TOP NMR Interview Questions

Introduction and Outcome: Nuclear Magnetic Resonance Spectroscopy (NMR)

Nuclear Magnetic Resonance Spectroscopy (NMR) plays a vital role in drug development because without it, elucidation of the structure of any pharmaceutical is impossible. It provides detailed information about the atomic and molecular structure of pharmaceuticals. In this article, you will learn about NMR interview questions and their answer.

What is the NMR?

Nuclear Magnetic Resonance Spectroscopy (NMR) is the study of molecules by recording the interaction of radio frequency electromagnetic radiations with the nuclei of the molecules placed in the strong magnetic field.

Note: It is a powerful and direct method for the characterization and analysis of pharmaceuticals. NMR spectroscopy can also be applied to 13C and other nuclei.

What is the Principle of NMR?

The working principle of NMR is based on the spin of atomic nuclei. Nuclei with odd masses or odd atomic numbers have “nuclear spin” (similar to the spin of electrons). Since the nucleus is a charged particle in motion, it will develop a magnetic field.

A nucleus with spin quantum number, I≠0, when placed in an external uniform constant magnetic field of strength, H0, is aligned with respect to the field in the (2I + 1) potential orientation. Thus, for nuclei with I = ½, which includes most isotopes of analytical importance, there are two possible orientations corresponding to two different energy states. Nuclear resonance is the transition between these states by absorption or emission of corresponding amounts of energy.

How is NMR helpful in drug development?

The following are the different applications of Nuclear Magnetic Resonance Spectroscopy (NMR) in the pharmaceutical development:

Structure Elucidation of Pharmaceuticals
pKa determination
Conformational analysis
Keto-enol tautomerism
Hydrogen bonding
Determination of reaction velocities
Quality Control and Formulation

Structure Elucidation of Pharmaceuticals

Chemical Structure Determination: Nuclear Magnetic Resonance Spectroscopy (NMR) is crucial for determining the chemical structure of pharmaceuticals by providing detailed information about the arrangement of atoms in a molecule. This allows synthetic chemists to confirm the molecular structure of newly synthesized compounds.
Isomer Identification: NMR can distinguish between different isomers of a molecule including stereoisomers (e.g., enantiomers or diastereomers), which is vital for selecting the most effective and safe drug candidates.

Conformational analysis /Studying Molecular Dynamics

Conformational Flexibility: Nuclear Magnetic Resonance Spectroscopy (NMR) can give insight into the dynamics of molecules, such as how a drug or a biomolecule changes conformation over time. This is critical in drug design, as the flexibility of a target protein or a drug molecule often impacts its efficacy and selectivity.
Conformational States of Drugs: Understanding the conformational states that a drug adopts in solution can inform its pharmacodynamics and pharmacokinetics, as well as guide the design of more stable or effective drug candidates.

Quality Control and Formulation

Drug Formulation Analysis: NMR helps in the analysis of drug formulations, ensuring consistency, purity, and stability of the final pharmaceutical product. NMR can detect impurities, degradation products, or contaminants, ensuring that the drug product meets regulatory standards.
Characterization of Polymorphs: Some drugs exist in multiple solid-state forms (polymorphs), and NMR can be used to characterize these forms, which can have different bioavailability or stability profiles. Identifying the most appropriate polymorph for clinical use can be a critical part of drug development.

Natural Product Discovery

Isolating Bioactive Compounds: NMR is used extensively in the isolation and characterization of bioactive compounds from natural sources (plants, microbes, etc.). These compounds can serve as starting points for drug development or be developed into drug candidates themselves.
Identifying Novel Compounds: NMR provides a detailed, non-destructive way to identify and characterize complex natural products, which are often challenging to study by other methods.

What are the different Key NMR Techniques Used in Drug Development?

1D and 2D NMR: Used for structure elucidation and molecular characterization.
3D NMR: For studying larger molecules, such as proteins and nucleic acids.
STD NMR: For studying protein-ligand interactions.
Diffusion-Ordered Spectroscopy (DOSY): Used for studying molecular size and aggregation states in solution.
NOE (Nuclear Overhauser Effect): Helps in determining the spatial proximity of atoms, especially useful for studying the structure of proteins and nucleic acids.

