at 60 MHz. Thus, the resonance proton in an applied field of; 14,100 G magnetic strength is at approximately 60 MHz where 100 MHz is at field strength of 23,500 G. The SI unit Telsa (T) is the unit of measurement for magnetic strength, B, replacing the term Gauss (G). i.e 1T = 10 4 G 49

Chemical shift in Hz... Therefore the ratio of the resonance frequency is the same as the ratio of the two field strength. 100 MHz 60 MHz 23,500 G = = 14,100 G 5 3 50

Dependence of chemical shift in Hz on magnetic strength Reich, Chem 345,Univ. Wisconsin, Madison 51

Chemical shift expressed in d (ppm) Usually chemical shifts are expressed in d unit, It is independent of the field strength (instrument used) due availability of instruments with varying frequency (60, 90, 300,... MHz). 52

Chemical shift in d d is a proportionality and thus dimensionless. Therefore, chemical shifts in Hz are converted into d unit as shown below: Chemical shift, d or ppm) = Observed shift from TMS in Hz Spectrometer frequency in Hz x 10 6 For 60 MHz instrument has an oscillation frequency 6 10 6 Hz. The factor 10 6 is included in the equation to avoid fractional values, since d is expressed in ppm. 53

Chemical shift in d... Independence of Chemical shift in d on B o a peak at 60 Hz (n 60) from TMS at an applied frequency of 60 MHz would be at d1.00 or 1.00 ppm. the same peak of the same proton at 100 MHz would be at 100 Hz but would still be at d1.00 or 1.00 ppm. d (or ppm) 60 = = 1.00 60 10 6 d (or ppm) 100 = = 1.00 100 10 6 54

Problem 1 What would be the chemical shift in d of a peak that occur 655.2 Hz downfield of TMS on a spectrum recorded using a 90 MHz spectrometer? Chemical shift, d or ppm) = Observed shift from TMS in Hz Spectrometer frequency in Hz x 10 6 Solution: d (or ppm) = [655.2 Hz/90x10 6 Hz] 10 6 = 7.28 ppm 55

Problem 2 A specific proton in an organic compound has a chemical shift of 3.4 ppm in a 60 MHz NMR spectrum. What will be the chemical shift in ppm if the spectrum is recorded using a 90 MHz instrument? 56

Solution The chemical shift in d unit express the amount by which a proton resonance is shifted from TMS in ppm of the spectrometer basic operating frequency. Hence the value of d for a given proton will always be the same irrespective of whether the measurement was made at 60 MHz or at 90 MHz. So the peak of that proton will be observed at 3.4 ppm. 57

Problem 3 (1) What does the term dispersion mean in NMR spectroscopy? (2)Two signals occur at 2.1 and 2.3 ppm in the proton spectrum in a spectrometer operating at 200 MHz for 1 H. (i) What is the frequency difference between the resonances in Hz? (ii) What is their frequency difference (in Hz) in a spectrometer operating at 600 MHz for 1 H observation? 58

Solution (1) Dispersion is a term used to express the notion of how well resonances in an NMR spectrum are separated from one another (in Hz) it is a qualitative term expressing the ease with which signals can be distinguished. 59

Solution...(2) i) Frequency difference between the resonances in Hz at 200 MHz Chemical shift = {Frequency of signal - Frequency of reference (Hz)} divide by spectrometer frequency (in Hz) At 200 MHz : 1ppm = 200 Hz, 0.1ppm = 20Hz Difference in Hz = 0.2 ppm = 40Hz 60

Solution... (ii) Frequency difference between the resonances in Hz at 600 MHz. At 600 MHz: 1 ppm = 600Hz, 0.2 ppm = 120 Hz 61

Steps to consider in interpreting 1 HNMR spectrum 1. How many type of H s? :This indicated by how many groups of signals are there in a spectrum 2. What type of H s?: Indicated by the chemical shift of each group. e.g shielded or deshielded, CH, CH 2, CH 3 with respect to chemical environment and multiplicity 3. How many H s of each type are there?: Indicated by integration (relative area of signal for each group) 4. What is the connectivity?: Look at the coupling patterns. This tells you what is next to each group 62

1 H NMR for Benzyl acetate and Phenylacetone 8 H (c) 7 H H 7 6 H H H 6 O CH 2 (b) H H 5 C 5 CH 3 (a) O H H 4 PPM 4 PPM O CH 2 3 3 CH 3 (a) 2 2 1 1 0 Similarity Both phenyl acetone and benzyl acetate have δ = 7.3 ppm. Methyl groups attached directly to a carbonyl have resonance at δ = 2.1 ppm. Aromatic protons characteris-tically have resonance near 7-8 ppm Acetyl groups (methyl group of this type) resonance ~2 ppm. 63 0

