Vibrational spectroscopy

A molecular vibration occurs when atoms in a molecule are in periodic motion while the
molecule as a whole has constant translational and rotational motion. The frequency of the
periodic motion is known as a vibration frequency. A molecular vibration is excited when the
molecule absorbs a quantum of energy, E, corresponding to the vibration's frequency, ν,
according to the relation E=hν. A fundamental vibration is excited when one such quantum
of energy is absorbed by the molecule in its ground state. When two quanta are absorbed
the first overtone is excited, and so on to the higher overtones. To a first approximation, the
motion in a normal vibration can be described as a kind of simple harmonic motion. In
reality, vibrations are anharmonic. Excitation of the higher overtones involves progressively
less and less additional energy and eventually leads to dissociation of the molecule.

The vibrational states of a molecule can be probed in a variety of ways. The most direct way
is through infrared spectroscopy, as vibrational transitions typically require an amount of
energy that corresponds to the infrared region of the spectrum. Raman spectroscopy, which
typically uses visible light, can also be used to measure vibration frequencies directly. Two
types are coordinates describe the vibrational states.

Vibrational coordinates: The coordinate of a normal vibration is a combination of changes

in the positions of atoms in the molecule. When the vibration is excited the coordinate
changes sinusoidally with a frequency ν, the frequency of the vibration.

Internal coordinates: These are of the following types, illustrated with reference to the
planar molecule ethylene,

 Stretching: a change in the length of a bond, such as C-H or C-C

 Bending: a change in the angle between two bonds, such as the HCH angle in a
methylene group
 Rocking: a change in angle between a group of atoms, such as a methylene group
and the rest of the molecule.
 Wagging: a change in angle between the plane of a group of atoms, such as a
methylene group and a plane through the rest of the molecule,
 Twisting: a change in the angle between the planes of two groups of atoms, such as
a change in the angle between the two methylene groups.
 Out-of-plane: Not present in ethene, but an example is in BF3 when the boron atom
moves in and out of the plane of the three fluorine atoms.
Physics:

In the harmonic approximation the potential energy is a quadratic function of the normal
coordinates. Solving the Schrödinger wave equation, the energy states for each normal
coordinate are given by

where n is a quantum number that can take values of 0, 1, 2 ... Knowing the wave functions,
certain selection rules can be formulated. For example, for a harmonic oscillator transitions
are allowed only when the quantum number n changes by one,

but this does not apply to an anharmonic oscillator; the observation of overtones is only
possible because vibrations are anharmonic. Another consequence of anharmonicity is that
transitions such as between states n=2 and n=1 have slightly less energy than transitions
between the ground state and first excited state. Such a transition gives rise to a hot band.

Infrared spectroscopy

Also known as IR spectroscopy, this is the subset of spectroscopy that deals with the
infrared region of the electromagnetic spectrum. It covers a range of techniques, the most
common being a form of absorption spectroscopy. As with all spectroscopic techniques, it
can be used to identify compounds and investigate sample composition. A common
laboratory instrument that uses this technique is an infrared spectrophotometer.

The infrared portion of the electromagnetic spectrum is usually divided into three regions;
the near-, mid- and far- infrared, named for their relation to the visible spectrum. The
names and classifications of these subregions are merely conventions.

 The far-infrared, approximately 400–10 cm−1 (1000–30 μm), lying adjacent to the
microwave region, has low energy and may be used for rotational spectroscopy.
 The mid-infrared, approximately 4000–400 cm−1 (30–2.5 μm) may be used to study
the fundamental vibrations and associated rotational-vibrational structure.
 The higher energy near-IR, approximately 14000–4000 cm−1 (2.5–0.8 μm) can excite
overtone or harmonic vibrations.

Theory

Infrared spectroscopy exploits the fact that molecules absorb specific frequencies that are
characteristic of their structure. These absorptions are resonant frequencies, i.e. the
frequency of the absorbed radiation matches the frequency of the bond or group that
vibrates. The energies are determined by the shape of the molecular potential energy
surfaces, the masses of the atoms, and the associated vibronic coupling.
Number of vibrational modes

In order for a vibrational mode in a molecule to be "IR active", it must be associated with
changes in the permanent dipole.

A molecule can vibrate in many ways, and each way is called a vibrational mode. Linear
molecules have 3N-5 degrees of vibrational modes whereas nonlinear molecules have 3N-6
degrees of vibrational modes (also called vibrational degrees of freedom). As an example
H2O, a non-linear molecule, will have 3*3-6 = 3 degrees of vibrational freedom, or modes.

Simple diatomic molecules have only one bond and only one vibrational band. If the
molecule is symmetrical, such as Nitrogen molecule, the band is not observed in the IR
spectrum, but only in the Raman spectrum. Unsymmetrical diatomic molecules, e.g. CO,
absorb in the IR spectrum. More complex molecules have many bonds, and their vibrational
spectra are correspondingly more complex, i.e. big molecules have many peaks in their IR
spectra.

