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However, in the 1920s, Prof. C.V.Raman and his colleagues, studying the scattering of light by various liquid samples discovered that although the majority of the scattered photons have the same frequency as the incident light, a few of them had frequency generally lower or higher than that of the incident light and the change in frequency depended on the chemical composition of the scattering material. This, an inelastic scattering process, came to be known as “Raman Effect” or “Raman Scattering”. Prof. Raman was awarded the 1930 Nobel Prize in Physics for this discovery.

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WHEN an object is illuminated by light, the light can be absorbed,

transmitted, refl ected or scattered by the object. Scattering means that the light deviates from its original direction. Generally the scattered light has the same frequency (or energy or wavelength) as the original light. This process is known as elastic scattering or Rayleigh scattering.

How does Raman Scattering Arise? The atoms and molecules in any chemical compound can have three types of motions in the X, Y, and Z directions: translational, rotational and vibrational motions. While the translational motion corresponds to the motion of the molecule as a whole, the other two depend upon the structure of the molecule and its atomic constituents.

While the rotational mode corresponds to the motion of the molecule and its constituent groups about the three axes, vibrational motion arises due to periodic stretching and bending of the chemical bonds in the molecule.

M.S.S. MURTHY Raman Effect, which started off as a simple observation of scattering of light by an object, has found immense applications over the entire cross-section of the industrial sector. On the occasion of the National Science Day on 28 February, celebrated to commemorate the discovery of the Raman Effect by Sir C.V. Raman on this day in 1928, let’s take a look at some of its multifarious applications.

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The frequency of this periodic motion is known as the vibrational frequency of the molecule. A molecule can exhibit various translational, rotational and vibrational energy states.

At room temperature most of the molecules will be in the lowest energy state – the ground state. However, the molecules can be excited to higher levels by supplying energy. Of these, changes in the vibrational frequencies of the molecules in gases, liquids and solids are more easily observable than changes in other energy states. Hence, Raman scattering refers to changes in the vibrational frequencies of the molecule.

Raman scattering arises when a photon incident on a molecule is absorbed by it. The molecule is excited to a higher virtual vibrational energy state. The molecule cannot remain in this state and therefore instantly de-excites (in less than 10-14 seconds), returning to a different vibrational energy level than the one it started from. In the process it emits another photon. The frequency of this photon is different from the frequency of the photon that caused excitation.

The difference in frequency between the incident and scattered photon is numerically equal to the frequency difference between the initial and fi nal vibrational levels of the molecule. Thus, while the frequency of the scattered light refers to one of the vibrational frequencies of the molecule, the intensity of the scattered light depends upon the number of scattering molecules in the path of the incident light.

A plot of the intensity of the scattered light as a function of its frequency shift from the exciting photon (generally expressed in terms of wave numbers) is known as the Raman spectrum of the molecular species. Each molecular species has its own unique set of molecular vibrational states. Hence, Raman spectrum consists of a series of peaks and bands, each shifted in frequency from that of the incident light by one of the characteristic vibrational frequencies of the molecule and thus becomes unique to that molecule.

Raman himself had stated in his Nobel Lecture, “the characteristics of scattered radiation enable us to obtain insight into the ultimate structure of the scattering substance”.

Intrinsically Raman scattering is very weak. Of all the scattered photons only 0.001% or less arise from Raman Effect, the rest being Rayleigh scattered. Furthermore, the detectors available at that time – photomultiplier tubes and photographic plates – were too slow. Therefore, Raman spectroscopy – though useful as an experimental tool in physics and chemistry – was very tedious and time consuming and no industrial application was possible.

However, for enhancing the Raman signal several innovative methods like Stimulated Raman Scattering, Surface-enhanced Raman Spectroscopy, Resonance Raman Spectroscopy, Offset Raman Spectroscopy, Multiple Angle Raman Spectroscopy, etc. have been developed in the subsequent years, which have greatly contributed to the ease with which Raman spectroscopy can be practiced.

