RAMAN SPECTROSCOPY

October 11, 2006 at 1:37 am | Posted in Research, Science & Technology | Leave a comment

spin-globe.gif

books-globe.gif

globe-purple.gif

raman.jpg

Raman Spectroscopy

Raman Spectroscopy is a spectroscopic

technique used in condensed matter physics

and chemistry 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. Phonons or other excitations in the system are absorbed

or emitted by the laser light, 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 and those in a certain spectral window away from the laser line are 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. Raman spectrometers typically use holographic diffraction gratings and multiple dispersion stages

to achieve a high degree of laser rejection. A photon-counting photomultiplier tube (PMT) or, more commonly, a CCD camera is used to detect the Raman scattered light.

Raman spectroscopy has a stimulated version, analogous to stimulated emission, called stimulated Raman scattering.

Basic theory

The Raman effect occurs when light impinges upon a molecule

and interacts with the electron cloud of the bonds of that molecule. The amount of

deformation of the electron cloud is the polarizability of the molecule. The amount of the

polarizability of the bond will determine the intensity and frequency of the Raman shift.

The molecule must be symmetric to observe the Raman shift. The photon

(light quantum), excites one of the electrons into a virtual state. When the photon is

released the molecule relaxes back into vibrational energy state. The molecule will

typically relax into the first vibration energy state, and this generates Stokes Raman

scattering. If the molecule was already in an elevated vibrational energy state, the Raman

scattering is then called Anti-Stokes Raman scattering.

Applications

Raman spectroscopy is commonly used in chemistry, since vibrational information is specific to the chemical bonds and symmetry of molecules. Therefore, it provides a fingerprint by which the molecule can be identified. For instance, the vibrational frequencies of SiO, Si2O2, and Si3O3 were identified and assigned on the basis of normal coordinate analyses using infrared and Raman spectra.[3] The fingerprint region of organic molecules is in the (wavenumber

) range 500–2000 cm−1. Another way that the technique is used 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 (to be specific, 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.[5] 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.[6]

Raman spectroscopy has also been used to confirm the prediction of existence of low-frequency phonons [7] in proteins and DNA (see, e.g., [8] [9] [10] [11] greatly stimulating the studies of low-frequency collective motion in proteins and DNA and their biological functions.[12][13]

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 Helium-Neon 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.[14]

Polarized analysis

The polarization of the Raman scattered light also contains useful information. This property can be measured using (plane) polarized laser excitation and a polarization analyzer. Spectra acquired with the analyzer set at both perpendicular and parallel to the excitation plane can be used to calculate the depolarization ratio. Study of the technique is useful in teaching the connections between group theory, symmetry, Raman activity, and peaks in the corresponding Raman spectra.

The spectral information arising from this analysis gives insight into molecular orientation and vibrational symmetry. In essence, it allows the user to obtain valuable information relating to the molecular shape, for example in synthetic chemistry or polymorph analysis. It is often used to understand macromolecular orientation in crystal lattices, liquid crystals or polymer samples.[15]

Variations

Several variations of Raman spectroscopy have been developed. The usual purpose is to enhance the sensitivity (e.g., surface-enhanced Raman), to improve the spatial resolution (Raman microscopy), or to acquire very specific information (resonance Raman).

References

1.      a b Gardiner, D.J. (1989). Practical Raman spectroscopy. Springer-Verlag. ISBN 978-0387502540.

2.      Placzek G.: “Rayleigh Streeung und Raman Effekt”, In: Hdb. der Radiologie, Vol. VI., 2, 1934, p. 209

3.      Khanna, R.K. (1981). “Raman-spectroscopy of oligomeric SiO species isolated in solid methane”. Journal of Chemical Physics 74 (4): 2108. doi:10.1063/1.441393.

4.      “Fake drugs caught inside the pack”. BBC News. 2007-01-31. http://news.bbc.co.uk/2/hi/health/6314287.stm. Retrieved 2008-12-08.

5.      Irish classic is still a hit (in calfskin, not paperback) – New York Times, nytimes.com

6.      Ben Vogel (29 August 2008). “Raman spectroscopy portends well for standoff explosives detection”. Jane’s. http://www.janes.com/news/transport/business/jar/jar080829_1_n.shtml. Retrieved 2008-08-29.