The atoms in a CH2 group, commonly found in organic compounds can vibrate in six
different ways: symmetrical and antisymmetrical stretching, scissoring, rocking, wagging
and twisting:

Practical IR spectroscopy

The infrared spectrum of a sample is recorded by passing a beam of infrared light through
the sample. Examination of the transmitted light reveals how much energy was absorbed at
each wavelength. This can be done with a monochromatic beam, which changes in
wavelength over time, or by using a Fourier transform instrument to measure all
wavelengths at once. From this, a transmittance or absorbance spectrum can be produced,
showing at which IR wavelengths the sample absorbs. Analysis of these absorption
characteristics reveals details about the molecular structure of the sample. When the
frequency of the IR is the same as the vibrational frequency of a bond, absorption occurs.

This technique works almost exclusively on samples with covalent bonds. Simple spectra are
obtained from samples with few IR active bonds and high levels of purity. More complex
molecular structures lead to more absorption bands and more complex spectra. The
technique has been used for the characterization of very complex mixtures.

Conventional apparatus

A beam of infrared light is produced and split into two separate beams. One is passed
through the sample, the other passed through a reference which is often the substance the
sample is dissolved in. The beams are both reflected back towards a detector, however first
they pass through a splitter which quickly alternates which of the two beams enters the
detector. The two signals are then compared and a printout is obtained.
Uses and applications

Infrared spectroscopy is widely used in both research and industry as a simple and reliable
technique for measurement, quality control and dynamic measurement. It is of especial use
in forensic analysis in both criminal and civil cases, enabling identification of polymer
degradation for example.

By measuring at a specific frequency over time, changes in the character or quantity of a

particular bond can be measured. This is especially useful in measuring the degree of
polymerization in polymer manufacture.

Infrared spectroscopy has been highly successful for applications in both organic and
inorganic chemistry. Infrared spectroscopy has also been successfully utilized in the field of
semiconductor microelectronics: for example, infrared spectroscopy can be applied to
semiconductors like silicon, gallium arsenide, gallium nitride, zinc selenide, amorphous
silicon, silicon nitride, etc.

Raman spectroscopy
Raman spectroscopy (named after C. V. Raman) is a spectroscopic technique used to study
vibrational, rotational, and other low-frequency modes in a system. It relies on inelastic
scattering, or Raman scattering, of monochromatic light, usually from a laser in the visible,
near infrared, or near ultraviolet range. The laser light interacts with phonons or other
excitations in the system, resulting in the energy of the laser photons being shifted up or
down. The shift in energy gives information about the phonon modes in the system. Infrared
spectroscopy yields similar, but complementary, information.

Typically, a sample is illuminated with a laser beam. Light from the illuminated spot is
collected with a lens and sent through a monochromator. Wavelengths close to the laser
line, due to elastic Rayleigh scattering, are filtered out while the rest of the collected light is
dispersed onto a detector.
Spontaneous Raman scattering is typically very weak, and as a result the main difficulty of
Raman spectroscopy is separating the weak inelastically scattered light from the intense
Rayleigh scattered laser light. Historically, Raman spectrometers used holographic gratings
and multiple dispersion stages to achieve a high degree of laser rejection. In the past,
photomultipliers were the detectors of choice for dispersive Raman setups, which resulted
in long acquisition times. However, modern instrumentation almost universally employs
notch or edge filters for laser rejection and spectrographs (either axial transmissive (AT),
Czerny-Turner (CT) monochromator) or FT (Fourier transform spectroscopy based), and CCD
detectors.

There are a number of advanced types of Raman spectroscopy, including surface-enhanced

Raman, tip-enhanced Raman, polarised Raman, stimulated Raman (analogous to stimulated
emission), transmission Raman, spatially-offset Raman, and hyper Raman.

Basic theory

The Raman effect occurs when light impinges upon a molecule and interacts with the
electron cloud and the bonds of that molecule. For the spontaneous Raman effect, a photon
excites the molecule from the ground state to a virtual energy state. When the molecule
relaxes it emits a photon and it returns to a different rotational or vibrational state. The
difference in energy between the original state and this new state leads to a shift in the
emitted photon's frequency away from the excitation wavelength.

If the final vibrational state of the molecule is more energetic than the initial state, then the
emitted photon will be shifted to a lower frequency in order for the total energy of the
system to remain balanced. This shift in frequency is designated as a Stokes shift. If the final
vibrational state is less energetic than the initial state, then the emitted photon will be
shifted to a higher frequency, and this is designated as an Anti-Stokes shift. Raman
scattering is an example of inelastic scattering because of the energy transfer between the
photons and the molecules during their interaction.

A change in the molecular polarization potential — or amount of deformation of the

electron cloud — with respect to the vibrational coordinate is required for a molecule to
exhibit a Raman effect. The amount of the polarizability change will determine the Raman
scattering intensity. The pattern of shifted frequencies is determined by the rotational and
vibrational states of the sample.
Energy level diagram showing the states involved in Raman signal. The line thickness is
roughly proportional to the signal strength from the different transitions.