These advancements, along with the availability of powerful lasers and effi cient detectors like the charge-coupled diodes (CCDs), have rendered Raman spectroscopy an extremely rapid method for obtaining molecular information of any sample.

Along with this, the chemical specifi city of the technique, non-contact, non-destructive nature of the measurement, small sample size, ease of sample preparation, ability to measure both organic and inorganic samples in gaseous, liquid or solid forms, adaptability of the technique for on-line, in situ, in vivo, and in vitro conditions have all made Raman spectroscopy one of the most sought after techniques in a wide range of industries for process control, quality assurance, optimising yield, etc. as well as in a variety of biomedical investigations, narcotic and explosives screening and so on. The list is endless.

What happens in Raman spectroscopy? Essentially, the sample is irradiated by a laser beam (in UV, visible or infrared range) and the Raman scattered photons are collected by a lens and passed through an interference fi lter or spectrometer to obtain the Raman spectrum of the sample. However, depending upon the application, the complexity of the equipment varies from a simple hand-held monitor to huge on-line systems used for real-time analysis from remote areas using optical fi bres for transmitting the exciting laser to the sample as well as the Raman signal to the spectroscope.

Some ApplicationsChemical industry: One of the main applications of Raman spectroscopy is in industries where catalysts are used. Catalysts can modify reaction pathways and lead to higher yields. They are extensively used in production of fuels, polymers, chemicals and pharmaceuticals. In all these areas Raman spectroscopy is used in-situ and in real-time to provide molecular level information on synthesis of novel catalysts, characterisation of catalysts, and information on the kinetics of transformation at the catalyst site, etc.

In the petrochemical industry Raman spectroscopy is used for in-situ monitoring of chemical purity and operation of distillation columns and processes such as cracking, refi ning,

Incident lightScattered light

Rayleigh scatter (same frequency as the incident light)Raman scatter (changed frequency)

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Schematic of a simple Raman spectrometer

Raman Effect, which started off as a simple observation of scattering of light by an object, has found immense applications over the entire cross-section of the industrial sector.

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solvent extraction, aromatic processing, blending, etc. Similarly, it is used to analyse chemical composition and control of polymerisation reaction in various polymers, oligomers, and polymer precursors like low density polyethylene, high density polyethylene, polypropylene, polyvinyl chloride and polystyrene since it is extremely sensitive to changes in the polymer C=C backbone seen during conversion of monomers to polymers. Other properties such as density, cross linking, curing can also be studied in-situ with great ease.

Nanotechnology and materials science: Raman spectroscopy is a standard tool for the characterisation and study of the physical properties of carbon-based nanomaterials. Fullerenes and nanotubes generate complex Raman spectra which can be used to obtain information on their length and radial size. Besides, modifi cations in the host structure and

properties by the introduction of dopants can also studied by Raman spectroscopy.

Researchers in nanotechnology need tools that enable them not only to image sub-microscopic structures but also to critically analyze them. For this purpose Raman spectroscopy has been successfully integrated with techniques such as electron microscopy, confocal microscopy, laser scanning microscopy, etc. to study materials at high spatial resolution and direct chemical analyses.

For example, a technique known as “Tip-enhanced Raman Spectroscopy (TERS)” combines atomic fi eld microscope (AFM) with surface-enhanced Raman spectroscopy. If the tip of the AFM is coated with SERS metal or metal nanoparticles then the SERS effect would be expected to occur only in the immediate vicinity of the tip. Since the tip has a diameter less than 100 nanometre, the spatial resolution will also be of that order.

Integrating Raman spectroscopy with optical microscope provides a powerful tool for chemical imaging – merging digital imaging with molecular specifi city. For example, in pharmaceutical industries it provides a visual representation of the ingredients in tablets with high spatial resolution, particle sizing, controlled release and polymorph analysis during drug development. Such systems can also be used for studying living cells at cellular and sub-cellular levels.

