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Raman Spectroscopy: Principle, Instrumentation, Applications,

Advantages
July 14, 2023 by Kabita Sharma

Raman spectroscopy is a type of vibrational spectroscopy that allows for the simple
interpretation and very sensitive structural identification of trace amounts of substances
based on their distinctive vibrational properties. Sir C.V Raman, an Indian physicist,
demonstrated Raman spectroscopy, which is based on the inelastic scattering of
monochromatic light with the sample. Because of the inelastic scattering, the resultant
light will have a different frequency than the incident light.

This method is commonly used to investigate vibrational, rotational, and other low-
frequency interactions in molecules. This is quite beneficial for identifying molecular
structure, identifying functional groups, etc.
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Because of its diagnostic potential, Raman spectroscopy has become a commonly

utilized instrument in biomedical engineering and life science. It is used in medical
research to examine the biochemical environment of single cells or to monitor cell
reactions to medications, as well as in the pharmaceutical sector for process and quality
control in drug manufacturing.

Table of Contents
Raman effect
Raman shift
Principle of Raman spectroscopy
How does Raman Spectroscopy work?
Instrumentation of Raman spectroscopy
Types of Raman spectroscopy
Surface-enhanced Raman spectroscopy (SERS)
Surface-enhanced hyper Raman Scattering (HR) scattering
Tip enhanced Raman spectroscopy (TRES)
Coherent anti-Stokes Raman Spectroscopy (CARS)
Resonance Roman spectroscopy (RRS)
Confocal Raman microscopy
Applications of Raman spectroscopy
Advantages of Raman Spectroscopy
Comparison between Raman and IR spectroscopy
Comparison between Raman spectroscopy and FTIR spectroscopy
References

Raman effect
Raman spectroscopy is founded on the Raman effect, which was discovered in 1928 by
Indian physicist Chandrasekhara Venkata Raman. The Raman effect is based on light
scattering, which comprises elastic (Rayleigh) scattering at the same wavelength as the
incident light as well as inelastic (Raman) scattering at various wavelengths caused by
molecular vibrations. Rayleigh scattering is a million times more intense than Raman
scattering. To acquire Raman spectra, Rayleigh scattering must be prevented from
overwhelming the weaker Raman scattering.

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Raman spectra are obtained by stimulating a sample with a high-intensity laser beam
and passing the scattered light through a spectrometer. The Raman shift is the
difference in energy between the incident and dispersed light. The vertical axis of the
resulting spectrum represents the intensity of the scattered light, and the horizontal
axis is the wavenumber of the Raman shift (cm-1).

Raman shift
The Raman shift corresponds to two distinct energy bands. Stokes scattering refers to
the shift at wavelengths greater than those of the incident light. Anti-Stokes scattering
refers to the shift at wavelengths shorter than those of the incident light.

Principle of Raman spectroscopy

Raman spectroscopy is based on the inelastic scattering of electromagnetic light by a
molecule. There are two forms of scattering: elastic and inelastic scattering.

Elastic scattering follows the Rayleigh law, which asserts that no energy or momentum
is lost between the incident and dispersed radiation. As a result, this scattering is also
referred to as Rayleigh scattering. However, there will be certain (1 in a million) cases
when the energy of the dispersed radiation differs from the energy of the incidence
radiation. This is known as inelastic scattering.

Raman spectroscopy uses inelastic scattered light to identify an analyte through

molecular bond vibrations. A limited number of photons are dispersed when a sample
is subjected to laser light. The majority of the scattering is elastically dispersed light
with the same frequency as the incident light (i.e., Rayleigh scattering). Approximately
one out of every 106-108 photons is inelastically scattered (i.e., Raman scattering),
causing frequency differences (i.e., Raman shifts) between incident and scattered
photons when energy is exchanged between photon and molecule. In this case,
incident light photon energy is acquired by transferring energy from molecules to
photons (anti-Stokes Raman scattering) or lost by transferring energy from photons to
molecules (Stokes Raman scattering).

