Nuclear magnetic resonance spectroscopy and mass spectrometry are analytical techniques used to determine molecular structure and composition. NMR spectroscopy uses radiofrequency pulses to analyze atomic nuclei and determine organic compound structures. Mass spectrometry ionizes molecules and sorts the resulting ions based on their mass-to-charge ratio to determine molecular mass and elemental composition. Both techniques provide essential information for applications in chemistry, biochemistry, medicine, and environmental analysis.

1. NUCLEAR MAGNETIC
RESONANCE SPECTROSCOPY

2. • Nuclear magnetic resonance spectroscopy, most commonly known as NMR
spectroscopy or magnetic resonance spectroscopy (MRS), is a spectroscopic
technique to observe local magnetic fields around atomic nuclei.
• It is a spectroscopy technique that is based on the absorption of electromagnetic
radiation in the radiofrequency region 4 to 900 MHz by nuclei of the atoms.
• Over the past fifty years, NMR has become the preeminent technique for
determining the structure of organic compounds.
• Of all the spectroscopic methods, it is the only one for which a complete analysis
and interpretation of the entire spectrum is normally expected.

3. Principle of Nuclear Magnetic Resonance (NMR)
Spectroscopy
• The principle behind NMR is that many nuclei have spin and all nuclei are
electrically charged. If an external magnetic field is applied, an energy transfer is
possible between the base energy to a higher energy level (generally a single
energy gap).
• The energy transfer takes place at a wavelength that corresponds to radio
frequencies and when the spin returns to its base level, energy is emitted at the
same frequency.
• The signal that matches this transfer is measured in many ways and processed in
order to yield an NMR spectrum for the nucleus concerned.

4. Instrumentation of Nuclear Magnetic Resonance
(NMR) Spectroscopy

5. • Sample holder
Glass tube with 8.5 cm long, 0.3 cm in diameter.
• Permanent magnet
It provides a homogeneous magnetic field at 60-100 MHZ
• Magnetic coils
These coils induce a magnetic field when current flows through them
• Sweep generator
To produce an equal amount of magnetic field pass through the sample

6. • Radio frequency transmitter
A radio transmitter coil transmitter that produces a short powerful pulse of
radio waves
• Radio frequency receiver
A radio receiver coil that detects radio frequencies emitted as nuclei relax to
a lower energy level
• Read out systems
A computer that analyses and records the data.

7. Applications of Nuclear Magnetic Resonance
(NMR) Spectroscopy
• Spectroscopy is the study of the interaction of electromagnetic radiation with
matter. NMR spectroscopy is the use of the NMR phenomenon to study the
physical, chemical, and biological properties of matter.
• It is an analytical chemistry technique used in quality control.
• It is used in research for determining the content and purity of a sample as well as
its molecular structure. For example, NMR can quantitatively analyze mixtures
containing known compounds.

8. • NMR spectroscopy is routinely used by chemists to study chemical structure using
simple one-dimensional techniques. Two-dimensional techniques are used to
determine the structure of more complicated molecules.
• These techniques are replacing x-ray crystallography for the determination of
protein structure.
• Time domain NMR spectroscopy techniques are used to probe molecular
dynamics in solution.
• Solid state NMR spectroscopy is used to determine the molecular structure of
solids.
• Other scientists have developed NMR methods-of measuring diffusion
coefficients.

9. Carbon-13 (C-13) nuclear
magnetic resonance

10. • 13C-NMR spectroscopy is a type of nuclear magnetic resonance spectroscopy.
• It is used to study number of nonequivalent proton present in unknown compound.
• Carbon NMR can used to determine the number of non-equivalent carbons and to
identify the types of carbon atoms(methyl, methylene , carbonyl..) which may
present in compound.
• It makes it easier to identify and count individual nuclei.

11. Principle of Carbon-13 (C13) nuclear magnetic
resonance
• The C-13 isotope of carbon has an odd mass number and thus an odd number
of neutrons i.e., 7 so it possesses a specific angular momentum value. The
13C isotope present in different organic compounds is always spinning about
its fixed axis.
• Under the influence of an external magnetic field , Zeeman splitting occurs
and the 13C nuclei occupy two different spin orientations. The nuclei flip their
spin orientation by absorbing energy equal to the energy gap between the
two spin states. This energy is called resonance energy and it is provided by
the radio wave frequency (25-100 MHz) of the electromagnetic spectrum. The
amount of energy absorbed is ultimately used to determine the chemical
environment and structural arrangement of 13C nuclei in the targeted sample
molecules.

12. Application of Carbon-13 (C13) nuclear magnetic
resonance
• C-13 NMR has elucidated and biochemical structure.
• C-13 NMR provides information about the backbone to molecule rather than
periphery.
• C-13 nuclei are stable isotopes and hence it is not danger to radiotracer.
• It is also used for quantification of drug purity to determination of the
composition of high molecular weight synthetic polymer.

13. MASS SPECTROMETRY

14. • Mass spectrometry, also called mass spectroscopy, analytic technique by which
chemical substances are identified by the sorting of gaseous ions in electric and
magnetic fields according to their mass-to-charge ratios.
• Mass spectroscopy is the accurate method for determining molecular mass of the
compound and its elemental composition.

15. Principle of Mass Spectroscopy
• When molecules are bombarded with an energetic electron beam, 1e- is
removed from the molecule. The removal of electron e- from a molecule
causes the molecule to become positively charged, resulting in the formation
of a molecular ion. Further fragmentation of molecular ions produces
daughter ions.
• Each ion has its particular mass-to-charge ratio (i.e. m/z). They are separated
according to their m/z ratio and give a mass spectrum. m/z vs abundance
gives the mass spectrum.

16. Instrumentation of Mass Spectrometry (MS)

17. • Ionizer
The bombarding of the sample is done by the electrons. These electrons move
between cathode and anode. When the sample passes through the electron stream
between the cathode and anode, electrons with high energy knock electrons out of
the sample and form ions.
• Accelerator
The ions placed between a set of charged parallel plates get attracted to one plate
and repel from the other plate. The acceleration speed can be controlled by adjusting
the charge on the plates.
• Deflector
Magnetic field deflects ions based on its charge and mass. If an ion is heavy or has
two or more positive charges, then it is least deflected. If an ion is light or has one
positive charge, then it is deflected the most.
• Detector
The ions with correct charge and mass move to the detector. the ratio of mass to
charge is analyzed through the ion that hits the detector.

18. Applications of Mass Spectrometry (MS)
• Environmental monitoring and analysis (soil, water, and air pollutants, water quality,
etc.)
• Geochemistry – age determination, soil, and rock composition, oil and gas surveying
• Chemical and Petrochemical industry – Quality control
• Identify structures of biomolecules, such as carbohydrates, nucleic acids
• Sequence biopolymers such as proteins and oligosaccharides
• Determination of the molecular mass of peptides, proteins, and oligonucleotides.
• Monitoring gases in patients’ breath during surgery.
• Identification of drug abuse and metabolites of drugs of abuse in blood, urine, and
saliva.
• Analyses of aerosol particles.
• Determination of pesticides residues in food.