Raman microscopy is also an established technique in characterising semiconductors. Strains caused by compression or elongation of silicon crystals induce frequency shifts in Raman lines. The band positions in the Raman spectrum give information on material composition, structure and stress in the silicon microcrystals and help in the development of microelectronic devices and novel photovoltaic cells.

Biomedical applications: The ability of Raman spectroscopy to study the chemical makeup of living tissues in a non-destructive way has been a boon in medical research and investigations. Some of these applications include in-vivo studies of the skin, transdermal drug transfer, cancer identifi cation, bone studies including its composition and disease state, information on atherosclerosis and so on. More recently a non-invasive in-vivo blood glucose monitor has been reported. In this system the patient simply places his fi nger in front of a lens and the system analyses the Raman scattered photons through the fi nger to estimate blood glucose to a sensitivity of less than 50 mg/decilitre.

Dr. Sanjiv Sam Gambhir, professor of radiology at Stanford University and his team are pioneering a way of detecting cancer by using the technique of Surface-enhanced Raman spectroscopy. They produced gold nanoparticles of less than the size of a virus and attached to them molecular latches like an antibody

A Raman microscope which can be adapted to various microscopy techniques like Atomic Force Microscopy, Scanning Tunnel Microscopy, Confocal Microscopy, Tip-enhanced Raman Spectroscopy and so on

Chemical imaging of a pharmaceutical tablet showing spatial distribution of different components and their Raman spectra Courtesy: Horiba Scientifi c

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fragment specifi c for a given type of tumour. When the nanoparticles are injected into the blood stream, they locate and bind to the cancer cells. When laser light is shone on the skin at the site of the tumour, the molecular latches produce a characteristic Raman signal, which is amplifi ed a thousand fold by the gold nanoparticles, strong enough to be detected outside the skin to mark the presence and location of the tumour.

Prof. Gambhir has found this method to be much more sensitive and faster than other methods of tumour detection like positron emission tomography which involves injecting radioactive material into the patient. Similar tumour detecting systems have been developed by scientists at the Emory-Georgia Tech Cancer Nanotechnology Centre in Atlanta, USA.

of viruses with a high degree of sensitivity and specifi city to the level of a single virus particle. This novel SERS assay can detect spectral differences between viruses, viral strains, and viruses with gene deletions. The method provides rapid diagnostics for detection and characterization of viruses generating reproducible spectra without the need for viral multiplication.

Since Raman spectroscopy is concerned with molecular specifi city, it has been successfully used in art and archaeology for determination of the real from fake objects – art restoration and pigment identifi cation, in gemmology to determine the origin of the gems, sorting out the fake from the real; and in other miscellaneous areas like establishing the composition of glass, ceramics, porcelain and similar objects.

Another key advantage of the technique is that by constructing tailor-made nanoparticles to specifi cally latch on to different types of tumour cells, more than one type of cancer can be detected in a single test. One of the major limitations of the approach is that it can work for tumours which are only as deep as the laser beam can penetrate the tissue under the skin, which is about 1.25 cm. However, by using fi bre optics, the researchers say, the test can be performed even at deeper locations. These are in experimental stages and human trials are expected to begin soon.

Based on Surface-Enhanced Raman Spectroscopy (SERS) using silver nanorod array substrates, S. Shanmukh of Athens University, Georgia (USA) has developed a method for rapid detection of trace levels

Using the Raman effect to detect cancer cellsScientists at the Stanford Center for Cancer Nanotechnology are pioneering a new way to scan for cancer tumors

Nanoparticle Molecular latch

Nanoparticles are specifi cally designed to target cancer cells and latch onto them

The nanoparticles are injected into the bloodstream, where they locate and then bind to cancer cells

Nanoparticles

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Healthy cells

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When laser light is beamed onto the skin, the nanoparticles refl ect a distinctive Raman signal, identifying the presence of the cancer cells

A hand-held Raman spectroscope for detecting narcotics

Dr. C. V. Raman

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principle of depth penetration, multiple scattering and diffusion of photons, it is possible to collect Raman scattered photons not just in the direction backward to that of the incident photon but over 360 degrees and in different planes. Such a system can easily be adopted to detect materials like narcotics or explosives hidden in containers made of high density polyethylene, thick paper, coloured glass or other non-metallic material and also to produce a 3-D image of the contents. The IISc scientists are working on a miniaturised system for fi eld application and have also applied for a patent.