Interpretation of NMR Spectra

Number of NMR Signals

The number of NMR signals is decided by equivalent and non-equivalent protons present in the molecule:

CH₃–CH₂-Cl: In this molecule will give two signals since the proton of the CH₃ group and CH₂ groups are not in the same environment and hence they are non equivalent

CH₃–CH₂-CH₂-Cl: This molecule contains protons in three different environments and hence this molecule will give three NMR signals

Splitting of Signals or Spin Spin Coupling

The multiplicity of the given group = (n+1), where n is the number of adjacent protons

Case study: CH₃–CH₂-OH

Multiplicity due to –CH₂ group will be 2 +1 = 3

Multiplicity due to –CH₃ group will be 3 +1 = 4

The intensity of the multiplet is given by (r+1)ⁿ, where is the number of protons on the adjacent atom.

For –CH₃ : (r+1)² = r² +2r + 1 = 1:2:1

For –CH₂ : (r+1)³ = r³ +3r² + 3r+1 = 1:3:3:1

How to read and interpret 1H NMR and 13C NMR spectra?

To read and interpret 1H NMR and 13C NMR spectra, look for the following key information: chemical shift values, peak multiplicity, coupling constants, and integration values. Analyze the spectra to determine the hydrogen and carbon environments present in the molecule. To read and interpret 1H NMR and 13C NMR spectra, look for the following key information: chemical shift values, peak multiplicity, coupling constants, and integration values. Analyze the spectra to determine the hydrogen and carbon environments present in the molecule.

How to interpret integration in NMR?

Integration in NMR spectroscopy measures the relative amounts of protons in a molecule. The area under each peak is proportional to the number of protons responsible for that peak. The ratio of the integrated peak areas corresponds to the ratio of the number of protons giving rise to those signals.

What is the proton NMR?

Proton Nuclear Magnetic Resonance (proton NMR or ¹H NMR) is a powerful analytical technique used to determine the structure of organic compounds. It provides information about the hydrogen atoms (protons) within a molecule. By analyzing the way protons in different chemical environments interact with an applied magnetic field, proton NMR can give insights into the molecular structure, functional groups, and the connectivity of atoms in a compound.

Key features of proton NMR include:

Chemical Shifts: This is the position of signals (peaks) in the spectrum, measured in parts per million (ppm). The chemical shift depends on the electronic environment of the hydrogen atoms. For example, protons attached to carbon atoms adjacent to electronegative atoms like oxygen or nitrogen will resonate at different frequencies than those attached to carbon atoms in alkyl groups.
Multiplicity (Splitting Patterns): This refers to the splitting of NMR signals due to the interaction between protons that are nearby (coupling). The number of peaks in a signal gives information about how many neighboring protons (n) there are, following the (n+1) rule. For example, a signal that splits into a doublet indicates one neighboring proton.
Integration: The area under each peak is proportional to the number of protons contributing to that signal. This helps determine the relative number of protons in different environments.
Coupling Constants: The distance between the split peaks in a signal, which provides information about the interaction between coupled protons. This can give clues about the spatial relationship and chemical environment of the protons.
Spin-Spin Coupling: Protons interact with nearby protons, leading to the splitting of NMR signals into multiple peaks. This coupling is influenced by factors like the distance and spatial arrangement of the protons.

Proton NMR is widely used in organic chemistry to determine the structure of organic compounds, identify functional groups, and determine the purity of a sample.

What is the Carbon NMR?

Carbon Nuclear Magnetic Resonance (Carbon NMR) is a type of spectroscopy that is used to determine the structure of organic compounds by analyzing the magnetic properties of carbon atoms in the molecule. It provides detailed information about the carbon environment within a molecule, helping chemists deduce the number and type of carbon atoms present, their connectivity, and their chemical environment.

In Carbon NMR, a sample is exposed to a magnetic field, and when it is subjected to radiofrequency radiation, the carbon nuclei in the sample absorb the radiation at frequencies characteristic of their environment. The resulting data produces a spectrum, where the chemical shifts on the x-axis represent the different environments of carbon atoms in the molecule, and the intensity of the peaks gives information about the number of equivalent carbons in each environment.