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Raman spectra are generated using the measured Raman shifts. Each Raman peak in
the spectrum corresponds to a single chemical bond, permitting molecular
identification of an analyte by producing a unique vibrational fingerprint.

Rayleigh Scattering

The molecule is stimulated to any virtual state.

The molecule returns to its former state by relaxing.
The photon is elastically scattered, retaining its initial energy.

Stokes Scattering

The molecule is stimulated to any virtual state.

The molecule returns to a higher vibrational state than it was before.
The photon has been scattered inelastically with energy hv- ΔE.

Anti-Stokescattering

In its first state, the molecule is vibrationally excited.

The molecule is stimulated to any virtual state.
The molecule returns to a lower vibrational state than it had previously.
The photon escapes with energy hv+ ΔE and is superelastically scattered.

How does Raman Spectroscopy work?

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Unlike FTIR spectroscopy, which examines variations in dipole moments, Raman

examines changes in the polarizability of a chemical bond. Light interaction with a
molecule can cause its electron cloud to distort. This deformation is known as
polarizability change. Molecular bonds have unique energy transitions that cause
polarizability to vary, resulting in Raman active modes. When photons interact with
molecules that include bonds between homonuclear atoms, such as carbon-carbon,
sulfur-sulfur, and nitrogen-nitrogen bonds, their polarizability changes. These are
instances of bonds that give rise to Raman active spectral bands but that are either
invisible or difficult to see in FTIR.

Because Raman is a weak effect by nature, the optical components of a Raman

Spectrometer must be carefully matched and optimized. Furthermore, as organic
molecules have a higher tendency to fluoresce when exposed to shorter wavelength
radiation, longer wavelength monochromatic excitation sources, such as solid-
state laser diodes emitting light at 785 nm, are commonly utilized.

Instrumentation of Raman spectroscopy

Fig: Instrumentation of Raman Spectroscopy

Image source: https://www.sciencedoze.com/2022/10/raman-spectroscopy-

principle.html

Source of excitation (laser)

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A laser is used as a light source in Raman spectroscopy. The bandwidth of the utilized
laser source determines the radiation’s spectrum. In general, a shorter wavelength
produces more Raman scattering. Because Raman scattering intensity varies as the
fourth power of frequency, argon, and krypton ion sources emitting in the blue and
green regions of the spectrum have an advantage over other sources.

Sample

Laser is irradiated on the sample present in the sample chamber. After interacting with
the laser beam it gives elastic and inelastic scattering before passing through the filter.

Liquid Samples:

A significant advantage of Raman spectroscopy sample handling over infrared is that

water is a poor Raman scatterer but a powerful absorber of

infrared radiation. As a result, aqueous solutions can be examined using Raman

spectroscopy but not infrared.

This benefit is especially essential for biological and inorganic systems, as well as
studies dealing with water pollution issues.

Solid samples

Solid-state Raman spectra are frequently obtained by filling a tiny cavity with the
sample after it has been ground to a fine powder. Polymers are typically evaluated
immediately without sample preprocessing.

Gas samples

Gas is often contained in glass tubes that are 1-2 cm in diameter and around Imm
thick. Small capillary tubes can also be used to seal gases.

Filter

In Raman spectroscopy, a filter is employed to separate Raman scattered light from

rayleigh scattered light. This is done in order to obtain high-quality Raman spectra.
Notch, long pass, and volume halogen filters are examples of filters.

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The detector

The detector aids in the detection of the scattered light signal. LCD array detectors are
commonly employed in current Raman spectrometers. They are designed to detect
signals of various wavelengths, as well as very weak signals.

Computer

A computer with relevant software aids in the creation of a final Raman spectroscopy
graph/ spectrum.