The Raman Effect, which started

off as a simple observation of scattering of light by an object, has found immense applications over the entire cross-section of the industrial sector. Even 80 years after the discovery it still remains a fountainhead for newer applications. A library of Raman spectra of thousands of molecules is available, making it convenient to quickly identify the contents of the test sample.

In fact, in 1998, the Raman Effect was designated as the National Historic Chemical Landmark by the American Chemical Society and the Indian Association for Cultivation of Sciences in recognition of its signifi cance as a tool for analysing the composition of liquids, solids and gases.

Mr M.S.S. Murthy, B-104, Terrace Garden Apartments, 2nd Main Road, BSK IIIrd Stage, Bangalore-560085

Detection of narcotics and explosives: An area of recent application of Raman Spectroscopy has been the detection of narcotics and explosives. Its inherent molecular specifi city and ability to detect picomole quantities in a non-contact way makes Raman Spectroscopy eminently suited for detecting narcotics and explosives.

Hand-held Raman scanners, weighing less than half-a-kilo and programmed to detect more than 100 narcotic substances like heroin, cocaine, amphetamine, etc. are already being used by the US Narcotic Squads at airports and other entry points. These scanners have a laser, a spectroscope, a data bank containing Raman spectra of the common narcotics and an electronic brain which would immediately compare the detected spectrum with the data bank to determine the nature of the sample.

Similarly, explosives like RDX and nitro-glycerine can be detected without opening the packet. Raman scanners can do the job in just less than 20 seconds.

More recently, scientists from the Defence Research and Development Organization, Government of India have also developed a handheld system to detect explosives. Using UV laser as the exciting beam, it can detect a variety of explosives such as TNT, TATP, DNT, RDX, HMX from a range of fi ve meters in 10-12 seconds.

In most of the above applications the analyses involves, for the sake of operational convenience, collection of Raman photons scattered in the direction backward (backscattering mode) with respect to the direction of the incident

beam. Such systems identify only the material on the surface layer and not the underlying substance. But there may be situations in which analysis has to be made on multiple layers of substances, liquids or solids contained in diffuse scattering polymer containers and so on. Two innovative modifi cations in the collection of Raman signals have been developed to meet such situations.

In the technique developed by Dr. P. Matousek of the STFC Rutherford Appleton Laboratory at Harwell Oxford called the Spatially Offset Raman Spectroscopy (SORS), Raman scattered light is collected from a region of the sample that is laterally offset from the excitation laser spot. The Raman spectra obtained this way contains information on both the surface and subsurface layers.

In practice, two Raman measurements are made – one in the conventional position and another at an offset position, a few millimetres away. The two spectra are further analysed to obtain the pure spectrum of the underlying sample.

Aircraft passengers are prohibited from carrying bottles containing any liquid, even water or shampoo in their cabin baggage since the screening X-rays cannot identify the chemical nature of their contents. Now, based on SORS simple, table-top equipment have been developed to screen and identify the contents in such bottles to the benefi t of the passengers.

More recently Siva Umapathy and Sanchita Sil of the Indian Institute of Science, Bengaluru have developed what is called Multiple Angle Raman Spectroscopy (UMARS). Based on the

Airport security: Spatially off-set Raman spectroscopy for screening bottles containing liquids Courtesy: http://www.wakeupnews.eu/wp content/uploads/2014/11/insught100.jpg

Spectrometer & Detector

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UMARS for detecting hidden substances like an explosive or narcotic. Laser light is shone from one side and the Raman scattered light collected over 360 degrees around the object. Raman spectra of both the surrounding material 1 (ammonium nitrate) and the hidden material 2 (t-Stilbene) can be detected. Courtesy: Prof. Sive Umapathy