Applications

Raman spectroscopy is commonly used in chemistry, since vibrational information is specific

to the chemical bonds and symmetry of molecules. It therefore provides a fingerprint by
which the molecule can be identified. For instance, the vibrational frequencies of SiO, Si 2O2,
and Si3O3 were identified and assigned on the basis of normal coordinate analyses using
infrared and Raman spectra. The fingerprint region of organic molecules is in the
(wavenumber) range 500–2000 cm−1. Another way that the technique is used is to study
changes in chemical bonding, e.g., when a substrate is added to an enzyme.

Raman gas analyzers have many practical applications. For instance, they are used in
medicine for real-time monitoring of anaesthetic and respiratory gas mixtures during
surgery.

In solid state physics, spontaneous Raman spectroscopy is used to, among other things,
characterize materials, measure temperature, and find the crystallographic orientation of a
sample. As with single molecules, a given solid material has characteristic phonon modes
that can help an experimenter identify it. In addition, Raman spectroscopy can be used to
observe other low frequency excitations of the solid, such as plasmons, magnons, and
superconducting gap excitations. The spontaneous Raman signal gives information on the
population of a given phonon mode in the ratio between the Stokes (downshifted) intensity
and anti-Stokes (upshifted) intensity.

Raman scattering by an anisotropic crystal gives information on the crystal orientation. The
polarization of the Raman scattered light with respect to the crystal and the polarization of
the laser light can be used to find the orientation of the crystal, if the crystal structure
(specifically, its point group) is known.

Raman active fibers, such as aramid and carbon, have vibrational modes that show a shift in
Raman frequency with applied stress. Polypropylene fibers also exhibit similar shifts. The
radial breathing mode is a commonly used technique to evaluate the diameter of carbon
nanotubes. In nanotechnology, a Raman microscope can be used to analyze nanowires to
better understand the composition of the structures.

Spatially-offset Raman spectroscopy (SORS), which is less sensitive to surface layers than
conventional Raman, can be used to discover counterfeit drugs without opening their
internal packaging, and for non-invasive monitoring of biological tissue.[4] Raman
spectroscopy can be used to investigate the chemical composition of historical documents
such as the Book of Kells and contribute to knowledge of the social and economic conditions
at the time the documents were produced. This is especially helpful because Raman
spectroscopy offers a non-invasive way to determine the best course of preservation or
conservation treatment for such materials.

Raman spectroscopy is being investigated as a means to detect explosives for airport

security.

Microspectroscopy

Raman spectroscopy offers several advantages for microscopic analysis. Since it is a

scattering technique, specimens do not need to be fixed or sectioned. Raman spectra can be
collected from a very small volume (< 1 µm in diameter); these spectra allow the
identification of species present in that volume. Water does not generally interfere with
Raman spectral analysis. Thus, Raman spectroscopy is suitable for the microscopic
examination of minerals, materials such as polymers and ceramics, cells and proteins. A
Raman microscope begins with a standard optical microscope, and adds an excitation laser,
a monochromator, and a sensitive detector (such as a charge-coupled device (CCD), or
photomultiplier tube (PMT)). FT-Raman has also been used with microscopes.

In direct imaging, the whole field of view is examined for scattering over a small range of
wavenumbers (Raman shifts). For instance, a wavenumber characteristic for cholesterol
could be used to record the distribution of cholesterol within a cell culture.

The other approach is hyperspectral imaging or chemical imaging, in which thousands of

Raman spectra are acquired from all over the field of view. The data can then be used to
generate images showing the location and amount of different components. Taking the cell
culture example, a hyperspectral image could show the distribution of cholesterol, as well as
proteins, nucleic acids, and fatty acids. Sophisticated signal- and image-processing
techniques can be used to ignore the presence of water, culture media, buffers, and other
interferents.

Raman microscopy, and in particular confocal microscopy, has very high spatial resolution.
For example, the lateral and depth resolutions were 250 nm and 1.7 µm, respectively, using
a confocal Raman microspectrometer with the 632.8 nm line from a He-Ne laser with a
pinhole of 100 µm diameter. Since the objective lenses of microscopes focus the laser beam
to several micrometres in diameter, the resulting photon flux is much higher than achieved
in conventional Raman setups. This has the added benefit of enhanced fluorescence
quenching. However, the high photon flux can also cause sample degradation, and for this
reason some setups require a thermally conducting substrate (which acts as a heat sink) in
order to mitigate this process.

By using Raman microspectroscopy, in vivo time- and space-resolved Raman spectra of

microscopic regions of samples can be measured. As a result, the fluorescence of water,
media, and buffers can be removed. Consequently in vivo time- and space-resolved Raman
spectroscopy is suitable to examine proteins, cells and organs.

Raman microscopy for biological and medical specimens generally uses near-infrared (NIR)
lasers (785 nm diodes and 1064 nm Nd:YAG are especially common). This reduces the risk of
damaging the specimen by applying higher energy wavelengths. However, the intensity of
NIR Raman is low (owing to the ω4 dependence of Raman scattering intensity), and most
detectors required very long collection times. Recently, more sensitive detectors have
become available, making the technique better suited to general use. Raman microscopy of
inorganic specimens, such as rocks and ceramics and polymers, can use a broader range of
excitation wavelengths.