Types of Raman spectroscopy

Surface-enhanced Raman spectroscopy (SERS)
Surface-enhanced Raman spectroscopy (SERS) is one of the most sensitive devices,
allowing for highly sensitive structural detection of low-concentration analytes via the
amplification of electromagnetic fields generated by the excitation of adsorbate
molecules’ localized surface plasmons (LSP) on a roughened metal surface.

Surface-enhanced hyper Raman Scattering (HR) scattering

Surface-enhanced hyper Raman scattering is an inelastic sum of frequency scatted from
two photons, whereas normal Raman scattering results from a single photon. SERS’s
total surface enhancement factors are expected to be in the order of 1014. The
amplification could be due to molecular electronic resonance, in which the molecular
transition corresponds with the sum frequency of the input photons.

Tip enhanced Raman spectroscopy (TRES)

Tip enhanced Raman spectroscopy (TERS) is a strong technology that combines two
techniques: scanning probe microscopy and Raman spectroscopy. TERS can give
topographical and spectral/chemical information at the same time by utilizing SPM and
Raman.

TERS uses a metal tip or metal nanoparticle instead of a metal film and has identical
instruments, material requirements, and enhancement principles as SERS.

Coherent anti-Stokes Raman Spectroscopy (CARS)

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Coherent anti-stoke Raman scattering spectroscopy is a nonlinear Raman technique

used to amplify the Raman signal. It employs coherent laser beams to generate a signal
with a frequency greater than the excitation frequency and is thus classified as an anti-
stoke frequency approach.

Resonance Roman spectroscopy (RRS)

Resonance Raman spectroscopy is a technique that measures the shift in photon
frequency when the energy of photons from incident light is approximately similar to
the energy required for electronic transition. Resonance excitation can increase electron
oscillation charge displacement. As a result, it will raise the induced dipole moment,
which instantly enhances Raman scattering efficiency.

Confocal Raman microscopy

Marvin Minsky invented the first confocal Raman microscopy in 1955. In Confocal
Raman Microscopy, the probe head focuses laser light on the sample via the
microscope objective. And the pinhole refocuses the backscattered Raman signal,
causing it to act as a spatial filter. The signal will then be gathered by a charge-coupled
device camera (detector) to generate a spectrum.

Applications of Raman spectroscopy

Raman spectra are similar to infrared spectra in that they consist of functional group
detection zones and fingerprint sections that allow specific compound
identification. Raman spectra provide more information about certain types of
organic molecules than infrared spectra.

Raman spectra are less populated with peaks than infrared spectra. As a result, peak
overlap in mixtures is less frequent, and quantitative measurements are more
straightforward. Furthermore, Raman sampling equipment is not affected by
moisture, and little amounts of water in a sample have no effect.
Since aqueous solutions can be used, the Raman technique is frequently superior to
infrared for the spectroscopy of inorganic systems.

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It is used to investigate the structure of CO2, N2O, mercurous salts, mercury chloro
complexes, and the nature of bonding.
It is useful for studying electrolytic dissociation, hydrolysis, and the transition from
crystalline to amorphous states in physical chemistry.
It is also used to determine the presence or absence of certain links in a molecule, as
well as the structure of simple compounds and the study of isomers.
Raman amplification is based on Raman scattering, in which lower-
frequency photons are pumped to a high-frequency zone with an excess of energy.
This method can be used in telecommunications.
Raman spectroscopy is based on the Raman effect and has applications in many
fields, including nanotechnology to study the structure of nanowires, biology, and
medicine to study low-frequency DNAs and proteins, and chemistry to study the
structure of molecules and their bonds.
Remote sensing and planetary exploration make use of Raman scattering.
When combined with optical fibers, Raman spectroscopy provides the added benefit
of remote sensing. The optical fibers are responsible for transporting Raman signals
by collecting scattered photons.
Raman spectroscopy has been employed in real-time monitoring systems to detect
illegal substances, harmful environmental materials, and chemical and biological
warfare weapons.