7.      Kuo-Chen Chou and Nian-Yi Chen (1977) The biological functions of low-frequency phonons. Scientia Sinica, 20, 447-457.

8.      Urabe, H., Tominaga, Y. and Kubota, K. (1983) Experimental evidence of collective vibrations in DNA double helix Raman spectroscopy. Journal of Chemical Physics, 78, 5937-5939.

9.      Chou, K.C. (1983) Identification of low-frequency modes in protein molecules. Biochemical Journal, 215, 465-469.

10.                         Chou, K.C. (1984) Low-frequency vibration of DNA molecules. Biochemical Journal, 221, 27-31.

11.                         Urabe, H., Sugawara, Y., Ataka, M. and Rupprecht, A. (1998) Low-frequency Raman spectra of lysozyme crystals and oriented DNA films: dynamics of crystal water. Biophys J, 74, 1533-1540.

12.                         Kuo-Chen Chou (1988) Review: Low-frequency collective motion in biomacromolecules and its biological functions. Biophysical Chemistry, 30, 3-48.

13.                         Chou, K.C. (1989) Low-frequency resonance and cooperativity of hemoglobin. Trends in Biochemical Sciences, 14, 212.

14.                         Ellis DI, Goodacre R (August 2006). “Metabolic fingerprinting in disease diagnosis: biomedical applications of infrared and Raman spectroscopy”. Analyst 131 (8): 875–85. doi:10.1039/b602376m. PMID 17028718.

15.                         Khanna, R.K. (1957). Evidence of ion-pairing in the polarized Raman spectra of a Ba2+CrO doped KI single crystal. John Wiley & Sons, Ltd. doi:10.1002/jrs.1250040104.

16.                         Jeanmaire DL, van Duyne RP (1977). “Surface Raman Electrochemistry Part I. Heterocyclic, Aromatic and Aliphatic Amines Adsorbed on the Anodized Silver Electrode”. Journal of Electroanalytical Chemistry (Elsevier Sequouia S.A.) 84: 1–20. doi:10.1016/S0022-0728(77)80224-6.

17.                         Lombardi JR, Birke RL (2008). “A Unified Approach to Surface-Enhanced Raman Spectroscopy”. [Journal of Physical Chemistry C] (American Chemical Society) 112: 5605–5617. doi:10.1021/jp800167+CCC.

18.                         Chao RS, Khanna RK, Lippincott ER (1974). “Theoretical and experimental resonance Raman intensities for the manganate ion”. J Raman Spectroscopy 3 (2-3): 121. doi:10.1002/jrs.1250030203.

19.                         Kneipp K, et al. (1999). “Surface-Enhanced Non-Linear Raman Scattering at the Single Molecule Level”. Chem. Phys. 247: 155–162. doi:10.1016/S0301-0104(99)00165-2.

20.                         Matousek P, Clark IP, Draper ERC, et al. (2005). “Subsurface Probing in Diffusely Scattering Media using Spatially Offset Raman Spectroscopy”. Applied Spectroscopy 59 (12): 393. doi:10.1366/000370205775142548. PMID 16390587.

21.                         Barron LD, Hecht L, McColl IH, Blanch EW (2004). “Raman optical activity comes of age”. Molec. Phys. 102 (8): 731–744. doi:10.1080/00268970410001704399.

22.                         B. Schrader, G. Bergmann, Fresenius. Z. (1967). Anal. Chem.: 225–230.

23.                         P. Matousek, A. W. Parker (2006). “Bulk Raman Analysis of Pharmaceutical Tablets”. Applied Spectroscopy 60 (12): 1353–1357. doi:10.1366/000370206779321463. PMID 17217583.

24.                         P. Matousek, N. Stone (2007). “Prospects for the diagnosis of breast cancer by noninvasive probing of calcifications using transmission Raman spectroscopy”. Journal of Biomedical Optics 12 (2): 024008. doi:10.1117/1.2718934. PMID 17477723.

External links

TrackBack URI


Entries and comments feeds.

%d bloggers like this: