The structure and purity of any compound can be determined through the spectroscopic analytical technique, Nuclear Magnetic Resonance (NMR). The basic principle of the NMR is the interaction between nuclei of different atomic isotopes with a static magnetic field. In medical science, magnetic resonance imaging (MRI), the same as NMR, is used to diagnose different diseases. In the food industry, NMR determines the ratio of water and fat in foods. NMR is also used to measure the corrosive fluids in pipes.
Basic knowledge is important to understand every topic. The first question in proton NMR is; why do we call it a proton NMR? The answer is simple because we use isotopes of hydrogen in proton NMR, mainly deuterium ²H. The other things that may confuse the students are the resonances of the nucleus and the magnet. So make the different resonances and their interaction clear to students.
It is difficult for the students to understand and analyze the proton NMR spectra at the school and college levels. So, practice is a must to analyze it; take some simple spectra to the classroom and make them clear about the different types of peaks like singlet, duplet, and triplet. Make them clear about the intensities of the peaks. Tell them how to differentiate the solvent peak from the sample peaks.
Understanding the process of NMR and its spectra interpretation by theoretical means is very challenging. In NMR shapes, the signals tell everything; the students must know what a doublet signal looks like. So to clear all these doubts through theoretical means is nearly impossible. Video graphics is the best option for understanding the spectra of NMR. Various NMR spectra analysis software are available online, like ‘Mnova’; you can use them to interpret any NMR spectra.
Magnetic resonance: The emission/absorption of electromagnetic radiation when a magnetic field is applied to any atom or nuclei is called magnetic resonance.
Isotopes: Atoms having similar atomic numbers but different atomic masses are called isotopes.
NMR: Nuclear Magnetic Resonance (NMR) is an analytical technique through which the structure and purity of any compound can be determined.
¹H NMR: When we use an isotope of hydrogen in NMR, it is called ¹H NMR or proton NMR.
¹³C NMR: When we use an isotope of carbon in NMR, it is called ¹³C NMR or carbon NMR.
Chemical shift: The resonance frequency of any atomic nucleus relative to a standard compound set at 0 ppm. Chemical shift is denoted by the symbol (δ) and is measured in ppm (parts per million).
There are different types of NMR: Solid state, liquid-state, and solution-state NMR. The way the sample is prepared will depend on the type of NMR. The most common is the solution state. For this, you will require a suitable solvent and the internal standard TMS - Tetramethyl silane.
Equipment: NMR tubes are used as they are designed specifically for the spectrometer. They are typically 5mm in diameter and have a small cap and an NMR adaptor which helps the tube stand upright. A glass pipette is used instead of a plastic pipette and micropipette because it is much thinner for a more precise transfer of the sample in the thin NMR tube.
How much solid to weigh out: The mass of solid compounds for carbon NMR is typically between 20-50 mg. This is much more than the mass required for proton NMR - which is 1-2 mg, and is to do with the difference in technique sensitivity.
Solvents: The solvent used is typically deuterated. This means that the hydrogens in the solvent molecules are replaced with the isotope of hydrogen named Deuterium. It helps with the locking and shimming process inside the machine.
Reference Compound: TMS is the chosen reference compound used because the carbons and protons are in the exact same chemical environment and causes a resonance peak at 0 ppm and so it lets us know where the chemical shift scale begins. Adding TMS directly to your sample will be too concentrated, so you must dissolve a small drop in some solvent before you dissolve your solid.
In the case of a solid sample, these are the steps to follow:
Weigh out 20-50 mg of solid
Add one drop of TMS to some deuterated solvent
Dissolve the solid in solvent mixed with TMS
Pipette the sample solution in the NMR tube
Place the tube in the NMR adaptor
Wipe the tube with tissue paper
Proton NMR, also notated as ¹H NMR, is a spectroscopic technique that is commonly used for structure elucidation and purity assessments of organic compounds. In proton NMR, which stands for proton Nuclear Magnetic Resonance, a sample is placed in a strong magnetic field and then hit with radiofrequency radiation. The protons will have aligned or anti-aligned their so-called spin state with the magnetic field, and the difference between these two states has resonance energy. When hit by the appropriate frequency, the lower energy protons can flip their alignment. This exact energy is emitted from the protons detected in this technique as they flip back to the lower energy state.
In NMR, the chemical shift represents the resonance frequency for a proton relative to a reference compound. Chemical shift is denoted by the symbol (δ) and is measured in ppm (parts per million). In a proton NMR spectrum, the chemical shift gives information on the chemical environment of the targeted proton. The chemical shift value is highly impacted by the structure of the analyzed compound, especially by electronegative elements or effects. One reason is that electronegative elements pull electron density away from the proton, increasing the chemical shift value. This effect is known as 'de-shielding' since it leaves the proton more exposed to the externally applied magnetic field.
Another way the chemical shift can be impacted is through protons of OH and NH groups, NMR spectra of compounds such as alcohols and amines have a broad peak representing the OH and NH protons. This is because these protons are especially labile and interact with the solvent, where the protons are exchanged at a faster rate than the NMR is probed—appearing as a broad peak on the spectrum, whereas the protons that do not undergo fast exchange appear sharp and defined.
Below is an overview of the chemical shift values of protons in common functional groups. Note that only the protons in a compound will give rise to a signal in proton NMR analysis.
Figure 2. Overview of Chemical shift ranges in ppm for common functional groups (Image source Labster theory).