Key points about Carbon NMR:

Chemical Shifts: The location of the peaks on the spectrum, measured in parts per million (ppm), reflects the electronic environment of the carbon atom.
Peak Splitting (Multiplicity): Similar to proton NMR, carbon NMR peaks can split into multiple peaks (multiplets) due to interactions with neighboring hydrogen atoms or other carbon atoms, which provides further structural insights.
DEPT (Distortionless Enhancement by Polarization Transfer): A technique used in Carbon NMR to enhance signals from specific carbon environments (such as CH, CH2, and CH3 groups) and help clarify the structure.

Carbon NMR is a powerful tool in organic chemistry for identifying and confirming the structure of unknown compounds.

Which solvent is used in NMR?

n Nuclear Magnetic Resonance (NMR) spectroscopy, the most commonly used solvents are deuterated solvents. These solvents contain deuterium (a hydrogen isotope) instead of regular hydrogen (protium) because deuterium has a much lower NMR resonance frequency than hydrogen, which minimizes interference with the sample’s NMR signals.

Common deuterated solvents include:

Deuterated chloroform (CDCl₃): One of the most widely used solvents in NMR.
Deuterated methanol (CD₃OD): Often used for polar compounds.
Deuterated dimethyl sulfoxide (DMSO-d₆): A common solvent for polar and high-boiling compounds.
Deuterated acetone (acetone-d₆): Used for polar organic compounds.
Deuterated water (D₂O): For water-soluble compounds and aqueous solutions.

These solvents are chosen not only for their deuterium content but also for their ability to dissolve the sample and their low background signals in NMR spectra.

What are the 4 types of NMR?

The 4 types of NMR are: proton NMR, carbon-13 NMR, fluorine-19 NMR, and phosphorus-31 NMR.

Why is DMSO used as solvent in NMR?

DMSO is commonly used as a solvent in NMR spectroscopy because it has a low viscosity, high polarity, and does not interfere with most NMR signals.

How do I know what NMR solvent to use?

Choosing the right NMR solvent depends on several factors, including the solubility of your sample, the type of NMR experiment you’re conducting, and whether you need to avoid interfering signals. The following steps are followed while selecting the NMR solvent:

Sample Solubility: Ensure the solvent dissolves your sample completely. If your compound is not soluble in a common solvent like deuterated chloroform (CDCl₃) or deuterated DMSO (D₆-DMSO), you might need to try alternative solvents.
Solvent Signal: The solvent must not interfere with the region of interest in your spectrum.
Nature of the Compound:
- Polar compounds: For polar compounds, DMSO-d₆ or acetone-d₆ are commonly used.
- Non-polar compounds: For non-polar compounds, CDCl₃, benzene-d₆, or toluene-d₈ are often good choices.
- Aqueous compounds: For water-soluble compounds, use D₂O, but be cautious of proton exchange.
Type of NMR Experiment:
- For 1H and 13C NMR, solvents like CDCl₃, D₂O, and DMSO-d₆ are commonly used.
- For 2D NMR experiments (like COSY, HSQC, or NOESY), solvents such as CDCl₃ or DMSO-d₆ are used depending on solubility.
For solvent suppression in 1H NMR experiments, a solvent like acetone-d₆ or DMSO-d₆ may be better because their signals are more easily suppressed.
Avoiding Interference:
- Choose a solvent with minimal impurities and with signals that do not overlap with your compound’s signals.
- If there’s a chance of proton exchange (e.g., if your sample contains alcohols, phenols, amines), D₂O or acetone-d₆ might be good choices as they do not have exchangeable protons in the same range.

Commonly Used Solvents:

CDCl₃ (Deuterated Chloroform): Common, good for non-polar compounds.
D₂O (Deuterated Water): Used for aqueous compounds, but protons may exchange.
DMSO-d₆ (Deuterated Dimethyl Sulfoxide): Good for polar, high-boiling compounds.
Acetone-d₆ (Deuterated Acetone): Suitable for polar compounds, used in solvent suppression experiments.

In summary, the solvent you choose should dissolve your sample, not interfere with the region of interest, and be compatible with the type of NMR experiment you’re performing.

What is J value in NMR?

The J value in NMR spectroscopy refers to the coupling constant, which represents the strength of the spin-spin interaction between nuclei.