1 H NMR for Benzyl acetate and Phenylacetone 8 H (c) 7 H H 7 H H 6 H 6 O CH 2 (b) 5 H H CH 3 (a) C O 4 PPM H O CH 2 H 5 4 PPM (a) 3 2 CH 3 3 2 1 1 0 0 Difference Resonance of the benzyl (- CH 2 -) protons comes at higher value of δ = 5.1 ppm in benzyl acetate than phenyl acetone δ = 3.6 ppm. Reason? Been attached to an electronegative element (oxygen atom) these electrons are more deshielding than those in phenyl acetone 64

Note: The higher the electron density the higher the shielding hence the slow the resonance. The higher the shielding the more the external energy is required. 65

Working out integration on Proton NMR The intensity of an 1 H-NMR signal is proportional to the number of proton of each type in the molecule. The integral measures the area of the peak and gives the relative ratio of the number of H s for each peak. An FT-NMR instrument (to be discussed later) digitally integrates each signal area and provides ratios of number of hydrogen for each signal. 66

Two ways to determine the # of H atoms under each signal from the measured heights. 1. For a known molecular formula Proton per unit is determined by taking total number of proton divide by total units. # of Hs in signal = height of integral total heights of all integrals total # of Hs 2. For unknown molecular formula Integrate by determine the ratio between the signals and round off to a nearest whole number or multiply by a factor to produce a whole number x 67

Example Known Molecular formula (C 11 H 16 ) signal 1 (8.8 units), signal 2 (2.9 units), signal 3 (3.8 units). Proton per unit is determined by taking total number of proton in a molecule divide by total units of all signals Number of protons per signal can be determined by multiplying the number of protons per unit by number of units per signal. #of H s in signal 1 = 1.03 H per unit 8.8 units 9.0 H. In the same way signal 2 = 3 H and signal 3 4H 68

Example Unknown molecular formula signal 1 (8.8 units), signal 2 (2.9 units), signal 3 (3.8 units). Assuming the previous case the molecular formula in not known then we need to use integral to determine the ratio between the signals and round off to a nearest whole number or multiply by a factor to produce a whole number i.e ratio of units in each signal to a nearest whole number ; 8.8:2.9:3.8 = 9.0:3.0:4.0 for signal 1, 2 and 3, respectively. 69

Example (Multiplicity will be discussed later) 70

Given formula C 5 H 10 O Solution Signal units (units total units) Total Hs # of Hs (1) 2 (2 /10) x 10 = 2H (2) 3 (3 /10) x 10 = 3H (3) 2 (2 /10) x 10 = 2H (4) 3 (3 /10) x 10 = 3H Total 10 10 H Calculating DBE = [(2C+2+N)-(H + h)]/2 =1; May be 1 double bond or a ring. Possible structure H 3 C (4) H 2 C 71 (3) C H 2 (1) O CH 3 (2) pentan-2-one

72

Problem 4 The line of integration of the two signals in the 1 H-NMR spectrum of a ketone of molecular formula C 7 H 14 O raises 62 and 10 chart divisions, respectively. Calculate the number of Hydrogen atoms giving rise to each signal and propose a structural formula for this ketone. 73

Problem-5 Following compound I and II are constitutional isomers of molecular formula C 6 H 12 O 2. O O I O Predict the number of signals in the 1 H-NMR spectrum of each isomer Predict the ratio of areas of the signals in each spectrum Show how you can distinguish these isomers in the basis of chemical shifts Draw and label the proposed 1 H-NMR for each. Predict IR absorption frequencies of major FG s and describe their spectrum O 74 II

Summary 1 H NMR chemical shift 1. Stronger magnetic fields B o cause the instrument to operate at higher frequency (v) 2. NMR field strength vs 1 H operating frequency 1.41T => 60 MHz; 2.36T => 100MHz; 7.06T => 300 MHz 3. The d unit has been criticized because d values increased in the downfield direction. These are really negative numbers. Other scales are expressed in t values; t = 10.00 - d. 4. d unit is treated as positive number. Shifts at higher fields than TMS are rare, that is, if d = -1.00. Then, t = 10.00- (-1.00) = 11.00. 3. Each ppm unit represents either a 1 ppm change in B o (magnetic field strength, Tesla) or a 1 ppm change in the processional frequency in (MHz) 75

Summary 1 H NMR chemical shift 6. The shift observed for a given proton in Hz also depends on the frequency of the instrument used. The Higher frequencies the larger shifts in Hz i.e 60, 100 and 300 MHz( Operating frequency) is 60, 100 and 300 Hz (equivalent to 1 ppm), respectively. n MHz x 10-6 = n Hz 76

Summary 1 H NMR chemical shift Larmor frequency is dependent upon MF strength, and the use of Hz position of the peaks are also dependent upon MF strength 77