# IS THE THING I MADE THE THING I WANT: METHODS OF CHARACTERISING AN UNKNOWN SAMPLE

Now I’m sure we’ve all been in the situation where, we’re in a lab and we have made our product with very high yield, and we’re happy. But how can we be certain that the product we have is the product we wanted to make in the first place?

Well, there are several spectroscopic methods that, when used in conjunction with one another, can paint a very accurate picture of the molecule, from its molecular structure, to the functional groups attached to the molecule. But how do these spectroscopic methods work?

## Molecular structure determination

In order to determine the molecular structure of our sample, we use Nuclear Magnetic Resonance (NMR) spectroscopy. This works based off the principle that some atomic nuclei have nuclear spin, and the presence of this spin makes these nuclei behave like a small magnet. The spinning charge of the nuclei can generate a magnetic dipole, which can interact with an applied external magnetic field.

In the absence of the magnetic field, the nuclei are randomly arranged and the energies of the magnetic moments are equal, which doesn’t produce an NMR signal. In the applied magnetic field, the magnetic moments have 1 of 2 possible energy states, one low and one high, and follows a Boltzmann distribution. (1)

Fig 1: Nuclei in the absence and presence of an applied magnetic field (2)

When the sample is irradiated with energy of a suitable frequency, this distribution will change. For example, 1H nuclei will flip from low to high energy states. This is the resonance part of NMR and involves absorption of radiation. The frequency of the irradiated energy depends on the type of nucleus we want to resonate and abides by the following resonance condition:

Where nu is the frequency of the absorbed radiation, gamma is the magnetogyric ratio, which is simply the ratio of the nucleus magnetic moment to its angular momentum (the ratio is unique to each nucleus) and  is the applied magnetic field.

From this, we can see that different nuclei absorb energy at different frequencies, and this is due to the electrons surrounding the nucleus “shielding” the nucleus from B0 (2). As the electrons shield the nucleus from the effects of B0, their local diamagnetic shielding can be reduced. This affects the positions of the atom under resonance, the local magnetic field that the nucleus feels and the frequency at which the nucleus resonates. The shielding of the electrons cause each nucleus to experience a different B0, and thus give different peaks for different nuclei, and even the same nuclei with different numbers of electrons, which can inform us about the structure of the molecule.

NMR spectroscopy is done multiple times on the same sample, with each successive NMR focusing on a different nucleus in the sample, and the results being used in conjunction with each other to determine the molecular structure. For example, for C3H6O, there are multiple different ways to structure the molecule, with just three being shown below.

Figure 2.a) One possible arrangement of C3H6 (a. Acetone)

Fig 2.b : Three possible arrangements of C3H6 ( b. (Z)-1-Propenol)

Fig 2.c : Three possible arrangements of C3H6 (c. (E)-1-Propenol)

If we obtained a 13C and a 1H NMR spectra for all three isomers, we could quickly determine which arrangement is held by our sample. For acetone, if we did 1H NMR, we would see that all the hydrogens are in the same chemical environment

(the electron shielding is the same for all hydrogens), and we would get a single very big peak in the spectrum. If we did 13C NMR, we should see that two of the carbons are in the same chemical environment, but one of the carbons would experience a greater B0 as the attached oxygen effects the electron shielding.

For (Z)-1-propenol and (E)-1-propenol, they would be very similar but for one difference in the 13C NMR, as the carbon at the end of the chain, in the (Z)-1-propenol, would experience a slightly greater B0 than the same carbon in the (E)-1-propenol, as the (Z)-1-propenol carbon is closer to the oxygen.

## Determination of functional groups

To determine what is attached to our molecule, we can use infrared (IR) absorption spectroscopy. This works on the principle that molecules absorb frequencies that are unique to their structure. These absorptions happen when the frequency of the absorbed radiation matches the vibrational frequency. The energy of the absorbed radiation is affected by the masses of the atoms, and the associated strength of the bond between the atoms.

The IR light is absorbed when the oscillating dipole moment (due to molecular vibrations) interacts with the oscillating electric IR beam, which tells us that the dipole moment at one end of the molecule must be different to the dipole moment at the other end (3). Hence, a molecule that is symmetrical is inactive in the infrared spectrum, for example, N2.

The vibrations of the molecule give rises to multiple absorptions that occur between 4000 cm-1 and 400 cm-1, with the region below 1500 cm-1 being known as the fingerprint region, which is generally unique to the molecule in question. When examining an IR spectrum, one would use an IR table, which would tell you approximately the region certain functional groups absorb light in, and by matching the absorption peaks in your spectrum to the IR table, you can determine what functional groups are in your molecule.

References

1. Gunnlaugsson (2022), Analytical and Computational Methods: Lecture 4 https://tcd.blackboard.com/bbcswebdav/pid-2070153-dt-content-rid-12142873_1/courses/CHU33405-202122/Lecture%20Notes%20No%204.%20Organic%20Spectroscopy%202021%20TG.pdf
2. Gunnlaugsson (2022), Analytical and Computational Methods: Lecture 5 https://tcd.blackboard.com/bbcswebdav/pid-2076008-dt-content-rid-12194659_1/courses/CHU33405-202122/Lecture%20Notes%20No%205.%20Organic%20Spectroscopy%202021%20TG.pdf
3. Gunnlaugsson (2022), Analytical and Computational Methods: Lecture 3 https://tcd.blackboard.com/bbcswebdav/pid-2070150-dt-content-rid-12142868_1/courses/CHU33405-202122/Lecture%20Notes%20No%203.%20Organic%20Spectroscopy%202021%20TG.pdf

Image references

Fig 2.a https://pubchem.ncbi.nlm.nih.gov/compound/Acetone#section=Structures

Fig.2.b https://pubchem.ncbi.nlm.nih.gov/compound/Z_-1-Propenol#section=2D-Structure

Fig 2.c https://pubchem.ncbi.nlm.nih.gov/compound/Prop-1-en-1-ol#section=2D-Structure

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