What is the N 1 rule in NMR?

The N+1 rule in NMR states that the number of signals observed for a given nucleus is one more than the number of equivalent neighboring nuclei.

What is the time scale of NMR?

The typical timescale of nuclear magnetic resonance (NMR) spectroscopy is on the order of milliseconds to seconds.

What is coupling in NMR?

Coupling in NMR refers to the interaction between neighboring nuclear spins, which can split the signals in the NMR spectrum.

What is shielding and deshielding in NMR?

In Nuclear Magnetic Resonance (NMR) spectroscopy, shielding and deshielding refer to the way the electron cloud around a nucleus affects its magnetic environment and, consequently, its resonance frequency when placed in a magnetic field.

Shielding:

Shielding occurs when the electron cloud around a nucleus generates a magnetic field that opposes the external magnetic field applied during NMR.
The presence of this opposing magnetic field reduces the effective magnetic field experienced by the nucleus, causing the resonance frequency to be lower (or the chemical shift to be smaller).
A nucleus in a shielded environment will appear at a lower chemical shift in the NMR spectrum. This typically happens when the nucleus is surrounded by more electrons, such as in electron-donating environments (e.g., alkyl groups).
Shielding is usually observed in more electron-rich environments, where the electron density around the nucleus is high.

Deshielding:

Deshielding is the opposite process, where the electron cloud around a nucleus is not able to counteract the applied external magnetic field effectively.
This leads to an increased effective magnetic field at the nucleus, causing the resonance frequency to be higher (or the chemical shift to be larger).
A nucleus in a deshielded environment will appear at a higher chemical shift in the NMR spectrum. This typically happens when the nucleus is in an electron-withdrawing environment, such as near electronegative atoms (e.g., oxygen, nitrogen) or groups that pull electron density away from the nucleus (e.g., halogens, carbonyl groups).
Deshielding is generally seen in environments where the electron density around the nucleus is lower.

Key Factors Influencing Shielding/Deshielding:

Electron density: More electron density around a nucleus typically shields it, while less electron density causes deshielding.
Electronegative substituents: Electronegative atoms (like fluorine, oxygen, etc.) tend to withdraw electrons, causing deshielding.
Inductive and resonance effects: Groups that donate electrons (such as alkyl groups) tend to shield, while those that withdraw electrons (like -NO2, -CN) tend to deshield.

These phenomena are crucial in interpreting the chemical shifts in NMR spectra, as they help identify the electronic environment of different atoms in a molecule.

Why is TMS used in NMR?

TMS, or tetramethylsilane, is used as a reference compound in nuclear magnetic resonance (NMR) spectroscopy. It provides a consistent and stable reference signal for calibrating the chemical shift scale.

Which detector is used in NMR?

RF detector is used in NMR. It helps in determining unabsorbed radio frequencies.

Recorder: It records the NMR signals which are received by the RF detector.

What are 5 uses of NMR spectroscopy?

Nuclear magnetic resonance (NMR) spectroscopy has numerous applications:Structural elucidation of organic compounds Identification and quantification of chemical compounds Monitoring chemical reactions and processes Studying molecular dynamics and interactions Imaging and analysis of biological samples

Which radiation is used in NMR?

Nuclear magnetic radiation.

Conclusion

Nuclear Magnetic Resonance Spectroscopy (NMR) is the backbone of pharmaceutical development since it provides insights into molecular structure. I hope this article has helped you understand Nuclear Magnetic Resonance Spectroscopy (NMR) in Pharmaceutical Development and its importance. You may also want to check out other articles on my blog, such as LCMS, GCMS and Pharmaceutical Structural Elucidation

Related topic:

Advantages

Very helpful in structure elucidation

Disadvantages

This is expensive equipment and may not be affordable for small companies
Dedicated and skilled scientists with knowledge of chemistry are needed to operate the equipment

References

Applications of NMR
Molecular Spectroscopy: P.R Singh and S.K Dikshit
Nuclear Magnetic resonance

Abbreviations

NMR: Nuclear magnetic resonance
DOSY: Diffusion-Ordered Spectroscopy
NOE:Nuclear Overhauser Effect
HTS: High-Throughput Screening
STD: Saturation Transfer Difference