Why don’t all protons resonate at the same frequency? In a ¹H NMR spectrum, each peak represents a different kind of chemical environment, where the protons have different resonance frequencies. In a molecule, each nucleus experiences a different magnetic field due to the surrounding electrons of other atoms. The magnetic fields of the surrounding atoms can oppose the magnetic field that is applied during an NMR experiment, and the protons become shielded - this reduces their resonance frequency and, thus their chemical shift.
While elements such as carbon have shielding effects, more electronegative elements such as fluorine, chlorine, and oxygen have deshielding effects. With protons that are in the proximity of these groups, the electron cloud is more polarised towards the electronegative elements, and the proton becomes less shielded from the applied magnetic field, contributing to a greater chemical shift. The chemical shift of protons changes for different functional groups due to these shielding effects. These effects can be attributed to the extent of the polarisation of electrons due to the various functional groups.
The proton environments with the largest chemical shift values are the ones that are more deshielded; this effect is useful in deducing NMR structures as it separates different proton resonances into distinct peaks over a wide range allowing for effective elucidation.
Equivalent protons, also termed chemical equivalence, is the concept of a group of protons being in the exact same chemical environment. As they are indistinguishable, this leads to the protons giving a single signal together in proton NMR.
In ethanol, the three protons in the methyl group are equivalent because each is in the same chemical environment (remember that there's free rotation around the single (σ) bonds). The protons in the methyl group are NOT equivalent to the other protons in the molecule, as their chemical environment is different.
When considering equivalent vs. non-equivalent protons, it's important to pay attention to symmetry. Check out the structure of the compound in the spectrum below. Because there's symmetry around the -CH group, the two methyl (-CH3) groups are in the exact same chemical environment, which means the combined 6 protons will give a combined signal in the spectrum (at 0.9 ppm). Note that this is due to the symmetry in the molecule, not because they are both methyl groups.
Figure 4. NMR spectrum and structure of 1-bromo-2-methylpropane (Image source Labster theory).
Peak splitting in proton NMR, or spin-spin coupling, is the phenomenon that neighboring protons will interact with each other's resonance and create a splitting pattern for the signal in the NMR spectrum. A signal will have a splitting pattern of n+1, where n is the number of neighboring protons (see example in Figure 1 below). This rule can be a very powerful tool for structure elucidation in deducing the molecular groups of a compound.
Figure 5. Example of NMR spectrum of ethanol with peak splitting. Notice the splitting pattern of each signal in relation to the number of each equivalent set of protons (a, b, c) (Image source Labster theory).
So if a set of equivalent protons have 3 neighboring protons, the signal will be split into 4, as you can see in Figure 1 for the "c" set. As a rule of thumb, this effect occurs only through 3 bonds and generally only through carbon-carbon bonds, not, e.g., carbon-oxygen. There are other types of information that can be extracted from the spin-spin coupling, e.g., via the distance between each peak, which is called the coupling constant.
A proton NMR spectrum will often include information on the relative peak integrals. These are the relative areas under each signal in the spectrum, and they are correlated with the number of protons that give rise to the signal. We can therefore use peak integrals to determine the relative number of protons in each signal. See the example here in Figure 6.
Figure 6. Examples of proton NMR spectrum include relative integral numbers above each peak (Image source Labster theory).
For a splitting pattern, the entire signal is usually included in the integral. It's key to realize that the integrals are relative and does not necessarily express the exact number of protons. From the integrals in Figure 6, one might conclude that each signal represents either 2 or 3 protons. But from the integrals alone, the reality might just as well be 4 and 6 protons or 6 and 9 protons. So while the integrals provide useful information, they can often not stand alone in elucidating a structure.
Proton and Carbon NMR are the spectra that differ in many ways.
In terms of spectroscopic sensitivity carbon-13, NMR is a much less sensitive technique because of two factors, the gyromagnetic ratio and natural abundance. The gyromagnetic ratio is another parameter that determines this value. It is represented by the constant gamma and is a characteristic of each isotope. The sensitivity of a nucleus in NMR depends on gamma. The higher the value of gamma, the higher the sensitivity, and vice versa.
The value of gamma for the carbon-13 nucleus is 9.6, and for hydrogen-1, it is 27.4. The natural abundance of carbon-13 is 1.1%, whereas, for Hydrogen-1, it is 99.9%. The theory behind it is simple, there are more hydrogen atoms contained in a sample and so easier to detect.
You don't see coupling in Carbon-13 NMR because of the low natural abundance. No carbons can come close enough to one another in proximity for coupling. This is why we don't see splitting in the spectrum. In the case of proton NMR, we see hydrogen coupling and hence splitting patterns in the peak. Figure 7 shows an example of peak splitting in a proton NMR spectrum.
Figure 7: Proton NMR spectrum of ethanol (Image source Labster theory).
With technological advances, it is much easier to explain complex and challenging processes with the help of simulations. Now, you can simulate experiments without the need for any valuable equipment.
In this regard, you can take help from Labster’s virtual lab simulations. These simulations engage students through interactive learning scenarios. Students dive into a 3D world, where they visually learn and apply their concepts to solving real-life problems.
In Labster’s interactive Proton NMR: Spectra interpretation Virtual Lab, students will learn the basic tools needed for interpreting a proton NMR spectrum and use them to elucidate the structure of an unknown compound.