Advantages of Raman Spectroscopy

Because of its better spectral resolution and narrower bandwidths, Raman
spectroscopy enables multiplex detection of various analytes.
Raman works with any sample composition. Analyte concentrations in solvents,
tissue, and solids (for example, drug composition) can all be measured.
There is no requirement for sample preparation or particular markers (but they can
be used in conjunction with Raman measurements).
Raman can sense individual analytes on a subcellular level and determine their
distribution in a cell when paired with microscopy.

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Raman can be easily combined with microfluidic devices to develop more

complicated and automated analytical measurement setups.
It is a non-invasive technology for subsurface sensing in tissue and measurements in
vivo.

Comparison between Raman and IR spectroscopy

Although Raman and infrared spectroscopy are both based on molecular vibrations, as
shown below.

IR spectroscopy

The absorption of light energy corresponding to the vibrational energy of molecules

is the basis for infrared spectroscopy.
A vibration is Raman active if it induces a change in polarisability.
The molecule does not require to have a constant dipole moment.
Water can be used as a solvent.
Sample preparation is simple; it can be in any state.
It Indicates the presence of covalent bonds in the molecule.
The cost of instrumentation is high.

Raman spectroscopy

Raman spectroscopy is based on the scattering of incident light at an energy altered

by the molecule’s vibrational energy (hv).
Vibration is IR active if the dipole moment changes.
The vibration in question should cause a change in the dipole moment.
Water cannot be utilized due to its high IR absorption.
The sample preparation is rigorous.
Gaseous samples are rarely useful.
Indicates the ionic nature of the molecule.
Relatively affordable

Comparison between Raman spectroscopy and FTIR spectroscopy

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Even though FTIR and Raman Spectroscopy provide complementary information and
are frequently interchangeable, several practical considerations affect which one is the
best choice for a particular experiment.

Raman spectroscopy is used

When the primary focus of research is on carbon bonding in aliphatic and aromatic
rings.
For the bonds that are difficult to see in FTIR (for example, O-O, S-H, C=S, N=N, C=C,
and so on)
Polymorphism, for example, requires the examination of particles in solution.
For lower frequency modes (for example, inorganic oxides) are relevant.
To the study of reactions in aqueous media
To investigate lower-frequency lattice modes.
For investigation of biphasic and colloidal reaction initiation, endpoint, and product
stability

FTIR is used

For investigating liquid-phase reactions

For Fluorescent reactions in which reactants, reagents, solvents, and reaction species
Bonds having large dipole changes (e.g., C=O, O-H, N=O) are important.
For reactions involving low concentrations of reagents and reactants
For reactions with strong Raman solvent bands that can obscure crucial species
signals
For reactions in which the intermediates produced are IR active

References
1. https://chem.libretexts.org/Bookshelves/Analytical_Chemistry/Physical_Methods_in_C
hemistry_and_Nano_Science_%28Barron%29/04%3A_Chemical_Speciation/4.03%3A_
Raman_Spectroscopy.
2. https://www.slideshare.net/BhaumikBavishi/raman-spectroscopy-54641528.
3. https://www.jasco-global.com/principle/1-what-is-raman-spectroscopy/.

https://scienceinfo.com/raman-spectroscopy-instrumentation/ 11/15
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4. https://www.researchgate.net/publication/309179824_Raman_Spectroscopy_a_review
.
5. https://www.sciencedoze.com/2022/10/raman-spectroscopy-principle.htm
6. https://byjus.com/physics/raman-scattering/.
7. https://www.mdpi.com/2079-6374/11/6/187

About Author

Kabita Sharma

Kabita Sharma, a Central Department of Chemistry graduate, is a young enthusiast

interested in exploring nature's intricate chemistry. Her focus areas include organic
chemistry, drug design, chemical biology, computational chemistry, and natural
products. Her goal is to improve the comprehension of chemistry among a diverse
audience through writing.

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